Journal: Animals : an Open Access Journal from MDPI

Article Title: Low Expression of Sirtuin 1 in the Dairy Cows with Mild Fatty Liver Alters Hepatic Lipid Metabolism

doi: 10.3390/ani10040560

Figure Lengend Snippet: Primers sequences qRT-PCR used in this experiment.

Article Snippet: Primary antibodies against ACO (YN0401), FAS (YM1224), ACCα (YT0074), p-ACCα (YP0620), ACSL1 (YN0827), LDLR (YN2236), RXRα (YN0018), Nrf1 (YT3188) and TFAM (YT2916) were purchased from ImmunoWay Biotechnology Company (Newark, DE, USA).

Techniques: Amplification

Journal: PPAR Research

Article Title: PPAR- γ Ligand Inhibits Nasopharyngeal Carcinoma Cell Proliferation and Metastasis by Regulating E2F2

doi: 10.1155/2019/8679271

Figure Lengend Snippet: Expressions of E2F2 and PPAR- γ in clinical samples of undifferentiated NPC and NPG and associations with clinicopathological characteristics of undifferentiated NPC samples.

Article Snippet: After three washes with phosphate-buffered saline (PBS), the sections were incubated in a blocking solution containing bovine serum albumin (BSA) for 20 min, followed by exposure to primary antibodies against E2F2 (YT1443, Immunoway, rabbit polyclonal, 1:200 dilution) and PPAR- γ (Novusbio, rabbit polyclonal, 1:100 dilution) at 4°C overnight.

Techniques: Expressing

Journal: PPAR Research

Article Title: PPAR- γ Ligand Inhibits Nasopharyngeal Carcinoma Cell Proliferation and Metastasis by Regulating E2F2

doi: 10.1155/2019/8679271

Figure Lengend Snippet: E2F2 and PPAR-γ protein expression in nonkeratinizing nasopharyngeal carcinoma (NPC) and nasopharyngitis (NPG) tissues . Immunohistochemistry was performed to detect E2F2 and PPAR- γ protein expression in nonkeratinizing NPC tissues (a, b) and NPG tissues (c, d). Scale bar, 50 μ m (e, f). The immunohistochemistry data were analyzed semiquantitatively to determine the E2F2 and PPAR- γ expression levels in nonkeratinizing NPC and NPG tissues. Data are shown as means ± standard deviations. ∗∗ P <0.01.

Article Snippet: After three washes with phosphate-buffered saline (PBS), the sections were incubated in a blocking solution containing bovine serum albumin (BSA) for 20 min, followed by exposure to primary antibodies against E2F2 (YT1443, Immunoway, rabbit polyclonal, 1:200 dilution) and PPAR- γ (Novusbio, rabbit polyclonal, 1:100 dilution) at 4°C overnight.

Techniques: Expressing, Immunohistochemistry

Journal: PPAR Research

Article Title: PPAR- γ Ligand Inhibits Nasopharyngeal Carcinoma Cell Proliferation and Metastasis by Regulating E2F2

doi: 10.1155/2019/8679271

Figure Lengend Snippet: E2F2 and PPAR-γ protein expression in nonkeratinizing nasopharyngeal carcinoma (NPC) and nasopharyngitis (NPG) tissue lysates . (a, b) RT-PCR and Western blotting were performed to detect the expression of E2F2, PPAR- γ , and GAPDH in lysates of nonkeratinizing NPC and NPG tissues. (c, d) Quantitative analyses of the E2F2 and PPAR- γ expression levels in nonkeratinizing NPC and NPG tissues. Data are expressed as means ± standard deviations. ∗ P <0.05; ∗∗ P <0.01.

Article Snippet: After three washes with phosphate-buffered saline (PBS), the sections were incubated in a blocking solution containing bovine serum albumin (BSA) for 20 min, followed by exposure to primary antibodies against E2F2 (YT1443, Immunoway, rabbit polyclonal, 1:200 dilution) and PPAR- γ (Novusbio, rabbit polyclonal, 1:100 dilution) at 4°C overnight.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot

Journal: PPAR Research

Article Title: PPAR- γ Ligand Inhibits Nasopharyngeal Carcinoma Cell Proliferation and Metastasis by Regulating E2F2

doi: 10.1155/2019/8679271

Figure Lengend Snippet: E2F2 and PPAR-γ expression in CNE1 and CNE2 nasopharyngeal carcinoma cells after treatment with PPAR-γ ligand . (a, b) Western blotting was performed to detect the expression of E2F2, PPAR- γ , and GAPDH in CNE1 and CNE2 cells after treatment with dose-dependent PPAR- γ agonist Rog. (c, d) Western blotting was performed to detect the expression of E2F2, PPAR- γ , and GAPDH in CNE1 and CNE2 cells after treatment with PPAR- γ agonist (rosiglitazone, Rog) and PPAR- γ antagonist (GW9662).

Article Snippet: After three washes with phosphate-buffered saline (PBS), the sections were incubated in a blocking solution containing bovine serum albumin (BSA) for 20 min, followed by exposure to primary antibodies against E2F2 (YT1443, Immunoway, rabbit polyclonal, 1:200 dilution) and PPAR- γ (Novusbio, rabbit polyclonal, 1:100 dilution) at 4°C overnight.

Techniques: Expressing, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: The Post-Translational Modifications, Localization, and Mode of Attachment of Non-Covalently Bound Glucanosyltransglycosylases of Yeast Cell Wall as a Key to Understanding their Functioning

doi: 10.3390/ijms21218304

Figure Lengend Snippet: Post-translational modifications of Bgl2 and Scw4 in T, G and L pools. Glut—glutathionylation, P—phosphorylation. Modified amino acid residues are underlined and highlighted in bold.

Article Snippet: Samples were stained with mouse primary polyclonal antibodies against Bgl2 (obtained as described in the ) and secondary polyclonal goat anti-mouse antibodies IgG labeled with Alexa-488 fluorophore (Invitrogen, Moscow, Russia) or with rabbit primary polyclonal antibodies against Gas1 and secondary polyclonal goat anti-rabbit antibodies IgG labeled with Alexa-647 fluorophore (Invitrogen, Moscow, Russia; Rostock, Germany).

Techniques: Modification

Journal: International Journal of Molecular Sciences

Article Title: The Post-Translational Modifications, Localization, and Mode of Attachment of Non-Covalently Bound Glucanosyltransglycosylases of Yeast Cell Wall as a Key to Understanding their Functioning

doi: 10.3390/ijms21218304

Figure Lengend Snippet: Analysis of extracts from Saccharomyces cerevisiae cell walls obtained with 0.1 M Tris for 3.5 h at 30 °C—T, with 6 M GuHCl for 2 h at 30 °C—G ( A – C ) and with water at 100 °C after removal of lipid component—L ( C , D ). PAGE stained with Coomassie G-250 ( A ) or with silver nitrate staining ( C ) and Western blot stained with antibodies against Bgl2 ( B , D ).

Article Snippet: Samples were stained with mouse primary polyclonal antibodies against Bgl2 (obtained as described in the ) and secondary polyclonal goat anti-mouse antibodies IgG labeled with Alexa-488 fluorophore (Invitrogen, Moscow, Russia) or with rabbit primary polyclonal antibodies against Gas1 and secondary polyclonal goat anti-rabbit antibodies IgG labeled with Alexa-647 fluorophore (Invitrogen, Moscow, Russia; Rostock, Germany).

Techniques: Staining, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: The Post-Translational Modifications, Localization, and Mode of Attachment of Non-Covalently Bound Glucanosyltransglycosylases of Yeast Cell Wall as a Key to Understanding their Functioning

doi: 10.3390/ijms21218304

Figure Lengend Snippet: Western blot analysis of Saccharomyces cerevisiae cell lysates obtained from WT-OE, N202-OE, and N284-OE strains. Samples were incubated for 15 min with (+) or without (−) endoglycosydase H (EndoH). Bgl2 bands are denoted by arrowheads; (*) indicates unglycosylated Bgl2. Staining with antibodies against Bgl2.

Article Snippet: Samples were stained with mouse primary polyclonal antibodies against Bgl2 (obtained as described in the ) and secondary polyclonal goat anti-mouse antibodies IgG labeled with Alexa-488 fluorophore (Invitrogen, Moscow, Russia) or with rabbit primary polyclonal antibodies against Gas1 and secondary polyclonal goat anti-rabbit antibodies IgG labeled with Alexa-647 fluorophore (Invitrogen, Moscow, Russia; Rostock, Germany).

Techniques: Western Blot, Incubation, Staining

Journal: International Journal of Molecular Sciences

Article Title: The Post-Translational Modifications, Localization, and Mode of Attachment of Non-Covalently Bound Glucanosyltransglycosylases of Yeast Cell Wall as a Key to Understanding their Functioning

doi: 10.3390/ijms21218304

Figure Lengend Snippet: Microscopy of structures formed in G pool extracted from Saccharomyces cerevisiae cell walls: TEM ( A – F ), immunofluorescence microscopy, staining with antibodies against Bgl2 ( G – I ). General view of jellyfish-like associates obtained from wt strain ( A , B , G , H ), fibrillar structure of their bodies ( D , E ). Control samples from bgl2Δ strain ( C , F , I ).

Article Snippet: Samples were stained with mouse primary polyclonal antibodies against Bgl2 (obtained as described in the ) and secondary polyclonal goat anti-mouse antibodies IgG labeled with Alexa-488 fluorophore (Invitrogen, Moscow, Russia) or with rabbit primary polyclonal antibodies against Gas1 and secondary polyclonal goat anti-rabbit antibodies IgG labeled with Alexa-647 fluorophore (Invitrogen, Moscow, Russia; Rostock, Germany).

Techniques: Microscopy, Immunofluorescence, Staining

Journal: International Journal of Molecular Sciences

Article Title: The Post-Translational Modifications, Localization, and Mode of Attachment of Non-Covalently Bound Glucanosyltransglycosylases of Yeast Cell Wall as a Key to Understanding their Functioning

doi: 10.3390/ijms21218304

Figure Lengend Snippet: Structural model of Bgl2 molecule without ( A ) and with glutathione ( B – D ). N-terminal is highlighted in green, C-terminal is highlighted in cyan. G32-S42 loop is shown in magenta. The glutathione molecule located in the cavity of Bgl2 greatly changes the structure ( B ). The external location of glutathione molecule does not contribute to significant changes in Bgl2 structure ( C , D ). Possible conformations of glutathione resulting from molecular dynamics simulation are shown as molecular surface (gray area). Axial ( A – C ) and frontal ( D ) views of the cartoon.

Article Snippet: Samples were stained with mouse primary polyclonal antibodies against Bgl2 (obtained as described in the ) and secondary polyclonal goat anti-mouse antibodies IgG labeled with Alexa-488 fluorophore (Invitrogen, Moscow, Russia) or with rabbit primary polyclonal antibodies against Gas1 and secondary polyclonal goat anti-rabbit antibodies IgG labeled with Alexa-647 fluorophore (Invitrogen, Moscow, Russia; Rostock, Germany).

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: The Post-Translational Modifications, Localization, and Mode of Attachment of Non-Covalently Bound Glucanosyltransglycosylases of Yeast Cell Wall as a Key to Understanding their Functioning

doi: 10.3390/ijms21218304

Figure Lengend Snippet: Immunofluorescence microscopy of Saccharomyces cerevisiae cells and cell walls stained with antibodies against Bgl2. Scale bars correspond to 2 µm. Cells ( A – C ). CW with ( D , E ) and without ( F ) EDC crosslinking and boiled in 3% SDS. CW before ( G – I ) and after ( J – K ) Tris extraction. Control samples from bgl2Δ strain ( C , I , L ).

Article Snippet: Samples were stained with mouse primary polyclonal antibodies against Bgl2 (obtained as described in the ) and secondary polyclonal goat anti-mouse antibodies IgG labeled with Alexa-488 fluorophore (Invitrogen, Moscow, Russia) or with rabbit primary polyclonal antibodies against Gas1 and secondary polyclonal goat anti-rabbit antibodies IgG labeled with Alexa-647 fluorophore (Invitrogen, Moscow, Russia; Rostock, Germany).

Techniques: Immunofluorescence, Microscopy, Staining

Journal: International Journal of Molecular Sciences

Article Title: The Post-Translational Modifications, Localization, and Mode of Attachment of Non-Covalently Bound Glucanosyltransglycosylases of Yeast Cell Wall as a Key to Understanding their Functioning

doi: 10.3390/ijms21218304

Figure Lengend Snippet: The hypothetical scheme of Saccharomyces cerevisiae cell wall segment with a microcompartment. We suppose that PTM-free Bgl2 enters the cell wall as a part of L pool. Depending on the degree of association with proteins in L pool, Bgl2 is directed to T or G pool. Probably, Bgl2, acquiring one or another set of PTMs, can migrate between T and G pools. In these two pools, at least some of the molecules are closely located, therefore, we suppose that they form the microcompartment. The localization of L pool could not be revealed. It is possible that proteins of L pool are dispersed rather than compactly located in the cell wall. Glut—glutathionylation, MultiP and MonoP—multi- and monophosphorylation, respectively.

Article Snippet: Samples were stained with mouse primary polyclonal antibodies against Bgl2 (obtained as described in the ) and secondary polyclonal goat anti-mouse antibodies IgG labeled with Alexa-488 fluorophore (Invitrogen, Moscow, Russia) or with rabbit primary polyclonal antibodies against Gas1 and secondary polyclonal goat anti-rabbit antibodies IgG labeled with Alexa-647 fluorophore (Invitrogen, Moscow, Russia; Rostock, Germany).

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: The Post-Translational Modifications, Localization, and Mode of Attachment of Non-Covalently Bound Glucanosyltransglycosylases of Yeast Cell Wall as a Key to Understanding their Functioning

doi: 10.3390/ijms21218304

Figure Lengend Snippet: Saccharomyces cerevisiae strains used in present research.

Article Snippet: Samples were stained with mouse primary polyclonal antibodies against Bgl2 (obtained as described in the ) and secondary polyclonal goat anti-mouse antibodies IgG labeled with Alexa-488 fluorophore (Invitrogen, Moscow, Russia) or with rabbit primary polyclonal antibodies against Gas1 and secondary polyclonal goat anti-rabbit antibodies IgG labeled with Alexa-647 fluorophore (Invitrogen, Moscow, Russia; Rostock, Germany).

Techniques:

Journal: Cell Biology and Toxicology

Article Title: Functional characterization of OR51B5 and OR1G1 in human lung epithelial cells as potential drug targets for non-type 2 lung diseases

doi: 10.1007/s10565-024-09935-9

Figure Lengend Snippet: Detection of OR51B5 and OR1G1 expression in A549 cells. RT-PCR demonstrated the presence of ( A ) OR51B5 PCR fragment at 227 bp and ( B ) OR1G1 at 203 bp in A549. ( C ) mRNA transcripts of OR1A1 and OR10S1 were not detected but the genomic DNA was detected for both. The expression of the corresponding proteins was confirmed by ( D , E ) Western Blot at the estimated molecular mass of about 35 kDa and ( F , G ) immunofluorescence staining, specific antibodies were used against the ORs (red) and the plasma membrane marker E-cadherin (green). The nucleus was stained using 4′,6-Diamidin-2-phenylindol (Dapi; blue). The overlay between the OR (red) and nucleus (blue) staining indicated as violet color in the merged picture. Scale bars: 50 μm and the squares delineate the magnified region. Representative of TN = 3 individual experiments

Article Snippet: Primary antibodies against OR51B5 (BIOZOL, USA, cat: A13766), OR1G1 (NOVUS, cat: NBP1-68970) and the membrane marker E-cadherin (Thermo Fisher Scientific, cat: 13–1700) were applied at dilutions of 1:100, 1:200 and 1:2000, respectively in PBST containing 2.5% goat serum and incubated o/n at 4 °C.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot, Immunofluorescence, Staining, Clinical Proteomics, Membrane, Marker

Journal: Cell Biology and Toxicology

Article Title: Functional characterization of OR51B5 and OR1G1 in human lung epithelial cells as potential drug targets for non-type 2 lung diseases

doi: 10.1007/s10565-024-09935-9

Figure Lengend Snippet: Farnesol-Isononyl alcohol (OR51B5 agonists)-triggered calcium mobilization is mediated by both cAMP/PLC . A549 cells were stimulated each measurement three times with ( A - C ) Farnesol (300 µM) or ( D , E ) Isononyl alcohol (1mM) in presence or absence of the inhibitors; ( A , D ) extracellular calcium chelator (EGTA 10mM), (B) Adenylate cyclase inhibitor (SQ 22536 10 µM) or ( C , E ) PLC inhibitor (U73122, 2 µM) in the second application, inhibitors were pre-incubated for three minutes before adding odorants in the second application. The bars of all experiments indicate the stimulus duration. Data are shown as mean ± SEM, TN = 7–9. Statistical comparison utilized paired student's t-test referring to the ratio of calcium amplitude between second to first applications in the experimental group (odorants-inhibitors application) vs. the corresponding control group (odorants alone). ** p ≤ 0.01; *** p ≤ 0.001

Article Snippet: Primary antibodies against OR51B5 (BIOZOL, USA, cat: A13766), OR1G1 (NOVUS, cat: NBP1-68970) and the membrane marker E-cadherin (Thermo Fisher Scientific, cat: 13–1700) were applied at dilutions of 1:100, 1:200 and 1:2000, respectively in PBST containing 2.5% goat serum and incubated o/n at 4 °C.

Techniques: Incubation, Comparison, Control

Journal: Cell Biology and Toxicology

Article Title: Functional characterization of OR51B5 and OR1G1 in human lung epithelial cells as potential drug targets for non-type 2 lung diseases

doi: 10.1007/s10565-024-09935-9

Figure Lengend Snippet: Stimulation of OR51B5 and OR1G1 reduces A549 cell viability. A549 cells were exposed for 24h to increasing concentrations of ( A , B ) Farnesol, ( C , D ) Isononyl alcohol or ( E , F ) Nonanal or solvent DMSO (0.1%). Proportions of viable, apoptotic, and necrotic cells were determined by flow cytometry. Relative cell counts were calculated in comparison to the total cell population. Cells staining positive for Annexin V (AV +) were indicative of early apoptosis and were denoted in the lower right quadrant of the chart (R3), represented by the dark grey portion in the accompanying graphs. AV/propidium iodide (PI) double-positive staining, (AV + , PI +), signified necrotic cells and was in the upper right quadrant of the chart (R2), depicted by the light grey section of the graphs. Viable cells, defined as those unstained by AV and PI (AV − , PI −), were found in the lower left quadrant of the chart and were represented by the black bars in the graphs. Data are shown as mean ± SEM, TN = 5–7. Statistical significance vs. DMSO (0.1%) utilized One-way ANOVA. test with post hoc “Two-stage-up method of Benjamini, Krieger, and Yekutieli,”. * p ≤ 0.05, ** p ≤ 0.01 and *** p ≤ 0.001

Article Snippet: Primary antibodies against OR51B5 (BIOZOL, USA, cat: A13766), OR1G1 (NOVUS, cat: NBP1-68970) and the membrane marker E-cadherin (Thermo Fisher Scientific, cat: 13–1700) were applied at dilutions of 1:100, 1:200 and 1:2000, respectively in PBST containing 2.5% goat serum and incubated o/n at 4 °C.

Techniques: Solvent, Flow Cytometry, Comparison, Staining

Journal: Cell Biology and Toxicology

Article Title: Functional characterization of OR51B5 and OR1G1 in human lung epithelial cells as potential drug targets for non-type 2 lung diseases

doi: 10.1007/s10565-024-09935-9

Figure Lengend Snippet: Expression and functional characterization of OR51B5 and OR1G1 ALI-cultured primary epithelial cell . RT-PCR revealed the presence of mRNA transcripts of ( A ) OR51B5 and ( B ) OR1G1. Validation of ( C ) OR51B5 and ( D ) OR1G1 protein expression by Western Blot. Evident bands for both receptors at the expected size of 35 kDa are shown. Representative of 3 donors. ALI-PBECs were stimulated with increasing concentrations of ( E ) Farnesol, ( F ) Isononyl alcohol or ( G ) Nonanal or solvent DMSO (0.1%). Proportions of viable, apoptotic, and necrotic cells were determined by flow cytometry using AV/ PI staining. Percentage of ( E – G ) necrotic cells were calculated versus total cell count of 5 individual experiments from five donors. ALI-PBECs were stimulated for 6 h with ( H , K ) Farnesol, ( I , L ) Isononyl alcohol and ( J , M ) Nonanal. After 6 h media was removed from apical to preserve cell integrity . Supernatant was collected from the basal part after 24 h. Concentrations are indicated in the graphs. IL-8 and IL-6 were measured by ELISA. Experiments were conducted with TN = 6–10 independent samples from six to tent different donors. Data were normalized to solvent controls (DMSO). ( E - M ) data are shown as mean ± SEM. Statistical significance vs. DMSO (0.1%) utilized Friedman test with post hoc “Two-stage-up method of Benjamini, Krieger, and Yekutieli,”. * p ≤ 0.05, ** p ≤ 0.01 and *** p ≤ 0.001, ns = non-significant

Article Snippet: Primary antibodies against OR51B5 (BIOZOL, USA, cat: A13766), OR1G1 (NOVUS, cat: NBP1-68970) and the membrane marker E-cadherin (Thermo Fisher Scientific, cat: 13–1700) were applied at dilutions of 1:100, 1:200 and 1:2000, respectively in PBST containing 2.5% goat serum and incubated o/n at 4 °C.

Techniques: Expressing, Functional Assay, Cell Culture, Reverse Transcription Polymerase Chain Reaction, Biomarker Discovery, Western Blot, Solvent, Flow Cytometry, Staining, Cell Counting, Enzyme-linked Immunosorbent Assay

Journal: The Journal of allergy and clinical immunology

Article Title: Nasal polyps 15-lipoxygenase 1 promotes CCL26/Eotaxin3 expression through ERK activation

doi: 10.1016/j.jaci.2019.06.037

Figure Lengend Snippet: 15LO1 mRNA and protein are higher in fresh NECs of CRSwNP subjects compared to HCs. Total RNA and protein from freshly brushed NECs were harvested for RT-PCR and Western blot analysis. NP and MT tissues from subjects with CRSwNP and HCs were fixed for Immunohistochemistry studies. (A) 15LO1 mRNA is higher in NECs of NP from CRSwNP subjects compared to HC. (B) 15LO1 protein is higher in NECs of NP from CRSwNP compared to NECs of MT from HCs. (C) Densitometry analysis of 15LO1 protein from CRSwNP subjects and HCs by WB. (D) 15LO1 protein is higher in NECs of NP compared to MT from same CRSwNP subjects. (E) Densitometry analysis of the paired MT and NP 15LO1 protein. (F) Representative immunostaining photomicrograph showed 15LO1 positive staining is upregulated in NECs from CRSwNP subjects than MT of CRSwNP and HCs. Abbreviations: NECs=nasal epithelial cells, HC=healthy controls, NP=nasal polyps, MT=middle turbinate, IT=inferior turbinate, and CRSwNP=chronic rhinosinusitis with nasal polyps.

Article Snippet: Tissue sections were blocked with 5% donkey serum and incubated with primary antibodies against 15LO1 (1: 200 dilution), CCL26 (1:100 dilution), and Cytokeratin-5 (CK5, 1: 200 dilution, Novus, USA).

Techniques: Reverse Transcription Polymerase Chain Reaction, Western Blot, Immunohistochemistry, Immunostaining, Staining

Journal: The Journal of allergy and clinical immunology

Article Title: Nasal polyps 15-lipoxygenase 1 promotes CCL26/Eotaxin3 expression through ERK activation

doi: 10.1016/j.jaci.2019.06.037

Figure Lengend Snippet: CCL26 mRNA and protein are higher in fresh NECs from CRSwNP compared to HCs and correlated with 15LO1 expression. Total RNA and protein from freshly brushed NECs were harvested for RT-PCR and ELISA analysis. (A) CCL26 mRNA is higher in NECs of NP from CRSwNP subjects compared to HCs. (B) CCL26 mRNA expression correlated with 15LO1 mRNA ex vivo. (C) CCL26 protein (by ELISA) is higher in NECs of MT and NP cell lysates from CRSwNP subjects, especially NP, compared to HCs. (D) CCL26 protein (by ELISA) expression correlates with 15LO1 protein expression (by WB) in NP cell lysates. Abbreviations: NECs=nasal epithelial cells, HC=healthy controls, NP=nasal polyps, MT=middle turbinate, and CRSwNP=chronic rhinosinusitis with nasal polyps.

Article Snippet: Tissue sections were blocked with 5% donkey serum and incubated with primary antibodies against 15LO1 (1: 200 dilution), CCL26 (1:100 dilution), and Cytokeratin-5 (CK5, 1: 200 dilution, Novus, USA).

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

Journal: The Journal of allergy and clinical immunology

Article Title: Nasal polyps 15-lipoxygenase 1 promotes CCL26/Eotaxin3 expression through ERK activation

doi: 10.1016/j.jaci.2019.06.037

Figure Lengend Snippet: Immunostaining results showed that 15LO1/CCL26 and CCL26/cytokeratin-5 positive cell numbers increased in MT and NP tissues from CRSwNP subjects, especially in NP tissues, compared to HCs. NP and MT tissues from subjects with CRSwNP and HCs were fixed for IF analysis. (A) Representative immunostaining photomicrograph showed CCL26 positive cells are expressed in MT and NP epithelial cells of CRSwNP subjects. CCL26 colocalized with 15LO1 in MT (n=6) and NP (n=6) epithelial cells of CRSwNP. (B) Representative immunostaining photomicrograph showed no positive 15LO1/CCL26 double staining in MT (n=5) tissues of HCs. (C) Quantitation of 15LO1/CCL26 positive signals in NP and MT epithelial cells from CRSwNP and HCs. (D) Representative immunostaining photomicrograph showed CCL26 colocalized with cytokeratin-5 in MT (n=3) and NP (n=3) epithelial cells from CRSwNP subjects. (E) Quantitation of CCL26/CK5 positive signals in NP and MT tissues from CRSwNP and HCs (n=3). Abbreviations: HC=healthy controls, NP=nasal polyps, MT=middle turbinate, and CRSwNP=chronic rhinosinusitis with nasal polyps.

Article Snippet: Tissue sections were blocked with 5% donkey serum and incubated with primary antibodies against 15LO1 (1: 200 dilution), CCL26 (1:100 dilution), and Cytokeratin-5 (CK5, 1: 200 dilution, Novus, USA).

Techniques: Immunostaining, Double Staining, Quantitation Assay

Journal: The Journal of allergy and clinical immunology

Article Title: Nasal polyps 15-lipoxygenase 1 promotes CCL26/Eotaxin3 expression through ERK activation

doi: 10.1016/j.jaci.2019.06.037

Figure Lengend Snippet: IL-13 induces 15LO1 and CCL26 expression in ALI cultured NECs and 15LO1 positively correlates with CCL26 expression in vitro. ALI cultured primary NECs were stimulated with IL-13 for 5 days. Cells were harvested for RT-PCR and Western blot analysis. Lower supernatants were harvested for CCL26 levels by ELISA. IL-13 increased (A) 15LO1 mRNA expression, (B) CCL26 mRNA expression and (C) 15LO1 mRNA correlates with CCL26 mRNA expression in vitro. (D) IL-13 induces intracellular 15LO1 and CCL26 protein in cell lysates. (E) IL-13 induced 15LO1 protein correlates with CCL26 protein in vitro. Abbreviations: ALI=air-liquid interface, NECs=nasal epithelial cells.

Article Snippet: Tissue sections were blocked with 5% donkey serum and incubated with primary antibodies against 15LO1 (1: 200 dilution), CCL26 (1:100 dilution), and Cytokeratin-5 (CK5, 1: 200 dilution, Novus, USA).

Techniques: Expressing, Cell Culture, In Vitro, Reverse Transcription Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: The Journal of allergy and clinical immunology

Article Title: Nasal polyps 15-lipoxygenase 1 promotes CCL26/Eotaxin3 expression through ERK activation

doi: 10.1016/j.jaci.2019.06.037

Figure Lengend Snippet: ALOX15 DsiRNA and 15LO1 specific inhibitor BLX2477 decreased IL-13 induced CCL26 expression. ALI cultured NECs were treated with ALOX15 DsiRNA for 5 days in the presence or absence of IL-13. ALI cultured NECs were treated with BLX2477 for 3 days in the presence of IL-13. Cells were harvested for RT-PCR and Western blot. Lower supernatants were harvested for CCL26 protein by ELISA. 15LO1 knockdown decreases (A) intracellular CCL26 mRNA and (B) intracellular CCL26 protein and (C) secreted CCL26 protein in lower supernatants. (D) Densitometry analysis of intracellular CCL26 protein in cell lysates. (E) The 15LO1 inhibitor decreases IL-13 induced CCL26 mRNA, (F) and CCL26 protein in cell lysates and (G) secreted CCL26 protein in lower supernatants. (H) Densitometry analysis of CCL26 protein in cell lysates. Abbreviations: ALI=air-liquid interface, NECs=nasal epithelial cells.

Article Snippet: Tissue sections were blocked with 5% donkey serum and incubated with primary antibodies against 15LO1 (1: 200 dilution), CCL26 (1:100 dilution), and Cytokeratin-5 (CK5, 1: 200 dilution, Novus, USA).

Techniques: Expressing, Cell Culture, Reverse Transcription Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay, Knockdown

Journal: The Journal of allergy and clinical immunology

Article Title: Nasal polyps 15-lipoxygenase 1 promotes CCL26/Eotaxin3 expression through ERK activation

doi: 10.1016/j.jaci.2019.06.037

Figure Lengend Snippet: ALOX15 DsiRNA and 15LO1 inhibitor BLX2477 decreased pERK expression. Blocking ERK pathway inhibits CCL26 production in ALI cultured NECs. (A) ALOX15 DsiRNA decreased pERK but not tERK by WB. (B) The 15LO1 specific inhibitor decreased pERK but not tERK by WB. (C) ALOX15 DsiRNA and 15LO1 specific inhibitor BLX2477 decreased pERK/tERK ratio. (D) ERK inhibitor PD98059 decreased pERK and CCL26 protein by WB. (E) ERK inhibitor U0126 decreased pERK and CCL26 protein by WB. (F) ERK pathway inhibitors PD98059 and U0126 decreased CCL26 mRNA in ALI cultured NECs. (G) The ERK pathway inhibitor PD98059 and U0126 decreased secreted CCL26 in supernatants. Abbreviations: ALI=air-liquid interface, NECs=nasal epithelial cells, WB=western blot.

Article Snippet: Tissue sections were blocked with 5% donkey serum and incubated with primary antibodies against 15LO1 (1: 200 dilution), CCL26 (1:100 dilution), and Cytokeratin-5 (CK5, 1: 200 dilution, Novus, USA).

Techniques: Expressing, Blocking Assay, Cell Culture, Western Blot

Journal: The Journal of allergy and clinical immunology

Article Title: Nasal polyps 15-lipoxygenase 1 promotes CCL26/Eotaxin3 expression through ERK activation

doi: 10.1016/j.jaci.2019.06.037

Figure Lengend Snippet: Increased pERK correlates with 15LO1 and CCL26 in fresh NECs. (A) Phosphorylated ERK is increased in fresh NECs of NP compared to HCs. (B) Densitometry analysis of pERK/tERK in fresh NECs of CRSwNP and HCs. (C) pERK/tERK ratio correlates with 15LO1 protein in fresh NECs of CRSwNP and HCs. (D) pERK/tERK ratio correlates with CCL26 protein (by ELISA) in fresh NECs of CRSwNP. Abbreviations: NECs=nasal epithelial cells, HC=healthy controls, NP=nasal polyps, MT=middle turbinate, and CRSwNP=chronic rhinosinusitis with nasal polyps.

Article Snippet: Tissue sections were blocked with 5% donkey serum and incubated with primary antibodies against 15LO1 (1: 200 dilution), CCL26 (1:100 dilution), and Cytokeratin-5 (CK5, 1: 200 dilution, Novus, USA).

Techniques: Enzyme-linked Immunosorbent Assay