non parenchymal cells npcs (Worthington Biochemical)

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Collagenase Type 2

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Prepared to contain higher clostripain activity Suggested for bone heart liver thyroid and salivary primary cell isolation Supplied as a dialyzed lyophilized powder

Catalog Number:

ls004174

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100 mg

Source:

Clostridium histolyticum

Cas Number:

9001.12.1

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Worthington Biochemical non parenchymal cells npcs
Microscope images of <t>HCs</t> and <t>NPCs</t> in culture. These pictures depict representative areas of untreated, PB in vitro as well as in vivo treated HCs and NPCs in culture extracted from microscopic images of equal magnification.

Microscope images of <t>HCs</t> and <t>NPCs</t> in culture. These pictures depict representative areas of untreated, PB in vitro as well as in vivo treated HCs and NPCs in culture extracted from microscopic images of equal magnification.

Prepared to contain higher clostripain activity Suggested for bone heart liver thyroid and salivary primary cell isolation Supplied as a dialyzed lyophilized powder
https://www.bioz.com/result/non parenchymal cells npcs/product/Worthington Biochemical
Average 85 stars, based on 3365 article reviews
Price from $9.99 to $1999.99

non parenchymal cells npcs - by Bioz Stars, 2021-01

85/100 stars

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Images

1) Product Images from "Phenobarbital Induces Alterations in the Proteome of Hepatocytes and Mesenchymal Cells of Rat Livers"

Article Title: Phenobarbital Induces Alterations in the Proteome of Hepatocytes and Mesenchymal Cells of Rat Livers

Journal: PLoS ONE

doi: 10.1371/journal.pone.0076137

Microscope images of HCs and NPCs in culture. These pictures depict representative areas of untreated, PB in vitro as well as in vivo treated HCs and NPCs in culture extracted from microscopic images of equal magnification.

Figure Legend Snippet: Microscope images of HCs and NPCs in culture. These pictures depict representative areas of untreated, PB in vitro as well as in vivo treated HCs and NPCs in culture extracted from microscopic images of equal magnification.

Techniques Used: Microscopy, In Vitro, In Vivo

$Proteome alterations induced by in vitro treatment of primary cells. Part A) shows schematic representations of a cell and her three sub-compartments, namely the supernatant, the cytoplasm and the nucleus. The intensity of red represents the degree of amount of the selected protein found in the respective compartment in contrast to the other experiments. The higher intensity of red corresponds to a higher occurrence. This allows an easy comparison of the expression levels of a protein in different experimental setups. NPCs induce the secretion of IL-1beta and TNF-alpha upon inflammatory stimulation with LPS. In vitro treatment with PB induced coronin-7 and ADP-ribosyl cyclase 1, which both are also induced by in vivo treatment. The expression of Hsp90, a stress response related protein, was increased upon LPS and PB treatment. Prostaglandin, a protein involved in promotion of proliferation in normal and preneoplastic cells, was induced upon LPS and in vivo PB treatment. HCs respond hardly to the in vitro treatment with PB. Treatment with IL-6 specifically induced the acute phase protein T-kininogen-2. UDP-glucuronosyltransferase 2B37 and the chaperone peptidyl-prolyl cis-trans isomerase D were induced by both in vitro stimulation experiments as well as by the in vivo treatment with PB. Carbamoyl-phosphate synthase is part of the urea cycle and has to be found in all four categories. Proteins in NPC: (1) O35828 Coronin-7, (2) P16599 Tumor necrosis factor, (3) P34058 Heat shock protein HSP 90-beta, (4) Q63264 Interleukin-1 beta, (5) Q63921 Prostaglandin G/H synthase 1, (6) Q64244 ADP-ribosyl cyclase 1. Proteins in HC: (1) P07756 Carbamoyl-phosphate synthase [ammonia], (2) P08932 T-kininogen 2, (3) P19488 UDP-glucuronosyltransferase 2B37, (4) Q6DGG0 Peptidyl-prolyl cis-trans isomerase D. Part B) demonstrates the distribution of distinct proteins within the three fractions, supernatant, cytoplasm and nuclear protein fractions, underneath the respective treatment of the cells, which gives an overview of the responsiveness of the cells. Abbr.: SN –proteome of the supernatant, Cyt – proteome of the cytoplasm, NE – proteome of the nuclear extract.$
Figure Legend Snippet: Proteome alterations induced by in vitro treatment of primary cells. Part A) shows schematic representations of a cell and her three sub-compartments, namely the supernatant, the cytoplasm and the nucleus. The intensity of red represents the degree of amount of the selected protein found in the respective compartment in contrast to the other experiments. The higher intensity of red corresponds to a higher occurrence. This allows an easy comparison of the expression levels of a protein in different experimental setups. NPCs induce the secretion of IL-1beta and TNF-alpha upon inflammatory stimulation with LPS. In vitro treatment with PB induced coronin-7 and ADP-ribosyl cyclase 1, which both are also induced by in vivo treatment. The expression of Hsp90, a stress response related protein, was increased upon LPS and PB treatment. Prostaglandin, a protein involved in promotion of proliferation in normal and preneoplastic cells, was induced upon LPS and in vivo PB treatment. HCs respond hardly to the in vitro treatment with PB. Treatment with IL-6 specifically induced the acute phase protein T-kininogen-2. UDP-glucuronosyltransferase 2B37 and the chaperone peptidyl-prolyl cis-trans isomerase D were induced by both in vitro stimulation experiments as well as by the in vivo treatment with PB. Carbamoyl-phosphate synthase is part of the urea cycle and has to be found in all four categories. Proteins in NPC: (1) O35828 Coronin-7, (2) P16599 Tumor necrosis factor, (3) P34058 Heat shock protein HSP 90-beta, (4) Q63264 Interleukin-1 beta, (5) Q63921 Prostaglandin G/H synthase 1, (6) Q64244 ADP-ribosyl cyclase 1. Proteins in HC: (1) P07756 Carbamoyl-phosphate synthase [ammonia], (2) P08932 T-kininogen 2, (3) P19488 UDP-glucuronosyltransferase 2B37, (4) Q6DGG0 Peptidyl-prolyl cis-trans isomerase D. Part B) demonstrates the distribution of distinct proteins within the three fractions, supernatant, cytoplasm and nuclear protein fractions, underneath the respective treatment of the cells, which gives an overview of the responsiveness of the cells. Abbr.: SN –proteome of the supernatant, Cyt – proteome of the cytoplasm, NE – proteome of the nuclear extract.

Techniques Used: In Vitro, Expressing, In Vivo

Distribution of distinct proteins, when comparing controls with PB-treatment from the in vitro and in vivo sample pools, respectively. This figure demonstrates the distribution of distinct proteins found in HCs and NPCs during the pooled A) in vitro and B) in vivo experiments, while including only proteins found with at least 2 peptides. The up- and down-regulation of proteins were neglected in this qualitative comparison.

Figure Legend Snippet: Distribution of distinct proteins, when comparing controls with PB-treatment from the in vitro and in vivo sample pools, respectively. This figure demonstrates the distribution of distinct proteins found in HCs and NPCs during the pooled A) in vitro and B) in vivo experiments, while including only proteins found with at least 2 peptides. The up- and down-regulation of proteins were neglected in this qualitative comparison.

Techniques Used: In Vitro, In Vivo

2) Product Images from "Fibroblast activation protein restrains adipogenic differentiation and regulates matrix-mediated mTOR signaling"

Article Title: Fibroblast activation protein restrains adipogenic differentiation and regulates matrix-mediated mTOR signaling

Journal: Matrix biology : journal of the International Society for Matrix Biology

doi: 10.1016/j.matbio.2019.07.007

FAP −/− mice display enhanced diet-induced weight gain with minimal changes to systemic metabolism. A) Timeline for diet-induced obesity model. B) Total body mass at time of euthanasia, 30 weeks of age (N=23-27 mice/group). C) Mass of total subcutaneous fat at 30 weeks of age (N=14-17 mice/group). D) Mass of total abdominal fat at 30 weeks of age (N=3-5 mice/group) E) Blood glucose levels following bolus insulin injection (1U/kg body mass; N=4-6 mice/group). F) Blood glucose levels following bolus glucose injection (p

Figure Legend Snippet: FAP −/− mice display enhanced diet-induced weight gain with minimal changes to systemic metabolism. A) Timeline for diet-induced obesity model. B) Total body mass at time of euthanasia, 30 weeks of age (N=23-27 mice/group). C) Mass of total subcutaneous fat at 30 weeks of age (N=14-17 mice/group). D) Mass of total abdominal fat at 30 weeks of age (N=3-5 mice/group) E) Blood glucose levels following bolus insulin injection (1U/kg body mass; N=4-6 mice/group). F) Blood glucose levels following bolus glucose injection (p

Techniques Used: Mouse Assay, Injection

FAP deletion promotes accumulation of collagen in non-fibrillar forms. A) Total collagen measured by aniline blue in subcutaneous fat (N=3-7 mice/group, 3 images/mouse). Scale bars=200 μm; statistical analysis by two-way ANOVA. B) Fibrillar collagen measured by SFIG in subcutaneous fat (N=3-7 mice/group, 5 images/mouse). Scale bars=200 μm; statistical analysis by two-way ANOVA. C) Fibrillar collagen imaged by picrosirius red stain under circular polarized light, where thin fibers appear green, intermediate fibers red, and thick fibers yellow (N=3-7 mice/group, 5 images/mouse). Scale bars=200 μm; statistical analysis by chi-square test.

Figure Legend Snippet: FAP deletion promotes accumulation of collagen in non-fibrillar forms. A) Total collagen measured by aniline blue in subcutaneous fat (N=3-7 mice/group, 3 images/mouse). Scale bars=200 μm; statistical analysis by two-way ANOVA. B) Fibrillar collagen measured by SFIG in subcutaneous fat (N=3-7 mice/group, 5 images/mouse). Scale bars=200 μm; statistical analysis by two-way ANOVA. C) Fibrillar collagen imaged by picrosirius red stain under circular polarized light, where thin fibers appear green, intermediate fibers red, and thick fibers yellow (N=3-7 mice/group, 5 images/mouse). Scale bars=200 μm; statistical analysis by chi-square test.

Techniques Used: Mouse Assay, Staining

3) Product Images from "Fli1-haploinsufficient dermal fibroblasts promote skin-localized transdifferentiation of Th2-like regulatory T cells"

Article Title: Fli1-haploinsufficient dermal fibroblasts promote skin-localized transdifferentiation of Th2-like regulatory T cells

Journal: Arthritis Research & Therapy

doi: 10.1186/s13075-018-1521-3

T helper type 2 cell (Th2)-like regulatory T cells (Tregs) are induced by coculture with Fli1 +/− dermal fibroblasts through interleukin (IL)-33. a Evaluation by flow cytometry of Th1-, Th2-, and Th17-like Treg induction by coculture with wild-type (WT) and Fli1 +/− dermal fibroblasts ( n = 6). b Evaluation by flow cytometry of the effect of IL-33-neutralizing antibody on Tregs cocultured with Fli1 +/− dermal fibroblasts ( n = 6). Representative plots of interferon (IFN)-γ-, IL-4-, and IL-17A-positive Tregs are shown in right upper panels of ( a ) and ( b ). Gating strategy for identification of CD4 + FoxP3 + Tregs is shown in the leftmost panels of ( a ) and ( b ). In each graph, the relative value compared with the control group is expressed as mean ± SEM. AU Arbitrary units, Fli1 Friend leukemia virus integration 1

Figure Legend Snippet: T helper type 2 cell (Th2)-like regulatory T cells (Tregs) are induced by coculture with Fli1 +/− dermal fibroblasts through interleukin (IL)-33. a Evaluation by flow cytometry of Th1-, Th2-, and Th17-like Treg induction by coculture with wild-type (WT) and Fli1 +/− dermal fibroblasts ( n = 6). b Evaluation by flow cytometry of the effect of IL-33-neutralizing antibody on Tregs cocultured with Fli1 +/− dermal fibroblasts ( n = 6). Representative plots of interferon (IFN)-γ-, IL-4-, and IL-17A-positive Tregs are shown in right upper panels of ( a ) and ( b ). Gating strategy for identification of CD4 + FoxP3 + Tregs is shown in the leftmost panels of ( a ) and ( b ). In each graph, the relative value compared with the control group is expressed as mean ± SEM. AU Arbitrary units, Fli1 Friend leukemia virus integration 1

Techniques Used: Flow Cytometry, Cytometry

4) Product Images from "Fli1-haploinsufficient dermal fibroblasts promote skin-localized transdifferentiation of Th2-like regulatory T cells"

Article Title: Fli1-haploinsufficient dermal fibroblasts promote skin-localized transdifferentiation of Th2-like regulatory T cells

Journal: Arthritis Research & Therapy

doi: 10.1186/s13075-018-1521-3

Techniques Used: Flow Cytometry, Cytometry

5) Product Images from "Fluid Shear Stress Alters the Hemostatic Properties of Endothelial Outgrowth Cells"

Article Title: Fluid Shear Stress Alters the Hemostatic Properties of Endothelial Outgrowth Cells

Journal: Tissue Engineering. Part A

doi: 10.1089/ten.tea.2010.0290

Thrombomodulin, nitric oxide synthase 3 (eNOS), tissue factor pathway inhibitor ( TFPI ), tissue factor ( F3 ), and vWF gene expression in ECs and EOCs was determined by quantitative real-time reverse transcriptase–polymerase chain reaction. mRNA

Figure Legend Snippet: Thrombomodulin, nitric oxide synthase 3 (eNOS), tissue factor pathway inhibitor ( TFPI ), tissue factor ( F3 ), and vWF gene expression in ECs and EOCs was determined by quantitative real-time reverse transcriptase–polymerase chain reaction. mRNA

Techniques Used: Expressing, Polymerase Chain Reaction

6) Product Images from "Phenobarbital Induces Alterations in the Proteome of Hepatocytes and Mesenchymal Cells of Rat Livers"

Article Title: Phenobarbital Induces Alterations in the Proteome of Hepatocytes and Mesenchymal Cells of Rat Livers

Journal: PLoS ONE

doi: 10.1371/journal.pone.0076137

Techniques Used: Microscopy, In Vitro, In Vivo

Techniques Used: In Vitro, Expressing, In Vivo

Techniques Used: In Vitro, In Vivo

7) Product Images from "A rigorous method to enrich for exosomes from brain tissue"

Article Title: A rigorous method to enrich for exosomes from brain tissue

Journal: Journal of Extracellular Vesicles

doi: 10.1080/20013078.2017.1348885

$Schematic of the exosome isolation protocol from solid brain tissue. Fresh frozen (−80°C) human frontal cortex was sliced with a razor blade on ice while frozen to generate 1–2 cm long, 2–3 mm wide sections. The cut sections are dissociated while partially frozen in 75 U/ml of collagenase type 3 in Hibernate-E at 37°C for a total of 20 min. The tissue is returned to ice immediately after incubation and protease and phosphatase inhibitors are added. The tissue is spun at 300 × g for 5 min at 4°C (pellet is used as the brain homogenate + collagenase control), the supernatant transferred to a fresh tube, spun at 2000 × g for 10 min at 4°C, then at 10,000 × g for 30 min at 4°C. The EV-containing supernatant is overlaid on a triple sucrose cushion (0.6 M, 1.3 M, 2.5 M) and ultracentrifuged for 3 h at 180,000 × g to separate vesicles based on density. The top of the gradient is discarded and fractions designated 1, 2 and 3 are collected and the refractive index is measured. Each fraction is further ultracentrifuged at 100,000 × g to pellet the vesicles contained in each fraction. Each preparation is validated by a combination of techniques including electron microscopy and RNA and protein analysis. Note – some tissue samples will not be amenable to this method. Post-mortem delay, storage time and the number of freeze-thaw cycles will negatively impact on tissue quality and result in contamination of the fractions with cellular debris and non-exosome vesicles.$
Figure Legend Snippet: Schematic of the exosome isolation protocol from solid brain tissue. Fresh frozen (−80°C) human frontal cortex was sliced with a razor blade on ice while frozen to generate 1–2 cm long, 2–3 mm wide sections. The cut sections are dissociated while partially frozen in 75 U/ml of collagenase type 3 in Hibernate-E at 37°C for a total of 20 min. The tissue is returned to ice immediately after incubation and protease and phosphatase inhibitors are added. The tissue is spun at 300 × g for 5 min at 4°C (pellet is used as the brain homogenate + collagenase control), the supernatant transferred to a fresh tube, spun at 2000 × g for 10 min at 4°C, then at 10,000 × g for 30 min at 4°C. The EV-containing supernatant is overlaid on a triple sucrose cushion (0.6 M, 1.3 M, 2.5 M) and ultracentrifuged for 3 h at 180,000 × g to separate vesicles based on density. The top of the gradient is discarded and fractions designated 1, 2 and 3 are collected and the refractive index is measured. Each fraction is further ultracentrifuged at 100,000 × g to pellet the vesicles contained in each fraction. Each preparation is validated by a combination of techniques including electron microscopy and RNA and protein analysis. Note – some tissue samples will not be amenable to this method. Post-mortem delay, storage time and the number of freeze-thaw cycles will negatively impact on tissue quality and result in contamination of the fractions with cellular debris and non-exosome vesicles.

Techniques Used: Isolation, Incubation, Electron Microscopy

8) Product Images from "The correlation of IRE1α oxidation with Nox4 activation in aging-associated vascular dysfunction"

Article Title: The correlation of IRE1α oxidation with Nox4 activation in aging-associated vascular dysfunction

Journal: Redox Biology

doi: 10.1016/j.redox.2020.101727

IRE1α physically interacts with Nox4 and controls NO bioavailability in D-gal-induced endothelial senescence. Lysates were immunoprecipitated and immunoblotted with anti -Nox4 antibody and immunoblotted with anti-IRE1α antibody for indicated times (A) and dose (B). The proximity ligation assay (PLA) assay with fluorescence images of HUVECs are shown (C). Quantification of the number of positive PLA dots per cell (Nox4 and IRE1α interactions) are shown for indicated samples. (D) Schematic of IRE1α structure and the mutants analyzed (TM; transmembrane domain). (E) Immunoprecipitation using anti-sulfonate antibody was conducted in cells with transiently transfected IRE1α wild type (WT) and IRE1α mutant plasmid tagging to flag (C715S/C762S, C762S, C715S, and K599A). (F) Western blots of anti -phospho-eNOS Ser1177 or total-eNOS antibody were performed in cells transiently transfected with IRE1α wild type (WT) and IRE1α mutant plasmid tagging to flag (C715S/C762S, C762S, C715S, and K599A). Scale bars, 20 μm. Data are mean ± standard deviation. # , p

Figure Legend Snippet: IRE1α physically interacts with Nox4 and controls NO bioavailability in D-gal-induced endothelial senescence. Lysates were immunoprecipitated and immunoblotted with anti -Nox4 antibody and immunoblotted with anti-IRE1α antibody for indicated times (A) and dose (B). The proximity ligation assay (PLA) assay with fluorescence images of HUVECs are shown (C). Quantification of the number of positive PLA dots per cell (Nox4 and IRE1α interactions) are shown for indicated samples. (D) Schematic of IRE1α structure and the mutants analyzed (TM; transmembrane domain). (E) Immunoprecipitation using anti-sulfonate antibody was conducted in cells with transiently transfected IRE1α wild type (WT) and IRE1α mutant plasmid tagging to flag (C715S/C762S, C762S, C715S, and K599A). (F) Western blots of anti -phospho-eNOS Ser1177 or total-eNOS antibody were performed in cells transiently transfected with IRE1α wild type (WT) and IRE1α mutant plasmid tagging to flag (C715S/C762S, C762S, C715S, and K599A). Scale bars, 20 μm. Data are mean ± standard deviation. # , p

Techniques Used: Immunoprecipitation, Proximity Ligation Assay, Fluorescence, Transfection, Mutagenesis, Plasmid Preparation, Western Blot, Standard Deviation

Nox4 deficiency in cultured endothelial cells improves vascular function in D-gal-induced endothelial senescence. (A) HUVECs were transiently transfected with Nox4 siRNA, followed by transfection with pcDNA-Nox4 or empty vector. (B) Representative senescence-associated β-galactosidase (SA-β-gal) staining in HUVECs. (C) Western blots analysis of anti -phospho-eNOS Ser1177 or total-eNOS antibody. (D) Representative fluorescent images of NO formation in HUVECs were shown using NO probe DAF-2DA. (E) Representative cells were stained for DHE. (F) Images showing cell transfected with ER-targeted HyPer-Red (HyPer Red ER). ER NADPH oxidase activity (G) and hydrogen peroxide level (H) were measured in HUVECs. (I) Irreversible sulfonation of IRE1α was analyzed with transiently transfected with non-specific-siRNA or Nox4 siRNA. Scale bars, 20 μm. Data are mean ± standard deviation. # , p

Figure Legend Snippet: Nox4 deficiency in cultured endothelial cells improves vascular function in D-gal-induced endothelial senescence. (A) HUVECs were transiently transfected with Nox4 siRNA, followed by transfection with pcDNA-Nox4 or empty vector. (B) Representative senescence-associated β-galactosidase (SA-β-gal) staining in HUVECs. (C) Western blots analysis of anti -phospho-eNOS Ser1177 or total-eNOS antibody. (D) Representative fluorescent images of NO formation in HUVECs were shown using NO probe DAF-2DA. (E) Representative cells were stained for DHE. (F) Images showing cell transfected with ER-targeted HyPer-Red (HyPer Red ER). ER NADPH oxidase activity (G) and hydrogen peroxide level (H) were measured in HUVECs. (I) Irreversible sulfonation of IRE1α was analyzed with transiently transfected with non-specific-siRNA or Nox4 siRNA. Scale bars, 20 μm. Data are mean ± standard deviation. # , p

Techniques Used: Cell Culture, Transfection, Plasmid Preparation, Staining, Western Blot, Activity Assay, Standard Deviation

Nox4 overexpression in cultured endothelial cells suppresses vascular function in endothelial senescence. (A)HUVECs were transiently transfected with non-specific siRNA or Nox4 siRNA, followed by transfection with pcDNA-Nox4 or empty vector. (B) Representative senescence-associated β-galactosidase (SA-β-gal) staining in HUVECs. (C) Western blots of anti -phospho-eNOS Ser1177 or total-eNOS antibody. (D) Representative fluorescent images of NO formation in HUVECs are shown using the NO probe DAF-2DA. (E) Representative aortic sections were stained for DHE. (F) Images showing cells transfected with ER-targeted HyPer-Red (HyPer Red ER). ER NADPH oxidase activity (G) and hydrogen peroxide levels (H) in HUVECs were measured. (I) Irreversible sulfonylation of IRE1α was analyzed by western blots. Scale bars, 20 μm. Data represent mean ± standard deviation. # , p

Figure Legend Snippet: Nox4 overexpression in cultured endothelial cells suppresses vascular function in endothelial senescence. (A)HUVECs were transiently transfected with non-specific siRNA or Nox4 siRNA, followed by transfection with pcDNA-Nox4 or empty vector. (B) Representative senescence-associated β-galactosidase (SA-β-gal) staining in HUVECs. (C) Western blots of anti -phospho-eNOS Ser1177 or total-eNOS antibody. (D) Representative fluorescent images of NO formation in HUVECs are shown using the NO probe DAF-2DA. (E) Representative aortic sections were stained for DHE. (F) Images showing cells transfected with ER-targeted HyPer-Red (HyPer Red ER). ER NADPH oxidase activity (G) and hydrogen peroxide levels (H) in HUVECs were measured. (I) Irreversible sulfonylation of IRE1α was analyzed by western blots. Scale bars, 20 μm. Data represent mean ± standard deviation. # , p

Techniques Used: Over Expression, Cell Culture, Transfection, Plasmid Preparation, Staining, Western Blot, Activity Assay, Standard Deviation

$Nox4 expression in vascular endothelial cells in aorta activates endoplasmic reticulum (ER) stress and unfolded protein response (UPR). Aortas from young (2-month) Nox4 WT, KO and aged (23- to 24-month) Nox4 WT, KO mice ( n = 7 animals per group). (A) Bioinformatic analysis was performed in aortas from aged Nox4 WT mice and young Nox4 WT mice by the heatmap. (B) KEGG enrichment analysis showing repressed signaling pathways from aged Nox4 WT mice and young Nox4 WT mice. (C) Western blots analysis of Nox1, Nox2, Nox3, Nox4, and β-actin expression in young and aged aorta tissues. (D) Lysates were processed to obtain subcellular fractions and analyzed by Western blot. (E) Confocal images (scale bars, 20 μm) show localization of Nox4 (red) to the ER, which was labeled with an anti -calnexin antibody (green). Yellow dots denote the co-localization of Nox4 and calnexin signals. Cell nuclei were stained with DAPI (blue). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)$
Figure Legend Snippet: Nox4 expression in vascular endothelial cells in aorta activates endoplasmic reticulum (ER) stress and unfolded protein response (UPR). Aortas from young (2-month) Nox4 WT, KO and aged (23- to 24-month) Nox4 WT, KO mice ( n = 7 animals per group). (A) Bioinformatic analysis was performed in aortas from aged Nox4 WT mice and young Nox4 WT mice by the heatmap. (B) KEGG enrichment analysis showing repressed signaling pathways from aged Nox4 WT mice and young Nox4 WT mice. (C) Western blots analysis of Nox1, Nox2, Nox3, Nox4, and β-actin expression in young and aged aorta tissues. (D) Lysates were processed to obtain subcellular fractions and analyzed by Western blot. (E) Confocal images (scale bars, 20 μm) show localization of Nox4 (red) to the ER, which was labeled with an anti -calnexin antibody (green). Yellow dots denote the co-localization of Nox4 and calnexin signals. Cell nuclei were stained with DAPI (blue). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Techniques Used: Expressing, Mouse Assay, Western Blot, Labeling, Staining

$Nox4 activates aging-associated oxidative stress and endothelial dysfunction. Aortas from young (2-month) Nox4 WT, KO and aged (23- to 24-month) Nox4 WT, KO mice. ( n = 7 animals per group) (A) Endothelial-dependent vasodilation to acetylcholine. Representative aortic sections (scale bars, 20 μm) were stained for DAF-2DA (B), p -eNOS (ser 1177) (C). Immunoblotting analysis of eNOS-dimers and -monomers in the young and aged aorta (D). Representative aortic sections(scale bars, 20 μm) were stained for DHE (E). Lysates from aorta tissues were analyzed for NADPH oxidase activity (E), hydrogen peroxide level (F), 4-HNE-malondialdehyde (MDA) level (G) in the aorta ER fraction. (H) Western blots analysis of p-IRE1α, IRE1α, p -eIF2α, eIF2α, ATF4, GRP78, CHOP, and β-actin expression in aorta tissues. ( n = 7 animals per experimental group). Data are mean ± standard deviation. # , p$
Figure Legend Snippet: Nox4 activates aging-associated oxidative stress and endothelial dysfunction. Aortas from young (2-month) Nox4 WT, KO and aged (23- to 24-month) Nox4 WT, KO mice. ( n = 7 animals per group) (A) Endothelial-dependent vasodilation to acetylcholine. Representative aortic sections (scale bars, 20 μm) were stained for DAF-2DA (B), p -eNOS (ser 1177) (C). Immunoblotting analysis of eNOS-dimers and -monomers in the young and aged aorta (D). Representative aortic sections(scale bars, 20 μm) were stained for DHE (E). Lysates from aorta tissues were analyzed for NADPH oxidase activity (E), hydrogen peroxide level (F), 4-HNE-malondialdehyde (MDA) level (G) in the aorta ER fraction. (H) Western blots analysis of p-IRE1α, IRE1α, p -eIF2α, eIF2α, ATF4, GRP78, CHOP, and β-actin expression in aorta tissues. ( n = 7 animals per experimental group). Data are mean ± standard deviation. # , p

Techniques Used: Mouse Assay, Staining, Activity Assay, Multiple Displacement Amplification, Western Blot, Expressing, Standard Deviation

Isolation:

Article Title: Exogenous Stimulation of Human Intervertebral Disc Cells in 3-Dimensional Alginate Bead Culture With BMP2 and L51P: Cytocompatibility and Effects on Cell Phenotype
Article Snippet: .. Briefly, cells were isolated by pronase (Roche, Basel, Switzerland) followed by collagenase type 2 (Worthington, London, UK) enzymatic digestion as previously reported. .. NPCs, AFCs and CEPCs were expanded in proliferation medium (low-glucose [1g/L] Dulbecco’s Modified Eagle Medium supplemented with 10% foetal bovine serum [FBS] and penicillin/streptomycin [P/S]).

Article Title: O-GlcNAc transferase inhibits visceral fat lipolysis and promotes diet-induced obesity
Article Snippet: .. For brown adipocyte isolation, freshly dissected interscapular BAT was cut into ~2 mm diameter pieces, digested in Krebs-Ringer buffer supplemented with 1% fatty acid free BSA, 0.1% HEPES (1 M, pH 7.3), 0.8 mg/mL Type 2 Collagenase (Worthington, LS004176), and 1.2 mM calcium chloride at 37 °C for 50 min. Digested tissue was then filtered through a 100 μm membrane and centrifuged at 300 g for 3 min. ..

Incubation:

Article Title: Fli1-haploinsufficient dermal fibroblasts promote skin-localized transdifferentiation of Th2-like regulatory T cells
Article Snippet: .. Dermis was minced and then incubated with 2 mg/ml collagenase type 2 (CLS-2; Worthington Biochemical, Lakewood, NJ, USA) in Tyrode’s solution for 60–90 minutes. ..

Mouse Assay:

Article Title: Cardiac O-GlcNAcylation blunts autophagic signaling in the diabetic heart
Article Snippet: .. Briefly, hearts were rapidly excised from heparinized and anesthetized mice, perfused retrogradely with Ca2+ -free perfusion buffer consisting of (in mM) 0.6 KH2 PO4 , 0.6 Na2 HPO4 , 1.2 MgSO4 • 7H2 O, 0.032 phenol red, 12 NaHCO3 , 10 KHCO3 , 10 HEPES, 30 taurine, 10 2,3-butanedione monoxime, and 5.5 glucose, pH 7.46, followed by buffer containing 12.5 μM CaCl2 and 0.4 mg/ml collagenase type 2 (Worthington). ..