alpha smooth muscle actin α sma antibody  (Cell Signaling Technology Inc)

Journal: International Journal of Molecular Medicine

Article Title: Orcinol glucoside ameliorates pulmonary fibrosis by suppressing hyaluronic acid synthesis and macrophage M2 polarization via targeting hyaluronic acid synthase 2

doi: 10.3892/ijmm.2026.5764

Figure Lengend Snippet: OG exerts an anti-pulmonary fibrosis effect by impairing myofibroblast-mediated macrophage M2 polarization. (A) Schematic workflow: Fibroblast treatment with OG during TGF-β1 stimulation, followed by conditioned medium collection and macrophage culture for downstream assays. (B) TGF-β1 release from macrophages measured by ELISA (n=3 for each group). (C) Flow cytometric analysis of CD206 and CD86 expression in macrophages cultured in conditioned medium. The bar graph quantifies the proportion of CD206 + CD86 − cells (M2-like macrophage subpopulation; n=3 for each group). (D) Animal experimental protocol. (E) A BLM-induced mouse model was established to assess the anti-fibrotic effects of OG. Histopathological alterations in lung tissues from different groups were assessed using H&E and Masson's trichrome staining. Scale bars: 500 μ m; magnification, ×40. (F) Western blotting analysis to measure the expression levels of fibrotic markers, COL1A1 and α-SMA, in lung tissues (n=4 for each group). (G) The levels of HA in mouse serum were quantified by ELISA. (H) The levels of TGF-β1 in mouse serum were quantified by ELISA (n=6 for each group). (I) OG concentrations in serum and lung tissues after 14-day oral administration. Left: serum concentrations of OG across treatment groups. Right: lung tissue concentrations of OG, expressed as a percentage of tissue weight (n=3 for each group). * P<0.05, ** P<0.01, *** P<0.001. OG, glucoside; BLM, bleomycin; COL1A1, Collagen type I α 1 chain; α-SMA, α-smooth muscle actin; H&E, hematoxylin and eosin.

Article Snippet: Antibodies for Collagen type I α 1 chain (COL1A1; cat. no. 72026; 1:1,000), Phospho-STAT6 (Tyr641; cat. no. 56554S; 1:1,000) and α-smooth muscle actin (α-SMA; cat. no. 19245; 1:1,000) were obtained from Cell Signaling Technology, Inc.

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

Journal: eBioMedicine

Article Title: USP25 regulates atherosclerosis by restricting RIPK1-mediated inflammatory responses

doi: 10.1016/j.ebiom.2026.106213

Figure Lengend Snippet: USP25 is downregulated in human and mouse atherosclerotic lesions. (A) Screening of USP family genes in human carotid artery plaques based on GSE100927 and GSE41571 datasets. In volcano plots, red and blue dots represent upregulated and downregulated USPs, respectively. (B) Transcriptional levels of selected USPs in aortas of ApoE −/− mice fed with ND and HFD were determined by qRT-PCR. ∗P < 0.05, ∗∗P < 0.01, ns, not significant, unpaired two-tailed Student's t- test (n = 5). (C) Representative immunoblots (upper panel) and densitometric quantification (lower panel) of USP25 in aortas of ND and HFD-fed ApoE −/− mice. ∗∗P < 0.01, unpaired two-tailed Student's t- test (n = 3). (D) Representative immunoblots (upper panel) and densitometric quantification (lower panel) of USP25 in mild and severe atherosclerotic lesions of human carotid arteries. ∗P < 0.05, unpaired two-tailed Student's t- test (n = 3). (E–G) Representative immunofluorescence staining of USP25 and F4/80 (E), α-SMA (F), as well as CD31 (G) in aortic roots. Scale bar: 15 μm. (H) Representative immunofluorescence staining of USP25 and F4/80 in aortas of ApoE −/− mice fed with HFD for indicated time. Scale bar: 100 μm ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001, two-way ANOVA with Bonferroni post hoc test (n = 5). (I) Representative immunofluorescence staining of USP25 and CD68 in carotid arteries from patients with mild and severe atherosclerosis. Scale bar: 100 μm.

Article Snippet: Antibodies against RIPK1 (Cat#: 3493, 1: 1000, RRID: AB_2305314 ), p-RIPK1 (Cat#: 31122, 1: 1000, RRID: AB_2799000 ), p-IKKα/β (Cat#: 2697, 1: 1000, RRID: AB_2079382 ), IKKα (Cat#: 2682, 1:1000, RRID: AB_331626 ), p-p65 (Cat#: 3033, 1: 1000 for Western bolt and 1: 200 for immunofluorescence, RRID: AB_331284 ), p65 (Cat#: 8242, 1: 1000, RRID: AB_10859369 ), p-p38 (Cat#: 4511, 1: 1000, RRID: AB_2139682 ), p38 (Cat#: 8690, 1: 1000, RRID: AB_10999090 ), p-JNK (Cat#: 4668S, 1:1000, RRID: AB_823588 ), JNK (Cat#: 9252S, 1:1000, RRID: AB_2250373 ), p-ERK1/2 (Cat#: 4370S, 1:1000, RRID: AB_2315112 ), ERK1/2 (Cat#: 4695S, 1:1000, RRID: AB_390779 ), GAPDH (Cat#: 5174, 1: 1000, RRID: AB_10622025 ), α-SMA (Cat#: 19245, 1: 200 for immunofluorescence, RRID: AB_2734735 ), and Ubiquitin (Cat#: 3936, 1: 1000, RRID: AB_331292 ) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Quantitative RT-PCR, Two Tailed Test, Western Blot, Immunofluorescence, Staining

Journal: eBioMedicine

Article Title: USP25 regulates atherosclerosis by restricting RIPK1-mediated inflammatory responses

doi: 10.1016/j.ebiom.2026.106213

Figure Lengend Snippet: USP25 deficiency exacerbates atherosclerosis in HFD-fed ApoE −/− mice. (A) Representative Oil Red O staining of aortas from HFD-fed Usp25 +/+ ApoE −/− and Usp25 −/− ApoE −/− mice. Scale bar: 5 mm. (B) Data show the percentage of plaque area/total vessel area. AA, aortic arch; TA, thoracic aorta. ∗∗∗P < 0.001, unpaired 2-tailed Student's t test (n = 6). (C) Representative H&E staining of aortic root sections. Scale bar: 200 μm. (D) Quantification of lesion area (left) and percentages of necrotic core (right). ∗∗P < 0.01, unpaired 2-tailed Student's t test (n = 10). (E) Representative Oil Red O (top), anti-F4/80 immunofluorescence (middle), and anti-α-SMA immunofluorescence (bottom) staining of aortic root sections. Scale bar: 100 μm. (F) Percentages of Oil Red O (top), F4/80 + (middle), and α-SMA + (bottom) area in aortic root sections. ∗∗P < 0.01 , ∗∗∗P < 0.001, unpaired 2-tailed Student's t test (n = 5). (G) Protein levels of IL-6 and TNF-α in atherosclerotic lesions were analysed using ELISA. ∗∗P < 0.01 , ∗∗∗P < 0.001, unpaired 2-tailed Student's t test (n = 6). (H) Transcriptional levels of indicated genes in aortas were determined by qRT-PCR. Target gene expression was normalized to the level of Actb mRNA. ∗P < 0.05, ∗∗P < 0.01, unpaired 2-tailed Student's t test (n = 5).

Article Snippet: Antibodies against RIPK1 (Cat#: 3493, 1: 1000, RRID: AB_2305314 ), p-RIPK1 (Cat#: 31122, 1: 1000, RRID: AB_2799000 ), p-IKKα/β (Cat#: 2697, 1: 1000, RRID: AB_2079382 ), IKKα (Cat#: 2682, 1:1000, RRID: AB_331626 ), p-p65 (Cat#: 3033, 1: 1000 for Western bolt and 1: 200 for immunofluorescence, RRID: AB_331284 ), p65 (Cat#: 8242, 1: 1000, RRID: AB_10859369 ), p-p38 (Cat#: 4511, 1: 1000, RRID: AB_2139682 ), p38 (Cat#: 8690, 1: 1000, RRID: AB_10999090 ), p-JNK (Cat#: 4668S, 1:1000, RRID: AB_823588 ), JNK (Cat#: 9252S, 1:1000, RRID: AB_2250373 ), p-ERK1/2 (Cat#: 4370S, 1:1000, RRID: AB_2315112 ), ERK1/2 (Cat#: 4695S, 1:1000, RRID: AB_390779 ), GAPDH (Cat#: 5174, 1: 1000, RRID: AB_10622025 ), α-SMA (Cat#: 19245, 1: 200 for immunofluorescence, RRID: AB_2734735 ), and Ubiquitin (Cat#: 3936, 1: 1000, RRID: AB_331292 ) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Staining, Immunofluorescence, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Targeted Gene Expression

Journal: eBioMedicine

Article Title: USP25 regulates atherosclerosis by restricting RIPK1-mediated inflammatory responses

doi: 10.1016/j.ebiom.2026.106213

Figure Lengend Snippet: Deficiency of USP25 in hematopoietic cells exacerbates atherosclerosis in HFD-fed ApoE −/− mice. (A) Representative Oil Red O staining of aortas from Usp25 +/+ → ApoE −/− and Usp25 −/− → ApoE −/− mice fed a HFD for 16 weeks. Scale bar: 5 mm. (B) Data show the percentage of plaque area/total vessel area. ∗∗∗P < 0.001, unpaired two-tailed Student's t test (n = 6). (C) H&E staining of aortic root sections. Scale bar: 200 μm. (D) Quantitative analysis of lesion area (left) and percentages of necrotic cores (right). ∗∗ P < 0.01, unpaired two-tailed Student's t test (n = 10). (E) Representative Oil Red O (top), anti-F4/80 immunofluorescence (middle), and anti-α-SMA immunofluorescence (bottom) staining of aortic root sections. Scale bar: 100 μm. (F) Percentages of Oil Red O (top), F4/80 + (middle), and α-SMA + (bottom) area in aortic root sections. ∗∗P < 0.01, unpaired two-tailed Student's t test (n = 5). (G) Protein levels of IL-6 and TNF-α in atherosclerotic lesions were analysed using ELISA. ∗∗P < 0.01 , ∗∗∗P < 0.001, unpaired 2-tailed Student's t test (n = 6). (H) Aortas were isolated from Usp25 +/+ → ApoE −/− and Usp25 −/− → ApoE −/− mice after 16 weeks of HFD feeding. Transcriptional levels of indicated genes in aortas were determined by qRT-PCR. ∗∗P < 0.01, ∗∗∗P < 0.001, unpaired two-tailed Student's t test (n = 6).

Article Snippet: Antibodies against RIPK1 (Cat#: 3493, 1: 1000, RRID: AB_2305314 ), p-RIPK1 (Cat#: 31122, 1: 1000, RRID: AB_2799000 ), p-IKKα/β (Cat#: 2697, 1: 1000, RRID: AB_2079382 ), IKKα (Cat#: 2682, 1:1000, RRID: AB_331626 ), p-p65 (Cat#: 3033, 1: 1000 for Western bolt and 1: 200 for immunofluorescence, RRID: AB_331284 ), p65 (Cat#: 8242, 1: 1000, RRID: AB_10859369 ), p-p38 (Cat#: 4511, 1: 1000, RRID: AB_2139682 ), p38 (Cat#: 8690, 1: 1000, RRID: AB_10999090 ), p-JNK (Cat#: 4668S, 1:1000, RRID: AB_823588 ), JNK (Cat#: 9252S, 1:1000, RRID: AB_2250373 ), p-ERK1/2 (Cat#: 4370S, 1:1000, RRID: AB_2315112 ), ERK1/2 (Cat#: 4695S, 1:1000, RRID: AB_390779 ), GAPDH (Cat#: 5174, 1: 1000, RRID: AB_10622025 ), α-SMA (Cat#: 19245, 1: 200 for immunofluorescence, RRID: AB_2734735 ), and Ubiquitin (Cat#: 3936, 1: 1000, RRID: AB_331292 ) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Staining, Two Tailed Test, Immunofluorescence, Enzyme-linked Immunosorbent Assay, Isolation, Quantitative RT-PCR

Journal: Inflammation Research

Article Title: Macrophage β-catenin-Ihh axis induces hepatic stellate cell activation and fibrosis in metabolic dysfunction-associated steatohepatitis

doi: 10.1007/s00011-026-02220-x

Figure Lengend Snippet: Deficiency of myeloid β-catenin ameliorates MASH-associated liver fibrosis. A Representative Sirius Red staining of liver sections was shown from β-catenin FL/FL and β-catenin M−KO mice. These mice were fed with either a normal chow diet (NCD) or a high-fat diet (HFD) for 28 weeks ( n = 5 samples/group). Scale bars were 100 μm. B Immunofluorescence images of α-SMA + myofibroblasts in liver sections were obtained from NCD-fed or HFD-fed β-catenin FL/FL and β-catenin M−KO mice ( n = 5 samples/group). Scale bars measured 50 μm. C Relative mRNA levels of fibrogenic genes such as Acta2 , Col1α1 , Col3α 1, and Timp 1 were detected in the livers of β-catenin FL/FL and β-catenin M−KO mice that were fed with NCD or HFD ( n = 5 samples/group). Data were expressed as the mean ± standard deviation (SD). ** P < 0.01

Article Snippet: The α-SMA immunofluorescence staining used rabbit monoclonal antibody α-SMA (Cat#: 19245, 1:200 dilution, Cell Signaling Technology).

Techniques: Staining, Immunofluorescence, Standard Deviation

Journal: Inflammation Research

Article Title: Macrophage β-catenin-Ihh axis induces hepatic stellate cell activation and fibrosis in metabolic dysfunction-associated steatohepatitis

doi: 10.1007/s00011-026-02220-x

Figure Lengend Snippet: Ihh signaling is crucial for macrophage β-catenin-mediated hepatic fibrosis in HFD-induced MASH. To restore Ihh expression in macrophages, mice were administered the Ihh plasmid via polyethylenimine nanoparticles, namely in vivo jetPEI-Man. A Representative Sirius Red staining images of liver sections from HFD-fed β-catenin FL/FL and β-catenin M−KO mice. These mice were either treated or not treated with jetPEI-Man-Ihh (jetPEI-Ihh) or jetPEI-Man-GFP (jetPEI-GFP) ( n = 5 mice per group). Scale bars are 100 μm. B Immunofluorescence images of α-SMA + myofibroblasts in liver sections from HFD-fed β-catenin FL/FL and β-catenin M−KO mice. These mice were either treated or not treated with jetPEI-Ihh or jetPEI-GFP ( n = 5 mice per group). Scale bars measure 50 μm. C Relative mRNA levels of fibrogenic genes such as Acta2 , Col1α1 , Col3α1 , and Timp1 in the livers of HFD-fed β-catenin FL/FL and β-catenin M−KO mice. These mice were either treated or not treated with jetPEI-Ihh or jetPEI-GFP ( n = 5 mice per group). Data are expressed as the mean ± standard deviation (SD). * P < 0.05, ** P < 0.01

Article Snippet: The α-SMA immunofluorescence staining used rabbit monoclonal antibody α-SMA (Cat#: 19245, 1:200 dilution, Cell Signaling Technology).

Techniques: Expressing, Plasmid Preparation, In Vivo, Staining, Immunofluorescence, Standard Deviation

Journal: Inflammation Research

Article Title: Macrophage β-catenin-Ihh axis induces hepatic stellate cell activation and fibrosis in metabolic dysfunction-associated steatohepatitis

doi: 10.1007/s00011-026-02220-x

Figure Lengend Snippet: Macrophage β-catenin-induced Ihh promotes HSC activation. A The schematic illustration of the co-culture model that utilizes bone marrow-derived macrophages (BMDMs) and primary murine hepatic stellate cells (HSCs). B The relative mRNA levels of fibrogenic genes such as Acta2 , Col1α1 , and Col3α1 in HSCs were measured after co-culturing with BMDMs from β-catenin FL/FL and β-catenin M−KO mice. C Immunofluorescence pictures of α-SMA expression in HSCs were obtained after co-culturing with BMDMs from β-catenin FL/FL and β-catenin M−KO mice. The scale bars represent 100 μm. D Immunofluorescence images of α-SMA expression in HSCs were captured after co-culturing with BMDMs transfected with the Lv-β-catenin plasmid or the control vector (Lv-GFP). The scale bars are 100 μm. E Immunofluorescence images of α-SMA expression in HSCs were taken after co-culturing with Lv-β-catenin-transfected BMDMs, either in the presence or absence of Ihh neutralizing antibody. The scale bars measure 100 μm. F The mRNA levels of Gli1 , Gli2 , and Gli3 in HSCs were determined after co-culturing with BMDMs from β-catenin FL/FL and β-catenin M−KO mice. The data are presented as the mean ± standard deviation (SD). * P < 0.05, ** P < 0.01

Article Snippet: The α-SMA immunofluorescence staining used rabbit monoclonal antibody α-SMA (Cat#: 19245, 1:200 dilution, Cell Signaling Technology).

Techniques: Activation Assay, Co-Culture Assay, Derivative Assay, Immunofluorescence, Expressing, Transfection, Plasmid Preparation, Control, Standard Deviation