tgf β1  (MedChemExpress)

Journal: Redox Biology

Article Title: Fibroblast circSamd4 promotes cardiac fibrosis via activating plasminogen activator inhibitor-1

doi: 10.1016/j.redox.2026.104018

Figure Lengend Snippet: CircSamd4 is enriched in cardiac fibroblasts and upregulated in human and mouse fibrotic hearts. (A) Validation of fibrosis and fibrotic gene expression in mouse hearts after TAC. (B) CircRNA array analysis of normal hearts and fibrotic hearts (n = 5). A total of 361 circRNAs were upregulated, whereas 143 were downregulated in fibrotic hearts. (|log 2 FC| ≥ 0.5, P < 0.05). (C) The top 10 overexpressed circRNAs in fibrotic hearts. (D) Differential analysis of the top three expressed circRNAs in normal and heart failure samples of humans (Normal, n = 3; Heart failure, n = 9). (E) qRT-PCR results showing circSamd4 upregulation in human failing hearts (n = 7–14). (F) Expression of the top three circRNAs in primary cardiomyocytes and mouse cardiac fibroblasts stimulated with TGF-β1 (n = 4). (G) Expression of circSamd4 in cardiac fibroblasts derived from the heart of the sham group and TAC group, respectively (n = 5). (H) Representative images and fluorescence intensity statistics of α-SMA in cardiac fibroblasts with or without TGF-β1 treatment for 24h (n = 12). (I) Expression of fibrotic genes Postn , Ccn2 , and circSamd4 in cardiac fibroblasts with or without TGF-β1 treatment (n = 6). (J) Sanger sequencing revealed the head-to-tail junction of circSamd4 . (K) The existence of circSamd4 was confirmed by RT-PCR and gel electrophoresis using convergent and divergent primers. Divergent primers amplified circSamd4 from cDNA but not from genomic DNA. (L) Comparison of the expression of circSamd4 in the cytoplasm and in the nucleus by RT‒qPCR, with Gapdh as an internal reference in the cytoplasm and U6 as an internal reference in the nucleus. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. Data A , G was analyzed by Mann–Whitney test, data in D , E , F , H and I was analyzed with Student's t -test.

Article Snippet: Before treatment, the CFs were serum-starved in serum-free DMEM for 4 h. Subsequently, they were treated with 10 ng/mL TGF-β1 (Sino Biological Inc. 80116-RNAH, China) for 24 h to induce activation.

Techniques: Biomarker Discovery, Gene Expression, Quantitative RT-PCR, Expressing, Derivative Assay, Fluorescence, Sequencing, Reverse Transcription Polymerase Chain Reaction, Nucleic Acid Electrophoresis, Amplification, Comparison, MANN-WHITNEY

Journal: Redox Biology

Article Title: Fibroblast circSamd4 promotes cardiac fibrosis via activating plasminogen activator inhibitor-1

doi: 10.1016/j.redox.2026.104018

Figure Lengend Snippet: circSamd4 regulates cardiac fibroblast proliferation and activation. (A) Expression of circSamd4 and its host gene Samd4 after transfection of si- circSamd4 in cardiac fibroblasts (n = 3). (B) The mRNA levels of the fibrotic marker genes Ccn2, Postn, and Acta2 in cardiac fibroblasts are reduced by circSamd4 knockdown (n = 3). (C) circSamd4 knockdown represses TGF-β1-induced α-SMA overexpression in cardiac fibroblasts (n = 3). (D) EdU staining showing circSamd4 knockdown represses TGF-β1-induced proliferation of cardiac fibroblasts (n = 12). (E) circSamd4 knockdown represses TGF-β1-induced deregulated contraction of cardiac fibroblasts (n = 3). (F) circSamd4 knockdown represses TGF-β1-induced migration of cardiac fibroblasts (n = 3). (G) Representative images staining of GFP and RT-qPCR results showing adenovirus-mediated circSamd4 expression in cardiac fibroblasts (n = 3). (H) circSamd4 overexpression promotes the mRNA levels of the fibrotic marker genes Ccn2 , Postn and Acta2 in TGF-β1-treated cardiac fibroblasts (n = 3). (I) circSamd4 overexpression upregulates protein levels of FN1, COL3A1, COL1A2, CTGF, and α-SMA in TGF-β1-treated cardiac fibroblasts. (J) circSamd4 overexpression upregulates protein levels of α-SMA in TGF-β1-treated cardiac fibroblasts (n = 3). ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. Data in A and G was analyzed with Student's t -test, data in B , C , D , E , F, H, I and J was analyzed by one-way ANOVA with Bonferroni post hoc multiple comparison test.

Article Snippet: Before treatment, the CFs were serum-starved in serum-free DMEM for 4 h. Subsequently, they were treated with 10 ng/mL TGF-β1 (Sino Biological Inc. 80116-RNAH, China) for 24 h to induce activation.

Techniques: Activation Assay, Expressing, Transfection, Marker, Knockdown, Over Expression, Staining, Migration, Quantitative RT-PCR, Comparison

Journal: Redox Biology

Article Title: Fibroblast circSamd4 promotes cardiac fibrosis via activating plasminogen activator inhibitor-1

doi: 10.1016/j.redox.2026.104018

Figure Lengend Snippet: Serpine1 is a downstream effector of circSamd4 in cardiac fibroblasts. (A) Bulk RNA sequencing was performed to analyze the transcriptome of cardiac fibroblasts (Control, TGF-β1_treated, TGF-β1_treated_plus_ circSamd4 _knockdown) (n = 3). (B) Trend analysis of the differentially expressed genes identified nine key clusters. (C) Heatmap of the top 20 genes in clusters #3 and #4. (D) Gene Ontology (GO) analysis results of genes in clusters #3 and #4 identify the enrichment of the coagulation-regulated pathway (negative regulation of the plasminogen activation). (E) Systemic analysis identifying the PAI-1 coding gene Serpine1 as a circSamd4 downstream effector. The schematic flow chart of the analysis of circSamd4 downstream target genes. Venn diagram showing the overlapping genes identified by the two methods. A total of 6 overlapping genes were identified, and Serpine1 ranks as the top one. (F) Construction of the circRNA-based ceRNA network based on the six overlapping genes identifies miR-1894-3p as a microRNA mediating the effects of circSamd4 on Serpine1 . (G) Single-cell RNA sequencing data (public databases) showed that SERPINE1 is primarily enriched in cardiac fibroblasts in failure human hearts. (H) Correlation analysis between SERPINE1 levels and fibrosis marker genes ( POSTN, CCN2, and FIBRONECTIN ) in human fibrotic hearts. (I) circSamd4 knockdown reduces the mRNA levels of Serpine1 in fibrotic hearts (n = 5–7). (J) circSamd4 knockdown reduces Serpine1 mRNA level in cardiac fibroblasts treated with TGF-β1 (n = 4). (K) circSamd4 knockdown reduces TGF-β1-induced expression of Serpine1- coding PAI-1 in cardiac fibroblasts (n = 3). ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. Data in I and J was analyzed by one-way ANOVA with Bonferroni post hoc multiple comparison test.

Article Snippet: Before treatment, the CFs were serum-starved in serum-free DMEM for 4 h. Subsequently, they were treated with 10 ng/mL TGF-β1 (Sino Biological Inc. 80116-RNAH, China) for 24 h to induce activation.

Techniques: RNA Sequencing, Control, Knockdown, Coagulation, Activation Assay, Marker, Expressing, Comparison

Journal: Redox Biology

Article Title: Fibroblast circSamd4 promotes cardiac fibrosis via activating plasminogen activator inhibitor-1

doi: 10.1016/j.redox.2026.104018

Figure Lengend Snippet: CircSamd4 regulates Serpine1 expression by sponging miR-1894-3p. (A) The expression of miR-1894-3p is reduced in fibrotic hearts induced by TAC (n = 5). (B) The expression of miR-1894-3p is reduced in activated fibroblasts induced by TGF-β1 (n = 3). (C) Validation of miR-1894-3p overexpression in cardiac fibroblasts transfected with mimics-miR-1894-3p (n = 3). (D) miR-1894-3p overexpression decreases the protein levels of the fibrotic markers FN1, POSTN, and CTGF in TGF-β1-treated cardiac fibroblasts (n = 3). (E) The expression of miR-1894-3p in cardiac fibroblasts is upregulated by circSamd4 knockdown (n = 3). (F) Prediction of the base pairing of circSamd4 and miR-1894-3p. (G) Verification of miR-1894-3p as a sponge target of circSamd4 via a dual-luciferase reporter assay (n = 4). (H) Relative mRNA expression of Serpine1 in cardiac fibroblasts is reduced by miR-1894-3p overexpression (n = 3). (I) The protein expression of PAI-1 in cardiac fibroblasts is reduced by miR-1894-3p overexpression (n = 3). (J) Prediction of the base pairing of miR-1894-3p and the Serpine1 3′-UTR. (K) Verification of Serpine1 as a target gene of miR-1894-3p via a dual-luciferase reporter assay (n = 4). (L) circSamd4 regulates Serpine1 expression in a miR-1894-3p-depedent manner in cardiac fibroblasts (n = 3). (M) Schematic showing circSamd4 serves as a sponge of miR-1894-3p to release its suppression of Serpine1 expression in cardiac fibroblasts. ns, P > 0.05, ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. Data in A, B, C, E, G, H, I and K was analyzed with Student's t -test, data in D and L was analyzed by one-way ANOVA with Bonferroni post hoc multiple comparison test.

Article Snippet: Before treatment, the CFs were serum-starved in serum-free DMEM for 4 h. Subsequently, they were treated with 10 ng/mL TGF-β1 (Sino Biological Inc. 80116-RNAH, China) for 24 h to induce activation.

Techniques: Expressing, Biomarker Discovery, Over Expression, Transfection, Knockdown, Luciferase, Reporter Assay, Comparison

Journal: Redox Biology

Article Title: Fibroblast circSamd4 promotes cardiac fibrosis via activating plasminogen activator inhibitor-1

doi: 10.1016/j.redox.2026.104018

Figure Lengend Snippet: circSamd4 regulates cardiac fibroblast activation via the miR-1894-3p- Serpine1 axis. (A–B) The efficiency of siRNA-mediated Serpine1 silencing in cardiac fibroblasts (n = 3). (C) The protein levels of fibrotic markers (COL3A1, COL1A2, and CTGF) were reduced by Serpine1 knockdown in TGF-β1-treated cardiac fibroblasts (n = 3). (D) Serpine1 knockdown reduces TGF-β1-induced α-SMA overexpression in cardiac fibroblasts (n = 3). (E) miR-1894-3p and Serpine1 are involved in circSamd4 regulation of COL3A1, COL1A2, and CTGF protein levels in cardiac fibroblasts (n = 3). (F) miR-1894-3p and Serpine1 are involved in circSamd4 regulation of α-SMA in cardiac fibroblasts (n = 3). (G) miR-1894-3p and Serpine1 are involved in circSamd4 regulation of the migration ability of cardiac fibroblasts (n = 3). (H) miR-1894-3p and Serpine1 are involved in circSamd4 regulation of contraction of cardiac fibroblasts (n = 3). ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. Data in A and B was analyzed with Student's t -test, data in C, D, F to H was analyzed by one-way ANOVA with Bonferroni post hoc multiple comparison test.

Article Snippet: Before treatment, the CFs were serum-starved in serum-free DMEM for 4 h. Subsequently, they were treated with 10 ng/mL TGF-β1 (Sino Biological Inc. 80116-RNAH, China) for 24 h to induce activation.

Techniques: Activation Assay, Knockdown, Over Expression, Migration, Comparison

Journal: Journal of Traditional and Complementary Medicine

Article Title: Dahuang Zhechong pill alleviates pulmonary fibrosis by inhibiting apoptosis and epithelial-mesenchymal transition of type II alveolar epithelial cells in vitro and in vivo

doi: 10.1016/j.jtcme.2025.06.002

Figure Lengend Snippet: DHZCP inhibits EMT via the TGF-β1/Hedgehog signalling pathway in RLE-6TN cells. (A) RLE-6TN cells were treated with DHZCP drug-containing serum of indicated concentrations for 48 h. Cell viability was assessed using the CCK-8 assay. For subsequent experiments, a serum concentration of 15 % containing the drug was selected. RLE-6TN cells were treated with the drug-containing serum (15 %) and/or SAG (0.25 nM), followed by stimulation with TGF-β1 (5 ng/ml). The cells were cultured for 48 h before further analysis. (B–E) Western blot analysis of Gli1, Shh, and Smo. (F–H) qRT-PCR analysis results show the mRNA expression levels of Gli1, Shh, and Smo in RLE-6TN cells. (I–K) Western blot analysis of E-cadherin and N-cadherin. Data are shown as mean ± SD, n = 3. Group details: the control group was treated with no TGF-β1 and 15 % drug-free serum; TGF-β1 group was treated with TGF-β1 and 15 % drug-free serum; DHZCP group was treated with TGF-β1 and 15 % DHZCP-containing serum; SAG group was treated with TGF-β1 and SAG; and DHZCP + SAG group was treated with TGF-β1, 15 % DHZCP medicated serum, and SAG. Statistical analysis was performed using one-way ANOVA with Tukey's multiple comparisons test. ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.005, and ∗∗∗∗p < 0.001 vs Control group. #p < 0.05; ##p < 0.01; ###p < 0.005, and ####p < 0.001 vs TGF-β1 group. $ p < 0.05 vs DHZCP group.

Article Snippet: DMOG (#HY-15893), TGF-β1 (#HY- P70543 ), and SAG (#HY-12848) were purchased from MCE, USA.

Techniques: CCK-8 Assay, Concentration Assay, Cell Culture, Western Blot, Quantitative RT-PCR, Expressing, Control

Journal: Biochemistry and Biophysics Reports

Article Title: LncRNA RMST knockout inhibits fibrosis by down-regulating Smad3 during mouse skin wound healing

doi: 10.1016/j.bbrep.2025.102386

Figure Lengend Snippet: Smad3-OE reversed the fibrosis suppression resulting from RMST-KO. (A) QPCR confirmed the upregulating effect of Smad3-OE lentivirus. (B) Representative blot of Smad3 under the effect of Smad3-OE and/or RMST-KO. (C) Statistical analysis of the immunoblot bands. (D) H&E and Masson staining of the wounds as indicated. (E–F) Statistical analysis of the (E) thickness and (F) area of neogenic fibrous tissue 21 dps. (G–I) qPCR detection of fibrosis markers and proteins in classical fibrosis pathways, including COL1, α-SMA, and TGF-β1. (J–M). Representative blot of COL1, α-SMA, and TGF-β1. (N–O) qPCR detection of inflammatory cytokines TNF-α and IL-1β. (P–R) Representative blot of TNF-α and IL-1β. Data were shown as mean ± SD, n = 6, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001.

Article Snippet: TGF-β1 (21898-1-AP) , 1:2000 , Proteintech.

Techniques: Western Blot, Staining

Journal: Biomolecules

Article Title: Notch Signaling Exacerbates Pulmonary Fibrosis by Regulating the Differentiation of CD4 + Tissue-Resident Memory T Cells

doi: 10.3390/biom16020328

Figure Lengend Snippet: Enhanced pro-inflammatory and pro-fibrotic function of lung CD4 + T RM Cells in PF. ( A ) Gene set enrichment analysis (GSEA) plot of the upregulated signature of T cell activation and cytokine production in lung CD4 + T RM compared to CD4 + non-T RM (using scRNA-seq data derived from GSE122960 ). ( B – I ) Male C57BL/6J mice (6–8 weeks old) were randomized to receive a single intratracheal dose of BLM (2 mg/kg) or sterile PBS. On day 14, lungs were collected for single-cell suspension preparation and flow cytometry analysis after euthanasia by anesthetic overdose. n = 8 in BLM group, and n = 4 in PBS group. ( B , C ) IL-17A expression in CD4 + CD69 + CD103 + T RM cells, CD4 + CD69 + CD103 − T cells, and CD4 + CD69 − CD103 − T cells from mice with BLM-induced PF (BLM group) was measured by flow cytometry. ( D , E ) Granzyme B (GZMB) expression in CD4 + CD69 + CD103 + T RM cells, CD4 + CD69 + CD103 − T cells, and CD4 + CD69 − CD103 − T cells from mice with BLM-induced PF (BLM group) was measured by flow cytometry. ( F , G ) IL-17A expression in CD4 + CD69 + CD103 + T RM cells from mice with or without BLM-induced PF was measured by flow cytometry. ( H , I ) GZMB expression in CD4 + CD69 + CD103 + T RM cells from mice with or without BLM-induced PF was measured by flow cytometry. ( J ) Schematic of CD4 + T RM and fibroblast co-culture. CD4 + T cells isolated from mouse spleens were stimulated with anti-CD3 (5 µg/mL), anti-CD28 (2 µg/mL), with or without TGF-β (10 ng/mL). After 3 days, TGF-β-induced CD4 + T RM and CD4 + non-T RM cells were co-cultured for 24 h with BLM (200 ng/mL)-pre-stimulated mouse pulmonary fibroblasts. ( K – M ) Fibrosis marker expressions were detected by qPCR. n = 4 for each group. Data were expressed as mean ± SEM. One-way ANOVA in panel ( C , E , K , L , M ), and Student’s t test in panel ( G , I ). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. ns: not significant.

Article Snippet: CD4 + T RM cell differentiation was induced by culturing activated cells in complete medium with 10 ng/mL TGF-β (Sino Biological, Beijing, China, Cat# 80116-RNAH-5) over a 3-day period, following previously reported methods [ ].

Techniques: Activation Assay, Derivative Assay, Sterility, Single Cell, Suspension, Flow Cytometry, Expressing, Co-Culture Assay, Isolation, Cell Culture, Marker

Journal: Biomolecules

Article Title: Notch Signaling Exacerbates Pulmonary Fibrosis by Regulating the Differentiation of CD4 + Tissue-Resident Memory T Cells

doi: 10.3390/biom16020328

Figure Lengend Snippet: Activation of Notch signaling in CD4 + T RM cells from fibrotic lungs. ( A , B ) scRNA-seq analysis of lung tissue from donors and PF patients ( GSE122960 ). ( A ) GSEA revealed enrichment of the positive regulation of Notch signaling in CD4 + T RM cells vs. CD4 + non-T RM cells ( p -value determined by clusterProfiler GSEA). ( B ) GSEA revealed enrichment in the Notch related-signaling in PF CD4 + T RM cells vs. donor CD4 + T RM cells ( p -value determined by clusterProfiler GSEA). ( C – H ) CD4 + T cells from control and BLM-treated mouse splenocytes were stimulated for 3 days with anti-CD3 (5 µg/mL), anti-CD28 (2 µg/mL), and TGF-β (10 ng/mL). n = 4 for each group. ( C , D ) Representative flow cytometry plots ( C ) and quantitative statistics ( D ) of Notch1 mean fluorescence intensity (MFI) in CD4 + T RM cells differentiated in vitro from splenic CD4 + T cells of the two mouse groups. ( E , F ) Representative flow cytometry plots ( E ) and quantitative statistics (F) of Notch1 MFI in in vitro-differentiated CD4 + CD69 + CD103 + T RM , CD4 + CD69 − CD103 − , and CD4 + CD69 + CD103 − T cell subsets from control mouse splenic CD4 + T cells. ( G , H ) Representative flow cytometry plots ( G ) and quantitative statistics ( H ) of Notch1 MFI in in vitro-differentiated CD4 + CD69 + CD103 + T RM , CD4 + CD69 − CD103 − , and CD4 + CD69 + CD103 − T cell subsets from BLM-induced PF mouse splenic CD4 + T cells. ( I , J ) Representative flow cytometry plots (I) and quantitative statistics ( J ) of Notch1 MFI in CD4 + CD69 + CD103 + T RM , CD4 + CD69 − CD103 − , and CD4 + CD69 + CD103 − T cell subsets from BLM-treated mouse lung tissues. n = 9 for each group. Data were expressed as mean ± SEM. Student’s t test in panel ( C ), and One-way ANOVA in panel ( F , H , J ). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. The grey line in panels ( C , E , G , I ) indicates the blank control in flow cytometry.

Article Snippet: CD4 + T RM cell differentiation was induced by culturing activated cells in complete medium with 10 ng/mL TGF-β (Sino Biological, Beijing, China, Cat# 80116-RNAH-5) over a 3-day period, following previously reported methods [ ].

Techniques: Activation Assay, Control, Flow Cytometry, Fluorescence, In Vitro