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R&D Systems elisa kits
Perineural high doses (20 μg/kg) of dexmedetomidine reduced IL-6 and <t>TNF-α</t> levels in the sciatic nerve. ( A , B ) RT-PCR showed that high doses of dexmedetomidine reduced the IL-6 and TNF-α mRNA levels in the sciatic nerve; ( C , D ) <t>ELISA</t> showed that high dose of dexmedetomidine reduced the IL-6 and TNF-α protein levels in the sciatic nerve; ( E – G ) Western blotting showed that high dose of dexmedetomidine reduced IL-6 and TNF-α protein levels in the sciatic nerve.
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1) Product Images from "Perineural Dexmedetomidine Attenuates Inflammation in Rat Sciatic Nerve via the NF-?B Pathway"

Article Title: Perineural Dexmedetomidine Attenuates Inflammation in Rat Sciatic Nerve via the NF-?B Pathway

Journal: International Journal of Molecular Sciences

doi: 10.3390/ijms15034049

Perineural high doses (20 μg/kg) of dexmedetomidine reduced IL-6 and TNF-α levels in the sciatic nerve. ( A , B ) RT-PCR showed that high doses of dexmedetomidine reduced the IL-6 and TNF-α mRNA levels in the sciatic nerve; ( C , D ) ELISA showed that high dose of dexmedetomidine reduced the IL-6 and TNF-α protein levels in the sciatic nerve; ( E – G ) Western blotting showed that high dose of dexmedetomidine reduced IL-6 and TNF-α protein levels in the sciatic nerve.
Figure Legend Snippet: Perineural high doses (20 μg/kg) of dexmedetomidine reduced IL-6 and TNF-α levels in the sciatic nerve. ( A , B ) RT-PCR showed that high doses of dexmedetomidine reduced the IL-6 and TNF-α mRNA levels in the sciatic nerve; ( C , D ) ELISA showed that high dose of dexmedetomidine reduced the IL-6 and TNF-α protein levels in the sciatic nerve; ( E – G ) Western blotting showed that high dose of dexmedetomidine reduced IL-6 and TNF-α protein levels in the sciatic nerve.

Techniques Used: Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Western Blot

PDTC attenuated the inflammation of the sciatic nerve via the NF-κB pathway. ( A , C , D ) Western blotting showed that PDTC decreased NF-κB translocation into the nucleus; ( B , E , F ) Western blotting showed that PDTC reduced the IL-6 and TNF-α protein levels in the sciatic nerve; ( G , H ) RT-PCR showed that PDTC reduced the IL-6 and TNF-α mRNA levels in the sciatic nerve; ( I , J ) ELISA showed that PDTC reduced the IL-6 and TNF-α protein level in the sciatic nerve.
Figure Legend Snippet: PDTC attenuated the inflammation of the sciatic nerve via the NF-κB pathway. ( A , C , D ) Western blotting showed that PDTC decreased NF-κB translocation into the nucleus; ( B , E , F ) Western blotting showed that PDTC reduced the IL-6 and TNF-α protein levels in the sciatic nerve; ( G , H ) RT-PCR showed that PDTC reduced the IL-6 and TNF-α mRNA levels in the sciatic nerve; ( I , J ) ELISA showed that PDTC reduced the IL-6 and TNF-α protein level in the sciatic nerve.

Techniques Used: Western Blot, Translocation Assay, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

2) Product Images from "C3a triggers formation of sub-retinal pigment epithelium deposits via the ubiquitin proteasome pathway"

Article Title: C3a triggers formation of sub-retinal pigment epithelium deposits via the ubiquitin proteasome pathway

Journal: Scientific Reports

doi: 10.1038/s41598-018-28143-0

C3a treatment results in increased MMP-2 activity by RPE cells. ( a ) mRNA expression of MMP-2 and TIMP-3 measured by qRT-PCR and normalized to GAPDH in cultures of hfRPE cells treated with different doses of C3a for two weeks. ( b ) Levels of TIMP-3 measured in conditioned media of the same cultures by ELISA. ( c ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with different doses of C3a for 2 weeks. ( d ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with or without C3aR antagonist for 2 weeks (ANOVA, n = 9/treatment. Data presented as mean ± SD. *p
Figure Legend Snippet: C3a treatment results in increased MMP-2 activity by RPE cells. ( a ) mRNA expression of MMP-2 and TIMP-3 measured by qRT-PCR and normalized to GAPDH in cultures of hfRPE cells treated with different doses of C3a for two weeks. ( b ) Levels of TIMP-3 measured in conditioned media of the same cultures by ELISA. ( c ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with different doses of C3a for 2 weeks. ( d ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with or without C3aR antagonist for 2 weeks (ANOVA, n = 9/treatment. Data presented as mean ± SD. *p

Techniques Used: Activity Assay, Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

Inhibition of the UPP results in abnormal ECM turnover. ( a ) mRNA expression of MMP-2 and TIMP-3 measured by qRT-PCR and normalized to GAPDH, ( b ) MMP-2 activity measured by zymography, and ( c ) TIMP-3 levels measured by ELISA in conditioned media of hfRPE cells treated with proteasome inhibitor (+Prot inh) for 2 weeks and controls (−Prot inh) (t-test, n = 5. Data presented as mean ± SD. ***p
Figure Legend Snippet: Inhibition of the UPP results in abnormal ECM turnover. ( a ) mRNA expression of MMP-2 and TIMP-3 measured by qRT-PCR and normalized to GAPDH, ( b ) MMP-2 activity measured by zymography, and ( c ) TIMP-3 levels measured by ELISA in conditioned media of hfRPE cells treated with proteasome inhibitor (+Prot inh) for 2 weeks and controls (−Prot inh) (t-test, n = 5. Data presented as mean ± SD. ***p

Techniques Used: Inhibition, Expressing, Quantitative RT-PCR, Activity Assay, Zymography, Enzyme-linked Immunosorbent Assay, T-Test

3) Product Images from "IL-18 Induces Airway Hyperresponsiveness and Pulmonary Inflammation via CD4+ T Cell and IL-13"

Article Title: IL-18 Induces Airway Hyperresponsiveness and Pulmonary Inflammation via CD4+ T Cell and IL-13

Journal: PLoS ONE

doi: 10.1371/journal.pone.0054623

Overproduction of IL-18 in lungs increases Th1 and Th2 cytokines, and airway hyperresponsiveness. (A) The concentrations of IFN-γ, IL-13, IL-1β, and eotaxin in the bronchoalveolar lavage fluids (BALFs) were measured by specific ELISA kits. (n = 4 to 6 per each group) *: p
Figure Legend Snippet: Overproduction of IL-18 in lungs increases Th1 and Th2 cytokines, and airway hyperresponsiveness. (A) The concentrations of IFN-γ, IL-13, IL-1β, and eotaxin in the bronchoalveolar lavage fluids (BALFs) were measured by specific ELISA kits. (n = 4 to 6 per each group) *: p

Techniques Used: Enzyme-linked Immunosorbent Assay

4) Product Images from "Asiatic acid inhibits left ventricular remodeling and improves cardiac function in a rat model of myocardial infarction"

Article Title: Asiatic acid inhibits left ventricular remodeling and improves cardiac function in a rat model of myocardial infarction

Journal: Experimental and Therapeutic Medicine

doi: 10.3892/etm.2015.2871

Expression levels of inflammatory cytokines in the infarct border zone were suppressed by AA after MI. Quantitative analysis of the protein expression levels of the inflammatory cytokines (A) TGF-β1, (B) NF-κB, (C) TNF-α and (D) IL-1β in four groups (n=6–8 per group) at 4 weeks after MI, as determined using ELISA. Values are presented as the mean ± standard deviation. *P
Figure Legend Snippet: Expression levels of inflammatory cytokines in the infarct border zone were suppressed by AA after MI. Quantitative analysis of the protein expression levels of the inflammatory cytokines (A) TGF-β1, (B) NF-κB, (C) TNF-α and (D) IL-1β in four groups (n=6–8 per group) at 4 weeks after MI, as determined using ELISA. Values are presented as the mean ± standard deviation. *P

Techniques Used: Expressing, Enzyme-linked Immunosorbent Assay, Standard Deviation

5) Product Images from "Pycnogenol Attenuates the Release of Proinflammatory Cytokines and Expression of Perilipin 2 in Lipopolysaccharide-Stimulated Microglia in Part via Inhibition of NF-κB and AP-1 Activation"

Article Title: Pycnogenol Attenuates the Release of Proinflammatory Cytokines and Expression of Perilipin 2 in Lipopolysaccharide-Stimulated Microglia in Part via Inhibition of NF-κB and AP-1 Activation

Journal: PLoS ONE

doi: 10.1371/journal.pone.0137837

PYC suppressed LPS-induced proinflammatory cytokine production in BV2 microglia. Cells were incubated with the indicated concentrations of PYC or vehicle for 1h before LPS treatment (500ng/mL). After 24 h incubation, the culture supernatants were collected, and the amount of TNF-α, IL-6, IL-1β and IL-10 were measured by ELISA (A-D). Data are represented as mean ± SD from at least 3 independent experiments. *P
Figure Legend Snippet: PYC suppressed LPS-induced proinflammatory cytokine production in BV2 microglia. Cells were incubated with the indicated concentrations of PYC or vehicle for 1h before LPS treatment (500ng/mL). After 24 h incubation, the culture supernatants were collected, and the amount of TNF-α, IL-6, IL-1β and IL-10 were measured by ELISA (A-D). Data are represented as mean ± SD from at least 3 independent experiments. *P

Techniques Used: Incubation, Enzyme-linked Immunosorbent Assay

6) Product Images from "Deletion of African swine fever virus interferon inhibitors from the genome of a virulent isolate reduces virulence in domestic pigs and induces a protective response"

Article Title: Deletion of African swine fever virus interferon inhibitors from the genome of a virulent isolate reduces virulence in domestic pigs and induces a protective response

Journal: Vaccine

doi: 10.1016/j.vaccine.2016.08.011

Immune responses in vaccinated pigs. Panels A and B show numbers of IFN-γ producing cells by ELISPOT assays using PBMC stimulated with ASFV Benin 97/1. PBMC isolated on day 46 post-inoculation (panel A) or day 17 post-challenge (panel B) with OURT88/3 or BeninΔMGF were stimulated ex vivo with either medium alone or Benin 97/1 virus. Results show mean and SD of numbers of IFN-γ producing cells per 10 6 lymphocytes ( y -axis) and type of stimulation ( x -axis) for the different pigs. Panel C shows the antibody response to ASFV post-immunisation and challenge. Sera were collected at different days post-inoculation ( x -axis) of pigs with OURT88/3 or BeninΔMGF strains. Levels of anti-VP72 antibodies were detected using a blocking ELISA assay. The percentage of blocking is shown ( y -axis). Results show mean and SEM for the different groups of pigs and asterisks indicate days on which statistically significant differences were observed between the groups of pigs ( ** P ⩽ 0.01; *** P ⩽ 0.001; **** P ⩽ 0.0001).
Figure Legend Snippet: Immune responses in vaccinated pigs. Panels A and B show numbers of IFN-γ producing cells by ELISPOT assays using PBMC stimulated with ASFV Benin 97/1. PBMC isolated on day 46 post-inoculation (panel A) or day 17 post-challenge (panel B) with OURT88/3 or BeninΔMGF were stimulated ex vivo with either medium alone or Benin 97/1 virus. Results show mean and SD of numbers of IFN-γ producing cells per 10 6 lymphocytes ( y -axis) and type of stimulation ( x -axis) for the different pigs. Panel C shows the antibody response to ASFV post-immunisation and challenge. Sera were collected at different days post-inoculation ( x -axis) of pigs with OURT88/3 or BeninΔMGF strains. Levels of anti-VP72 antibodies were detected using a blocking ELISA assay. The percentage of blocking is shown ( y -axis). Results show mean and SEM for the different groups of pigs and asterisks indicate days on which statistically significant differences were observed between the groups of pigs ( ** P ⩽ 0.01; *** P ⩽ 0.001; **** P ⩽ 0.0001).

Techniques Used: Enzyme-linked Immunospot, Isolation, Ex Vivo, Blocking Assay, Enzyme-linked Immunosorbent Assay

7) Product Images from "Deoxycholic acid activates epidermal growth factor receptor and promotes intestinal carcinogenesis by ADAM17‐dependent ligand release, et al. Deoxycholic acid activates epidermal growth factor receptor and promotes intestinal carcinogenesis by ADAM17‐dependent ligand release"

Article Title: Deoxycholic acid activates epidermal growth factor receptor and promotes intestinal carcinogenesis by ADAM17‐dependent ligand release, et al. Deoxycholic acid activates epidermal growth factor receptor and promotes intestinal carcinogenesis by ADAM17‐dependent ligand release

Journal: Journal of Cellular and Molecular Medicine

doi: 10.1111/jcmm.13709

Deoxycholic acid stimulated the release of amphiregulin ( AREG ), but not transforming growth factor ( TGF )‐α or heparin‐binding ( HB )‐ EGF in intestinal tumour cells. A‐C,E, The concentration of epidermal growth factor receptor ligand TGF ‐α, HB ‐ EGF and AREG by ELISA assay in Immorto‐Min colonic epithelial cell line ( IMCE ) and human colorectal cancer cell line ( HCT ‐116) cell lines treated with deoxycholic acid ( DCA ) along with time variation. D,F, Real‐time PCR results showed that AREG mRNA levels peaked at 2 h of incubation with DCA in HCT ‐116 cells, whereas no change significantly occurred after DCA exposure in IMCE cells. DCA , deoxycholic acid. *, P
Figure Legend Snippet: Deoxycholic acid stimulated the release of amphiregulin ( AREG ), but not transforming growth factor ( TGF )‐α or heparin‐binding ( HB )‐ EGF in intestinal tumour cells. A‐C,E, The concentration of epidermal growth factor receptor ligand TGF ‐α, HB ‐ EGF and AREG by ELISA assay in Immorto‐Min colonic epithelial cell line ( IMCE ) and human colorectal cancer cell line ( HCT ‐116) cell lines treated with deoxycholic acid ( DCA ) along with time variation. D,F, Real‐time PCR results showed that AREG mRNA levels peaked at 2 h of incubation with DCA in HCT ‐116 cells, whereas no change significantly occurred after DCA exposure in IMCE cells. DCA , deoxycholic acid. *, P

Techniques Used: Binding Assay, Concentration Assay, Enzyme-linked Immunosorbent Assay, Real-time Polymerase Chain Reaction, Incubation

8) Product Images from "Regulation and biological role of the peptide/histidine transporter SLC15A3 in Toll-like receptor-mediated inflammatory responses in macrophage"

Article Title: Regulation and biological role of the peptide/histidine transporter SLC15A3 in Toll-like receptor-mediated inflammatory responses in macrophage

Journal: Cell Death & Disease

doi: 10.1038/s41419-018-0809-1

SLC15A3 expression was increased in mice with E. coli -induced peritonitis, and positively related to inflammation. Mice were intraperitoneally injected with PBS or E.coli (1 × 10 7 cfu/mL, 0.5 mL/ 25 g), and the serum or peritoneal macrophages collected after 24 h. a IL-6 and TNF-α protein levels in mouse serum were determined by ELISA assays. b , c Slc15a3 mRNA ( b ) and protein expression ( c ) in PMs was determined by real-time PCR and Western blotting. Quantification of protein (i.e., SLC15A3/GAPDH ratio) is shown in the right side of each western blot figure. d mRNA expression of Il-6 and Tnf-α in peritoneal macrophages as a function of time. e Correlation analysis of Slc15a3 with Il-6 or Tnf-α was performed by linear regression, in which Slc15a3 mRNA expression in x-axis and Il-6 (left) or Tnf-α (right) mRNA expression in y -axis. Statistical analyses were performed using an unpaired t test. * P
Figure Legend Snippet: SLC15A3 expression was increased in mice with E. coli -induced peritonitis, and positively related to inflammation. Mice were intraperitoneally injected with PBS or E.coli (1 × 10 7 cfu/mL, 0.5 mL/ 25 g), and the serum or peritoneal macrophages collected after 24 h. a IL-6 and TNF-α protein levels in mouse serum were determined by ELISA assays. b , c Slc15a3 mRNA ( b ) and protein expression ( c ) in PMs was determined by real-time PCR and Western blotting. Quantification of protein (i.e., SLC15A3/GAPDH ratio) is shown in the right side of each western blot figure. d mRNA expression of Il-6 and Tnf-α in peritoneal macrophages as a function of time. e Correlation analysis of Slc15a3 with Il-6 or Tnf-α was performed by linear regression, in which Slc15a3 mRNA expression in x-axis and Il-6 (left) or Tnf-α (right) mRNA expression in y -axis. Statistical analyses were performed using an unpaired t test. * P

Techniques Used: Expressing, Mouse Assay, Injection, Enzyme-linked Immunosorbent Assay, Real-time Polymerase Chain Reaction, Western Blot

Knockdown or overexpression of SLC15A3 influenced the TLR4-dependent proinflammatory cytokine production. a mRNA and protein expression of SLC15A3 was determined in THP-1 cells transfected with 30 nM control siRNA (Ctrl) or two different SLC15A3 siRNA (#1 and #2) after 24 h transfection. Quantification of protein (i.e., SLC15A3/GAPDH ratio) is shown in the right side of each Western blot figure. b THP-1 cells transfected with 30 nM control siRNA (Ctrl) or SLC15A3 siRNA (#1). Twenty-four hours after transfection, the cells were treated with 100 ng/mL LPS for 12 h. The levels of IL-6 and TNF-α in the cell culture supernatants were determined by ELISA assays. c , d A549 cells transfected with 1 μg pcDNA3.1 or human SLC15A3 plasmid. Twenty-four hours after transfection, the cells were treated with 1 μg/mL LPS for 8 h. The mRNA ( c ) and protein ( d ) levels of IL-6 in cells and cell culture supernatants were determined by real-time PCR and ELISA assays. Statistical analyses were performed using an unpaired t test. * P
Figure Legend Snippet: Knockdown or overexpression of SLC15A3 influenced the TLR4-dependent proinflammatory cytokine production. a mRNA and protein expression of SLC15A3 was determined in THP-1 cells transfected with 30 nM control siRNA (Ctrl) or two different SLC15A3 siRNA (#1 and #2) after 24 h transfection. Quantification of protein (i.e., SLC15A3/GAPDH ratio) is shown in the right side of each Western blot figure. b THP-1 cells transfected with 30 nM control siRNA (Ctrl) or SLC15A3 siRNA (#1). Twenty-four hours after transfection, the cells were treated with 100 ng/mL LPS for 12 h. The levels of IL-6 and TNF-α in the cell culture supernatants were determined by ELISA assays. c , d A549 cells transfected with 1 μg pcDNA3.1 or human SLC15A3 plasmid. Twenty-four hours after transfection, the cells were treated with 1 μg/mL LPS for 8 h. The mRNA ( c ) and protein ( d ) levels of IL-6 in cells and cell culture supernatants were determined by real-time PCR and ELISA assays. Statistical analyses were performed using an unpaired t test. * P

Techniques Used: Over Expression, Expressing, Transfection, Western Blot, Cell Culture, Enzyme-linked Immunosorbent Assay, Plasmid Preparation, Real-time Polymerase Chain Reaction

9) Product Images from "Inflammatory Effects of Menthol vs. Non-menthol Cigarette Smoke Extract on Human Lung Epithelial Cells: A Double-Hit on TRPM8 by Reactive Oxygen Species and Menthol"

Article Title: Inflammatory Effects of Menthol vs. Non-menthol Cigarette Smoke Extract on Human Lung Epithelial Cells: A Double-Hit on TRPM8 by Reactive Oxygen Species and Menthol

Journal: Frontiers in Physiology

doi: 10.3389/fphys.2017.00263

Responses of intracellular Ca 2+ and IL-8 to M-CSE and Non-M-CSE in HEK293 cells transfected with human TRPM8 (hTRPM8). (A) Cells were transfected with control (pCMV6) or Flag-tagged hTRPM8 vector for 24 h. Anti-Flag and anti-TRPM8 antibodies were used to perform Western blot to confirm successful transfection. (B) Control cells or hTRPM8-expressing cells were exposed to 1% Non-M-CSE or M-CSE for 1, 2, 5, 10, and 30 min. Intracellular Ca 2+ levels were measured by Fluo-8 fluorescent probe assay. The hTRPM8-expressing cells were exposed to medium alone, 1% Non-M-CSE, 1% M-CSE, or menthol for 2 min ( C) and 24 h (D) with or without pretreatment with AMTB. IL-8 levels were analyzed by ELISA. Data from each group are means ± SEM from four independent experiments. * p
Figure Legend Snippet: Responses of intracellular Ca 2+ and IL-8 to M-CSE and Non-M-CSE in HEK293 cells transfected with human TRPM8 (hTRPM8). (A) Cells were transfected with control (pCMV6) or Flag-tagged hTRPM8 vector for 24 h. Anti-Flag and anti-TRPM8 antibodies were used to perform Western blot to confirm successful transfection. (B) Control cells or hTRPM8-expressing cells were exposed to 1% Non-M-CSE or M-CSE for 1, 2, 5, 10, and 30 min. Intracellular Ca 2+ levels were measured by Fluo-8 fluorescent probe assay. The hTRPM8-expressing cells were exposed to medium alone, 1% Non-M-CSE, 1% M-CSE, or menthol for 2 min ( C) and 24 h (D) with or without pretreatment with AMTB. IL-8 levels were analyzed by ELISA. Data from each group are means ± SEM from four independent experiments. * p

Techniques Used: Transfection, Plasmid Preparation, Western Blot, Expressing, Enzyme-linked Immunosorbent Assay

10) Product Images from "Cannabinoid Type 2 (CB2) Receptors Activation Protects against Oxidative Stress and Neuroinflammation Associated Dopaminergic Neurodegeneration in Rotenone Model of Parkinson's Disease"

Article Title: Cannabinoid Type 2 (CB2) Receptors Activation Protects against Oxidative Stress and Neuroinflammation Associated Dopaminergic Neurodegeneration in Rotenone Model of Parkinson's Disease

Journal: Frontiers in Neuroscience

doi: 10.3389/fnins.2016.00321

The pro-inflammatory cytokines: IL-1β, IL-6, and TNF-α were measured by enzyme linked immunosorbent assay (ELISA) in the midbrain . Values are expressed as mean ± SEM ( n = 6–8). The level of IL-1β (A) , IL-6 (B) , and TNF-α (C) was significantly (* p
Figure Legend Snippet: The pro-inflammatory cytokines: IL-1β, IL-6, and TNF-α were measured by enzyme linked immunosorbent assay (ELISA) in the midbrain . Values are expressed as mean ± SEM ( n = 6–8). The level of IL-1β (A) , IL-6 (B) , and TNF-α (C) was significantly (* p

Techniques Used: Enzyme-linked Immunosorbent Assay

11) Product Images from "Comparative analysis of the growth and biological activity of a respiratory and atheroma isolate of Chlamydia pneumoniae reveals strain-dependent differences in inflammatory activity and innate immune evasion"

Article Title: Comparative analysis of the growth and biological activity of a respiratory and atheroma isolate of Chlamydia pneumoniae reveals strain-dependent differences in inflammatory activity and innate immune evasion

Journal: BMC Microbiology

doi: 10.1186/s12866-015-0569-3

Cytokine induction in the lungs of AO3 or AR39 infected mice. Group mice were infected with AO3 or AR39, or mock infected, and euthanized at designated time points as described in figure four and the Methods section. Removed lung were homogenized and the lung homogenates were assayed for cytokines IL-6 ( a ), TNF-α ( b ), IL-1β ( c ), IL-10 ( d ), IP-10 ( e ) and IFN-β ( f ) by ELISA. Each data point represents one mouse. Horizontal bar represents the mean. Significance: NS, not significant; *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001. Data is representative of 2 independent experiments
Figure Legend Snippet: Cytokine induction in the lungs of AO3 or AR39 infected mice. Group mice were infected with AO3 or AR39, or mock infected, and euthanized at designated time points as described in figure four and the Methods section. Removed lung were homogenized and the lung homogenates were assayed for cytokines IL-6 ( a ), TNF-α ( b ), IL-1β ( c ), IL-10 ( d ), IP-10 ( e ) and IFN-β ( f ) by ELISA. Each data point represents one mouse. Horizontal bar represents the mean. Significance: NS, not significant; *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001. Data is representative of 2 independent experiments

Techniques Used: Infection, Mouse Assay, Enzyme-linked Immunosorbent Assay

12) Product Images from "Cranial irradiation induces transient microglia accumulation, followed by long-lasting inflammation and loss of microglia"

Article Title: Cranial irradiation induces transient microglia accumulation, followed by long-lasting inflammation and loss of microglia

Journal: Oncotarget

doi: 10.18632/oncotarget.12929

Experimental design Reporter mice, CX3CR1 GFP/+ and CCR2 RFP/+ , expressing GFP in resident microglia and RFP in monocyte-derived macrophages, were subjected to irradiation (IR) or sham procedures (SH). Hippocampal tissue was subjected to immunohistochemistry and stereological quantification, or analyzed for CCL2 and IL-1ß protein using ELISA. Microglia were isolated from the hippocampus using MACS and further analyzed using either RT-PCR or FACS ( A). All CX3CR1 GFP/+- labeled cells expressed Iba-1 in sham control brains, and all Iba-1-positive cells were positive for CX3CR1 GFP/+ . Scale bar, 10 μm ( B).
Figure Legend Snippet: Experimental design Reporter mice, CX3CR1 GFP/+ and CCR2 RFP/+ , expressing GFP in resident microglia and RFP in monocyte-derived macrophages, were subjected to irradiation (IR) or sham procedures (SH). Hippocampal tissue was subjected to immunohistochemistry and stereological quantification, or analyzed for CCL2 and IL-1ß protein using ELISA. Microglia were isolated from the hippocampus using MACS and further analyzed using either RT-PCR or FACS ( A). All CX3CR1 GFP/+- labeled cells expressed Iba-1 in sham control brains, and all Iba-1-positive cells were positive for CX3CR1 GFP/+ . Scale bar, 10 μm ( B).

Techniques Used: Mouse Assay, Expressing, Derivative Assay, Irradiation, Immunohistochemistry, Enzyme-linked Immunosorbent Assay, Isolation, Magnetic Cell Separation, Reverse Transcription Polymerase Chain Reaction, FACS, Labeling

13) Product Images from "Circulating HLA-DR+CD4+ effector memory T cells resistant to CCR5 and PD-L1 mediated suppression compromise regulatory T cell function in tuberculosis"

Article Title: Circulating HLA-DR+CD4+ effector memory T cells resistant to CCR5 and PD-L1 mediated suppression compromise regulatory T cell function in tuberculosis

Journal: PLoS Pathogens

doi: 10.1371/journal.ppat.1007289

PTB Teff containing HLA-DR + subset are compromised in CCR5 and PD-L1 mediated signalling. Sorted PTB total and HLA-DR - Teff cells were treated with 50 ng/ml each of CCL3 and CCL4 (A), 5 μg/ml anti-CCR5 (A), 5 μg/ml anti-PD-L1 (B) or 10 μg/ml recombinant PD-L1 (B). Cells were activated with anti-CD3/anti-CD28 beads (beads:cell ratio of 1:1). Anti-CCR5, anti-PD-L1 and recombinant PD-L1 were added 20 minutes prior to addition of mitogenic anti-CD3/anti-CD28. Unstimulated cells were used as control. NFκB activation was measured by ELISA after 180 minutes of stimulation and was expressed as absorbance at 450 nm. (A) shows data pertaining to CCR5 mediated signaling and (B) shows data pertaining to PD-L1 mediated signaling. Data shows median and maximum and minimum values from N = 6 PTB donors. Paired Wilcoxon matched-pairs signed rank test with Bonferroni correction was used to determine P value. *p ≤ 0.03.
Figure Legend Snippet: PTB Teff containing HLA-DR + subset are compromised in CCR5 and PD-L1 mediated signalling. Sorted PTB total and HLA-DR - Teff cells were treated with 50 ng/ml each of CCL3 and CCL4 (A), 5 μg/ml anti-CCR5 (A), 5 μg/ml anti-PD-L1 (B) or 10 μg/ml recombinant PD-L1 (B). Cells were activated with anti-CD3/anti-CD28 beads (beads:cell ratio of 1:1). Anti-CCR5, anti-PD-L1 and recombinant PD-L1 were added 20 minutes prior to addition of mitogenic anti-CD3/anti-CD28. Unstimulated cells were used as control. NFκB activation was measured by ELISA after 180 minutes of stimulation and was expressed as absorbance at 450 nm. (A) shows data pertaining to CCR5 mediated signaling and (B) shows data pertaining to PD-L1 mediated signaling. Data shows median and maximum and minimum values from N = 6 PTB donors. Paired Wilcoxon matched-pairs signed rank test with Bonferroni correction was used to determine P value. *p ≤ 0.03.

Techniques Used: Recombinant, Activation Assay, Enzyme-linked Immunosorbent Assay

PTB HLA-DR - but not HLA-DR + total Teff are susceptible to Treg suppression via CCR5 and PD-L1. (A, E) RNA was isolated from sorted PTB total and HLA-DR - Teff cells activated with anti-CD3/anti-CD28 for 2 hrs and converted to cDNA. Expression of CCL3L3 , CCL4 and PD-L1 was measured by qRT-PCR. Expression is shown as relative copy number relative to GAPDH. (B) CCL3, CCL4 and CCL5 levels were measured in supernatants from cultures of PTB total and HLA-DR - cells activated with anti-CD3/anti-CD28 for 24 hrs by ELISA. CCR5 (C) and PD-L1 (F) expression was measured on unstimulated and anti-CD3/anti-CD28 activated PTB total and HLA-DR - Teff cells after 24 hrs and MFI of CCR5 (C) and PD-L1 (F) from multiple PTB donors was plotted for comparison between total and HLA-DR - Teff cells. CFSE labelled sorted PTB and IGRA-ve total and HLA-DR - Teff cells were co-cultured with autologous Treg cells at a ratio of 1:1 in the presence of either 5 μg/ml anti-CCR5 (D) or 5 μg/ml anti-PD-L1 (G). Appropriate 5 μg/ml isotype antibody was used as control in both cases and cells were activated with anti-CD3/anti-CD28 beads (beads:cell ratio of 1:1). Proliferation was measured by CFSE dilution after 4 days and percentage suppression was calculated (D and G). A total of 6–8 PTB and 6 IGRA-ve donors were used. Upper box shows data pertaining to CCR5 and lower box shows data pertaining to PD-L1. Paired Wilcoxon matched-pairs signed rank test with Bonferroni correction was used to determine P value. *p ≤ 0.03.
Figure Legend Snippet: PTB HLA-DR - but not HLA-DR + total Teff are susceptible to Treg suppression via CCR5 and PD-L1. (A, E) RNA was isolated from sorted PTB total and HLA-DR - Teff cells activated with anti-CD3/anti-CD28 for 2 hrs and converted to cDNA. Expression of CCL3L3 , CCL4 and PD-L1 was measured by qRT-PCR. Expression is shown as relative copy number relative to GAPDH. (B) CCL3, CCL4 and CCL5 levels were measured in supernatants from cultures of PTB total and HLA-DR - cells activated with anti-CD3/anti-CD28 for 24 hrs by ELISA. CCR5 (C) and PD-L1 (F) expression was measured on unstimulated and anti-CD3/anti-CD28 activated PTB total and HLA-DR - Teff cells after 24 hrs and MFI of CCR5 (C) and PD-L1 (F) from multiple PTB donors was plotted for comparison between total and HLA-DR - Teff cells. CFSE labelled sorted PTB and IGRA-ve total and HLA-DR - Teff cells were co-cultured with autologous Treg cells at a ratio of 1:1 in the presence of either 5 μg/ml anti-CCR5 (D) or 5 μg/ml anti-PD-L1 (G). Appropriate 5 μg/ml isotype antibody was used as control in both cases and cells were activated with anti-CD3/anti-CD28 beads (beads:cell ratio of 1:1). Proliferation was measured by CFSE dilution after 4 days and percentage suppression was calculated (D and G). A total of 6–8 PTB and 6 IGRA-ve donors were used. Upper box shows data pertaining to CCR5 and lower box shows data pertaining to PD-L1. Paired Wilcoxon matched-pairs signed rank test with Bonferroni correction was used to determine P value. *p ≤ 0.03.

Techniques Used: Isolation, Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Cell Culture

14) Product Images from "CB2 Cannabinoid Receptors Contribute to Bacterial Invasion and Mortality in Polymicrobial Sepsis"

Article Title: CB2 Cannabinoid Receptors Contribute to Bacterial Invasion and Mortality in Polymicrobial Sepsis

Journal: PLoS ONE

doi: 10.1371/journal.pone.0006409

CB 2 receptor deficiency decreases IL-10, IL-6, and MIP-2 levels in the plasma and peritoneal lavage fluid of mice subjected to CLP. IL-10 (a,b), IL-6 (c,d) and MIP-2 (e,f) concentrations were measured at 16 hours after surgery using ELISA. Data are the mean±SEM of n = 6–9 mice per group. *** p
Figure Legend Snippet: CB 2 receptor deficiency decreases IL-10, IL-6, and MIP-2 levels in the plasma and peritoneal lavage fluid of mice subjected to CLP. IL-10 (a,b), IL-6 (c,d) and MIP-2 (e,f) concentrations were measured at 16 hours after surgery using ELISA. Data are the mean±SEM of n = 6–9 mice per group. *** p

Techniques Used: Mouse Assay, Enzyme-linked Immunosorbent Assay

15) Product Images from "IL-31 plays dual roles in lung inflammation in an OVA-induced murine asthma model"

Article Title: IL-31 plays dual roles in lung inflammation in an OVA-induced murine asthma model

Journal: Biology Open

doi: 10.1242/bio.036244

IL-31RA KO mice exhibit exacerbated lung inflammation following challenge with OVA. IL-31RA KO mice were generated to delete the fourth exon of IL-31RA by homologous recombination. Ten IL-31RA KO mice were sensitized intraperitoneally with 100 μg OVA in the presence of aluminum hydroxide at days 0, 7 and 14, and an intranasal challenge with 5% OVA started on day 21 for 7 consecutive days. (A) Paraffin sections of lungs from OVA challenged mice were HE stained. (B) Serum was assayed for total IgE ( n =10). (C) BALF was collected for analysis of cell infiltrates ( n =10). (D) IL-6 and OSM in BALF were detected by ELISA ( n =8). Data expressed as the mean±s.d. Significance was determined by two-tailed Student's t -test, *** P
Figure Legend Snippet: IL-31RA KO mice exhibit exacerbated lung inflammation following challenge with OVA. IL-31RA KO mice were generated to delete the fourth exon of IL-31RA by homologous recombination. Ten IL-31RA KO mice were sensitized intraperitoneally with 100 μg OVA in the presence of aluminum hydroxide at days 0, 7 and 14, and an intranasal challenge with 5% OVA started on day 21 for 7 consecutive days. (A) Paraffin sections of lungs from OVA challenged mice were HE stained. (B) Serum was assayed for total IgE ( n =10). (C) BALF was collected for analysis of cell infiltrates ( n =10). (D) IL-6 and OSM in BALF were detected by ELISA ( n =8). Data expressed as the mean±s.d. Significance was determined by two-tailed Student's t -test, *** P

Techniques Used: Mouse Assay, Generated, Homologous Recombination, Staining, Enzyme-linked Immunosorbent Assay, Two Tailed Test

16) Product Images from "RNA editing enzyme APOBEC3A promotes pro-inflammatory M1 macrophage polarization"

Article Title: RNA editing enzyme APOBEC3A promotes pro-inflammatory M1 macrophage polarization

Journal: Communications Biology

doi: 10.1038/s42003-020-01620-x

Knockdown of A3A impairs pro-inflammatory phenotype of M1 macrophages. a A3A KD decreases TNF-α secretion by M1 macrophages from normal donors compared to controls (SC). P = 0.0049, Paired t test, two-tailed, n = 5 donors. b A3A KD decreases IL-1β secretion by M1 macrophages from normal donors compared to controls (SC). P = 0.0103, Paired t test, two-tailed, n = 6 donors. c A3A KD decreases IL6 secretion by M1 macrophages from normal donors compared to controls (SC). P = 0.0192, Paired t test, two-tailed, n = 6 donors. d A3A KD reduces CD86 surface protein expression in M1-macrophages by flow cytometry. Cytokines are measured by ELISA (see the “Methods” section). e The quantification of CD86 ( P = 0.0395, Paired t test, two-tailed, n = 5 donors) is made using flow cytometry (see the “Methods” section) following M1 polarization in CD33-positive cells. MFI = mean (geometric) Fluorescent Intensity. M2 macrophage data from a smaller number of donors ( n = 2–4) highlight low expression levels of inflammatory markers in M2. Data in scatter dot plots with mean ± SD.
Figure Legend Snippet: Knockdown of A3A impairs pro-inflammatory phenotype of M1 macrophages. a A3A KD decreases TNF-α secretion by M1 macrophages from normal donors compared to controls (SC). P = 0.0049, Paired t test, two-tailed, n = 5 donors. b A3A KD decreases IL-1β secretion by M1 macrophages from normal donors compared to controls (SC). P = 0.0103, Paired t test, two-tailed, n = 6 donors. c A3A KD decreases IL6 secretion by M1 macrophages from normal donors compared to controls (SC). P = 0.0192, Paired t test, two-tailed, n = 6 donors. d A3A KD reduces CD86 surface protein expression in M1-macrophages by flow cytometry. Cytokines are measured by ELISA (see the “Methods” section). e The quantification of CD86 ( P = 0.0395, Paired t test, two-tailed, n = 5 donors) is made using flow cytometry (see the “Methods” section) following M1 polarization in CD33-positive cells. MFI = mean (geometric) Fluorescent Intensity. M2 macrophage data from a smaller number of donors ( n = 2–4) highlight low expression levels of inflammatory markers in M2. Data in scatter dot plots with mean ± SD.

Techniques Used: Two Tailed Test, Expressing, Flow Cytometry, Enzyme-linked Immunosorbent Assay

17) Product Images from "Autophagy protein ATG7 is a critical regulator of endothelial cell inflammation and permeability"

Article Title: Autophagy protein ATG7 is a critical regulator of endothelial cell inflammation and permeability

Journal: Scientific Reports

doi: 10.1038/s41598-020-70126-7

ATG7 knockdown prevents RelA/p65 nuclear translocation and DNA binding, but not IκBα degradation or RelA/p65 phosphorylation. ( A ) HPAEC were transfected for 48 h with si-Con or si-ATG7 and then treated with thrombin (5 U/ml) for 1 h. Nuclear extracts were obtained and processed according to an ELISA-based DNA binding assay kit as described in the “ Materials and Methods ” section. Error bars represent mean ± S.E. (n = 4 for each condition). ( B C ) HPAEC were transfected with si-Con or si-ATG7 for 48 h and treated with thrombin (5 U/ml) for 1 h. Total cell lysates were analyzed by Western blot for ( B ) IκBα levels and ( C ) RelA/p65 phosphorylation. RelA/p65 was used as a loading control. Error bars represent mean ± S.E. (n = 3–4 for each condition). ( D ) HPAEC were transfected for 48 h with si-Con or si-ATG7 and treated with thrombin (5 U/ml) for 1 h. Nuclear extracts were obtained and analyzed by Western blot to measure the level of RelA/p65, and TATA-binding protein (TBP) was used as a loading control for nuclear protein. Error bars represent mean ± S.E. (n = 3–4 for each condition).
Figure Legend Snippet: ATG7 knockdown prevents RelA/p65 nuclear translocation and DNA binding, but not IκBα degradation or RelA/p65 phosphorylation. ( A ) HPAEC were transfected for 48 h with si-Con or si-ATG7 and then treated with thrombin (5 U/ml) for 1 h. Nuclear extracts were obtained and processed according to an ELISA-based DNA binding assay kit as described in the “ Materials and Methods ” section. Error bars represent mean ± S.E. (n = 4 for each condition). ( B C ) HPAEC were transfected with si-Con or si-ATG7 for 48 h and treated with thrombin (5 U/ml) for 1 h. Total cell lysates were analyzed by Western blot for ( B ) IκBα levels and ( C ) RelA/p65 phosphorylation. RelA/p65 was used as a loading control. Error bars represent mean ± S.E. (n = 3–4 for each condition). ( D ) HPAEC were transfected for 48 h with si-Con or si-ATG7 and treated with thrombin (5 U/ml) for 1 h. Nuclear extracts were obtained and analyzed by Western blot to measure the level of RelA/p65, and TATA-binding protein (TBP) was used as a loading control for nuclear protein. Error bars represent mean ± S.E. (n = 3–4 for each condition).

Techniques Used: Translocation Assay, Binding Assay, Transfection, Enzyme-linked Immunosorbent Assay, DNA Binding Assay, Western Blot

ATG7 knockdown inhibits thrombin-mediated expression of inflammatory proteins and NF-κB activity. HPAEC were transfected with si-Con or si-ATG7 for 48 h and then treated with thrombin (5 U/ml) for 6 h. ( A , B ) Conditioned media was collected from the cells and ELISAs were performed for ( A ) IL-6, ( B ) MCP-1. Error bars represent mean ± S.E. (n = 5 for each condition). ( C D ) Cell lysates were analyzed by ELISA for ( C ) ICAM-1 and ( D ) VCAM-1 levels. Error bars represent mean ± S.E. (n = 5 for each condition). ( E ) HPAEC were transfected with si-Con or si-ATG7 for 24 h, then transfected with NF-κBLUC and Renilla LUC constructs as described in the “ Materials and Methods ”. The cells were then treated with thrombin (5 U/ml) for 6 h and cell extracts were assayed for Firefly and Renilla luciferase activities. Renilla luciferase was used as an internal control for transfection efficiency. Error bars represent mean ± S.E. (n = 6 for each condition).
Figure Legend Snippet: ATG7 knockdown inhibits thrombin-mediated expression of inflammatory proteins and NF-κB activity. HPAEC were transfected with si-Con or si-ATG7 for 48 h and then treated with thrombin (5 U/ml) for 6 h. ( A , B ) Conditioned media was collected from the cells and ELISAs were performed for ( A ) IL-6, ( B ) MCP-1. Error bars represent mean ± S.E. (n = 5 for each condition). ( C D ) Cell lysates were analyzed by ELISA for ( C ) ICAM-1 and ( D ) VCAM-1 levels. Error bars represent mean ± S.E. (n = 5 for each condition). ( E ) HPAEC were transfected with si-Con or si-ATG7 for 24 h, then transfected with NF-κBLUC and Renilla LUC constructs as described in the “ Materials and Methods ”. The cells were then treated with thrombin (5 U/ml) for 6 h and cell extracts were assayed for Firefly and Renilla luciferase activities. Renilla luciferase was used as an internal control for transfection efficiency. Error bars represent mean ± S.E. (n = 6 for each condition).

Techniques Used: Expressing, Activity Assay, Transfection, Enzyme-linked Immunosorbent Assay, Construct, Luciferase

18) Product Images from "Erythropoietin Exacerbates Inflammation and Increases the Mortality of Histoplasma capsulatum-Infected Mice"

Article Title: Erythropoietin Exacerbates Inflammation and Increases the Mortality of Histoplasma capsulatum-Infected Mice

Journal: Mediators of Inflammation

doi: 10.1155/2015/786319

EPO pretreatment increases proinflammatory cytokines and chemokines in lungs from Hc -infected mice. Lungs were removed from uninfected mice treated with 45 U EPO (EPO) or from untreated ( Hc ) or EPO pretreated ( EPO + Hc ) mice infected with Hc (5 × 10 5 yeasts/100 μ L/mouse). Lungs were removed 14 days after infection and homogenized, and the concentrations of IFN- γ (a), IL-6 (b), MIP-1 α (c), and MCP-1 (d) were assessed in the supernatants using ELISA. Means ± SEM values from one representative experiment of two independent experiments are shown ( n = 4–6). Newman-Keuls multiple comparison test was used and the differences were considered significant when p
Figure Legend Snippet: EPO pretreatment increases proinflammatory cytokines and chemokines in lungs from Hc -infected mice. Lungs were removed from uninfected mice treated with 45 U EPO (EPO) or from untreated ( Hc ) or EPO pretreated ( EPO + Hc ) mice infected with Hc (5 × 10 5 yeasts/100 μ L/mouse). Lungs were removed 14 days after infection and homogenized, and the concentrations of IFN- γ (a), IL-6 (b), MIP-1 α (c), and MCP-1 (d) were assessed in the supernatants using ELISA. Means ± SEM values from one representative experiment of two independent experiments are shown ( n = 4–6). Newman-Keuls multiple comparison test was used and the differences were considered significant when p

Techniques Used: Infection, Mouse Assay, Enzyme-linked Immunosorbent Assay

19) Product Images from "Toll-Like Receptor 4 Mediates Acute Lung Injury Induced by High Mobility Group Box-1"

Article Title: Toll-Like Receptor 4 Mediates Acute Lung Injury Induced by High Mobility Group Box-1

Journal: PLoS ONE

doi: 10.1371/journal.pone.0064375

The effect of TLR4 inhibition on HMGB-1-induced ALI. RNAi was used to inhibit the expression of TLR4, and the animals received 100 µg of rhHMGB-1. Lung histological observation was performed after transfection and 24 h after HMGB-1 treatment. The negative and shTLR4 groups displayed normal lung structure similar to the control (A, B, and D) Stronger ALI effects were seen in the HMGB-1 group (C), but weaker ALI effects were observed in the shTLR4+HMGB-1 group (F).The anti-HMGB-1 and HMGB-1+anti-HMGB-1 groups also showed normal lung structure(E, G). Lung histopathological scores showed the degree of interstitial accumulation of neutrophils and edema: the HMGB-1 group was the highest, and the shTLR4+HMGB-1 was clearly lower than the shNT+HMGB-1 group, but still higher than the other three groups (H). The levels of TNF-α and IL-1β in lung were analyzed by ELISA after transfection and 24 h after HMGB-1 administration. The levels of IL-1β in lung were not significantly different between the control, negative, shTLR4, and anti-HMGB-1 groups. After treatment with HMGB-1, the levels of IL-1β and TNF-α increased dramatically in the HMGB-1 group and increased slightly in the shTLR4+HMGB-1 group, but did not change in the HMGB-1+anti-HMGB-1 group (J, K). TLR4 protein expression was measured by western blot analysis and TLR4 mRNA expression was measured by real-time quantitative PCR after transfection and 24 h after HMGB-1 administration. The TLR4 protein and mRNA levels were very low in the shTLR4 group, but dramatically increased in the HMGB-1 group. The levels of TLR4 expression in the other five groups were equal to each other (I, L). Data are shown as the mean±standard deviation (SD), n = 6, * P
Figure Legend Snippet: The effect of TLR4 inhibition on HMGB-1-induced ALI. RNAi was used to inhibit the expression of TLR4, and the animals received 100 µg of rhHMGB-1. Lung histological observation was performed after transfection and 24 h after HMGB-1 treatment. The negative and shTLR4 groups displayed normal lung structure similar to the control (A, B, and D) Stronger ALI effects were seen in the HMGB-1 group (C), but weaker ALI effects were observed in the shTLR4+HMGB-1 group (F).The anti-HMGB-1 and HMGB-1+anti-HMGB-1 groups also showed normal lung structure(E, G). Lung histopathological scores showed the degree of interstitial accumulation of neutrophils and edema: the HMGB-1 group was the highest, and the shTLR4+HMGB-1 was clearly lower than the shNT+HMGB-1 group, but still higher than the other three groups (H). The levels of TNF-α and IL-1β in lung were analyzed by ELISA after transfection and 24 h after HMGB-1 administration. The levels of IL-1β in lung were not significantly different between the control, negative, shTLR4, and anti-HMGB-1 groups. After treatment with HMGB-1, the levels of IL-1β and TNF-α increased dramatically in the HMGB-1 group and increased slightly in the shTLR4+HMGB-1 group, but did not change in the HMGB-1+anti-HMGB-1 group (J, K). TLR4 protein expression was measured by western blot analysis and TLR4 mRNA expression was measured by real-time quantitative PCR after transfection and 24 h after HMGB-1 administration. The TLR4 protein and mRNA levels were very low in the shTLR4 group, but dramatically increased in the HMGB-1 group. The levels of TLR4 expression in the other five groups were equal to each other (I, L). Data are shown as the mean±standard deviation (SD), n = 6, * P

Techniques Used: Inhibition, Expressing, Transfection, Enzyme-linked Immunosorbent Assay, Western Blot, Real-time Polymerase Chain Reaction, Standard Deviation

Lung inflammatory response after HMGB-1 treatment. Normal lung architecture was observed in the control group and 0 µg HMGB-1 group (A, B). Intense interstitial accumulation of neutrophils and edema was observed in 50 µg and 100 µg HMGB-1 groups (C, D). Rats exposed to 0 µg HMGB1 showed low lung histological scores. Intratracheal exposure of 50 µg and 100 µg of HMGB1 increased lung histological scores significantly (E). IL-1β and TNF-α levels in lungs were evaluated by ELISA. Compared with the control group, inflammatory mediators in the 0 µg HMGB-1 group were not significantly different, and marked increases were observed in 50 µg and 100 µg HMGB-1 groups (F, G). The increases in the 100 µg HMGB-1 group were higher than those in the 50 µg HMGB-1 group (F, G). Data are shown as the mean± SD, n = 6, * P
Figure Legend Snippet: Lung inflammatory response after HMGB-1 treatment. Normal lung architecture was observed in the control group and 0 µg HMGB-1 group (A, B). Intense interstitial accumulation of neutrophils and edema was observed in 50 µg and 100 µg HMGB-1 groups (C, D). Rats exposed to 0 µg HMGB1 showed low lung histological scores. Intratracheal exposure of 50 µg and 100 µg of HMGB1 increased lung histological scores significantly (E). IL-1β and TNF-α levels in lungs were evaluated by ELISA. Compared with the control group, inflammatory mediators in the 0 µg HMGB-1 group were not significantly different, and marked increases were observed in 50 µg and 100 µg HMGB-1 groups (F, G). The increases in the 100 µg HMGB-1 group were higher than those in the 50 µg HMGB-1 group (F, G). Data are shown as the mean± SD, n = 6, * P

Techniques Used: Enzyme-linked Immunosorbent Assay

The effect of TLR4 inhibition on the inflammatory response by HMGB-1-treated NR8383 cells. RNAi was used to inhibit TLR4 expression in NR8383 cells. The levels of IL-1β and TNF-α in the culture supernatants were determined by ELISA after transfection and 24 h after HMGB-1 administration. The levels of IL-1β and TNF-α in negative, shTLR4, and anti-HMGB-1 groups were comparable to the control group. After treatment with HMGB-1, the IL-1β and TNF-α concentrations in the HMGB-1 group were higher than those in the shTLR4+HMGB-1 group, while there were no changes in these concentrations in the HMGB-1+anti-HMGB-1 group (A, B). The expression of TLR4 was assessed by western lot and real-time quantitative PCR after transfection and 24 h after HMGB-1 administration. After transfection, The TLR4 protein and mRNA levels were very low in the shTLR4 group, decreasing by approximately 60%. After treatment with HMGB-1, the levels of TLR4 protein and mRNA in the shTLR4+HMGB-1 group returned to the levels of the control. In addition, the levels of TLR4 protein and mRNA remained the same in the anti-HMGB-1 and HMGB-1+anti-HMGB-1 groups (C, D). Data are shown as the mean±SD, n = 6, ** P
Figure Legend Snippet: The effect of TLR4 inhibition on the inflammatory response by HMGB-1-treated NR8383 cells. RNAi was used to inhibit TLR4 expression in NR8383 cells. The levels of IL-1β and TNF-α in the culture supernatants were determined by ELISA after transfection and 24 h after HMGB-1 administration. The levels of IL-1β and TNF-α in negative, shTLR4, and anti-HMGB-1 groups were comparable to the control group. After treatment with HMGB-1, the IL-1β and TNF-α concentrations in the HMGB-1 group were higher than those in the shTLR4+HMGB-1 group, while there were no changes in these concentrations in the HMGB-1+anti-HMGB-1 group (A, B). The expression of TLR4 was assessed by western lot and real-time quantitative PCR after transfection and 24 h after HMGB-1 administration. After transfection, The TLR4 protein and mRNA levels were very low in the shTLR4 group, decreasing by approximately 60%. After treatment with HMGB-1, the levels of TLR4 protein and mRNA in the shTLR4+HMGB-1 group returned to the levels of the control. In addition, the levels of TLR4 protein and mRNA remained the same in the anti-HMGB-1 and HMGB-1+anti-HMGB-1 groups (C, D). Data are shown as the mean±SD, n = 6, ** P

Techniques Used: Inhibition, Expressing, Enzyme-linked Immunosorbent Assay, Transfection, Western Blot, Real-time Polymerase Chain Reaction

IL-1β and TNF-α levels in culture media change after HMGB-1 stimulation in NR8383. NR8383 cells were stimulated with rhHMGB-1at different concentrations and then the IL-1β and TNF-α levels in the culture media were evaluated by ELISA assay at 24 h post-treatment. After treatment, the IL-1β levels increased slightly in the 10 ng/ml HMGB-1 group. In the 50 and 100 ng/ml HMGB-1 groups, the levels of IL-1β increased dramatically (A). Compared with the control group, TNF-α levels did not change in the 10 ng/ml HMGB-1 group, but clearly increased in the 50 and 100 ng/ml HMGB-1 groups (B). Data are shown as the mean±SD, n = 6, * P
Figure Legend Snippet: IL-1β and TNF-α levels in culture media change after HMGB-1 stimulation in NR8383. NR8383 cells were stimulated with rhHMGB-1at different concentrations and then the IL-1β and TNF-α levels in the culture media were evaluated by ELISA assay at 24 h post-treatment. After treatment, the IL-1β levels increased slightly in the 10 ng/ml HMGB-1 group. In the 50 and 100 ng/ml HMGB-1 groups, the levels of IL-1β increased dramatically (A). Compared with the control group, TNF-α levels did not change in the 10 ng/ml HMGB-1 group, but clearly increased in the 50 and 100 ng/ml HMGB-1 groups (B). Data are shown as the mean±SD, n = 6, * P

Techniques Used: Enzyme-linked Immunosorbent Assay

20) Product Images from "Activation of TRPV1 Prevents OxLDL-Induced Lipid Accumulation and TNF-α-Induced Inflammation in Macrophages: Role of Liver X Receptor α"

Article Title: Activation of TRPV1 Prevents OxLDL-Induced Lipid Accumulation and TNF-α-Induced Inflammation in Macrophages: Role of Liver X Receptor α

Journal: Mediators of Inflammation

doi: 10.1155/2013/925171

Knockdown of LXR α diminishes the protective effect of TRPV1 agonists against TNF- α -induced inflammation in macrophages. (a) BMDMs were pretreated with capsazepine (10 μ M) for 1 h or (b) transfected with control siRNA (50 nmol/L) or LXR α siRNA (50 nmol/L) for 24 h, then incubated with vehicle (DMSO), evodiamine (500 nM), and capsaicin (10 μ M) with or without TNF- α (10 ng/mL) for an additional 18 h. ELISA of levels of MCP-1, IL-6, and MIP-2 in the culture medium. Data are mean ± SD from 5 independent experiments. * P
Figure Legend Snippet: Knockdown of LXR α diminishes the protective effect of TRPV1 agonists against TNF- α -induced inflammation in macrophages. (a) BMDMs were pretreated with capsazepine (10 μ M) for 1 h or (b) transfected with control siRNA (50 nmol/L) or LXR α siRNA (50 nmol/L) for 24 h, then incubated with vehicle (DMSO), evodiamine (500 nM), and capsaicin (10 μ M) with or without TNF- α (10 ng/mL) for an additional 18 h. ELISA of levels of MCP-1, IL-6, and MIP-2 in the culture medium. Data are mean ± SD from 5 independent experiments. * P

Techniques Used: Transfection, Incubation, Enzyme-linked Immunosorbent Assay

21) Product Images from "Age-Related CD4+CD25+Foxp3+ Regulatory T-Cell Responses During Plasmodium berghei ANKA Infection in Mice Susceptible or Resistant to Cerebral Malaria"

Article Title: Age-Related CD4+CD25+Foxp3+ Regulatory T-Cell Responses During Plasmodium berghei ANKA Infection in Mice Susceptible or Resistant to Cerebral Malaria

Journal: The Korean Journal of Parasitology

doi: 10.3347/kjp.2013.51.3.289

Level of IFN-γ and TNF-α in young and middle-aged mice with P. berghei ANKA infection. ELISA was performed to detect the level of IFN-γ and TNF-α in supernatants of cultured spleen cells from infected mice. Results are presented as the arithmetic mean of 9 mice per group±SE. a P
Figure Legend Snippet: Level of IFN-γ and TNF-α in young and middle-aged mice with P. berghei ANKA infection. ELISA was performed to detect the level of IFN-γ and TNF-α in supernatants of cultured spleen cells from infected mice. Results are presented as the arithmetic mean of 9 mice per group±SE. a P

Techniques Used: Mouse Assay, Infection, Enzyme-linked Immunosorbent Assay, Cell Culture

22) Product Images from "Role of promoting inflammation of Krüppel-like factor 6 in acute kidney injury"

Article Title: Role of promoting inflammation of Krüppel-like factor 6 in acute kidney injury

Journal: Renal Failure

doi: 10.1080/0886022X.2020.1793353

The expression of inflammation cytokines in IR model. (A–D) Quantitative analysis of ICAM-1, MCP-1, TNF-α, and IL-10 released in the serum and kidney tissue using ELISA. (E–H) Quantitative analysis of ICAM-1, MCP-1, TNF-α, and IL-10 mRNA expression in the kidneys by RT-PCR. * p
Figure Legend Snippet: The expression of inflammation cytokines in IR model. (A–D) Quantitative analysis of ICAM-1, MCP-1, TNF-α, and IL-10 released in the serum and kidney tissue using ELISA. (E–H) Quantitative analysis of ICAM-1, MCP-1, TNF-α, and IL-10 mRNA expression in the kidneys by RT-PCR. * p

Techniques Used: Expressing, Enzyme-linked Immunosorbent Assay, Reverse Transcription Polymerase Chain Reaction

KLF6 expressed in the kidney during IR in the rat model. (A) Quantitative analysis of KLF6 released in the serum and kidney tissue using ELISA. (B) The expression of KLF6 was measured by RT-PCR, and the ratio of KLF6/Actin was normalized to the sham group. (C) The expression of KLF6 was measured by Western blot, and the ratio of KLF6/Actin was normalized to the sham group. (D) Representative photomicrographs of kidney sections Immunohistochemical staining for KLF6 in mice after sham or IR treatments . * p
Figure Legend Snippet: KLF6 expressed in the kidney during IR in the rat model. (A) Quantitative analysis of KLF6 released in the serum and kidney tissue using ELISA. (B) The expression of KLF6 was measured by RT-PCR, and the ratio of KLF6/Actin was normalized to the sham group. (C) The expression of KLF6 was measured by Western blot, and the ratio of KLF6/Actin was normalized to the sham group. (D) Representative photomicrographs of kidney sections Immunohistochemical staining for KLF6 in mice after sham or IR treatments . * p

Techniques Used: Enzyme-linked Immunosorbent Assay, Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot, Immunohistochemistry, Staining, Mouse Assay

23) Product Images from "Reversing established sepsis with antagonists of endogenous high-mobility group box 1"

Article Title: Reversing established sepsis with antagonists of endogenous high-mobility group box 1

Journal: Proceedings of the National Academy of Sciences of the United States of America

doi: 10.1073/pnas.2434651100

A box antagonizes HMGB1 in vitro . Subconfluent RAW 264.7 cells in 24-well dishes were treated with HMGB1 or IL-1β and various concentrations of A box (as indicated) for 16 h in Opti-MEM I medium in the presence of polymyxin B. The TNF- or IL-1β-stimulating activity was expressed as percent of maximal response, and inhibition by A box was expressed as percent of HMGB1 or IL-1β alone. ( A ) HMGB1 (1 μg/ml) induced the release of 13,344 ± 2,335 pg of TNF per ml. A box dose-dependently inhibited HMGB1-induced TNF release with an apparent EC 50 of 7.5 μg/ml. ( B ) Similarly, HMGB1-induced IL-1β was dose-dependently inhibited by the addition of A box. ( C ) In contrast, A box did not have any significant effect on IL-1β-induced TNF release. TNF release stimulated by IL-1β (100 ng/ml) was 640 ± 160 pg of TNF per ml. ( D ) RAW 267.4 cells were stimulated with LPS and A box, B box, or GST vector, or LPS at the concentration indicated for 16 h, and IL-10 released in the conditioned medium was measured with an ELISA kit. Data represent three or four separate experiments.
Figure Legend Snippet: A box antagonizes HMGB1 in vitro . Subconfluent RAW 264.7 cells in 24-well dishes were treated with HMGB1 or IL-1β and various concentrations of A box (as indicated) for 16 h in Opti-MEM I medium in the presence of polymyxin B. The TNF- or IL-1β-stimulating activity was expressed as percent of maximal response, and inhibition by A box was expressed as percent of HMGB1 or IL-1β alone. ( A ) HMGB1 (1 μg/ml) induced the release of 13,344 ± 2,335 pg of TNF per ml. A box dose-dependently inhibited HMGB1-induced TNF release with an apparent EC 50 of 7.5 μg/ml. ( B ) Similarly, HMGB1-induced IL-1β was dose-dependently inhibited by the addition of A box. ( C ) In contrast, A box did not have any significant effect on IL-1β-induced TNF release. TNF release stimulated by IL-1β (100 ng/ml) was 640 ± 160 pg of TNF per ml. ( D ) RAW 267.4 cells were stimulated with LPS and A box, B box, or GST vector, or LPS at the concentration indicated for 16 h, and IL-10 released in the conditioned medium was measured with an ELISA kit. Data represent three or four separate experiments.

Techniques Used: In Vitro, Activity Assay, Inhibition, Plasmid Preparation, Concentration Assay, Enzyme-linked Immunosorbent Assay

Anti-HMGB1 antibodies specifically inhibit HMGB1-induced cytokine release. Murine macrophage-like RAW 264.7 cells were stimulated with HMGB1, IL-1β, or TNF as indicated for 16 h in serum-free Opti-MEM medium in the absence or presence of purified rabbit anti-HMGB1 IgG antibodies or nonimmune rabbit IgG (as control). Conditioned media were collected and assayed for TNF and IL-6 levels by use of ELISA kits. Data represent mean ± SEM of three to six independent experiments, each done in duplicate. * , P
Figure Legend Snippet: Anti-HMGB1 antibodies specifically inhibit HMGB1-induced cytokine release. Murine macrophage-like RAW 264.7 cells were stimulated with HMGB1, IL-1β, or TNF as indicated for 16 h in serum-free Opti-MEM medium in the absence or presence of purified rabbit anti-HMGB1 IgG antibodies or nonimmune rabbit IgG (as control). Conditioned media were collected and assayed for TNF and IL-6 levels by use of ELISA kits. Data represent mean ± SEM of three to six independent experiments, each done in duplicate. * , P

Techniques Used: Purification, Enzyme-linked Immunosorbent Assay

24) Product Images from "Valproic acid attenuates nitric oxide and interleukin-1β production in lipopolysaccharide-stimulated iron-rich microglia"

Article Title: Valproic acid attenuates nitric oxide and interleukin-1β production in lipopolysaccharide-stimulated iron-rich microglia

Journal: Biomedical Reports

doi: 10.3892/br.2018.1062

Effect of VPA on (A) NO 2 − and (B) IL-1β (B) levels in iron-rich cultures of activated microglia. Microglial cells were plated in 24-well plates. After 24 h of the indicated treatments, the supernatants were collected and assayed for NO and IL-1β production using Griess reagent and ELISA, respectively. **P
Figure Legend Snippet: Effect of VPA on (A) NO 2 − and (B) IL-1β (B) levels in iron-rich cultures of activated microglia. Microglial cells were plated in 24-well plates. After 24 h of the indicated treatments, the supernatants were collected and assayed for NO and IL-1β production using Griess reagent and ELISA, respectively. **P

Techniques Used: Enzyme-linked Immunosorbent Assay

25) Product Images from "Host Gene Expression Profiling of Dengue Virus Infection in Cell Lines and Patients"

Article Title: Host Gene Expression Profiling of Dengue Virus Infection in Cell Lines and Patients

Journal: PLoS Neglected Tropical Diseases

doi: 10.1371/journal.pntd.0000086

Culture supernatants of A549 and HepG2 cells infected with TSV01 (MOI 10) or heat-inactivated TSV-01 (HI-TSV01; MOI 10) for 72 hrs were analyzed for IP-10 (A) and I-TAC (B) by ELISA. Results are expressed as mean±s.e.m. where n = 3–5. Significance (P
Figure Legend Snippet: Culture supernatants of A549 and HepG2 cells infected with TSV01 (MOI 10) or heat-inactivated TSV-01 (HI-TSV01; MOI 10) for 72 hrs were analyzed for IP-10 (A) and I-TAC (B) by ELISA. Results are expressed as mean±s.e.m. where n = 3–5. Significance (P

Techniques Used: Infection, Enzyme-linked Immunosorbent Assay

26) Product Images from "Regulation and biological role of the peptide/histidine transporter SLC15A3 in Toll-like receptor-mediated inflammatory responses in macrophage"

Article Title: Regulation and biological role of the peptide/histidine transporter SLC15A3 in Toll-like receptor-mediated inflammatory responses in macrophage

Journal: Cell Death & Disease

doi: 10.1038/s41419-018-0809-1

SLC15A3 expression was increased in mice with E. coli -induced peritonitis, and positively related to inflammation. Mice were intraperitoneally injected with PBS or E.coli (1 × 10 7 cfu/mL, 0.5 mL/ 25 g), and the serum or peritoneal macrophages collected after 24 h. a IL-6 and TNF-α protein levels in mouse serum were determined by ELISA assays. b , c Slc15a3 mRNA ( b ) and protein expression ( c ) in PMs was determined by real-time PCR and Western blotting. Quantification of protein (i.e., SLC15A3/GAPDH ratio) is shown in the right side of each western blot figure. d mRNA expression of Il-6 and Tnf-α in peritoneal macrophages as a function of time. e Correlation analysis of Slc15a3 with Il-6 or Tnf-α was performed by linear regression, in which Slc15a3 mRNA expression in x-axis and Il-6 (left) or Tnf-α (right) mRNA expression in y -axis. Statistical analyses were performed using an unpaired t test. * P
Figure Legend Snippet: SLC15A3 expression was increased in mice with E. coli -induced peritonitis, and positively related to inflammation. Mice were intraperitoneally injected with PBS or E.coli (1 × 10 7 cfu/mL, 0.5 mL/ 25 g), and the serum or peritoneal macrophages collected after 24 h. a IL-6 and TNF-α protein levels in mouse serum were determined by ELISA assays. b , c Slc15a3 mRNA ( b ) and protein expression ( c ) in PMs was determined by real-time PCR and Western blotting. Quantification of protein (i.e., SLC15A3/GAPDH ratio) is shown in the right side of each western blot figure. d mRNA expression of Il-6 and Tnf-α in peritoneal macrophages as a function of time. e Correlation analysis of Slc15a3 with Il-6 or Tnf-α was performed by linear regression, in which Slc15a3 mRNA expression in x-axis and Il-6 (left) or Tnf-α (right) mRNA expression in y -axis. Statistical analyses were performed using an unpaired t test. * P

Techniques Used: Expressing, Mouse Assay, Injection, Enzyme-linked Immunosorbent Assay, Real-time Polymerase Chain Reaction, Western Blot

Knockdown or overexpression of SLC15A3 influenced the TLR4-dependent proinflammatory cytokine production. a mRNA and protein expression of SLC15A3 was determined in THP-1 cells transfected with 30 nM control siRNA (Ctrl) or two different SLC15A3 siRNA (#1 and #2) after 24 h transfection. Quantification of protein (i.e., SLC15A3/GAPDH ratio) is shown in the right side of each Western blot figure. b THP-1 cells transfected with 30 nM control siRNA (Ctrl) or SLC15A3 siRNA (#1). Twenty-four hours after transfection, the cells were treated with 100 ng/mL LPS for 12 h. The levels of IL-6 and TNF-α in the cell culture supernatants were determined by ELISA assays. c , d A549 cells transfected with 1 μg pcDNA3.1 or human SLC15A3 plasmid. Twenty-four hours after transfection, the cells were treated with 1 μg/mL LPS for 8 h. The mRNA ( c ) and protein ( d ) levels of IL-6 in cells and cell culture supernatants were determined by real-time PCR and ELISA assays. Statistical analyses were performed using an unpaired t test. * P
Figure Legend Snippet: Knockdown or overexpression of SLC15A3 influenced the TLR4-dependent proinflammatory cytokine production. a mRNA and protein expression of SLC15A3 was determined in THP-1 cells transfected with 30 nM control siRNA (Ctrl) or two different SLC15A3 siRNA (#1 and #2) after 24 h transfection. Quantification of protein (i.e., SLC15A3/GAPDH ratio) is shown in the right side of each Western blot figure. b THP-1 cells transfected with 30 nM control siRNA (Ctrl) or SLC15A3 siRNA (#1). Twenty-four hours after transfection, the cells were treated with 100 ng/mL LPS for 12 h. The levels of IL-6 and TNF-α in the cell culture supernatants were determined by ELISA assays. c , d A549 cells transfected with 1 μg pcDNA3.1 or human SLC15A3 plasmid. Twenty-four hours after transfection, the cells were treated with 1 μg/mL LPS for 8 h. The mRNA ( c ) and protein ( d ) levels of IL-6 in cells and cell culture supernatants were determined by real-time PCR and ELISA assays. Statistical analyses were performed using an unpaired t test. * P

Techniques Used: Over Expression, Expressing, Transfection, Western Blot, Cell Culture, Enzyme-linked Immunosorbent Assay, Plasmid Preparation, Real-time Polymerase Chain Reaction

27) Product Images from "Identification of small‐molecule elastase inhibitors as antagonists of IL‐36 cytokine activation"

Article Title: Identification of small‐molecule elastase inhibitors as antagonists of IL‐36 cytokine activation

Journal: FEBS Open Bio

doi: 10.1002/2211-5463.12406

IL‐36γ is processed and activated by NE. HeLa IL‐36R cells were either left untreated or were treated with the indicated concentrations of full‐length recombinant human IL‐36γ (ranging from 5 to 0.3 n m ), or the same amounts of IL‐36γ that had been pre‐incubated for 2 h at 37 °C with purified HNE (50 n m ). Twenty‐four hours after incubation with either full‐length or elastase‐processed IL‐36γ preparations, cytokine concentrations in the culture SNs were determined by ELISA. The following cytokines were measured: (A) IL‐6, (B) IL‐8 and (C) CXCL1. Results shown are representative of at least three independent experiments. Error bars represent the mean ± SEM of triplicate determinations from a representative experiment.
Figure Legend Snippet: IL‐36γ is processed and activated by NE. HeLa IL‐36R cells were either left untreated or were treated with the indicated concentrations of full‐length recombinant human IL‐36γ (ranging from 5 to 0.3 n m ), or the same amounts of IL‐36γ that had been pre‐incubated for 2 h at 37 °C with purified HNE (50 n m ). Twenty‐four hours after incubation with either full‐length or elastase‐processed IL‐36γ preparations, cytokine concentrations in the culture SNs were determined by ELISA. The following cytokines were measured: (A) IL‐6, (B) IL‐8 and (C) CXCL1. Results shown are representative of at least three independent experiments. Error bars represent the mean ± SEM of triplicate determinations from a representative experiment.

Techniques Used: Recombinant, Incubation, Purification, Enzyme-linked Immunosorbent Assay

Inhibition of elastase‐mediated IL‐36γ activation by LCB016 and derivatives. (A) Schematic overview of the IL‐36 activity bioassay. (B) Purified recombinant IL‐36γ was incubated for 2 h at 37 °C in the presence or absence of elastase (50 n m ), either alone or in combination with the indicated small‐molecule elastase inhibitors. Reaction products were then added to HeLa IL‐36R cells, such that the final concentration of IL‐36γ was 500 p m . After 24 h, IL‐6 cytokine concentrations in cell culture SNs were determined by ELISA. Results shown are representative of at least three independent experiments. Error bars represent the mean ± SEM of triplicate determinations from a representative experiment.
Figure Legend Snippet: Inhibition of elastase‐mediated IL‐36γ activation by LCB016 and derivatives. (A) Schematic overview of the IL‐36 activity bioassay. (B) Purified recombinant IL‐36γ was incubated for 2 h at 37 °C in the presence or absence of elastase (50 n m ), either alone or in combination with the indicated small‐molecule elastase inhibitors. Reaction products were then added to HeLa IL‐36R cells, such that the final concentration of IL‐36γ was 500 p m . After 24 h, IL‐6 cytokine concentrations in cell culture SNs were determined by ELISA. Results shown are representative of at least three independent experiments. Error bars represent the mean ± SEM of triplicate determinations from a representative experiment.

Techniques Used: Inhibition, Activation Assay, Activity Assay, Purification, Recombinant, Incubation, Concentration Assay, Cell Culture, Enzyme-linked Immunosorbent Assay

Novel elastase inhibitors suppress IL‐36γ activation by proteases released from human neutrophil degranulate SNs. (A) Schematic representation of PMA‐induced neutrophil degranulation. (B) Assessment of elastase and CatG activity in SNs from untreated versus PMA‐activated human neutrophils. Protease activity was assessed through hydrolysis of FLF‐sBzl (CatG activity) or AAPV‐AMC (elastase/PR3 activity), as before. (C) Purified recombinant human IL‐36γ was incubated for 2 h at 37 °C in the presence or absence of control (unstimulated) or PMA‐activated neutrophil SNs (1 : 8), either alone or in combination with the indicated small‐molecule elastase inhibitors. Neutrophil cathepsin G inhibitor I (CGi) and elastase inhibitor IV (NEi) served as controls. Reaction products were then added to HeLa IL‐36R cells, such that the final concentration of IL‐36γ was 500 p m . After 24 h, IL‐6 cytokine concentrations in cell culture SNs were determined by ELISA. (D) Purified recombinant IL‐36β was incubated for 2 h at 37 °C in the presence or absence of control (unstimulated) or PMA‐activated neutrophil SNs (1 : 8), either alone or in combination with the indicated small‐molecule elastase inhibitors. Neutrophil cathepsin G inhibitor I (CGi) and elastase inhibitor IV (NEi) served as controls. Reaction products were then added to HeLa IL‐36R cells, such that the final concentration of IL‐36β was 500 p m . After 24 h, IL‐6 cytokine concentrations in cell culture SNs were determined by ELISA. Results shown are representative of at least three independent experiments. Error bars represent the mean ± SEM of triplicate determinations from a representative experiment.
Figure Legend Snippet: Novel elastase inhibitors suppress IL‐36γ activation by proteases released from human neutrophil degranulate SNs. (A) Schematic representation of PMA‐induced neutrophil degranulation. (B) Assessment of elastase and CatG activity in SNs from untreated versus PMA‐activated human neutrophils. Protease activity was assessed through hydrolysis of FLF‐sBzl (CatG activity) or AAPV‐AMC (elastase/PR3 activity), as before. (C) Purified recombinant human IL‐36γ was incubated for 2 h at 37 °C in the presence or absence of control (unstimulated) or PMA‐activated neutrophil SNs (1 : 8), either alone or in combination with the indicated small‐molecule elastase inhibitors. Neutrophil cathepsin G inhibitor I (CGi) and elastase inhibitor IV (NEi) served as controls. Reaction products were then added to HeLa IL‐36R cells, such that the final concentration of IL‐36γ was 500 p m . After 24 h, IL‐6 cytokine concentrations in cell culture SNs were determined by ELISA. (D) Purified recombinant IL‐36β was incubated for 2 h at 37 °C in the presence or absence of control (unstimulated) or PMA‐activated neutrophil SNs (1 : 8), either alone or in combination with the indicated small‐molecule elastase inhibitors. Neutrophil cathepsin G inhibitor I (CGi) and elastase inhibitor IV (NEi) served as controls. Reaction products were then added to HeLa IL‐36R cells, such that the final concentration of IL‐36β was 500 p m . After 24 h, IL‐6 cytokine concentrations in cell culture SNs were determined by ELISA. Results shown are representative of at least three independent experiments. Error bars represent the mean ± SEM of triplicate determinations from a representative experiment.

Techniques Used: Activation Assay, Activity Assay, Purification, Recombinant, Incubation, Concentration Assay, Cell Culture, Enzyme-linked Immunosorbent Assay

28) Product Images from "Daumone fed late in life improves survival and reduces hepatic inflammation and fibrosis in mice"

Article Title: Daumone fed late in life improves survival and reduces hepatic inflammation and fibrosis in mice

Journal: Aging Cell

doi: 10.1111/acel.12224

Effects of daumone on hepatic inflammation and fibrosis in aged mice. (A) Macrophage infiltration was detected by F4/80 immunohistochemical staining. Brown, F4/80; blue, hematoxylin. (B) Collagen was stained with Masson’s trichrome staining. The positive area was quantitated using Image-Pro Plus and presented as the mean±SE of 4 mice/group. (C, E) mRNA levels were determined by real-time qRT–PCR. (D, F) Immunoblots of the liver are presented and quantitated. Transforming growth factor-β1 (TGF-β1) was measured by ELISA. mRNA and protein levels were presented as the mean±SE of 7–17 mice/group. * P
Figure Legend Snippet: Effects of daumone on hepatic inflammation and fibrosis in aged mice. (A) Macrophage infiltration was detected by F4/80 immunohistochemical staining. Brown, F4/80; blue, hematoxylin. (B) Collagen was stained with Masson’s trichrome staining. The positive area was quantitated using Image-Pro Plus and presented as the mean±SE of 4 mice/group. (C, E) mRNA levels were determined by real-time qRT–PCR. (D, F) Immunoblots of the liver are presented and quantitated. Transforming growth factor-β1 (TGF-β1) was measured by ELISA. mRNA and protein levels were presented as the mean±SE of 7–17 mice/group. * P

Techniques Used: Mouse Assay, Immunohistochemistry, Staining, Quantitative RT-PCR, Western Blot, Enzyme-linked Immunosorbent Assay

29) Product Images from "Post-Translational Inhibition of IP-10 Secretion in IEC by Probiotic Bacteria: Impact on Chronic Inflammation"

Article Title: Post-Translational Inhibition of IP-10 Secretion in IEC by Probiotic Bacteria: Impact on Chronic Inflammation

Journal: PLoS ONE

doi: 10.1371/journal.pone.0004365

The inhibition of IP-10 secretion is independent of the signal-specific activation of IEC. (A) Mode-K cells were stimulated for 24 h with IFNγ (100 ng/ml) and L. casei (moi 20). Cell culture supernatants were analyzed by ELISA and (B) intracellular IP-10 protein levels by Western blot. (C) Mode-K cells were transfected with an IP-10 overexpression (pIP-10-DsRed)- or a ctr-DsRed-plasmid (transfection control) for 6 h and were then treated with L. casei for 24 h. IP-10 levels in the supernatants were determined by ELISA and intracellular DsRed-tagged IP-10 (IP-10-DsRed) or DsRed levels (transfection control) were determined by Western blot. Bars in A/C represent mean values (+/− SD) of triplicate samples and all figures are representative for three independent experiments.
Figure Legend Snippet: The inhibition of IP-10 secretion is independent of the signal-specific activation of IEC. (A) Mode-K cells were stimulated for 24 h with IFNγ (100 ng/ml) and L. casei (moi 20). Cell culture supernatants were analyzed by ELISA and (B) intracellular IP-10 protein levels by Western blot. (C) Mode-K cells were transfected with an IP-10 overexpression (pIP-10-DsRed)- or a ctr-DsRed-plasmid (transfection control) for 6 h and were then treated with L. casei for 24 h. IP-10 levels in the supernatants were determined by ELISA and intracellular DsRed-tagged IP-10 (IP-10-DsRed) or DsRed levels (transfection control) were determined by Western blot. Bars in A/C represent mean values (+/− SD) of triplicate samples and all figures are representative for three independent experiments.

Techniques Used: Inhibition, Activation Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Western Blot, Transfection, Over Expression, Plasmid Preparation

30) Product Images from "MircroRNA-19a promotes vascular inflammation and foam cell formation by targeting HBP-1 in atherogenesis"

Article Title: MircroRNA-19a promotes vascular inflammation and foam cell formation by targeting HBP-1 in atherogenesis

Journal: Scientific Reports

doi: 10.1038/s41598-017-12167-z

Inflammatory factor production and lipid uptake is regulated by miR-19a. ( A , D ) The expression of miR-19a was examined by qRT-PCR after transfection of miR-19a mimic, inhibitor, and control (100 nM) into macrophages that had been previously differentiated from THP-1 cells. ( B , E ) THP-1-derived macrophages were transfected with miR-19a mimic or its inhibitor at the indicated dose, the concentration of IL-6 in macrophages supernatant were measured by ELISA Kit. ( C , F ) THP-1-derived macrophages were transfected with miR-19a mimic or inhibitor at the indicated dose, the concentration of TNF-α in macrophages supernatant were measured by ELISA Kit. ( G , H ) THP-1-derived macrophages were transfected with miR-19a inhibitor at the indicated dose for 24h, and then incubated with Dil-oxLDL for 6h. Lipid uptake of macrophages was assessed by fluorescent imaging and fluorescence activated cell sorting (magnification 200×). ( I ) The quantitative results of the lipid uptake. * P
Figure Legend Snippet: Inflammatory factor production and lipid uptake is regulated by miR-19a. ( A , D ) The expression of miR-19a was examined by qRT-PCR after transfection of miR-19a mimic, inhibitor, and control (100 nM) into macrophages that had been previously differentiated from THP-1 cells. ( B , E ) THP-1-derived macrophages were transfected with miR-19a mimic or its inhibitor at the indicated dose, the concentration of IL-6 in macrophages supernatant were measured by ELISA Kit. ( C , F ) THP-1-derived macrophages were transfected with miR-19a mimic or inhibitor at the indicated dose, the concentration of TNF-α in macrophages supernatant were measured by ELISA Kit. ( G , H ) THP-1-derived macrophages were transfected with miR-19a inhibitor at the indicated dose for 24h, and then incubated with Dil-oxLDL for 6h. Lipid uptake of macrophages was assessed by fluorescent imaging and fluorescence activated cell sorting (magnification 200×). ( I ) The quantitative results of the lipid uptake. * P

Techniques Used: Expressing, Quantitative RT-PCR, Transfection, Derivative Assay, Concentration Assay, Enzyme-linked Immunosorbent Assay, Incubation, Imaging, Fluorescence, FACS

31) Product Images from "Isoflurane attenuates murine lupus nephritis by inhibiting NLRP3 inflammasome activation"

Article Title: Isoflurane attenuates murine lupus nephritis by inhibiting NLRP3 inflammasome activation

Journal: International Journal of Clinical and Experimental Medicine

doi:

ISO suppressed NLRP3 inflammasome activation in the kidney of MRL/lpr mice. A. Western blot was performed to analyze the expression of NLRP3, ASC, and caspase-1 p20. b-actin was used as the endogenous control. B. Quantitative expression of NLRP3, ASC, and caspase-1 p20 was normalized against b-actin. C and D. Renal and serum IL-1β levels were measured by ELISA. Each experiment was performed in triplicate. Values are presented as mean ± SD. ** P
Figure Legend Snippet: ISO suppressed NLRP3 inflammasome activation in the kidney of MRL/lpr mice. A. Western blot was performed to analyze the expression of NLRP3, ASC, and caspase-1 p20. b-actin was used as the endogenous control. B. Quantitative expression of NLRP3, ASC, and caspase-1 p20 was normalized against b-actin. C and D. Renal and serum IL-1β levels were measured by ELISA. Each experiment was performed in triplicate. Values are presented as mean ± SD. ** P

Techniques Used: Activation Assay, Mouse Assay, Western Blot, Expressing, Enzyme-linked Immunosorbent Assay

ISO treatment resulted in the reduction of Thl7/Treg cell ratio, macrophage infiltration, and IL-17 and TNF-α production in MRL/lpr mice. A. Thl7/Treg cell ratio was determined by flow cytometry analysis. B. ELISA analysis of serum IL-17 level. C. Macrophage accumulation was assessed based on the number of CD68-positive cells. D and E. Renal and serum TNF-α levels were measured by ELISA. Each experiment was performed in triplicate. Values are presented as mean ± SD. ** P
Figure Legend Snippet: ISO treatment resulted in the reduction of Thl7/Treg cell ratio, macrophage infiltration, and IL-17 and TNF-α production in MRL/lpr mice. A. Thl7/Treg cell ratio was determined by flow cytometry analysis. B. ELISA analysis of serum IL-17 level. C. Macrophage accumulation was assessed based on the number of CD68-positive cells. D and E. Renal and serum TNF-α levels were measured by ELISA. Each experiment was performed in triplicate. Values are presented as mean ± SD. ** P

Techniques Used: Mouse Assay, Flow Cytometry, Cytometry, Enzyme-linked Immunosorbent Assay

32) Product Images from "Neuroprotective effects of intrastriatal injection of rapamycin in a mouse model of excitotoxicity induced by quinolinic acid"

Article Title: Neuroprotective effects of intrastriatal injection of rapamycin in a mouse model of excitotoxicity induced by quinolinic acid

Journal: Journal of Neuroinflammation

doi: 10.1186/s12974-017-0793-x

Effect of rapamycin on the levels of neurotrophic factors in the striatum after 8 h of QA administration. Rapamycin was injected 15 min before QA injection into the striatum, and the quantification of BDNF ( n = 6–7 animals/group) ( a ) and NGF ( n = 5–7 animals/group) ( b ) was determined after 8 h of the QA injection using ELISA. Results are expressed as mean ± SEM. There are no statistically significant differences among treatment groups (one-way ANOVA followed by Newman-Keuls test)
Figure Legend Snippet: Effect of rapamycin on the levels of neurotrophic factors in the striatum after 8 h of QA administration. Rapamycin was injected 15 min before QA injection into the striatum, and the quantification of BDNF ( n = 6–7 animals/group) ( a ) and NGF ( n = 5–7 animals/group) ( b ) was determined after 8 h of the QA injection using ELISA. Results are expressed as mean ± SEM. There are no statistically significant differences among treatment groups (one-way ANOVA followed by Newman-Keuls test)

Techniques Used: Injection, Enzyme-linked Immunosorbent Assay

33) Product Images from "Preconditioning with Endoplasmic Reticulum Stress Ameliorates Endothelial Cell Inflammation"

Article Title: Preconditioning with Endoplasmic Reticulum Stress Ameliorates Endothelial Cell Inflammation

Journal: PLoS ONE

doi: 10.1371/journal.pone.0110949

SubAB inhibits NF-κB mediated proinflammatory gene expression. HPAEC were treated with 0.1 µg/ml of SubAB or mutant SubA A272 B for 6 h, followed by treatment with ( A C E ) thrombin (5 U/ml) or ( B D F ) TNFα (100 U/ml) for 6 h. Total cell lysates were immunoblotted with ( A B ) anti-ICAM-1 antibody and anti -VCAM-1 antibody. Actin was used to monitor loading . The conditioned media were subjected to ELISA to determine the levels of ( C D ) IL-8 or ( E F ) MCP-1. Data are means ± S.E. (n = 6–9 for each condition). ### p
Figure Legend Snippet: SubAB inhibits NF-κB mediated proinflammatory gene expression. HPAEC were treated with 0.1 µg/ml of SubAB or mutant SubA A272 B for 6 h, followed by treatment with ( A C E ) thrombin (5 U/ml) or ( B D F ) TNFα (100 U/ml) for 6 h. Total cell lysates were immunoblotted with ( A B ) anti-ICAM-1 antibody and anti -VCAM-1 antibody. Actin was used to monitor loading . The conditioned media were subjected to ELISA to determine the levels of ( C D ) IL-8 or ( E F ) MCP-1. Data are means ± S.E. (n = 6–9 for each condition). ### p

Techniques Used: Expressing, Mutagenesis, Enzyme-linked Immunosorbent Assay

34) Product Images from "Prediction of sarcopenia using a combination of multiple serum biomarkers"

Article Title: Prediction of sarcopenia using a combination of multiple serum biomarkers

Journal: Scientific Reports

doi: 10.1038/s41598-018-26617-9

Comparison of serum protein levels between normal and sarcopenic aged subjects. IL-6 ( a ), SPARC ( b ), MIF ( c ) and IGF-1 ( d ) protein levels in human serum were measured using sandwich ELISA. Box plots were used to visualize distribution of each serum protein level. P -values were obtained with two sample t-tests.
Figure Legend Snippet: Comparison of serum protein levels between normal and sarcopenic aged subjects. IL-6 ( a ), SPARC ( b ), MIF ( c ) and IGF-1 ( d ) protein levels in human serum were measured using sandwich ELISA. Box plots were used to visualize distribution of each serum protein level. P -values were obtained with two sample t-tests.

Techniques Used: Sandwich ELISA

35) Product Images from "A role for NF-?B-dependent gene transactivation in sunburn"

Article Title: A role for NF-?B-dependent gene transactivation in sunburn

Journal: Journal of Clinical Investigation

doi:

Impact of NF-κB decoy ODNs on UV-triggered cytokine production. ( a ) Pam 212 keratinocytes, NS47 fibroblasts, and XS106 Langerhans cells were transfected with the NF-κB reporter construct (3× κB luc) or the AP-1 reporter construct (4× AP-1 luc). After 6 hours of preincubation with 10 μM NF-κB decoy or scrambled ODNs, cells were washed and exposed to 50 J/m 2 UVB radiation (FS20 sunlamp) in PBS, cultured for an additional 24 hours in the continuous presence of fresh ODNs, and then examined for Luc activity. Data shown are representative of four independent experiments, showing the mean ± SD from triplicate samples. ( b and c ) Cells (1 × 10 6 cells/mL) were incubated for 6 hours with 10 μM NF-κB decoy or scrambled ODNs, washed, and then exposed to the indicated fluences of UV radiation using either FS20 sunlamps ( b ) or a Xenon arc solar simulator ( c ). Subsequently, cells were cultured for an additional 24 hours in complete RPMI 1640 in the presence of fresh ODNs, and culture supernatants were examined for the indicated cytokines by ELISA. Data shown are representative of three independent experiments, showing the mean ± SD from triplicate samples. A Statistically significant differences ( P
Figure Legend Snippet: Impact of NF-κB decoy ODNs on UV-triggered cytokine production. ( a ) Pam 212 keratinocytes, NS47 fibroblasts, and XS106 Langerhans cells were transfected with the NF-κB reporter construct (3× κB luc) or the AP-1 reporter construct (4× AP-1 luc). After 6 hours of preincubation with 10 μM NF-κB decoy or scrambled ODNs, cells were washed and exposed to 50 J/m 2 UVB radiation (FS20 sunlamp) in PBS, cultured for an additional 24 hours in the continuous presence of fresh ODNs, and then examined for Luc activity. Data shown are representative of four independent experiments, showing the mean ± SD from triplicate samples. ( b and c ) Cells (1 × 10 6 cells/mL) were incubated for 6 hours with 10 μM NF-κB decoy or scrambled ODNs, washed, and then exposed to the indicated fluences of UV radiation using either FS20 sunlamps ( b ) or a Xenon arc solar simulator ( c ). Subsequently, cells were cultured for an additional 24 hours in complete RPMI 1640 in the presence of fresh ODNs, and culture supernatants were examined for the indicated cytokines by ELISA. Data shown are representative of three independent experiments, showing the mean ± SD from triplicate samples. A Statistically significant differences ( P

Techniques Used: Transfection, Construct, Cell Culture, Activity Assay, Incubation, Enzyme-linked Immunosorbent Assay

36) Product Images from "ERBB signaling attenuates proinflammatory activation of nonclassical monocytes"

Article Title: ERBB signaling attenuates proinflammatory activation of nonclassical monocytes

Journal: American Journal of Physiology - Heart and Circulatory Physiology

doi: 10.1152/ajpheart.00486.2016

Phosphoinositide 3-kinase (PI3K) mediates the effect of GGF2 on LPS-induced TNF-α production in human monocytes. Analysis of PI3K activity and Akt phosphorylation (pAkt) was performed using human PB MNCs obtained from control subjects with documented expression of ERBB3 on monocytes. A : representative dot plots showing percentages of CD14 pos monocytes before ( left , pre-sort) and after ( middle , post-sort) enrichment of monocytes using negative selection (depletion of CD3-, CD19-, and CD56-expressing cells). Also shown is a dot plot showing percentages of CD14 hi CD16 neg and CD14 low CD16 pos after enrichment of monocytes ( right , post-sort). An enriched population of monocytes was used to determine PI3K activity and pAkt. B : phospholipids were isolated from monocytes incubated in the absence (basal) or presence of 30 ng/ml GGF2 for 10 min, and the level of PIP3 was determined using ELISA. Data represent means ± SE from 3 independent isolations of monocytes (unpaired t -test). C : effect of GGF2 on expression levels of the active form of Akt kinase (pAkt, Ser 437 ; top ) and total Akt ( bottom ) was determined using Western blot analysis. Purified peripheral blood monocytes were stimulated for 15 min; molecular weight, Precision Plus Protein Kaleidoscope standard (Bio-Rad Laboratories). D : representative flow cytometric histograms demonstrating the level of pAkt (open histograms) in CD14 hi ( left ) and CD14 low ( right ) HLA-DR + monocytes incubated in the absence (basal) or presence of 100 ng/ml insulin (positive control) or GGF2 (30 ng/ml); gray-shaded histograms, isotype-matched antibody. E and F : graphical representation of data from flow cytometry analysis of pAkt in CD14 hi HLA-DR + ( E ) and CD14 low HLA-DR + ( F ) monocytes. W, wortmannin; ns, not significant. Data represent means ± SE from 3 independent experiments performed in duplicate. P values are indicated (unpaired t -test). G : graphical representation of data from flow cytometry analysis of TNF-α production in CD14 low HLA-DR + monocytes incubated in the absence (basal) or presence of 100 nM wortmannin (Wtm) alone, 10 ng/ml LPS alone (LPS), and a combination of LPS and 30 ng/ml GGF2 and wortmannin (LPS+G+W). Data represent means ± SE from 3 independent experiments. The one-way ANOVA test was significant ( P ≤ 0.0001) for between-group differences; Bonferroni’s multiple-comparison posttest was used. P values between groups are shown.
Figure Legend Snippet: Phosphoinositide 3-kinase (PI3K) mediates the effect of GGF2 on LPS-induced TNF-α production in human monocytes. Analysis of PI3K activity and Akt phosphorylation (pAkt) was performed using human PB MNCs obtained from control subjects with documented expression of ERBB3 on monocytes. A : representative dot plots showing percentages of CD14 pos monocytes before ( left , pre-sort) and after ( middle , post-sort) enrichment of monocytes using negative selection (depletion of CD3-, CD19-, and CD56-expressing cells). Also shown is a dot plot showing percentages of CD14 hi CD16 neg and CD14 low CD16 pos after enrichment of monocytes ( right , post-sort). An enriched population of monocytes was used to determine PI3K activity and pAkt. B : phospholipids were isolated from monocytes incubated in the absence (basal) or presence of 30 ng/ml GGF2 for 10 min, and the level of PIP3 was determined using ELISA. Data represent means ± SE from 3 independent isolations of monocytes (unpaired t -test). C : effect of GGF2 on expression levels of the active form of Akt kinase (pAkt, Ser 437 ; top ) and total Akt ( bottom ) was determined using Western blot analysis. Purified peripheral blood monocytes were stimulated for 15 min; molecular weight, Precision Plus Protein Kaleidoscope standard (Bio-Rad Laboratories). D : representative flow cytometric histograms demonstrating the level of pAkt (open histograms) in CD14 hi ( left ) and CD14 low ( right ) HLA-DR + monocytes incubated in the absence (basal) or presence of 100 ng/ml insulin (positive control) or GGF2 (30 ng/ml); gray-shaded histograms, isotype-matched antibody. E and F : graphical representation of data from flow cytometry analysis of pAkt in CD14 hi HLA-DR + ( E ) and CD14 low HLA-DR + ( F ) monocytes. W, wortmannin; ns, not significant. Data represent means ± SE from 3 independent experiments performed in duplicate. P values are indicated (unpaired t -test). G : graphical representation of data from flow cytometry analysis of TNF-α production in CD14 low HLA-DR + monocytes incubated in the absence (basal) or presence of 100 nM wortmannin (Wtm) alone, 10 ng/ml LPS alone (LPS), and a combination of LPS and 30 ng/ml GGF2 and wortmannin (LPS+G+W). Data represent means ± SE from 3 independent experiments. The one-way ANOVA test was significant ( P ≤ 0.0001) for between-group differences; Bonferroni’s multiple-comparison posttest was used. P values between groups are shown.

Techniques Used: Activity Assay, Expressing, Selection, Isolation, Incubation, Enzyme-linked Immunosorbent Assay, Western Blot, Purification, Molecular Weight, Flow Cytometry, Positive Control, Cytometry

37) Product Images from "FTY720 inhibited proinflammatory cytokine release and osteoclastogenesis induced by Aggregatibacter actinomycetemcomitans"

Article Title: FTY720 inhibited proinflammatory cytokine release and osteoclastogenesis induced by Aggregatibacter actinomycetemcomitans

Journal: Lipids in Health and Disease

doi: 10.1186/s12944-015-0057-7

FTY720 dose-dependently inhibited IL-1β, IL-6, and TNF-α expressions induced by A. actinomycetemcomitans in BMMs. Murine BMMs were treated with vehicle (ethanol) or FTY720 (2 to 8 μM) for 30 min. Then the cells were either unstimulated or stimulated for 6 h with A. actinomycetemcomitans ( Aa ) (1.5 CFU/cell). a IL-1β, ( b ) IL-6, and ( c ) TNF-α protein levels in the cell culture media of BMMs were analyzed by ELISA. d Cell viability was tested in BMMs treated with vehicle or FTY720 (2 to 8 μM) for 8 h. Data are expressed as mean ± SEM ( n = 3, * p
Figure Legend Snippet: FTY720 dose-dependently inhibited IL-1β, IL-6, and TNF-α expressions induced by A. actinomycetemcomitans in BMMs. Murine BMMs were treated with vehicle (ethanol) or FTY720 (2 to 8 μM) for 30 min. Then the cells were either unstimulated or stimulated for 6 h with A. actinomycetemcomitans ( Aa ) (1.5 CFU/cell). a IL-1β, ( b ) IL-6, and ( c ) TNF-α protein levels in the cell culture media of BMMs were analyzed by ELISA. d Cell viability was tested in BMMs treated with vehicle or FTY720 (2 to 8 μM) for 8 h. Data are expressed as mean ± SEM ( n = 3, * p

Techniques Used: Cell Culture, Enzyme-linked Immunosorbent Assay

38) Product Images from "Structural Deformation of MTX Induced by Nanodrug Conjugation Dictate Intracellular Drug Transport and Drug Efficacy"

Article Title: Structural Deformation of MTX Induced by Nanodrug Conjugation Dictate Intracellular Drug Transport and Drug Efficacy

Journal: International Journal of Nanomedicine

doi: 10.2147/IJN.S317231

Anti-inflammatory efficacy. Low-dose therapeutic efficacy of covalent methotrexate (MTX)-carbon nanotube (CNT) compared to non-covalent MTX-CNT and free MTX in TNF-α-stimulated human fibroblast-like synovial cells (FLS) Suppression of ( A ) TNF-α, ( C ) IL-1-β, and ( E ) IL-6 mRNA levels in TNF-α-stimulated FLS. The data show that covalently conjugated MTX on nanotubes treatment led to a greater suppression of inflammatory cytokines than non-covalently conjugated MTX on nanotubes in FLSs. Suppression of protein levels was detected by ELISA in TNF-α-stimulated FLSs after treatment with nanodrugs for 24 h. ( B ) TNF-α, ( D ) IL-1-β, and ( F ) IL-6 protein levels were significantly suppressed by covalent MTX-CNTs compared to polyethylene glycolated (PEGylated) MTX-CNTs. All data are presented as mean ± SEM (n = 3). *** p
Figure Legend Snippet: Anti-inflammatory efficacy. Low-dose therapeutic efficacy of covalent methotrexate (MTX)-carbon nanotube (CNT) compared to non-covalent MTX-CNT and free MTX in TNF-α-stimulated human fibroblast-like synovial cells (FLS) Suppression of ( A ) TNF-α, ( C ) IL-1-β, and ( E ) IL-6 mRNA levels in TNF-α-stimulated FLS. The data show that covalently conjugated MTX on nanotubes treatment led to a greater suppression of inflammatory cytokines than non-covalently conjugated MTX on nanotubes in FLSs. Suppression of protein levels was detected by ELISA in TNF-α-stimulated FLSs after treatment with nanodrugs for 24 h. ( B ) TNF-α, ( D ) IL-1-β, and ( F ) IL-6 protein levels were significantly suppressed by covalent MTX-CNTs compared to polyethylene glycolated (PEGylated) MTX-CNTs. All data are presented as mean ± SEM (n = 3). *** p

Techniques Used: Enzyme-linked Immunosorbent Assay

39) Product Images from "Effects of Deacetylasperulosidic Acid on Atopic Dermatitis through Modulating Immune Balance and Skin Barrier Function in HaCaT, HMC-1, and EOL-1 Cells"

Article Title: Effects of Deacetylasperulosidic Acid on Atopic Dermatitis through Modulating Immune Balance and Skin Barrier Function in HaCaT, HMC-1, and EOL-1 Cells

Journal: Molecules

doi: 10.3390/molecules26113298

Effect of DAA and AA on the viability of EOL-1 cells and AD-related cytokines and chemokines secreted from EOL-1 cells. ( A ) Viability of EOL-1 cells treated with DAA and AA. ( B ) Inhibitory effects of DAA and AA on the levels of MCP-1 secreted from HDM-treated EOL-1 cells. ( C ) Inhibitory effects of DAA and AA on the levels of IL-5 secreted from HDM-treated EOL-1 cells. The levels of cytokines and chemokines in the supernatant of EOL-1 cells were determined by ELISA. The results are expressed as the mean ± SD (n = 4). ### p
Figure Legend Snippet: Effect of DAA and AA on the viability of EOL-1 cells and AD-related cytokines and chemokines secreted from EOL-1 cells. ( A ) Viability of EOL-1 cells treated with DAA and AA. ( B ) Inhibitory effects of DAA and AA on the levels of MCP-1 secreted from HDM-treated EOL-1 cells. ( C ) Inhibitory effects of DAA and AA on the levels of IL-5 secreted from HDM-treated EOL-1 cells. The levels of cytokines and chemokines in the supernatant of EOL-1 cells were determined by ELISA. The results are expressed as the mean ± SD (n = 4). ### p

Techniques Used: Enzyme-linked Immunosorbent Assay

Effect of DAA and AA on HMC-1 cell viability and the levels of AD-related histamine and IL-4 in HMC-1 cells. ( A ) Viability of HMC-1 cells treated with DAA and AA. ( B ) Inhibitory effects of DAA and AA on the levels of histamine secreted by PMACI-treated HMC-1 cells. ( C ) Inhibitory effects of DAA and AA on the levels of IL-4 secreted by PMACI-treated HMC-1 cells. The levels of cytokines and chemokines in the supernatant of HMC-1 cells were determined by ELISA. The results are expressed as the mean ± SD (n = 4). ### p
Figure Legend Snippet: Effect of DAA and AA on HMC-1 cell viability and the levels of AD-related histamine and IL-4 in HMC-1 cells. ( A ) Viability of HMC-1 cells treated with DAA and AA. ( B ) Inhibitory effects of DAA and AA on the levels of histamine secreted by PMACI-treated HMC-1 cells. ( C ) Inhibitory effects of DAA and AA on the levels of IL-4 secreted by PMACI-treated HMC-1 cells. The levels of cytokines and chemokines in the supernatant of HMC-1 cells were determined by ELISA. The results are expressed as the mean ± SD (n = 4). ### p

Techniques Used: Enzyme-linked Immunosorbent Assay

Effect of DAA on AD-related cytokines and chemokines secreted by HMC-1cells. The level of IL-1β ( A ), IL-6 ( B ), IL-8 ( C ), TNF-α ( D ), TSLP ( E ), and MCP-1 ( F ) in the supernatant of HMC-1 cells was determined by ELISA. The results are expressed as the mean ± SD (n = 4). ### p
Figure Legend Snippet: Effect of DAA on AD-related cytokines and chemokines secreted by HMC-1cells. The level of IL-1β ( A ), IL-6 ( B ), IL-8 ( C ), TNF-α ( D ), TSLP ( E ), and MCP-1 ( F ) in the supernatant of HMC-1 cells was determined by ELISA. The results are expressed as the mean ± SD (n = 4). ### p

Techniques Used: Enzyme-linked Immunosorbent Assay

Effect of DAA and AA on HaCaT cell viability and the levels of AD-related cytokines and chemokines in HaCaT cells. ( A ) The viability of HaCaT cells treated with DAA and AA. ( B ) DAA and AA inhibited the secretion of TSLP by HaCaT cells treated with both TNF-α and IFN-γ. ( C ) DAA and AA inhibited the secretion of IL-33 by HaCaT cells treated with both TNF-α and IFN-γ. The production of cytokines and chemokines in the supernatant of HaCaT cells was analyzed by ELISA. The results are expressed as the mean ± SD (n = 4). ### p
Figure Legend Snippet: Effect of DAA and AA on HaCaT cell viability and the levels of AD-related cytokines and chemokines in HaCaT cells. ( A ) The viability of HaCaT cells treated with DAA and AA. ( B ) DAA and AA inhibited the secretion of TSLP by HaCaT cells treated with both TNF-α and IFN-γ. ( C ) DAA and AA inhibited the secretion of IL-33 by HaCaT cells treated with both TNF-α and IFN-γ. The production of cytokines and chemokines in the supernatant of HaCaT cells was analyzed by ELISA. The results are expressed as the mean ± SD (n = 4). ### p

Techniques Used: Enzyme-linked Immunosorbent Assay

Effect of DAA on AD-related cytokines and chemokines secreted by HaCaT cells. The levels of IL-1β ( A ), IL-6 ( B ), IL-8 ( C ), TNF-α ( D ), and MCP-1 ( E ) in the supernatant of HaCaT cells were evaluated by ELISA. The results are expressed as the mean ± SD (n = 4). ### p
Figure Legend Snippet: Effect of DAA on AD-related cytokines and chemokines secreted by HaCaT cells. The levels of IL-1β ( A ), IL-6 ( B ), IL-8 ( C ), TNF-α ( D ), and MCP-1 ( E ) in the supernatant of HaCaT cells were evaluated by ELISA. The results are expressed as the mean ± SD (n = 4). ### p

Techniques Used: Enzyme-linked Immunosorbent Assay

Effect of DAA on AD-related cytokine and chemokine secretion in EOL-1cells. The levels of IL-6 ( A ), IL-8 ( B ), and TNF-α ( C ) in the supernatant of EOL-1 cells were analyzed by ELISA. The results are expressed as the mean ± SD (n = 4). ### p
Figure Legend Snippet: Effect of DAA on AD-related cytokine and chemokine secretion in EOL-1cells. The levels of IL-6 ( A ), IL-8 ( B ), and TNF-α ( C ) in the supernatant of EOL-1 cells were analyzed by ELISA. The results are expressed as the mean ± SD (n = 4). ### p

Techniques Used: Enzyme-linked Immunosorbent Assay

Effect of DAA on the levels of AD-related cytokines and chemokines known to activate Th2 cells secreted by HaCaT cells. The level of cytokine IL-25 ( A ) and chemokines TARC ( B ), RANTES ( C ), and MDC ( D ) in the supernatant of HaCaT cells was evaluated by ELISA. The results are expressed as the mean ± SD (n = 4). ### p
Figure Legend Snippet: Effect of DAA on the levels of AD-related cytokines and chemokines known to activate Th2 cells secreted by HaCaT cells. The level of cytokine IL-25 ( A ) and chemokines TARC ( B ), RANTES ( C ), and MDC ( D ) in the supernatant of HaCaT cells was evaluated by ELISA. The results are expressed as the mean ± SD (n = 4). ### p

Techniques Used: Enzyme-linked Immunosorbent Assay

40) Product Images from "Elamipretide (SS-31) Improves Functional Connectivity in Hippocampus and Other Related Regions Following Prolonged Neuroinflammation Induced by Lipopolysaccharide in Aged Rats"

Article Title: Elamipretide (SS-31) Improves Functional Connectivity in Hippocampus and Other Related Regions Following Prolonged Neuroinflammation Induced by Lipopolysaccharide in Aged Rats

Journal: Frontiers in Aging Neuroscience

doi: 10.3389/fnagi.2021.600484

Effects of lipopolysaccharide (LPS) exposure and elamipretide (SS-31) treatment. ELISA showed differences in (A,D) the systemic and (B,E) the hippocampus IL-1β and TNFα expression. RT-PCR showed differences in mRNA expression for (C) IL-1β and (F) TNFα Data are presented as mean ± SEMs ( n = 5). Comparisons were made by two-way ANOVA followed by the Bonferroni post-hoc analysis. # p
Figure Legend Snippet: Effects of lipopolysaccharide (LPS) exposure and elamipretide (SS-31) treatment. ELISA showed differences in (A,D) the systemic and (B,E) the hippocampus IL-1β and TNFα expression. RT-PCR showed differences in mRNA expression for (C) IL-1β and (F) TNFα Data are presented as mean ± SEMs ( n = 5). Comparisons were made by two-way ANOVA followed by the Bonferroni post-hoc analysis. # p

Techniques Used: Enzyme-linked Immunosorbent Assay, Expressing, Reverse Transcription Polymerase Chain Reaction

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    R&D Systems murine opg elisa kit
    Vasoactive intestinal peptide (VIP) modulates the pattern of expression of the <t>RANK/RANKL/OPG</t> system in joints from mice with collagen-induced arthritis (CIA). (a) Expression of mRNA for receptor activator of nuclear factor-κB (RANK), receptor activator of nuclear factor-κB ligand (RANKL) or osteoprotegerin (OPG) in the hind paws was measured by quantitative real time PCR and corrected by mRNA expression for β-actin in each sample (see Materials and methods). (b) Serum levels of OPG in control, CIA or VIP-treated CIA mice were determined by <t>ELISA.</t> On day 10 of VIP treatment, differences between the arthritic group and the CIA group treated with VIP were statistically significant (**p
    Murine Opg Elisa Kit, supplied by R&D Systems, used in various techniques. Bioz Stars score: 80/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Infections with C. difficile in sDMDMm2 mice resemble infections in antibiotic-treated mice . Cohorts of germ-free (black circles, n = 6), sDMDMm2 (red diamonds, n = 5), clindamycin-treated SPF (purple inverted triangles, n = 5), streptomycin-treated SPF (green triangles, n = 5), and SPF control (gray squares, n = 5) mice were gavaged with 10 3 CFU C. difficile DH1916 (filled symbols) or PBS vehicle as control (open symbols). (A) Comparison of CFU of C. difficile in fecal pellets (FP) or cecal contents (CC) at different time points over course of infection. (B) Comparison of <t>lipocalin-2</t> measured by <t>ELISA</t> in cecal content after 72 h of infection. (C) Comparison of calprotectin from cecal tissue after 72 h of infection measured by qPCR. Values show the ΔC t derived from measured values of calprotectin with β-actin as control. (D) Histopathological evaluation of H E stained sections of cecum tissue at time of necropsy (72 h). Arrows indicate the representative mice depicted in (E) . (E) Representative images of H E stained sections of cecum tissue at time of necropsy (72 h). L, Lumen; Scale bars: 200 μm. Statistical analyses in (B–D) used Mann-Whitney-U tests to compare uninfected with infected animals at individual timepoints. Each symbol represents one individual. Bars indicate medians. Dotted lines indicate lower limit of detection. ns, not statistically significant ( p ≥ 0.05); * p
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    C3a treatment results in increased MMP-2 activity by RPE cells. ( a ) mRNA expression of MMP-2 and <t>TIMP-3</t> measured by qRT-PCR and normalized to GAPDH in cultures of hfRPE cells treated with different doses of C3a for two weeks. ( b ) Levels of TIMP-3 measured in conditioned media of the same cultures by <t>ELISA.</t> ( c ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with different doses of C3a for 2 weeks. ( d ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with or without C3aR antagonist for 2 weeks (ANOVA, n = 9/treatment. Data presented as mean ± SD. *p
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    Plasma <t>TPO</t> levels and platelet Mpl expression. (A) TPO levels determined by <t>ELISA</t> on peripheral blood plasma from WT, mpl −/− , and Tg ( mpl ) mice; n = 3 to 4 mice for each strain. * P ≤ .003, Tg ( mpl ) versus WT and Tg ( mpl ) versus mpl −/− . Means plus or minus SE are shown. (B) Expression of Mpl on platelet lysates analyzed by Western blot probed with Mpl antiserum (top panel). The blot was stripped and reprobed with anti–pan-actin antibody to ensure equal amount of protein in each lane (bottom panel). Levels of Mpl in Tg ( mpl ) platelets are 9-fold lower than that in WT.
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    Vasoactive intestinal peptide (VIP) modulates the pattern of expression of the RANK/RANKL/OPG system in joints from mice with collagen-induced arthritis (CIA). (a) Expression of mRNA for receptor activator of nuclear factor-κB (RANK), receptor activator of nuclear factor-κB ligand (RANKL) or osteoprotegerin (OPG) in the hind paws was measured by quantitative real time PCR and corrected by mRNA expression for β-actin in each sample (see Materials and methods). (b) Serum levels of OPG in control, CIA or VIP-treated CIA mice were determined by ELISA. On day 10 of VIP treatment, differences between the arthritic group and the CIA group treated with VIP were statistically significant (**p

    Journal: Arthritis Research & Therapy

    Article Title: Protective effect of vasoactive intestinal peptide on bone destruction in the collagen-induced arthritis model of rheumatoid arthritis

    doi: 10.1186/ar1779

    Figure Lengend Snippet: Vasoactive intestinal peptide (VIP) modulates the pattern of expression of the RANK/RANKL/OPG system in joints from mice with collagen-induced arthritis (CIA). (a) Expression of mRNA for receptor activator of nuclear factor-κB (RANK), receptor activator of nuclear factor-κB ligand (RANKL) or osteoprotegerin (OPG) in the hind paws was measured by quantitative real time PCR and corrected by mRNA expression for β-actin in each sample (see Materials and methods). (b) Serum levels of OPG in control, CIA or VIP-treated CIA mice were determined by ELISA. On day 10 of VIP treatment, differences between the arthritic group and the CIA group treated with VIP were statistically significant (**p

    Article Snippet: Determination of osteoprotegerin in serum Mouse OPG in serum was assayed using a commercial murine OPG ELISA kit (mouse OPG/TNFSRSF11B immunoassay, R & D Systems).

    Techniques: Expressing, Mouse Assay, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    Infections with C. difficile in sDMDMm2 mice resemble infections in antibiotic-treated mice . Cohorts of germ-free (black circles, n = 6), sDMDMm2 (red diamonds, n = 5), clindamycin-treated SPF (purple inverted triangles, n = 5), streptomycin-treated SPF (green triangles, n = 5), and SPF control (gray squares, n = 5) mice were gavaged with 10 3 CFU C. difficile DH1916 (filled symbols) or PBS vehicle as control (open symbols). (A) Comparison of CFU of C. difficile in fecal pellets (FP) or cecal contents (CC) at different time points over course of infection. (B) Comparison of lipocalin-2 measured by ELISA in cecal content after 72 h of infection. (C) Comparison of calprotectin from cecal tissue after 72 h of infection measured by qPCR. Values show the ΔC t derived from measured values of calprotectin with β-actin as control. (D) Histopathological evaluation of H E stained sections of cecum tissue at time of necropsy (72 h). Arrows indicate the representative mice depicted in (E) . (E) Representative images of H E stained sections of cecum tissue at time of necropsy (72 h). L, Lumen; Scale bars: 200 μm. Statistical analyses in (B–D) used Mann-Whitney-U tests to compare uninfected with infected animals at individual timepoints. Each symbol represents one individual. Bars indicate medians. Dotted lines indicate lower limit of detection. ns, not statistically significant ( p ≥ 0.05); * p

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Functional Intestinal Bile Acid 7α-Dehydroxylation by Clostridium scindens Associated with Protection from Clostridium difficile Infection in a Gnotobiotic Mouse Model

    doi: 10.3389/fcimb.2016.00191

    Figure Lengend Snippet: Infections with C. difficile in sDMDMm2 mice resemble infections in antibiotic-treated mice . Cohorts of germ-free (black circles, n = 6), sDMDMm2 (red diamonds, n = 5), clindamycin-treated SPF (purple inverted triangles, n = 5), streptomycin-treated SPF (green triangles, n = 5), and SPF control (gray squares, n = 5) mice were gavaged with 10 3 CFU C. difficile DH1916 (filled symbols) or PBS vehicle as control (open symbols). (A) Comparison of CFU of C. difficile in fecal pellets (FP) or cecal contents (CC) at different time points over course of infection. (B) Comparison of lipocalin-2 measured by ELISA in cecal content after 72 h of infection. (C) Comparison of calprotectin from cecal tissue after 72 h of infection measured by qPCR. Values show the ΔC t derived from measured values of calprotectin with β-actin as control. (D) Histopathological evaluation of H E stained sections of cecum tissue at time of necropsy (72 h). Arrows indicate the representative mice depicted in (E) . (E) Representative images of H E stained sections of cecum tissue at time of necropsy (72 h). L, Lumen; Scale bars: 200 μm. Statistical analyses in (B–D) used Mann-Whitney-U tests to compare uninfected with infected animals at individual timepoints. Each symbol represents one individual. Bars indicate medians. Dotted lines indicate lower limit of detection. ns, not statistically significant ( p ≥ 0.05); * p

    Article Snippet: Lipocalin-2 quantification Lipocalin-2 was measured from terminally collected cecal content using a commercial ELISA kit (R & D Systems, USA).

    Techniques: Mouse Assay, Infection, Enzyme-linked Immunosorbent Assay, Real-time Polymerase Chain Reaction, Derivative Assay, Staining, MANN-WHITNEY

    Addition of C. scindens to sDMDMm2 can partially protect from C. difficile infection . sDMDMm2 animals were pre-colonized by gavage with 10 9 CFU C. scindens 96 h before infection with 10 3 CFU C. difficile DH1916 (blue circles). sDMDMm2 animals (red diamonds) served as control. Data is combined from two independent experiments with endpoints at 24 h ( n = 5 sDMDMm2 and n = 5 sDMDM + C. scindens , only data from endpoint shown) and 72 h ( n = 6 sDMDMm2 and n = 5 sDMDM + C. scindens ). (A) CFU of C. scindens over the course of the infection in pre-colonized mice. (B) Comparison of C. difficile CFU between sDMDMm2 and sDMDMm2 + C. scindens over the course of infection. Uninfected controls are not depicted. (C) Lipocalin-2 concentration measured by ELISA in cecal content at 24 and 72 h post infection, respectively. (D) Mucosal calprotectin expression in cecal tissue measured by qPCR at 24 and 72 h post infection, respectively. Values show the ΔC t derived from measured values of calprotectin with β-actin as control. (E) Histopathological evaluation of H E stained sections of cecum tissue at time of necropsy (24 or 72 h). Uninfected controls are not depicted. Arrows indicate the representative mice depicted in (F) . (F) H E stained histological sections of cecum sampled at 24 and 72 h post infection. FP, Fecal pellet; CC, Cecal content; L, Lumen; Scale bars: 200 μm. Statistical analyses in (B–E) used Mann-Whitney-U tests; Each symbol represents one individual. Bars indicate medians. Dotted lines indicate lower limit of detection. ns, not statistically significant ( p ≥ 0.05); * p

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Functional Intestinal Bile Acid 7α-Dehydroxylation by Clostridium scindens Associated with Protection from Clostridium difficile Infection in a Gnotobiotic Mouse Model

    doi: 10.3389/fcimb.2016.00191

    Figure Lengend Snippet: Addition of C. scindens to sDMDMm2 can partially protect from C. difficile infection . sDMDMm2 animals were pre-colonized by gavage with 10 9 CFU C. scindens 96 h before infection with 10 3 CFU C. difficile DH1916 (blue circles). sDMDMm2 animals (red diamonds) served as control. Data is combined from two independent experiments with endpoints at 24 h ( n = 5 sDMDMm2 and n = 5 sDMDM + C. scindens , only data from endpoint shown) and 72 h ( n = 6 sDMDMm2 and n = 5 sDMDM + C. scindens ). (A) CFU of C. scindens over the course of the infection in pre-colonized mice. (B) Comparison of C. difficile CFU between sDMDMm2 and sDMDMm2 + C. scindens over the course of infection. Uninfected controls are not depicted. (C) Lipocalin-2 concentration measured by ELISA in cecal content at 24 and 72 h post infection, respectively. (D) Mucosal calprotectin expression in cecal tissue measured by qPCR at 24 and 72 h post infection, respectively. Values show the ΔC t derived from measured values of calprotectin with β-actin as control. (E) Histopathological evaluation of H E stained sections of cecum tissue at time of necropsy (24 or 72 h). Uninfected controls are not depicted. Arrows indicate the representative mice depicted in (F) . (F) H E stained histological sections of cecum sampled at 24 and 72 h post infection. FP, Fecal pellet; CC, Cecal content; L, Lumen; Scale bars: 200 μm. Statistical analyses in (B–E) used Mann-Whitney-U tests; Each symbol represents one individual. Bars indicate medians. Dotted lines indicate lower limit of detection. ns, not statistically significant ( p ≥ 0.05); * p

    Article Snippet: Lipocalin-2 quantification Lipocalin-2 was measured from terminally collected cecal content using a commercial ELISA kit (R & D Systems, USA).

    Techniques: Infection, Mouse Assay, Concentration Assay, Enzyme-linked Immunosorbent Assay, Expressing, Real-time Polymerase Chain Reaction, Derivative Assay, Staining, MANN-WHITNEY

    C3a treatment results in increased MMP-2 activity by RPE cells. ( a ) mRNA expression of MMP-2 and TIMP-3 measured by qRT-PCR and normalized to GAPDH in cultures of hfRPE cells treated with different doses of C3a for two weeks. ( b ) Levels of TIMP-3 measured in conditioned media of the same cultures by ELISA. ( c ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with different doses of C3a for 2 weeks. ( d ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with or without C3aR antagonist for 2 weeks (ANOVA, n = 9/treatment. Data presented as mean ± SD. *p

    Journal: Scientific Reports

    Article Title: C3a triggers formation of sub-retinal pigment epithelium deposits via the ubiquitin proteasome pathway

    doi: 10.1038/s41598-018-28143-0

    Figure Lengend Snippet: C3a treatment results in increased MMP-2 activity by RPE cells. ( a ) mRNA expression of MMP-2 and TIMP-3 measured by qRT-PCR and normalized to GAPDH in cultures of hfRPE cells treated with different doses of C3a for two weeks. ( b ) Levels of TIMP-3 measured in conditioned media of the same cultures by ELISA. ( c ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with different doses of C3a for 2 weeks. ( d ) MMP-2 activity measured in apical and basal conditioned media of hfRPE cells treated with or without C3aR antagonist for 2 weeks (ANOVA, n = 9/treatment. Data presented as mean ± SD. *p

    Article Snippet: The fraction over 3 kDa was used to quantify TIMP-3 and IL-6 using ELISA kits from R & D Systems (Minneapolis, MN) following manufacturer’s instructions.

    Techniques: Activity Assay, Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

    Inhibition of the UPP results in abnormal ECM turnover. ( a ) mRNA expression of MMP-2 and TIMP-3 measured by qRT-PCR and normalized to GAPDH, ( b ) MMP-2 activity measured by zymography, and ( c ) TIMP-3 levels measured by ELISA in conditioned media of hfRPE cells treated with proteasome inhibitor (+Prot inh) for 2 weeks and controls (−Prot inh) (t-test, n = 5. Data presented as mean ± SD. ***p

    Journal: Scientific Reports

    Article Title: C3a triggers formation of sub-retinal pigment epithelium deposits via the ubiquitin proteasome pathway

    doi: 10.1038/s41598-018-28143-0

    Figure Lengend Snippet: Inhibition of the UPP results in abnormal ECM turnover. ( a ) mRNA expression of MMP-2 and TIMP-3 measured by qRT-PCR and normalized to GAPDH, ( b ) MMP-2 activity measured by zymography, and ( c ) TIMP-3 levels measured by ELISA in conditioned media of hfRPE cells treated with proteasome inhibitor (+Prot inh) for 2 weeks and controls (−Prot inh) (t-test, n = 5. Data presented as mean ± SD. ***p

    Article Snippet: The fraction over 3 kDa was used to quantify TIMP-3 and IL-6 using ELISA kits from R & D Systems (Minneapolis, MN) following manufacturer’s instructions.

    Techniques: Inhibition, Expressing, Quantitative RT-PCR, Activity Assay, Zymography, Enzyme-linked Immunosorbent Assay, T-Test

    Plasma TPO levels and platelet Mpl expression. (A) TPO levels determined by ELISA on peripheral blood plasma from WT, mpl −/− , and Tg ( mpl ) mice; n = 3 to 4 mice for each strain. * P ≤ .003, Tg ( mpl ) versus WT and Tg ( mpl ) versus mpl −/− . Means plus or minus SE are shown. (B) Expression of Mpl on platelet lysates analyzed by Western blot probed with Mpl antiserum (top panel). The blot was stripped and reprobed with anti–pan-actin antibody to ensure equal amount of protein in each lane (bottom panel). Levels of Mpl in Tg ( mpl ) platelets are 9-fold lower than that in WT.

    Journal: Blood

    Article Title: Incomplete restoration of Mpl expression in the mpl−/− mouse produces partial correction of the stem cell–repopulating defect and paradoxical thrombocytosis

    doi: 10.1182/blood-2007-11-124859

    Figure Lengend Snippet: Plasma TPO levels and platelet Mpl expression. (A) TPO levels determined by ELISA on peripheral blood plasma from WT, mpl −/− , and Tg ( mpl ) mice; n = 3 to 4 mice for each strain. * P ≤ .003, Tg ( mpl ) versus WT and Tg ( mpl ) versus mpl −/− . Means plus or minus SE are shown. (B) Expression of Mpl on platelet lysates analyzed by Western blot probed with Mpl antiserum (top panel). The blot was stripped and reprobed with anti–pan-actin antibody to ensure equal amount of protein in each lane (bottom panel). Levels of Mpl in Tg ( mpl ) platelets are 9-fold lower than that in WT.

    Article Snippet: TPO assays were performed on 1:5 diluted platelet-poor plasma from inferior cava–drawn blood using a TPO ELISA kit (R & D Systems, Minneapolis, MN) per the manufacturer's instructions.

    Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Mouse Assay, Western Blot