Journal: Heliyon

Article Title: Triptolide attenuates cardiac remodeling by inhibiting pyroptosis and EndMT via modulating USP14/Keap1/Nrf2 pathway

doi: 10.1016/j.heliyon.2024.e24010

Figure Lengend Snippet: TP inhibits EndMT in cardiac remodeling models. (A – D) Western blot analysis of CD31, VE-Cadherin and α-SMA protein expression in mouse heart tissue (n = 6). (E – H) Western blot analysis of Slug, Snail and Twist protein expression in mouse heart tissue (n = 6). (I – L) Western blot analysis of CD31, VE-Cadherin and α-SMA in EN (HCAEC) (n = 6). (M – P) Western blot analysis of Slug, Snail and Twist in EN (HCAEC) (n = 6). All the values are presented as mean ± SD. # P < 0.05, ## P < 0.01 compared with Sham or Ctl group; *P < 0.05, **P < 0.01 compared with TAC or TGF-β1 group. The original blots were provided in supplementary data as .

Article Snippet: In our study, recombinant human TGF‐β1 (10 ng/ml, MCE) was added to stimulate the cells for 6d to induce EndMT.

Techniques: Western Blot, Expressing

Journal: Heliyon

Article Title: Triptolide attenuates cardiac remodeling by inhibiting pyroptosis and EndMT via modulating USP14/Keap1/Nrf2 pathway

doi: 10.1016/j.heliyon.2024.e24010

Figure Lengend Snippet: TP inhibits EndMT via Nrf2 pathway. (A – B) Western blot analysis of Nrf2 in EN (HCAEC) ( n = 6). (C – D) Western blot analysis of Nrf2 expression in the presence or absence of Nrf2 siRNA in EN (HCAEC) ( n = 6 ) . (E – H) CD31, VE-Cadherin and α-SMA expression elicited by TGF-β1 in the presence or absence of Nrf2 siRNA in EN (HCAEC) (n = 6) (I – L) Slug, Snail and Twist expression elicited by TGF-β1 in the presence or absence of Nrf2 siRNA in EN (HCAEC) (n = 6). All the values are presented as mean ± SD. # P < 0.05, ## P < 0.01 compared with Ctl group; *P < 0.05, **P < 0.01 compared with siRNA-NC group or TGF-β1 group; & P < 0.05, && P < 0.01 compared with TGF-β1+TP + scRNA group. The original blots were provided in supplementary data as .

Article Snippet: In our study, recombinant human TGF‐β1 (10 ng/ml, MCE) was added to stimulate the cells for 6d to induce EndMT.

Techniques: Western Blot, Expressing

Journal: Journal of Asthma and Allergy

Article Title: Inhibition of LOXL2 Suppresses Nasal Mucosal Inflammation and Remodeling in Allergic Rhinitis

doi: 10.2147/JAA.S535065

Figure Lengend Snippet: Increased LOXL2 is associated with the expression of tissue remodeling markers in AR patients. ( A and B ) WB and ( C and D ) RT-qPCR for E-cadherin and α-SMA between the two groups (n=30); ( E and F ) The correlations between LOXL2 mRNA, E-cadherin and α-SMA mRNA in AR patients (n=30). **P<0.01, ***P<0.001, ****P<0.0001.

Article Snippet: Upon reaching 70% confluence, the cells were treated with varying concentrations of IL-4 recombinant protein, and a combination of 100nM LOXL2 inhibitor (2-Chloropyridin-4-yl methanamine hydrochloride) (HY-101771A, MCE, New Jersey, USA) and 40 ng/mL IL-4 recombinant protein (MCE, New Jersey, USA) to observe their effects on cell function and related signaling pathways.

Techniques: Expressing, Quantitative RT-PCR

Journal: Journal of Asthma and Allergy

Article Title: Inhibition of LOXL2 Suppresses Nasal Mucosal Inflammation and Remodeling in Allergic Rhinitis

doi: 10.2147/JAA.S535065

Figure Lengend Snippet: LOXL2 drives IL-4-induced remodeling via TGF-β1/Smad signaling. ( A ) WB showing LOX2 and EMT markers expression in nasal epithelial cells treated with different concentrations of IL-4 (n=6). ( B ) LOXL2 inhibitor reverses IL-4-induced EMT marker expression (n=6); ( C ) IL-4 dose-dependently activates Smad2/3 phosphorylation (n=6). ( D ) LOXL2 inhibition blocks IL-4-induced Smad2/3 phosphorylation (n=6). *P<0.05, **P<0.01, ****P<0.0001.

Article Snippet: Upon reaching 70% confluence, the cells were treated with varying concentrations of IL-4 recombinant protein, and a combination of 100nM LOXL2 inhibitor (2-Chloropyridin-4-yl methanamine hydrochloride) (HY-101771A, MCE, New Jersey, USA) and 40 ng/mL IL-4 recombinant protein (MCE, New Jersey, USA) to observe their effects on cell function and related signaling pathways.

Techniques: Expressing, Marker, Phospho-proteomics, Inhibition

Journal: Journal of Asthma and Allergy

Article Title: Inhibition of LOXL2 Suppresses Nasal Mucosal Inflammation and Remodeling in Allergic Rhinitis

doi: 10.2147/JAA.S535065

Figure Lengend Snippet: LOXL2 inhibitor alleviates nasal mucosal inflammation and tissue remodeling in murine AR model. ( A ) Serum IgE and OVA-sIgE levels (n=10). ( B ) IL-4 and IL-13 concentrations in nasal lavage fluid (n=10). ( C ) Representative HE and immunofluorescence images of nasal mucosal in control, AR and AR+inhibitor groups (n=5); ( D ) Relative fluorescence intensity of LOXL2 and EMT marker among the three groups (n=5); ( E ) Relative fluorescence intensity of LOXL2 and EMT marker among the three groups (n=5); *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.

Article Snippet: Upon reaching 70% confluence, the cells were treated with varying concentrations of IL-4 recombinant protein, and a combination of 100nM LOXL2 inhibitor (2-Chloropyridin-4-yl methanamine hydrochloride) (HY-101771A, MCE, New Jersey, USA) and 40 ng/mL IL-4 recombinant protein (MCE, New Jersey, USA) to observe their effects on cell function and related signaling pathways.

Techniques: Immunofluorescence, Control, Fluorescence, Marker

Journal: Biomolecules

Article Title: Activin A Secreted by Cancer-Associated Fibroblasts Reduces the Sensitivity of Breast Cancer Cells to Ixazomib via Inhibition of Proteasome Activity

doi: 10.3390/biom15091318

Figure Lengend Snippet: The screening of crucial secretory proteins for CAFs reduce to contribute the resistance of MCF7 cells to ixazomib. ( A ) A histogram showing the number of total and DEGs for CAFs not compared to CAFs reduce . ( B , C ) Top 15 Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analyses of DEGs. ( D ) GSEA of protein export signaling pathway between CAFs not and CAFs reduce . ( E ) The screening process of candidate DEGs between CAFs not and CAFs reduce that caused differences in ixazomib sensitivity of cancer cells ( left ). The intersection of four groups of differentially up-regulated genes ((CAF2# vs. CAF1#) ∩ (CAF2# vs. CAF10#) ∩ (CAF14# vs. CAF1#) ∩ (CAF14# vs. CAF10#)) ( Right ). ( F ) A heatmap of average expression trends of PLAU, INHBA and SCUBE3 genes in CAFs not and CAFs reduce . ( G ) INHBA expression levels in CAFs not and CAFs reduce were determined by RT-qPCR analysis. n = 3, ** p < 0.01 by Student’s t -test. ( H ) The protein secretion levels of Activin A in CAFs not and CAFs reduce were measured by ELISA. n = 3, ** p < 0.01 by Student’s t -test. ( I ) The UMAP plot of single-cell RNA sequencing (scRNA-Seq) data displaying CAF subclusters of breast tumor tissues, with color by sample type. ( J ) Aridge plot of INHBA expression in CAF subclusters based on scRNA-Seq data, with color by sample type. ( K ) Differences in INHBA expression in CAFs derived from each patient, with color coding (gray to blue) for expression of INHBA.

Article Snippet: Cells were cultured into 96-well plates at 2 × 10 3 /well and incubated in DMEM/F12 containing 10% FBS for 24 h. Then, after treatment with ixazomib (Selleck, Houston, TX, USA), Activin A (MCE, Monmouth Junction, NJ, USA), or its antagonist FST (MCE), the old medium was replaced by fresh medium containing 10% CCK8 solution (TransGen), following 2 h of incubation at 37 °C.

Techniques: Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Single Cell, RNA Sequencing, Derivative Assay

Journal: Biomolecules

Article Title: Activin A Secreted by Cancer-Associated Fibroblasts Reduces the Sensitivity of Breast Cancer Cells to Ixazomib via Inhibition of Proteasome Activity

doi: 10.3390/biom15091318

Figure Lengend Snippet: The blockade of CAF-secreted Activin A restores the sensitivity of MCF7 cells to ixazomib. ( A ) Comparison of effects of ixazomib on MCF7 cell viability when it was cultured in medium with or without recombinant Activin A. n = 5, ** p < 0.01 vs. control by one-way ANOVA. ( B ) Effects of FST on MCF7 relative cell viability (%). n = 4, ns, non-significant by Student’s t -test. ( C ) Effect of ixazomib treatment on MCF7 relative cell viability, when cells were cultured in regular medium, CAF-CM, or CAF-CM+FST. n = 3, ** p < 0.01 by one-way ANOVA. Regular medium: medium.

Article Snippet: Cells were cultured into 96-well plates at 2 × 10 3 /well and incubated in DMEM/F12 containing 10% FBS for 24 h. Then, after treatment with ixazomib (Selleck, Houston, TX, USA), Activin A (MCE, Monmouth Junction, NJ, USA), or its antagonist FST (MCE), the old medium was replaced by fresh medium containing 10% CCK8 solution (TransGen), following 2 h of incubation at 37 °C.

Techniques: Comparison, Cell Culture, Recombinant, Control

Journal: Biomolecules

Article Title: Activin A Secreted by Cancer-Associated Fibroblasts Reduces the Sensitivity of Breast Cancer Cells to Ixazomib via Inhibition of Proteasome Activity

doi: 10.3390/biom15091318

Figure Lengend Snippet: CAF reduce -secreted Activin A reduces the sensitivity of MCF7 cells to ixazomib by inhibiting proteasome activity. ( A ) Relative proteasome activity of MCF7 measured by proteasome activity ELISA kits. Cells were cultured in medium with or without Activin A. n = 3; ** p < 0.01 by Student’s t -test. ( B ) MCF7 cells were cultured in medium with or without Activin A, and the protein expression of p-ERK1/2, ERK1/2, p-AKT, and AKT was detected by Western blotting (original images can be found in ). n = 3, ns, non-significant, ** p < 0.01 by Student’s t -test. ( C ) MCF7 cells were cultured in normal medium, CAF not -CM (with or without FST), and CAF reduce -CM (with or without FST), respectively, and the protein expression of p-ERK1/2, ERK1/2, p-AKT, and AKT in the cells was detected by Western blotting (original images can be found in ). n = 3, ns, non-significant, ** p < 0.01 by one-way ANOVA. ( D ) Comparison of relative cell viability (%) of ixazomib on MCF7 when they were cultured in medium with or without ERK inhibitor (PD98059). n = 5, ** p < 0.01 by Student’s t -test. ( E ) Relative proteasome activity of MCF7 measured by proteasome activity ELISA kits. Cells were cultured in medium with or without ERK inhibitor ( PPD98059 ). n = 3, ** p < 0.01 by Student’s t -test.

Article Snippet: Cells were cultured into 96-well plates at 2 × 10 3 /well and incubated in DMEM/F12 containing 10% FBS for 24 h. Then, after treatment with ixazomib (Selleck, Houston, TX, USA), Activin A (MCE, Monmouth Junction, NJ, USA), or its antagonist FST (MCE), the old medium was replaced by fresh medium containing 10% CCK8 solution (TransGen), following 2 h of incubation at 37 °C.

Techniques: Activity Assay, Enzyme-linked Immunosorbent Assay, Cell Culture, Expressing, Western Blot, Comparison

Journal: Biomolecules

Article Title: Activin A Secreted by Cancer-Associated Fibroblasts Reduces the Sensitivity of Breast Cancer Cells to Ixazomib via Inhibition of Proteasome Activity

doi: 10.3390/biom15091318

Figure Lengend Snippet: Up-regulation of INHBA in CAFs reduces ixazomib sensitivity of MCF7 in vivo and in vitro. ( A ) INHBA expression levels in Vector-BJ and INHBA-BJ determined by RT-qPCR analysis. n = 3, ** p < 0.01 by Student’s t -test. ( B ) Protein secretion levels of INHBA in Vector-BJ and INHBA-BJ measured by ELISA. n = 3, ** p < 0.01 by Student’s t -test. ( C ) Effect of INHBA overexpression on BJ relative cell viability assayed by CCK-8 kit. n = 6, ** p < 0.01 by Student’s t -test. ( D ) Effect of INHBA overexpression on BJ cell migration evaluated by scratch assay. n = 4, ** p < 0.01 by Student’s t -test. ( E ) Effect of INHBA overexpression on relative cell viability of BJ cell induced by ixazomib. n = 4, ns, non-significant. ( F ) CCK-8 assay evaluating effect of ixazomib on MCF7 cell viability. MCF7 cells were cultured in conditioned medium from BJ cells overexpressing Activin A (INHBA-BJ-CM) or vector control medium (Vector-BJ-CM) ( Left ); or INHBA-BJ-CM either treated with neutralizing antibody FST (neutralization group) or untreated (non-neutralized group) ( Right ), to assess ixazomib’s effect on MCF7 cell activity under different conditions. n = 4, ** p < 0.01 by Student’s t -test. ( G ) A flow chart of animal experiments. Ixazomib was administered intraperitoneal at drug concentration of 7 mg/kg once weekly after average tumor volume reached size of 100 mm 3 . ( H ) Tumor growth curves after tumor formation at cell injection sites of nude mice before tumor volume reaches to 100 mm 3 . n = 6. ** p < 0.01 by Student’s t -test. ( I ) Curves of tumor volume changes in nude mice within 28 days after 1st administration of ixazomib. n = 6. ** p < 0.01 by Student’s t -test. ( J ) Photographs of extracted tumor and mice ( left ) and statistical analysis of mice tumor weights ( right ). n = 6, ** p < 0.01 by Student’s t -test. ( K ) Representative images of H&E and Ki-67 staining of xenograft tissue sections.

Article Snippet: Cells were cultured into 96-well plates at 2 × 10 3 /well and incubated in DMEM/F12 containing 10% FBS for 24 h. Then, after treatment with ixazomib (Selleck, Houston, TX, USA), Activin A (MCE, Monmouth Junction, NJ, USA), or its antagonist FST (MCE), the old medium was replaced by fresh medium containing 10% CCK8 solution (TransGen), following 2 h of incubation at 37 °C.

Techniques: In Vivo, In Vitro, Expressing, Plasmid Preparation, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Over Expression, CCK-8 Assay, Migration, Wound Healing Assay, Cell Culture, Control, Neutralization, Activity Assay, Concentration Assay, Injection, Staining