anti lamc2  (Proteintech)

Journal: International Journal of Chronic Obstructive Pulmonary Disease

Article Title: LAMC2 Drives Airway Remodeling in COPD via EMT Regulation Through the AKT Pathway

doi: 10.2147/COPD.S580964

Figure Lengend Snippet: Airway remodeling and LAMC2 upregulation are observed in both COPD patients and mouse models. ( A ) Representative Masson’s trichrome-stained lung sections from non-smokers (n=5), smokers without COPD (n=5), and COPD patients (n=5). Scale bar: 250μm. ( B ) Quantitative staining of Masson content in non-smoker (n=5), smoker (n=5), and COPD (n=5). ( C ) Representative Masson’s trichrome staining of lung sections from air-exposed control mice (n=5) and COPD model mice (n=5). Scale bar: 100μm. ( D ) Quantitative staining of Masson content in air-control (n=5) and COPD model mice (n=5). ( E ) LAMC2 expression levels in airway epithelial cells from non-smokers (n=12), smokers (n=12), and COPD patients (n=6) based on the GSE5058 dataset. ( F ) Representative airway LAMC2 immunohistochemical staining on lung sections of non-smoker (n=5), smoker (n=5), and COPD (n=5). Scale bar: 50μm. ( G ) Quantitative staining of LAMC2 intensity on airway epithelium of non-smoker (n=5), smoker (n=5), and COPD (n=5). ( H ) LAMC2 expression levels in lung tissues from air-control (n=30) and COPD model mice (n=34) based on the GSE87292 dataset. ( I ) Western blot and ( J ) quantitative analysis of LAMC2 protein expression in the lung tissue from mice from different groups (n=3 mice/group). ( K ) Representative airway LAMC2 immunohistochemical staining on lung sections of air-control (n=5) and COPD model mice (n=5).Scale bar: 100μm. ( L ) Quantitative staining of LAMC2 intensity on airway epithelium of air-control (n=5) and COPD model mice (n=5). Data presented as the means ± SD. * P < 0.05, ** P < 0.01.

Article Snippet: The membranes were blocked with 5% bovine serum albumin for 1 hour at room temperature and then incubated overnight at 4°C with the following primary antibodies: GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA), LAMC2 (1:1000; Proteintech, China), E-Cadherin (1:1000; Cell Signaling Technology), N-Cadherin (1:1000; Cell Signaling Technology), Fibronectin (1:1000; HUABIO, China), AKT (1:1000; Cell Signaling Technology), and phospho-AKT (p-AKT, 1:1000; Cell Signaling Technology).

Techniques: Staining, Control, Expressing, Immunohistochemical staining, Western Blot

Journal: International Journal of Chronic Obstructive Pulmonary Disease

Article Title: LAMC2 Drives Airway Remodeling in COPD via EMT Regulation Through the AKT Pathway

doi: 10.2147/COPD.S580964

Figure Lengend Snippet: Knockdown of LAMC2 attenuates airway remodeling and EMT in a COPD model. ( A ) Schematic overview of the animal experimental design and grouping. ( B ) Representative Masson staining on mice lung sections from different groups (n = 5 per group). ( C ) Quantitative staining of Masson content in different groups (n = 5 per group). ( D ) Representative airway Fibronectin immunohistochemical staining and ( E ) quantitative staining of Fibronectin intensity on lung sections of different groups (n = 5 per group). ( F ) Representative airway N-cadherin immunohistochemical staining and ( G ) quantitative staining of N-cadherin intensity on lung sections of different groups (n=5 per group). ( H ) Representative airway E-cadherin immunohistochemical staining and ( I ) quantitative staining of E-cadherin intensity on lung sections of different groups (n=5 per group). ( J – M ) Western blot analysis and quantification of EMT-related proteins (Fibronectin, N-cadherin, and E-cadherin) in lung tissues from the different treatment groups. * P < 0.05, ** P < 0.01, *** P < 0.001. Scale bars: 100μm.

Article Snippet: The membranes were blocked with 5% bovine serum albumin for 1 hour at room temperature and then incubated overnight at 4°C with the following primary antibodies: GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA), LAMC2 (1:1000; Proteintech, China), E-Cadherin (1:1000; Cell Signaling Technology), N-Cadherin (1:1000; Cell Signaling Technology), Fibronectin (1:1000; HUABIO, China), AKT (1:1000; Cell Signaling Technology), and phospho-AKT (p-AKT, 1:1000; Cell Signaling Technology).

Techniques: Knockdown, Staining, Immunohistochemical staining, Western Blot

Journal: International Journal of Chronic Obstructive Pulmonary Disease

Article Title: LAMC2 Drives Airway Remodeling in COPD via EMT Regulation Through the AKT Pathway

doi: 10.2147/COPD.S580964

Figure Lengend Snippet: LAMC2 expression is upregulated in bronchial epithelial cells following TGF-β1 stimulation. ( A and B ) Volcano plots illustrating DEGs from ( A ) GSE104908 and ( B ) GSE40374 datasets, with corresponding LAMC2 expression levels. Significantly upregulated genes (red) and downregulated genes (blue) are indicated (fold change >1.5, adjusted P < 0.05). ( C ) The mRNA levels of Fibronectin, N-cadherin, E-cadherin, and LAMC2 after treatment with different concentrations of TGF-β1 for 24 hours. ( D ) Western blot and ( E – H ) the relative protein level of Fibronectin, N-cadherin, E-cadherin, and LAMC2 after treatment with different concentrations of TGF-β1 for 24 hours. The data are the mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The membranes were blocked with 5% bovine serum albumin for 1 hour at room temperature and then incubated overnight at 4°C with the following primary antibodies: GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA), LAMC2 (1:1000; Proteintech, China), E-Cadherin (1:1000; Cell Signaling Technology), N-Cadherin (1:1000; Cell Signaling Technology), Fibronectin (1:1000; HUABIO, China), AKT (1:1000; Cell Signaling Technology), and phospho-AKT (p-AKT, 1:1000; Cell Signaling Technology).

Techniques: Expressing, Western Blot

Journal: International Journal of Chronic Obstructive Pulmonary Disease

Article Title: LAMC2 Drives Airway Remodeling in COPD via EMT Regulation Through the AKT Pathway

doi: 10.2147/COPD.S580964

Figure Lengend Snippet: Knockdown of LAMC2 reverses TGF-β1-induced EMT marker expression and inhibits 16HBE cell migration. ( A ) LAMC2 mRNA expression levels following transfection with different siLAMC2 constructs. ( B ) mRNA expression of Fibronectin, N-cadherin, and E-cadherin after siLAMC2 transfection and subsequent TGF-β1 treatment. ( C ) Western blot and ( D – F ) the relative protein level of Fibronectin, N-cadherin, E-cadherin, and LAMC2 after siLAMC2 transfection and TGF-β1 treatment. ( G ) Representative images of wound healing and transwell migration assays under different treatment conditions. Scale bar: 20 μm. ( H ) Quantification of migration rate (%) from wound healing assay and ( I ) Cell migration numbers counted from transwell assays. The data are the mean ± SD. ** P < 0.01, *** P < 0.001.

Article Snippet: The membranes were blocked with 5% bovine serum albumin for 1 hour at room temperature and then incubated overnight at 4°C with the following primary antibodies: GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA), LAMC2 (1:1000; Proteintech, China), E-Cadherin (1:1000; Cell Signaling Technology), N-Cadherin (1:1000; Cell Signaling Technology), Fibronectin (1:1000; HUABIO, China), AKT (1:1000; Cell Signaling Technology), and phospho-AKT (p-AKT, 1:1000; Cell Signaling Technology).

Techniques: Knockdown, Marker, Expressing, Migration, Transfection, Construct, Western Blot, Wound Healing Assay

Journal: International Journal of Chronic Obstructive Pulmonary Disease

Article Title: LAMC2 Drives Airway Remodeling in COPD via EMT Regulation Through the AKT Pathway

doi: 10.2147/COPD.S580964

Figure Lengend Snippet: Overexpression of LAMC2 promotes TGF-β1-induced EMT and enhances 16HBE cell migration. ( A ) LAMC2 mRNA expression levels following transfection with LAMC2 plasmid and TGF-β1treatment. ( B ) mRNA expression of Fibronectin, N-cadherin, and E-cadherin after LAMC2 plasmid vector transfection and subsequent TGF-β1 treatment. ( C ) Western blot and ( D – F ) the relative protein level of Fibronectin, N-cadherin, E-cadherin, and LAMC2 after LAMC2 plasmid vector transfection and TGF-β1 treatment. ( G ) Representative images of wound healing and transwell migration assays under different treatment conditions. Scale bar: 20 μm. ( H ) Quantification of migration rate (%) from wound healing assay and ( I ) Cell migration numbers counted from transwell assays. The data are the mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The membranes were blocked with 5% bovine serum albumin for 1 hour at room temperature and then incubated overnight at 4°C with the following primary antibodies: GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA), LAMC2 (1:1000; Proteintech, China), E-Cadherin (1:1000; Cell Signaling Technology), N-Cadherin (1:1000; Cell Signaling Technology), Fibronectin (1:1000; HUABIO, China), AKT (1:1000; Cell Signaling Technology), and phospho-AKT (p-AKT, 1:1000; Cell Signaling Technology).

Techniques: Over Expression, Migration, Expressing, Transfection, Plasmid Preparation, Western Blot, Wound Healing Assay

Journal: International Journal of Chronic Obstructive Pulmonary Disease

Article Title: LAMC2 Drives Airway Remodeling in COPD via EMT Regulation Through the AKT Pathway

doi: 10.2147/COPD.S580964

Figure Lengend Snippet: LAMC2 facilitates EMT in 16HBE cells through activation of the AKT signaling pathway. ( A ) GO enrichment analysis of down-regulated DEGs from RNA-seq data, showing the top three most significantly enriched terms in each category: Biological Process (BP), Cellular Component (CC), and Molecular Function (MF) (adjusted P < 0.05). Terms are ranked by gene count. ( B ) KEGG pathway analysis of down-regulated DEGs. Statistically significant pathways (adjusted P < 0.05) are visualized in a dot plot. ( C ) Western Blot and ( D ) Immunofluorescence detection, and ( E – G ) the relative protein level of AKT and p-AKT after si-NC/LAMC2 transfection and TGF-β1 treatment. ( H ) Western Blot and ( I ) Immunofluorescence detection, and ( J – L ) the relative protein level of AKT and p-AKT after LAMC2 plasmid vector transfection and TGF-β1 treatment. Scale bar: 50 μm. The data are the mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The membranes were blocked with 5% bovine serum albumin for 1 hour at room temperature and then incubated overnight at 4°C with the following primary antibodies: GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA), LAMC2 (1:1000; Proteintech, China), E-Cadherin (1:1000; Cell Signaling Technology), N-Cadherin (1:1000; Cell Signaling Technology), Fibronectin (1:1000; HUABIO, China), AKT (1:1000; Cell Signaling Technology), and phospho-AKT (p-AKT, 1:1000; Cell Signaling Technology).

Techniques: Activation Assay, RNA Sequencing, Western Blot, Immunofluorescence, Transfection, Plasmid Preparation

Journal: bioRxiv

Article Title: Cell-of-Origin, not Oncogenic Effect, Determines Desmoplastic Immune Exclusion in KRAS-Driven Liver Cancer

doi: 10.64898/2026.03.24.711280

Figure Lengend Snippet: Multi-omics integration identifies LAMC2, uPA and BSSP-4 as CCA-specific secreted factors. (A) Schematic of the experimental workflow to collect material for secretome and proteome analysis. (B) Venn diagram showing the overlap of PK-induced transcriptional and secretory responses in chol-orgs. Intersections include PK-upregulated genes in chol-orgs from RNA-seq (chol-PK vs chol-WT, bottom-left), PK-upregulated proteins in chol-orgs from secretome (CCM, bottom-right), genes whose PK-induced expression is stronger in interaction effects (interaction contrast, upper-right), and proteins predicted to be secreted according to the Human Protein Atlas (upper-left). (C) Bubble plot showing cross-dataset support for candidate PK-associated genes identified by the integrative analysis in (B). Rows indicate 15 selected genes from (B), and columns indicate the indicated in-house murine result and public human datasets. Dot color encodes z-scored log2 FC (z(LFC). Filled circles indicate significant differences (padj < 0.1) and open circles indicate non-significant results. Dot size reflects statistical strength-log10(padj).

Article Snippet: The antibodies used in this study were: Lamc2 (Proteintech, #19698-1-AP), uPA (Proteintech, #17968-1-AP), BSSP4 (ThermoFisher Scientific, #PA5-47245), and rabbit-IgG (Millipore, #pp64).

Techniques: Biomarker Discovery, RNA Sequencing, Expressing

Journal: bioRxiv

Article Title: Cell-of-Origin, not Oncogenic Effect, Determines Desmoplastic Immune Exclusion in KRAS-Driven Liver Cancer

doi: 10.64898/2026.03.24.711280

Figure Lengend Snippet: LAMC2 and uPA affect HSC activation and impair CCA formation in vivo . (A) Schematic overview of the in vitro fibroblast activation assay, where murine HSCs were treated with CCM from chol-PK and neutralizing antibodies. (B) Scatter plot indicating mRNA expression of Acta2 and Col1a1 in mHSCs treated with Chol-PK CCM and indicated neutralizing antibodies or control IgG. (n=5 from 2 independent experiments). (C) Schematic of scratch assay with mHSCs treated with low FBS (2%) media supplemented with respective recombinant protein. (D) Plot depicting mean relative wound coverage upon scratch injury over time with indicated recombinant protein at different dosage (n≥17 from 3 experiments). (E) Schematic of in vivo validation model: Chol-PK organoids were edited by CRISPR/Cas9 to generate Prss22 -, Lamc2 -, or Plau -deleted lines, which were orthotopically implanted into the liver and harvested at the same time point. (F) Macroscopic images of liver tumors 7 weeks after implantation of indicated lines. (G) Bar plot with respective tumor penetrance per line. (H) Representative histopathology images of liver tumors obtained upon orthotopic implantation of chol-PK and indicated KO lines. Stains as indicated. Red dotted line demarcates the boundary between non-tumor liver (N) and tumor (T). All scale bars are 100 μm. (I) Scatter plot depicting the total CD3 + and CD8 + T cell density per area per tumor for the indicated lines.

Article Snippet: The antibodies used in this study were: Lamc2 (Proteintech, #19698-1-AP), uPA (Proteintech, #17968-1-AP), BSSP4 (ThermoFisher Scientific, #PA5-47245), and rabbit-IgG (Millipore, #pp64).

Techniques: Activation Assay, In Vivo, In Vitro, Expressing, Control, Wound Healing Assay, Recombinant, Biomarker Discovery, CRISPR, Histopathology

Journal: bioRxiv

Article Title: Integrin beta 1 and mannose receptor 2 are involved in the antifungal activity of bronchial epithelial cells through Aspergillus fumigatus lectin FleA interactions

doi: 10.64898/2026.02.26.708144

Figure Lengend Snippet: ( A ) mRNA Laminin B3 and C2 expression after BECs transfection with siRNA-LAMB3 or siRNA-LAMC2. Data are expressed as percentage of control (siCtl) and presented as mean ± SD, n = 7 independent experiments. * p < 0.05, ** p < 0.01 (ANOVA test). ( B) Microscopic score of A. fumigatus filament formation after infection of BECs transfected with siRNA-LAMB3 and siRNA-LAMC2. BECs were transfected 48 h then infected for 15 h. Data are expressed as percentage of control (siCtl) and presented as mean ± SD, n = 10 independent experiments. *** p < 0.001 (ANOVA test). ( C ) Measure of galactomannan released in supernatant of infected BECs transfected with siRNA-LAMB3 or siRNA-LAMC2. Data are expressed as percentage of control (siCtl) and presented as mean ± SD, n= 8 independent experiments. * p < 0.05 (Student’s t-test). ( D ) Measure of galactomannan released in supernatant of BECs infected with A. fumigatus in presence of different concentrations of exogenous laminin-332. Data are expressed as percentage of control (0 µg/mL laminin-332) and presented as mean ± SEM, n = 3 independent experiments. * p < 0.05 (ANOVA test followed by Bonferroni’s multiple comparison test). (E) Conidia adhesion on BECs cells after 1 h of pre-incubation with laminin-332 (5 µg/mL) or FleA lectin (2 µM) and infection with 300 conidia/well during 2 h or 4 h. Control: Conidia adhesion to non-treated BECs. Data are expressed as % of control and presented as mean ± SD, n = 4 independent experiments. ** p < 0.01, *** p < 0.001 (ANOVA test followed by Bonferroni’s multiple comparison test).

Article Snippet: Reverse transcriptionwas was performed from 1μg RNA using a 2X RT Master Mix on a ProFlex thermocycler (Applied Biosystems, ThermoFischer), with the following program: 25°C for 10 min; 37°C for 2 h and 85°C for 5 min. Quantitative PCR was carried out using 2 μL of cDNA, 5 μL 2× TaqMan mix (Low ROX), 2.5 μL of RNase-free H 2 0 and 0.5 μL of the TaqMan probe: GAPDH (Hs02786624_g1); ITGB1 (Hs01127536_m1); MRC2 (Hs00195862_m1); LAMP1 (Hs00931461_m1); LAMB3 (Hs00165078_m1); LAMC2 (Hs01043717_m1) (Applied Biosystem).

Techniques: Expressing, Transfection, Control, Infection, Comparison, Incubation

Journal: Journal of Cell Communication and Signaling

Article Title: Structural maintenance of chromosome protein 1A exacerbates liver fibrosis by enhancing hepatic stellate cell activation and extracellular matrix synthesis via laminin subunit gamma 2 activation

doi: 10.1002/ccs3.70067

Figure Lengend Snippet: LAMC2 is significantly overexpressed in the liver tissues of cirrhotic mice. (A) Heatmap of differentially expressed genes in the GSE77627 dataset. (B) Heatmap of differentially expressed genes in the GSE33258 dataset. (C) The intersection of differentially expressed genes in the two GEO datasets and hub genes related to liver cirrhosis in the GeneCards database. (D) KEGG enrichment analysis of 373 intersecting genes. The expression of the profibrotic genes ACTA2 (E), TIMP1 (F), and PDGFRA (G) in the liver tissues of mice treated with DMSO (normal) or CCl4 (model), as examined by RT‐qPCR ( n = 3). (H–J) Western blot analysis and relative protein expression levels of LRAT and GFAP in the liver tissues of mice ( n = 3). (K) Analysis of fibrosis in the mouse liver lobule by HE staining ( n = 5). (L, M) Analysis of fibrosis in the mouse liver lobule by Masson's trichrome staining ( n = 5). (N, O) The LAMC2‐positive area in mouse liver samples was assessed by IHC ( n = 5). (P) LAMC2 mRNA expression in mouse liver samples was assessed by RT‐qPCR ( n = 3). Graphical data are presented as means ± SDs. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. An unpaired t ‐test was used for statistical analysis. DMSO, dimethyl sulfoxide; IHC, immunohistochemistry; LAMC2, laminin subunit gamma 2; PDGFRA, platelet‐derived growth factor receptor alpha; RT‐qPCR, reverse transcription quantitative PCR; TIMP1, TIMP metallopeptidase inhibitor 1.

Article Snippet: Tissues were incubated with antibodies against LAMC2 (1:100, PK08311, Abmart Shanghai Co., Ltd.), α‐smooth muscle actin (α‐SMA, 1:500, GTX100034, GeneTex, Inc.), and collagen I (1:100, GTX20292, GeneTex) overnight at 4°C and with goat anti‐rabbit IgG (1:2500, ab97051, Abcam) at 37°C for 1 h. Then, immunoassays were performed using a DAB horseradish peroxidase color development kit (P0202, Beyotime Biotechnology Co., Ltd.).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Staining, Immunohistochemistry, Derivative Assay, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Journal of Cell Communication and Signaling

Article Title: Structural maintenance of chromosome protein 1A exacerbates liver fibrosis by enhancing hepatic stellate cell activation and extracellular matrix synthesis via laminin subunit gamma 2 activation

doi: 10.1002/ccs3.70067

Figure Lengend Snippet: Poor expression of LAMC2 inhibits liver fibrosis and suppresses HSC activation in vivo. (A) Analysis of fibrosis in mouse liver tissues by HE staining ( n = 5). (B, C) Analysis of fibrosis in mouse liver tissues by Masson's trichrome staining ( n = 5). (D–F) The collagen I‐ and α‐SMA‐positive areas in mouse liver tissues were identified by IHC staining ( n = 5). (G) The mRNA expression of LAMC2 in the liver tissues of mice injected with sh‐NC or sh‐LAMC2 was examined via RT‐qPCR ( n = 3). The levels of the inflammatory factors TNF‐α (H), IL‐1β (I), and IL‐6 (J) in serum were measured using ELISAs ( n = 5). (K‒M) The protein expression of LRAT and GFAP, which are markers of HSC activation, in mouse liver tissues was analyzed by Western blot ( n = 3). Graphical data are presented as means ± SDs. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. An unpaired t ‐test was used for statistical analysis. ELISAs, enzyme‐linked immunosorbent assays; HSC, hepatic stellate cell; IHC, immunohistochemistry; LAMC2, laminin subunit gamma 2; RT‐qPCR, reverse transcription quantitative PCR.

Article Snippet: Tissues were incubated with antibodies against LAMC2 (1:100, PK08311, Abmart Shanghai Co., Ltd.), α‐smooth muscle actin (α‐SMA, 1:500, GTX100034, GeneTex, Inc.), and collagen I (1:100, GTX20292, GeneTex) overnight at 4°C and with goat anti‐rabbit IgG (1:2500, ab97051, Abcam) at 37°C for 1 h. Then, immunoassays were performed using a DAB horseradish peroxidase color development kit (P0202, Beyotime Biotechnology Co., Ltd.).

Techniques: Expressing, Activation Assay, In Vivo, Staining, Immunohistochemistry, Injection, Quantitative RT-PCR, Western Blot, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Journal of Cell Communication and Signaling

Article Title: Structural maintenance of chromosome protein 1A exacerbates liver fibrosis by enhancing hepatic stellate cell activation and extracellular matrix synthesis via laminin subunit gamma 2 activation

doi: 10.1002/ccs3.70067

Figure Lengend Snippet: PI3K/Akt signaling activation exacerbates liver fibrosis in mice with sh‐LAMC2. (A–C) The total protein content and phosphorylation of PI3K and Akt in the liver tissues of normal and model mice and mice treated with sh‐NC or sh‐LAMC2 were examined using Western blot assays ( n = 3). (D–H) The extent of PI3K and Akt phosphorylation and the protein expression of LRAT and GFAP in the liver tissues of mice treated with sh‐LAMC2 + DMSO or sh‐LAMC2 + recilisib were examined using Western blot assays ( n = 3). (I) Analysis of fibrosis in mouse liver tissues via HE staining ( n = 5). (J–L) The collagen I‐ and α‐SMA‐positive areas in mouse liver tissues were identified by IHC staining ( n = 5). (M, N) Fibrosis in mouse liver tissues was analyzed via Masson's trichrome staining ( n = 5). The levels of the inflammatory factors TNF‐α (O), IL‐1β (P), and IL‐6 (Q) in serum were measured using ELISAs ( n = 5). Graphical data are presented as means ± SDs. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. An unpaired t ‐test or one‐way ANOVA was used for statistical analysis. ANOVA, analysis of variance; DMSO, dimethyl sulfoxide; ELISAs, enzyme‐linked immunosorbent assays; IHC, immunohistochemistry.

Article Snippet: Tissues were incubated with antibodies against LAMC2 (1:100, PK08311, Abmart Shanghai Co., Ltd.), α‐smooth muscle actin (α‐SMA, 1:500, GTX100034, GeneTex, Inc.), and collagen I (1:100, GTX20292, GeneTex) overnight at 4°C and with goat anti‐rabbit IgG (1:2500, ab97051, Abcam) at 37°C for 1 h. Then, immunoassays were performed using a DAB horseradish peroxidase color development kit (P0202, Beyotime Biotechnology Co., Ltd.).

Techniques: Activation Assay, Phospho-proteomics, Western Blot, Expressing, Staining, Immunohistochemistry

Journal: Journal of Cell Communication and Signaling

Article Title: Structural maintenance of chromosome protein 1A exacerbates liver fibrosis by enhancing hepatic stellate cell activation and extracellular matrix synthesis via laminin subunit gamma 2 activation

doi: 10.1002/ccs3.70067

Figure Lengend Snippet: The transcription factor SMC1A promotes LAMC2 expression to induce HSC activation in vitro. (A) The transcription factors regulating LAMC2 in the hTFtarget database were intersected with differentially expressed genes in the two GEO datasets. (B) The binding peaks of SMC1A in the promoter region of LAMC2 were analyzed using the ChIP‐seq database. (C, D) The mRNA expression of SMC1A and LAMC2 in TGF‐β1‐stimulated LX‐2 cells and those preinfected with sh‐NC or sh‐SMC1A was examined via RT‐qPCR ( n = 3). The expression of the profibrotic genes PDGFRA (E), TIMP1 (F), and ACTA2 (G) in LX‐2 cells was examined via RT‐qPCR ( n = 3). The localization (H) and expression (I) of SMC1A in LX‐2 cells were examined via immunofluorescence experiments ( n = 3). (J) The enrichment of SMC1A in the LAMC2 promoter region in LX‐2 cells was examined via ChIP‐PCR; immunoprecipitation was performed using an anti‐SMC1A antibody ( n = 3). (K) Dual‐luciferase assay of the promoter transcriptional activity of LAMC2 in LX‐2 cells in the presence of sh‐SMC1A ( n = 3). Representative immunofluorescence images and quantification of collagen I (L, M) and α‐SMA (N, O) in LX‐2 cells ( n = 3). Graphical data are presented as means ± SDs. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. An unpaired t ‐test or one‐way ANOVA was used for statistical analysis. ACTA2, actin alpha 2, smooth muscle; ANOVA, analysis of variance; ChIP, chromatin immunoprecipitation; HSC, hepatic stellate cell; LAMC2, laminin subunit gamma 2; PDGFRA, platelet‐derived growth factor receptor alpha; RT‐qPCR, reverse transcription quantitative PCR; SMC1A, structural maintenance of chromosome protein 1A; TIMP1, TIMP metallopeptidase inhibitor 1.

Article Snippet: Tissues were incubated with antibodies against LAMC2 (1:100, PK08311, Abmart Shanghai Co., Ltd.), α‐smooth muscle actin (α‐SMA, 1:500, GTX100034, GeneTex, Inc.), and collagen I (1:100, GTX20292, GeneTex) overnight at 4°C and with goat anti‐rabbit IgG (1:2500, ab97051, Abcam) at 37°C for 1 h. Then, immunoassays were performed using a DAB horseradish peroxidase color development kit (P0202, Beyotime Biotechnology Co., Ltd.).

Techniques: Expressing, Activation Assay, In Vitro, Binding Assay, ChIP-sequencing, Quantitative RT-PCR, Immunofluorescence, Immunoprecipitation, Luciferase, Activity Assay, Chromatin Immunoprecipitation, Derivative Assay, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Journal of Cell Communication and Signaling

Article Title: Structural maintenance of chromosome protein 1A exacerbates liver fibrosis by enhancing hepatic stellate cell activation and extracellular matrix synthesis via laminin subunit gamma 2 activation

doi: 10.1002/ccs3.70067

Figure Lengend Snippet: LAMC2 overexpression supports the profibrotic phenotype of HSCs in the presence of sh‐SMC1A. (A) The mRNA expression of LAMC2 in LX‐2 cells infected with sh‐SMC1A + NC‐oe or sh‐SMC1A + LAMC2‐oe was examined via RT‐qPCR ( n = 3). The expression of the profibrotic genes ACTA2 (B), TIMP1 (C), and PDGFRA (D) in LX‐2 cells was examined via RT‐qPCR ( n = 3). (E–G) The total protein content and phosphorylation of PI3K and Akt in LX‐2 cells were examined using Western blot assays ( n = 3). (H–J) Representative immunofluorescence images (H) and quantification of collagen I (I) and α‐SMA (J) in LX‐2 cells ( n = 3). (K‒M) The protein expression of LRAT and GFAP, which are markers of HSC activation, in LX‐2 cells was analyzed by Western blot assays ( n = 3). Graphical data are presented as means ± SDs. * p < 0.05, ** p < 0.01, and *** p < 0.001. An unpaired t ‐test or one‐way ANOVA was used for statistical analysis. ACTA2, actin alpha 2, smooth muscle; ANOVA, analysis of variance; HSCs, hepatic stellate cells; LAMC2, laminin subunit gamma 2; PDGFRA, platelet‐derived growth factor receptor alpha; RT‐qPCR, reverse transcription quantitative PCR; SMC1A, structural maintenance of chromosome protein 1A; TIMP1, TIMP metallopeptidase inhibitor 1.

Article Snippet: Tissues were incubated with antibodies against LAMC2 (1:100, PK08311, Abmart Shanghai Co., Ltd.), α‐smooth muscle actin (α‐SMA, 1:500, GTX100034, GeneTex, Inc.), and collagen I (1:100, GTX20292, GeneTex) overnight at 4°C and with goat anti‐rabbit IgG (1:2500, ab97051, Abcam) at 37°C for 1 h. Then, immunoassays were performed using a DAB horseradish peroxidase color development kit (P0202, Beyotime Biotechnology Co., Ltd.).

Techniques: Over Expression, Expressing, Infection, Quantitative RT-PCR, Phospho-proteomics, Western Blot, Immunofluorescence, Activation Assay, Derivative Assay, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Journal of Cell Communication and Signaling

Article Title: Structural maintenance of chromosome protein 1A exacerbates liver fibrosis by enhancing hepatic stellate cell activation and extracellular matrix synthesis via laminin subunit gamma 2 activation

doi: 10.1002/ccs3.70067

Figure Lengend Snippet: LAMC2 overexpression accentuates liver fibrosis in mice treated with sh‐SMC1A. (A) HE analysis of fibrosis in liver tissues from mice treated with sh‐SMC1A alone or sh‐SMC1A + NC‐oe/LAMC2‐oe ( n = 5). (B, C) Analysis of fibrosis in mouse liver tissues via Masson's trichrome staining ( n = 5). (D–F) The collagen I‐ and α‐SMA‐positive areas in mouse liver tissues were identified by IHC staining ( n = 5). The levels of the inflammatory factors TNF‐α (G), IL‐1β (H), and IL‐6 (I) in serum were measured by ELISAs ( n = 5). The mRNA expression of SMC1A (J) and LAMC2 (K) in liver tissues was examined via RT‐qPCR ( n = 3). (L–O) The protein expression of SMC1A and LAMC2 in activated HSCs (α‐SMA + ) was examined via immunofluorescence staining ( n = 3). (P–R) The protein expression of LRAT and GFAP, which are markers of HSC activation, in mouse liver tissues was analyzed by Western blot ( n = 3). Graphical data are presented as means ± SDs. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. A one‐way ANOVA was used for statistical analysis. ANOVA, analysis of variance; ELISAs, enzyme‐linked immunosorbent assays; HSC, hepatic stellate cell; IHC, immunohistochemistry; LAMC2, laminin subunit gamma 2; RT‐qPCR, reverse transcription quantitative PCR; SMC1A, structural maintenance of chromosome protein 1A.

Article Snippet: Tissues were incubated with antibodies against LAMC2 (1:100, PK08311, Abmart Shanghai Co., Ltd.), α‐smooth muscle actin (α‐SMA, 1:500, GTX100034, GeneTex, Inc.), and collagen I (1:100, GTX20292, GeneTex) overnight at 4°C and with goat anti‐rabbit IgG (1:2500, ab97051, Abcam) at 37°C for 1 h. Then, immunoassays were performed using a DAB horseradish peroxidase color development kit (P0202, Beyotime Biotechnology Co., Ltd.).

Techniques: Over Expression, Staining, Immunohistochemistry, Expressing, Quantitative RT-PCR, Immunofluorescence, Activation Assay, Western Blot, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Journal of Cell Communication and Signaling

Article Title: Structural maintenance of chromosome protein 1A exacerbates liver fibrosis by enhancing hepatic stellate cell activation and extracellular matrix synthesis via laminin subunit gamma 2 activation

doi: 10.1002/ccs3.70067

Figure Lengend Snippet: Overview diagram of the mechanism. SMC1A activates the PI3K/Akt signaling pathway by promoting LAMC2 transcription, which facilitates HSC activation and ECM deposition, leading to liver fibrosis progression. This figure was drawn using Figdraw ( https://www.figdraw.com/#/ ). ECM, extracellular matrix; HSC, hepatic stellate cell; LAMC2, laminin subunit gamma 2; SMC1A, structural maintenance of chromosome protein 1A.

Article Snippet: Tissues were incubated with antibodies against LAMC2 (1:100, PK08311, Abmart Shanghai Co., Ltd.), α‐smooth muscle actin (α‐SMA, 1:500, GTX100034, GeneTex, Inc.), and collagen I (1:100, GTX20292, GeneTex) overnight at 4°C and with goat anti‐rabbit IgG (1:2500, ab97051, Abcam) at 37°C for 1 h. Then, immunoassays were performed using a DAB horseradish peroxidase color development kit (P0202, Beyotime Biotechnology Co., Ltd.).

Techniques: Activation Assay