Journal: Cell Death & Disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: A Workflow for comprehensive identification of transcriptionally active lncRNAs in the gastric mucosa of patients with GC. B Pie chart depicting gene loci harboring H3K4me3 peaks at transcription start sites (TSSs). These loci were categorized as protein-coding genes, lncRNA genes, or other genes. Peaks were categorized as patient-specific, healthy individual-specific, and common. C H3K4me3 peaks of representative genes in each category. D Summarized qRT-PCR results for 15 selected lncRNAs from eight GC cell lines. E qRT-PCR analysis of TCONS_00006264 (TM4SF1-AS1) and TM4SF1 in GC cell lines and normal stomach tissue. The locations of these two genes are shown at the top. ( n = 3). F Expression of TM4SF1-AS1 in primary GCs ( n = 374) and normal stomach tissues ( n = 32) from The Cancer Genome Atlas (TCGA) dataset. * P < 0.05.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1-AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Quantitative RT-PCR, Expressing

Journal: Cell Death & Disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: A qRT-PCR analysis of TM4SF1-AS1 in the indicated GC cell lines transfected with siRNAs targeting TM4SF1-AS1 (si-1 and 2) or a control siRNA (si-Ctrl). ( n = 3). B Results of cell viability assays with GC cell lines transfected with the indicated siRNAs. ( n = 8). C qRT-PCR analysis of TM4SF1-AS1 in HSC-45 cells with inducible shRNAs targeting TM4SF1-AS1 (sh-1 and 2) or a control shRNA (sh-Ctrl). Cells were incubated for 8 days with or without doxycycline (Dox). ( n = 3). D Colony formation assays using HSC-45 cells with inducible shRNAs. Cells were incubated for 8 days with or without Dox. Summarized results are shown on the right; error bars represent SDs. ( n = 3). E Tumor growth in mice injected with HSC-45 cells inducibly expressing the indicated shRNAs. Mice were treated with or without Dox. Growth curves (left), resected tumors (middle), and tumor weights (right) are shown. F qRT-PCR analysis of TM4SF1-AS1 in the resected tumors in E . ( n = 4). G Tumor growth in mice injected with SNU638 cells stably expressing GFP or TM4SF1-AS1. Growth curves are shown on the left, and resected tumors are shown on the right. ( n = 4). H qRT-PCR analysis of TM4SF1-AS1 in the resected tumors in G . ( n = 4). * P < 0.05, ** P < 0.01, *** P < 0.001, NS not significant.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1-AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Quantitative RT-PCR, Transfection, Control, shRNA, Incubation, Injection, Expressing, Stable Transfection

Journal: Cell Death & Disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: A qRT-PCR analysis of TM4SF1-AS1 in the indicated cellular fractions from HSC-45 cells. U6 snRNA and ACTB served as endogenous controls. ( n = 3). B Association between TM4SF1-AS1 and Pur-α in the indicated subcellular fractions from HSC-45 cells. Pur-α was detected by western blotting in proteins pulled-down with BrU-labeled TM4SF1-AS1, antisense of TM4SF1-AS1 or beads. C Association between Pur-α and YB-1. Immunoprecipitated YB-1 or Pur-α from HSC-45 cell extracts were probed for co-precipitating proteins. D YB-1 was immunoprecipitated from HSC-45 cells with or without RNase A treatment, after which Pur-α was detected by western blotting. E YB-1 or Pur-α in the indicated subcellular fractions from HSC-45 cells were immunoprecipitated with or without RNase A treatment and then probed for co-precipitating proteins. F Results of RIP assays. Pur-α (upper) or YB-1 (lower) was immunoprecipitated from HSC-45 cells, after which co-precipitated TM4SF1-AS1 was detected with qRT-PCR. IgG served as a negative control. ( n = 3). G qRT-PCR analysis of Pur-α (upper) and YB-1 (lower) in HSC-45 cells transfected with the indicated siRNAs. ( n = 3). H Results of cell viability assays with HSC-45 cells transfected with the indicated siRNAs. ( n = 3). * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1-AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Quantitative RT-PCR, Western Blot, Labeling, Immunoprecipitation, Negative Control, Transfection

Journal: Cell Death & Disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: A Heatmap showing expression of genes suppressed by TM4SF1-AS1 knockdown in HSC-45 cells. B GO analysis using the genes in A . C qRT-PCR analysis of representative IRDS genes in HSC-45 cells transfected with the indicated siRNAs. ( n = 3). D GSEA analysis using the microarray data. Gene sets with an FDR < 0.01 are shown on the left. Enrichment plots of indicated gene sets are shown on the right. E Western blot analysis of total and phosphorylated STAT1 in HSC-45 cells transfected with the indicated siRNAs. (F, G) qRT-PCR analysis of interferon genes in HSC-45 cells transfected with the indicated siRNAs ( F ) or in SNU638 cells stably transfected with the indicated genes ( G ). ( n = 3). H Western blot analysis of total and phosphorylated STAT1 in SNU638, SNU638-GFP, and SNU638-TM4SF1-AS1 cells. I qRT-PCR analysis of IRDS genes in SNU638-GFP and SNU638-TM4SF1-AS1 cells. ( n = 3). J Western blot analysis of STAT1 and Pur-α in SNU638-TM4SF1-AS1 cells transfected with the indicated siRNAs targeting Pur-α. K qRT-PCR analysis of IRDS genes in SNU638-TM4SF1-AS1 cells transfected with the indicated siRNAs. ( n = 3). * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1-AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Expressing, Knockdown, Quantitative RT-PCR, Transfection, Microarray, Western Blot, Stable Transfection

Journal: Cell Death & Disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: A Workflow of ChIRP coupled with mass spectrometry (ChIRP-MS) or RNA-seq (ChIRP-RNA-seq) to identify molecules associated with TM4SF1-AS1. B qRT-PCR confirming enrichment of TM4SF1-AS1 in ChIRP products derived from SNU638-TM4SF1-AS1 cells. ( n = 3). C Protein-protein interaction (PPI) network among the proteins identified by ChIRP-MS. Functional categories of the proteins are indicated by node colors. D RIP-qPCR assays validating the ChIRP-MS results. The indicated proteins in HSC-45 cells were immunoprecipitated, and co-precipitated TM4SF1-AS1 was detected by qRT-PCR. IgG served as a negative control. ( n = 3). E Immunofluorescence images showing staining of the SG marker G3PB2 in SNU638, SNU638-GFP, and SNU638-TM4SF1-AS1 cells treated with or without sodium arsenite (SA). Summarized results are shown on the right ( n = 5). Scale bars = 10 μm. F Immunofluorescence indicating G3BP2 (green), TIA1 (white), and inducible MS2-tagged TM4SF1-AS1 (red) in SNU638 cells. Cells were transfected with a MS2 coat protein (MCP)-RFP plasmid and incubated for 8 days with or without Dox. Magnified views of the respective markers are shown on the right. Scale bars = 10 μm. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1-AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Mass Spectrometry, RNA Sequencing Assay, Quantitative RT-PCR, Derivative Assay, Functional Assay, Immunoprecipitation, Negative Control, Immunofluorescence, Staining, Marker, Transfection, Plasmid Preparation, Incubation

Journal: Cell Death & Disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: A Immunofluorescence indicating RACK1 (green), G3BP1 (white), and inducible MS2-tagged TM4SF1-AS1 (red) in SNU638 cells. The cells were transfected with a MS2 coat protein (MCP)-RFP plasmid and incubated for 8 days with or without Dox. Magnified views of the respective markers are shown on the right. Scale bars = 10 μm. B Immunofluorescence images of G3PB2 in HSC-45 cells incubated for 8 days with or without Dox and expressing the indicated inducible shRNAs. Representative images are shown on the left. Summarized results are on the right. ( n = 5). C , D Apoptosis ( C ) and cell cycle ( D ) assays in HSC-45 cells incubated with or without Dox for 8 days and expressing the indicated inducible shRNAs. Representative results are shown on the left. Summarized results are shown on the right. ( n = 3). E , F Western blot analysis of PARP, caspase-3 ( E ) and p38 ( F ) in HSC-45 cells incubated with or without Dox for 8 days and expressing the indicated inducible shRNAs. Cleaved PARP and cleaved caspase-3 are indicated by arrows ( E ). Phosphorylated p38 is indicated by an arrow ( F ). ** P < 0.01, *** P < 0.001.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1-AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Immunofluorescence, Transfection, Plasmid Preparation, Incubation, Expressing, Western Blot

Journal: Cell Death & Disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: A , B Immunofluorescence images showing Pur-α (A) or YB-1 (B), TIA1 and inducible MS2-tagged TM4SF1-AS1 in SNU638 cells incubated for 8 days with or without Dox. Scale bars = 10 μm. C Immunofluorescent staining of G3PB2 in SNU638-TM4SF1-AS1 cells transfected with the indicated siRNAs. Representative results are shown on the left. Summarized results are shown on the right. ( n = 5). Scale bars = 10 μm. D Localization of GFP-tagged Pur-α or YB-1, G3BP1 and TIA1 immunofluorescence in SNU638 cells. Cells were transfected with vectors encoding GFP (upper), GFP-tagged Pur-α (middle) or GFP-tagged YB1 (bottom). Magnified views of the respective markers are shown on the right. Scale bars = 10 μm. E Cell cycle analysis of HSC-45 cells expressing the indicated siRNAs. ( n = 3). F , G Western blot analysis of PARP and caspase-3 ( F ) and total and phosphorylated p38 ( G ) in HSC-45 cells transfected with the indicated siRNAs. Cleaved PARP, cleaved caspase-3 and phosphorylated p38 are indicated by arrows. ** P < 0.01, *** P < 0.001.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1-AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Immunofluorescence, Incubation, Staining, Transfection, Cell Cycle Assay, Expressing, Western Blot

Journal: Cell Death & Disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: In the early steps of gastric tumorigenesis, TM4SF1-AS1 is transcriptionally activated. TM4SF1-AS1 sequesters RACK1 within SGs, which leads to suppression of stress-responsive MAPK signaling and inhibition of apoptosis (created with BioRender.com).

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1-AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Inhibition

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: Bioinformatics analysis of scRNA-seq and bulk-seq databases identified TM4SF1 as a key gene in HSC activation during HF. ( A ) TSNE plot of annotation of interstitial liver cells. ( B ) Volcano plot showing DEGs between qHSCs and fibroblasts. ( C ) Pseudotiming trajectories of qHSCs and fibroblasts. ( D ) Three clusters of branch-dependent genes are functionally enriched for fibrosis and other biologic processes. ( E ) UMAP plot of annotation of HSCs. ( F ) Volcano plot illustrating DEGs between qHSCs and activated HSCs. ( G ) Pseudotiming trajectories of qHSCs and activated HSCs. ( H ). Heatmap showing expression of the 3 clusters of the branch-dependent genes over pseudotiming. ( I ) Venn diagram showing gene intersection of differences and pseudotiming analysis. ( J ) The plot of module-trait relationships obtained by WGCNA. ( K, L ) Volcano plot exhibiting DEGs in cirrhosis group. ( M ) Venn diagram showing gene intersection of WGCNA and differential analysis. ( N ) Venn diagram of gene intersection of scRNA-seq and bulk-seq. ( O, P ) TSNE, violin, and pseudotiming diagram showed TM4SF1 expression in HSCS. ( Q ) Histogram showing TM4SF1 expression in patients with HF. Data are presented as mean ± standard error of the mean. ∗ P < .05, ∗∗ P < .01, ∗∗∗∗ P < .001.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Activation Assay, Expressing

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: Expression analysis of the primary human HSCs database and enrichment analysis related to TM4SF1. ( A ) The expression of the 10 genes in the primary human HSCs database GSE119606 . ( B ) Venn diagram showing the intersection of PDGF-up-regulated genes and the top 10 genes. ( C ) The signal pathways related to TM4SF1 obtained from the Gene Set Enrichment Analysis (GSEA) of the dataset GSE171904 . ( D ) The GSEA result for the PI3K/AKT pathway.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Expressing

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: TM4SF1 expression was up-regulated in the liver tissue of patients with HF, and in activated HSCs. ( A ) Images of HF tissue samples stained with H&E, Masson, and Sirius Red, with scale bars indicating 50 μm. ( B, C ) The protein levels of TM4SF1, α-SMA, and Collagen 1 in a patient with HF (n = 4). ( D-F ) The mRNA expression of TM4SF1 , Acta2 , and Col1A1 in liver tissues (n = 5). ( G ) Immunofluorescence analysis illustrating the protein expression levels of TM4SF1 and α-SMA in primary mouse HSCs. ( H-J ) The protein levels of TM4SF1 and α-SMA in primary mouse HSCs (n = 3). ( K-M ) The protein levels of TM4SF1 and α-SMA in the human HSC line LX-2 (n = 3). Data are presented as mean ± standard error of the mean. ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001. ns, not significant.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Expressing, Staining, Immunofluorescence

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: TM4SF1 expression was up-regulated in liver tissues of mouse models with HF induced by BDL and CCL4. ( A ) Schematic representation of the mouse model of HF induced by BDL. ( B ) Representative macroscopic images of mouse liver tissue. ( C ) Histopathologic evaluation using H&E, Masson, and Sirius Red staining. Scale bars: 50 μm. ( D ) Comparison of serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) between sham and BDL groups (n = 5). ( E ) TM4SF1 expression in sham and BDL groups (n = 5). ( F ) The protein levels of α-SMA, Collagen I, and TM4SF1 in sham and BDL groups (n = 4). ( G ) Schematic diagram illustrating the induction of HF by CCL4. ( H ) Representative macroscopic images of mouse liver tissue. ( I ) Histopathologic evaluation using H&E, Masson, and Sirius Red staining. Scale bars: 50 μm. ( J ) Comparison of serum levels of ALT and AST between vehicle and CCL4 groups (n = 5). ( K ) TM4SF1 expression in vehicle and CCL4 groups (n = 5). ( L ) The protein levels of α-SMA, Collagen I, and TM4SF1 in vehicle and CCL4 groups (n = 4). Data are presented as mean ± standard error of the mean. ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001. ns, not significant.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Expressing, Staining, Comparison

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: TM4SF1 enhanced the proliferation, migration, and activation of the human HSC line LX-2. ( A ) The silencing effects of 3 siRNA sequences. ( B ) CCK8 assay assessing the impact of TM4SF1 silencing on LX-2 proliferation (n = 5). ( C ) Scratch assay evaluating the effect of TM4SF1 silencing on LX-2 migration (n = 4). ( D ) The protein levels of Cyclin D1, CDK4, BCL2, and p-ERK1/2 after TM4SF1 silencing. ( E ) The protein levels of TM4SF1 and α-SMA following TGF-β1 stimulation and TM4SF1 silencing (n = 3). ( F ) The mRNA levels of Acta2 and Col1A1 following TGFβ1 stimulation and TM4SF1 silencing. ( G ) The overexpression effects of plasmid. ( H ) CCK8 assay assessing the effect of TM4SF1 overexpression on LX-2 proliferation (n = 5). ( I ) Scratch assay evaluating the impact of TM4SF1 overexpression on LX-2 migration (n = 4). ( J ) The protein levels of Cyclin D1, CDK4, BCL2, and p-ERK1/2 after TM4SF1 overexpression. ( K ) The protein levels of TM4SF1 and α-SMA following TGFβ1 stimulation and TM4SF1 overexpression. ( L ) The mRNA levels of Acta2 and Col1A1 following TGFβ1 stimulation and TM4SF1 overexpression. Data are presented as mean ± standard error of the mean. ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001. ns, not significant.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Migration, Activation Assay, CCK-8 Assay, Wound Healing Assay, Over Expression, Plasmid Preparation

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: TM4SF1, in synergy with TGFβ1, inhibited the activation, proliferation, and migration of HSCs. ( A ) Western blot analysis of the effects of individual and dual knockdown of TM4SF1 and TGFβ1 on TM4SF1, TGFβ1, and α-SMA expression. ( B, C ) Scratch assays to assess the effects of individual and dual knockdown of TM4SF1 and TGFβ1 on cell migration. ( D ) CCK-8 assays to evaluate the effects of individual and dual knockdown of TM4SF1 and TGFβ1 on cell proliferation. ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Activation Assay, Migration, Western Blot, Knockdown, Expressing, CCK-8 Assay

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: TM4SF1 promoted the activation of the PI3K/AKT pathway in LX-2 cells. ( A ) The bubble plot illustrating the pathways linked to TM4SF1. ( B ) The GSEA plot showing enrichment scores of the pathway. ( C-F ) The protein levels of the PI3K/AKT pathway in HF mouse models (n = 4). ( G, H ) The protein levels of the PI3K/AKT pathway after TM4SF1 silencing in LX-2 cells (n = 3). ( I, J ) The protein levels of the PI3K/AKT pathway after TM4SF1 overexpression in LX-2 cells (n = 3). Data are presented as mean ± standard error of the mean. ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001. ns, not significant.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Activation Assay, Over Expression

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: TM4SF1 bound and activated c-Src to up-regulate the PI3K/AKT pathway in LX-2 cells. ( A, B ) Coimmunoprecipitation (Co-IP) assays showing TM4SF1 binding to phosphorylated c-Src (p-Src Try416). ( C, D ) P-c-Src and Total c-Src protein levels in HF mouse models (n = 4). ( E ) P-c-Src protein levels after TM4SF1 silencing in LX-2 cells (n = 3). ( F ) P-c-Src protein levels after TM4SF1 overexpression in LX-2 cells (n = 3). ( G ) Protein levels of the c-Src/PI3K/AKT pathway after treatment with the c-Src inhibitor saracatinib. ( H, I ) CCK8 proliferation assay and scratch wound assay after saracatinib treatment (n = 4). Data are presented as mean ± standard error of the mean. ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001. ns, not significant.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Co-Immunoprecipitation Assay, Binding Assay, Over Expression, Proliferation Assay, Scratch Wound Assay Assay

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: Saracatinib inhibited the up-regulation of the c-Src/PI3K/AKT signaling and the aggravation of HF caused by overexpression of TM4SF1 in HSCs. ( A ) Schematic of viral injection, drug treatment, and model establishment in mice (n = 4). ( B ) Pathologic staining indicative of liver injury and fibrosis in mice: H&E, Sirius red, Masson trichrome, and immunohistochemistry of collagen type 1 (n = 4). ( C ) Western blot analysis of the c-Src/PI3K/AKT pathway and α-SMA expression in mouse liver tissue. ( D ) Western blot analysis of the c-Src/PI3K/AKT pathway and α-SMA expression in primary mouse HSCs.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Over Expression, Injection, Staining, Immunohistochemistry, Western Blot, Expressing

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: Reproduction and identification of HSC-specific TM4SF1 knockout mice. ( A ) The principal mechanism diagram for generating HSC-specific knockout mice by mating flox/flox mice with Gfap mice. ( B ) Genotypes identified by agarose gel electrophoresis. ( C ) Primers for loxp sites and Gfap-Cre.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Knock-Out, Agarose Gel Electrophoresis

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: HSC-specific knockout of TM4SF1 alleviated BDL-induced HF in mice. ( A ) Schematic diagram illustrating the design of the animal experiment (n = 6). ( B ) Representative images of liver morphology and histologic analyses, including H&E, Sirius Red, Masson staining, and α-SMA immunohistochemistry. ( C-E ) Quantitative analysis of pathology and immunohistochemistry (n = 6). ( F, G ) α-SMA protein levels (n = 6). ( H, I ) Acta2 and Col1A1 expression (n = 5). Data are presented as mean ± standard error of the mean. ∗∗ P < .01, ∗∗∗ P < .001, ∗∗∗∗ P < .0001. ns, not significant.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Knock-Out, Staining, Immunohistochemistry, Expressing

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Hepatic Stellate Cell TM4SF1 Accelerates Hepatic Fibrosis Progression via Interacting With the Tyrosine Kinase c-Src

doi: 10.1016/j.jcmgh.2025.101559

Figure Lengend Snippet: HSC-specific knockout of TM4SF1 in mice inhibited the c-Src/PI3K/AKT pathway and HSCs activation. ( A ) Immunofluorescence assay showing the protein expression of α-SMA and TM4SF1. ( B ) The protein levels of TM4SF1 α-SMA, and the c-Src/PI3K/AKT pathway. ( C-G ) Quantitative analysis of the protein levels (n = 3). ( H ) Mechanism diagram: TM4SF1 accelerates hepatic fibrosis progression via activating the c-Src-mediated PI3K/AKT pathway. Data are presented as mean ± SEM. ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001, ∗∗∗∗ P < .0001. ns, not significant.

Article Snippet: HSC-specific TM4SF1 knockout mice were generated by breeding TM4SF1 flox mice with glial fibrillary acidic protein (GFAP) Cre mice (Cyagen Biotechnology Co, Ltd, Suzhou, China).

Techniques: Knock-Out, Activation Assay, Immunofluorescence, Expressing

Journal: Nature cardiovascular research

Article Title: A single-cell comparison of adult and fetal human epicardium defines the age-associated changes in epicardial activity.

doi: 10.1038/s44161-022-00183-w

Figure Lengend Snippet: Fig. 6 | EPDCs were found in sub-epicardium and myocardium. a, Spatial validation of this cluster as EPDCs was carried out using a pseudostain targeting strategy from UMAP of scRNA-seq data, with gene expression displayed in RGB color space and a constant blue channel value of 0.2 for heuristic DAPI simulation. b, Staining of fetal hearts using double stains for TM4SF1 and POSTN (bi), PRG4

Article Snippet: The primary antibodies used in this study for immunofluorescent imaging included: UPK3B, PA552696 (Themo Fisher Scientific); MSLN, sc33672 (Santa Cruz Biotechnology); KRT19, sc6278 (Santa Cruz Biotechnology); POSTN, MAB3548 (R&D Systems); DCN, AF143 (R&D Systems); PRG4, MABT400 (Sigma-Aldrich); and TM4SF1, MAB8164 (R&D Systems).

Techniques: Biomarker Discovery, Gene Expression, Staining

Journal: Journal of Gastrointestinal Oncology

Article Title: Analyzing the role of TM4SF1 expression in pancreatic adenocarcinoma: understanding prognostic implications and therapeutic opportunities

doi: 10.21037/jgo-24-564

Figure Lengend Snippet: Differential gene expression and functional enrichment analysis of PAAD. (A) Volcano plot highlighting DEGs between PAAD and normal tissues. The most significant upregulated gene in the tumor tissues was TM4SF1 . Red dots signify genes that are highly expressed in the tumor group. Blue dots indicate genes with low expression in the tumor group. Gray dots, labeled as ‘not’, represent genes deemed non-differentially expressed due to not meeting the screening threshold. (B) Bubble plot depicting the GO and KEGG functional enrichment of the up- and downregulated DEGs. (C) GSEA analysis results illustrating the signaling pathways significantly enriched with DEGs. BP, biological process; CC, cellular component; MF, molecular function; GO, Gene Ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes; NES, normalized enrichment score; PAAD, pancreatic adenocarcinoma; DEGs, differentially expressed genes.

Article Snippet: Primary antibodies against β-actin (#AF0003, Beyotime) and TM4SF1 (#NBP1-76549, NOVUS, Colorado, USA) were incubated on the membranes overnight at 4 °C.

Techniques: Gene Expression, Functional Assay, Expressing, Labeling, Protein-Protein interactions

Journal: Journal of Gastrointestinal Oncology

Article Title: Analyzing the role of TM4SF1 expression in pancreatic adenocarcinoma: understanding prognostic implications and therapeutic opportunities

doi: 10.21037/jgo-24-564

Figure Lengend Snippet: Immune cell infiltration analysis between high- and low-TM4SF1 expression groups. (A) Scatter plot correlating TM4SF1 expression with levels of immune cell infiltration. (B) Correlation analysis reinforcing the findings in (A). (C) Differential immune cell infiltration based on ssGSEA analysis between the high- and low-TM4SF1 expression groups. (D) High- and low-risk immune features. *, P<0.05; **, P<0.01; ***, P<0.001. MDSC, myeloid-derived suppressor cell; ssGSEA, single-sample gene set enrichment analysis; L, low; H, high.

Article Snippet: Primary antibodies against β-actin (#AF0003, Beyotime) and TM4SF1 (#NBP1-76549, NOVUS, Colorado, USA) were incubated on the membranes overnight at 4 °C.

Techniques: Expressing, Derivative Assay

Journal: Journal of Gastrointestinal Oncology

Article Title: Analyzing the role of TM4SF1 expression in pancreatic adenocarcinoma: understanding prognostic implications and therapeutic opportunities

doi: 10.21037/jgo-24-564

Figure Lengend Snippet: Gene mutation analysis of the five key prognostic genes. (A) Chromosomal positions of the five key genes. (B) Forest plot illustrating the hazard ratios of the five genes concerning PAAD prognosis. (C) Correlation heatmap of the five key genes. (D) Distribution of the mutation frequencies across patients. (E) Waterfall plot of the genes ranked by mutation count. (F) Tissue samples from 10 patients were obtained to examine the mRNA expression differences of TM4SF1, BPIFB4, CPTP, DVL1, PLEKHN1 , and DDR1 between cancer and normal tissues. **, P<0.01. SNP, single nucleotide polymorphism; INS, insertion; DEL, deletion; SNV, single-nucleotide variant; PAAD, pancreatic adenocarcinoma; mRNA, messenger RNA; HR, hazard ratio.

Article Snippet: Primary antibodies against β-actin (#AF0003, Beyotime) and TM4SF1 (#NBP1-76549, NOVUS, Colorado, USA) were incubated on the membranes overnight at 4 °C.

Techniques: Mutagenesis, Expressing, Variant Assay

Journal: Journal of Gastrointestinal Oncology

Article Title: Analyzing the role of TM4SF1 expression in pancreatic adenocarcinoma: understanding prognostic implications and therapeutic opportunities

doi: 10.21037/jgo-24-564

Figure Lengend Snippet: TM4SF1 expression and its functional effects on pancreatic cancer cell behavior. (A) Box plot depicting the relative expression of TM4SF1 mRNA in pancreatic tumor tissues compared to normal tissues, demonstrating a significant upregulation in tumors (*, P<0.05). (B) Kaplan-Meier survival curves stratifying patients into quartiles based on TM4SF1 expression levels, showing an inverse relationship between high TM4SF1 expression and overall survival. (C) Western blot analysis of TM4SF1 protein levels in normal (N1–N5) and tumor (T1–T5) pancreatic tissue samples, with tumors exhibiting higher expression levels. (D) Western blot comparison of the TM4SF1 protein across a panel of pancreatic cancer cell lines, identifying varying expression levels. (E) Bar graph quantifying TM4SF1 mRNA expression in various pancreatic cancer cell lines, with PANC-1 cells showing the highest expression. (F,G) Bar graph showing TM4SF1 mRNA levels in PANC-1 cells after treatment with NC or TM4SF1 -targeting siRNAs (si TM4SF1 #1, #2 and #3), indicating effective knockdown. Proliferation assay (H) and colony formation assay (1×) (stained with 0.1% crystal violet) (I) of PANC-1 cells following si TM4SF1 treatment, revealing reduced cell growth and colony number compared to NC. (J) Cell viability assay assessing the sensitivity of PANC-1 cells to Gemcitabine post-si TM4SF1 treatment, showing increased drug sensitivity. Transwell migration, invasion (10×) (stained with 0.1% crystal violet) (K) and cellular wound healing (5×) (L) assays in si TM4SF1 -treated PANC-1 cells demonstrated decreased migratory and invasive capacities. (M) Western blot confirming overexpression of TM4SF1 in CFPAC-1 cells transfected with oe TM4SF1 versus NC. Cell proliferation assay (N) and colony formation assay (1×) (stained with 0.1% crystal violet) (O) in CFPAC-1 cells with oe TM4SF1 showed enhanced growth and colony-forming ability. Transwell migration, invasion (10×) (stained with 0.1% crystal violet) (P) and cellular wound healing (5×) (Q) assays in oe TM4SF1 -expressing CFPAC-1 cells indicated increased migration and invasion potential. *, P<0.05; **, P<0.01; ***, P<0.001. TPM, transcripts per million; PAAD, pancreatic adenocarcinoma; HR, hazard ratio; NC, control; IC 50 , half-maximal inhibitory concentration; oeTM4SF1, overexpression of TM4SF1; mRNA, messenger RNA.

Article Snippet: Primary antibodies against β-actin (#AF0003, Beyotime) and TM4SF1 (#NBP1-76549, NOVUS, Colorado, USA) were incubated on the membranes overnight at 4 °C.

Techniques: Expressing, Functional Assay, Western Blot, Comparison, Knockdown, Proliferation Assay, Colony Assay, Staining, Viability Assay, Migration, Over Expression, Transfection, Control, Concentration Assay

Journal: Cell death & disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells.

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: Fig. 1 Screening for lncRNAs transcriptionally activated in the gastric mucosa of patients with gastric cancer (GC). A Workflow for comprehensive identification of transcriptionally active lncRNAs in the gastric mucosa of patients with GC. B Pie chart depicting gene loci harboring H3K4me3 peaks at transcription start sites (TSSs). These loci were categorized as protein-coding genes, lncRNA genes, or other genes. Peaks were categorized as patient-specific, healthy individual-specific, and common. C H3K4me3 peaks of representative genes in each category. D Summarized qRT-PCR results for 15 selected lncRNAs from eight GC cell lines. E qRT-PCR analysis of TCONS_00006264 (TM4SF1- AS1) and TM4SF1 in GC cell lines and normal stomach tissue. The locations of these two genes are shown at the top. (n = 3). F Expression of TM4SF1-AS1 in primary GCs (n = 374) and normal stomach tissues (n = 32) from The Cancer Genome Atlas (TCGA) dataset. *P < 0.05.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Quantitative RT-PCR, Expressing

Journal: Cell death & disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells.

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: Fig. 2 TM4SF1-AS1 promotes GC cell growth. A qRT-PCR analysis of TM4SF1-AS1 in the indicated GC cell lines transfected with siRNAs targeting TM4SF1-AS1 (si-1 and 2) or a control siRNA (si-Ctrl). (n = 3). B Results of cell viability assays with GC cell lines transfected with the indicated siRNAs. (n = 8). C qRT-PCR analysis of TM4SF1-AS1 in HSC-45 cells with inducible shRNAs targeting TM4SF1-AS1 (sh-1 and 2) or a control shRNA (sh-Ctrl). Cells were incubated for 8 days with or without doxycycline (Dox). (n = 3). D Colony formation assays using HSC-45 cells with inducible shRNAs. Cells were incubated for 8 days with or without Dox. Summarized results are shown on the right; error bars represent SDs. (n = 3). E Tumor growth in mice injected with HSC-45 cells inducibly expressing the indicated shRNAs. Mice were treated with or without Dox. Growth curves (left), resected tumors (middle), and tumor weights (right) are shown. F qRT-PCR analysis of TM4SF1-AS1 in the resected tumors in E. (n = 4). G Tumor growth in mice injected with SNU638 cells stably expressing GFP or TM4SF1-AS1. Growth curves are shown on the left, and resected tumors are shown on the right. (n = 4). H qRT-PCR analysis of TM4SF1-AS1 in the resected tumors in G. (n = 4). *P < 0.05, **P < 0.01, ***P < 0.001, NS not significant.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Quantitative RT-PCR, Transfection, Control, shRNA, Incubation, Injection, Expressing, Stable Transfection

Journal: Cell death & disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells.

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: Fig. 4 TM4SF1-AS1 activates interferon signaling in GC cells. A Heatmap showing expression of genes suppressed by TM4SF1-AS1 knockdown in HSC-45 cells. B GO analysis using the genes in A. C qRT-PCR analysis of representative IRDS genes in HSC-45 cells transfected with the indicated siRNAs. (n = 3). D GSEA analysis using the microarray data. Gene sets with an FDR < 0.01 are shown on the left. Enrichment plots of indicated gene sets are shown on the right. E Western blot analysis of total and phosphorylated STAT1 in HSC-45 cells transfected with the indicated siRNAs. (F, G) qRT-PCR analysis of interferon genes in HSC-45 cells transfected with the indicated siRNAs (F) or in SNU638 cells stably transfected with the indicated genes (G). (n = 3). H Western blot analysis of total and phosphorylated STAT1 in SNU638, SNU638-GFP, and SNU638-TM4SF1-AS1 cells. I qRT-PCR analysis of IRDS genes in SNU638-GFP and SNU638-TM4SF1-AS1 cells. (n = 3). J Western blot analysis of STAT1 and Pur-α in SNU638-TM4SF1-AS1 cells transfected with the indicated siRNAs targeting Pur-α. K qRT-PCR analysis of IRDS genes in SNU638-TM4SF1-AS1 cells transfected with the indicated siRNAs. (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Expressing, Knockdown, Quantitative RT-PCR, Transfection, Microarray, Western Blot, Stable Transfection

Journal: Cell death & disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells.

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: Fig. 6 TM4SF1-AS1 inhibits apoptosis by suppressing stress-responsive MAPK signaling in GC cells. A Immunofluorescence indicating RACK1 (green), G3BP1 (white), and inducible MS2-tagged TM4SF1-AS1 (red) in SNU638 cells. The cells were transfected with a MS2 coat protein (MCP)-RFP plasmid and incubated for 8 days with or without Dox. Magnified views of the respective markers are shown on the right. Scale bars = 10 μm. B Immunofluorescence images of G3PB2 in HSC-45 cells incubated for 8 days with or without Dox and expressing the indicated inducible shRNAs. Representative images are shown on the left. Summarized results are on the right. (n = 5). C, D Apoptosis (C) and cell cycle (D) assays in HSC-45 cells incubated with or without Dox for 8 days and expressing the indicated inducible shRNAs. Representative results are shown on the left. Summarized results are shown on the right. (n = 3). E, F Western blot analysis of PARP, caspase-3 (E) and p38 (F) in HSC-45 cells incubated with or without Dox for 8 days and expressing the indicated inducible shRNAs. Cleaved PARP and cleaved caspase-3 are indicated by arrows (E). Phosphorylated p38 is indicated by an arrow (F). **P < 0.01, ***P < 0.001.

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Transfection, Plasmid Preparation, Incubation, Expressing, Western Blot

Journal: Cell death & disease

Article Title: TM4SF1-AS1 inhibits apoptosis by promoting stress granule formation in cancer cells.

doi: 10.1038/s41419-023-05953-3

Figure Lengend Snippet: Fig. 8 Model for the TM4SF1-AS1-induced stress adaptation. In the early steps of gastric tumorigenesis, TM4SF1-AS1 is transcriptionally activated. TM4SF1-AS1 sequesters RACK1 within SGs, which leads to suppression of stress-responsive MAPK signaling and inhibition of apoptosis (created with BioRender.com).

Article Snippet: pCMV6 encoding GFP-fused TM4SF1 was purchased from OriGene Technologies (Rockville, MD, USA). pIDTSMART-AMP encoding TM4SF1AS1 (ENST496491.1) was purchased from Codex DNA Inc. (San Diego, CA, USA).

Techniques: Inhibition