Journal: Molecular Medicine Reports

Article Title: COL1A1 expression induced by overexpression of both a 15-amino acid peptide from the fibrinogen domain of tenascin-X and integrin α11 in LX-2 cells

doi: 10.3892/mmr.2022.12846

Figure Lengend Snippet: Primers used for quantitative PCR analyses.

Article Snippet: To investigate the possible signaling pathway involved in the induction of COL1A1 expression by overexpression of both hTNX-FGFFFF and integrin α11 in LX-2 cells, a TGF-β receptor type 1 (TGFBRI) inhibitor (SB525334) (MedChemExpress) and a YAP inhibitor (verteporfin) (Cayman Chemical) were used.

Techniques: Real-time Polymerase Chain Reaction, Sequencing

Journal: Molecular Medicine Reports

Article Title: COL1A1 expression induced by overexpression of both a 15-amino acid peptide from the fibrinogen domain of tenascin-X and integrin α11 in LX-2 cells

doi: 10.3892/mmr.2022.12846

Figure Lengend Snippet: Narrowing down of the domain involved in the induction of COL1A1 expression in LX-2 cells. (A) Overexpression of both full-length fibrinogen domain of TNX [TNX-FG] and ITGA11 failed to induce the expression of fibrosis marker genes, including ACTA2, COL1A1 and TGFB1 . LX-2 cells were transfected with expression vectors for hTNX-FG (lane 2), ITGA11 (lane 3) and hTNX-FG and ITGA11 (lane 4) in DMEM/0.5% FBS. (B) Induction of COL1A1 expression by overexpression of both hTNX-FGF and ITGA11. LX-2 cells were transfected with expression vectors for hTNX-FGF (lane 2), hTNX-FGL (lane 3), ITGA11 (lane 4), hTNX-FGF and ITGA11 (lane 5) and hTNX-FGL and ITGA11 (lane 6) in DMEM/0.5% FBS. (C) Induction of COL1A1 expression by overexpression of both hTNX-FGFF and ITGA11. LX-2 cells were transfected with expression vectors for hTNX-FGFF (lane 2), hTNX-FGFL (lane 3), ITGA11 (lane 4), hTNX-FGFF and ITGA11 (lane 5) and hTNX-FGFL and ITGA11 (lane 6) in DMEM/0.5% FBS. (D) Induction of COL1A1 expression by overexpression of both hTNX-FGFFF and ITGA11. LX-2 cells were transfected with expression vectors for hTNX-FGFFF (lane 2), hTNX-FGFFL (lane 3), ITGA11 (lane 4), hTNX-FGFFF and ITGA11 (lane 5) and hTNX-FGFFL and ITGA11 (lane 6) in DMEM/0.5% FBS. (A-D) As a control, RNA from the cells without transfection (lane 1) was used. The cell lysate was prepared 48 h after transfection and then RNA was purified. Subsequently, the expression levels of ACTA2, COL1A1 and TGFB1 were examined by reverse transcription-quantitative PCR. The expression level of each gene in the control was set to 1.0, and the relative expression level of each gene compared with that of the control is shown (n=3). Data are presented as the mean ± SD. *P<0.05, **P<0.01 vs. control (lane 1), one-way ANOVA with Dunnett's post hoc test. COL1A1 , type I collagen α1 chain; TNX, tenascin-X; ITGA11, integrin α11; ACTA2 , α-smooth muscle actin; hTNX-FG, fibrinogen-related domain of human tenascin-X; hTNX-FGF, first half of hTNX-FG; hTNX-FGL, latter half of hTNX-FG; hTNX-FGFF, first half of hTNX-FGF; hTNX-FGFL, latter half of hTNX-FGF; hTNX-FGFFF, first half of hTNX-FGFF; hTNX-FGFFL, latter half of hTNX-FGFF.

Article Snippet: To investigate the possible signaling pathway involved in the induction of COL1A1 expression by overexpression of both hTNX-FGFFFF and integrin α11 in LX-2 cells, a TGF-β receptor type 1 (TGFBRI) inhibitor (SB525334) (MedChemExpress) and a YAP inhibitor (verteporfin) (Cayman Chemical) were used.

Techniques: Expressing, Over Expression, Marker, Transfection, Control, Purification, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Molecular Medicine Reports

Article Title: COL1A1 expression induced by overexpression of both a 15-amino acid peptide from the fibrinogen domain of tenascin-X and integrin α11 in LX-2 cells

doi: 10.3892/mmr.2022.12846

Figure Lengend Snippet: Identification of the minimal sequence responsible for induction of COL1A1 expression in LX-2 cells. (A) Induction of COL1A1 expression by overexpression of both hTNX-FGFFFF and ITGA11. LX-2 cells were transfected with expression vectors for hTNX-FGFFFF (lane 2), hTNX-FGFFFL (lane 3), ITGA11 (lane 4), hTNX-FGFFFF and ITGA11 (lane 5) and hTNX-FGFFFL and ITGA11 (lane 6) in DMEM/0.5% FBS. (B) Overexpression of both hTNX-FGpeptide2-5 and ITGA11 did not cause induction of COL1A1 expression. LX-2 cells were transfected with expression vectors for hTNX-FGFFFF (lane 2), hTNX-FGpeptide2-5 (lane 3), ITGA11 (lane 4), hTNX-FGFFFF and ITGA11 (lane 5) and hTNX-FGpeptide2-5 and ITGA11 (lane 6) in DMEM/0.5% FBS. (C) Overexpression of both hTNX-FGFFFM and ITGA11 did not cause induction of COL1A1 expression. LX-2 cells were transfected with expression vectors for hTNX-FGFFFF (lane 2), hTNX-FGFFFM (lane 3), ITGA11 (lane 4), hTNX-FGFFFF and ITGA11 (lane 5) and hTNX-FGFFFM and ITGA11 (lane 6) in DMEM/0.5% FBS. After transfection followed by cell culture, cell lysate extraction and RNA purification, the expression levels of ACTA2, COL1A1 and TGFB1 were examined by reverse transcription-quantitative PCR. (A-C) The expression level of each gene in the control (lane 1, RNA from cells without transfection) was set to 1.0, and the relative expression level of each gene compared with that of the control is shown (n=3). Data are presented as the mean ± SD. *P<0.05, **P<0.01 vs. control (lane 1), one-way ANOVA with Dunnett's post hoc test. COL1A1 , type I collagen α1 chain; TNX, tenascin-X; ITGA11, integrin α11; ACTA2 , α-smooth muscle actin; hTNX-FGFFFF, GGLRIPFPRDCGEEM peptide from fibrinogen-related domain of human tenascin-X (hTNX-FG); hTNX-FGFFFM, PRDCGEEMQNGAGAS peptide from hTNX-FG; hTNX-FGFFFL, QNGAGASRTSTIFL peptide from hTNX-FG; hTNX-FGpeptide2-5, GGLRIPF peptide from hTNX-FG.

Article Snippet: To investigate the possible signaling pathway involved in the induction of COL1A1 expression by overexpression of both hTNX-FGFFFF and integrin α11 in LX-2 cells, a TGF-β receptor type 1 (TGFBRI) inhibitor (SB525334) (MedChemExpress) and a YAP inhibitor (verteporfin) (Cayman Chemical) were used.

Techniques: Sequencing, Expressing, Over Expression, Transfection, Cell Culture, Extraction, Purification, Reverse Transcription, Real-time Polymerase Chain Reaction, Control

Journal: Molecular Medicine Reports

Article Title: COL1A1 expression induced by overexpression of both a 15-amino acid peptide from the fibrinogen domain of tenascin-X and integrin α11 in LX-2 cells

doi: 10.3892/mmr.2022.12846

Figure Lengend Snippet: Induction of COL1A1 expression by overexpression of both hTNX-FGFFFF and ITGA11 in addition to inhibitors. (A) Induction of COL1A1 expression by overexpression of both hTNX-FGFFFF and ITGA11 with a TGFBRI inhibitor (SB525334). DMSO (lanes 1, 2 and 3) and SB525334 (lane 3) were added to the culture medium (DMEM/0.5% FBS) after the transfection of expression plasmids for both hTNX-FGFFFF and ITGA11 in LX-2 cells (lanes 2 and 3). (B) Induction of COL1A1 expression by overexpression of both hTNX-FGFFFF and ITGA11 with a YAP inhibitor (verteporfin). DMSO (lanes 1, 2 and 3) and vertepofin (lane 3) were added to the culture medium (DMEM/0.5% FBS) after the transfection of expression plasmids for both hTNX-FGFFFF and ITGA11 in LX-2 cells (lanes 2 and 3). Subsequently, the cells were cultured followed by cell lysate extraction, RNA purification and reverse transcription-quantitative PCR. (A and B) The expression level of each gene [ ACTA2, COL1A1 and TGFB1 for (A) and ACTA2, COL1A1 and YAP1 for (B)] in the control (lane 1) was set to 1.0, and the relative expression level of each gene compared with that of the control (lane 1) is shown (n=3). Data are presented as the mean ± SD. *P<0.05, **P<0.01 vs. control (lane 1); ## P<0.01 vs. lane 2, one-way ANOVA with the Bonferroni post hoc test. COL1A1 , type I collagen α1 chain; TNX, tenascin-X; ITGA11, integrin α11; ACTA2 , α-smooth muscle actin; YAP, Yes-associated protein; hTNX-FGFFFF, GGLRIPFPRDCGEEM peptide from fibrinogen-related domain of human tenascin-X (hTNX-FG).

Article Snippet: To investigate the possible signaling pathway involved in the induction of COL1A1 expression by overexpression of both hTNX-FGFFFF and integrin α11 in LX-2 cells, a TGF-β receptor type 1 (TGFBRI) inhibitor (SB525334) (MedChemExpress) and a YAP inhibitor (verteporfin) (Cayman Chemical) were used.

Techniques: Expressing, Over Expression, Transfection, Cell Culture, Extraction, Purification, Reverse Transcription, Real-time Polymerase Chain Reaction, Control

Journal: Molecular Medicine Reports

Article Title: COL1A1 expression induced by overexpression of both a 15-amino acid peptide from the fibrinogen domain of tenascin-X and integrin α11 in LX-2 cells

doi: 10.3892/mmr.2022.12846

Figure Lengend Snippet: Effect of YAP1 knockdown on induction of COL1A1 expression. YAP1 was knocked down with YAP1 siRNA prior to overexpression of both hTNX-FGFFFF and ITGA11, and then the expression levels of ACTA2 and COL1A1 were analyzed by reverse transcription-quantitative PCR in LX-2 cells. RNA from cells treated with transfection of control siRNA only (lane 1), transfection of control siRNA followed by co-transfection with both hTNX-FGFFFF and ITGA11 expression plasmids (lane 2) and transfection of YAP1 siRNA followed by co-transfection with both hTNX-FGFFFF and ITGA11 expression plasmids (lane 3) was used. The expression level of each gene ( ACTA2, COL1A1 and YAP1 ) in the control (control siRNA only) (lane 1) was set to 1.0, and the relative expression level of each gene compared with that of the control (control siRNA) is shown (n=3). Data are presented as the mean ± SD. **P<0.01 vs. control (lane 1); # P<0.05, ## P<0.01 vs. lane 2, one-way ANOVA with the Bonferroni post hoc test. At the bottom of the figure, the relative expression levels of YAP1, hTNX-FGFFFF and ITGA11 are also shown, setting lane 2 to 1.0, since hTNX-FGFFFF expression was not detected in lane 1. COL1A1 , type I collagen α1 chain; TNX, tenascin-X; ITGA11, integrin α11; ACTA2 , α-smooth muscle actin; YAP1, Yes-associated protein 1; siRNA, small interfering RNA; hTNX-FGFFFF, GGLRIPFPRDCGEEM peptide from fibrinogen-related domain of human tenascin-X (hTNX-FG).

Article Snippet: To investigate the possible signaling pathway involved in the induction of COL1A1 expression by overexpression of both hTNX-FGFFFF and integrin α11 in LX-2 cells, a TGF-β receptor type 1 (TGFBRI) inhibitor (SB525334) (MedChemExpress) and a YAP inhibitor (verteporfin) (Cayman Chemical) were used.

Techniques: Knockdown, Expressing, Over Expression, Reverse Transcription, Real-time Polymerase Chain Reaction, Transfection, Control, Cotransfection, Small Interfering RNA

Journal: bioRxiv

Article Title: Metformin induces lipogenic differentiation in myofibroblasts to reverse mouse and human lung fibrosis

doi: 10.1101/401265

Figure Lengend Snippet: (A) Schematic representation of the experimental setup. (B-D) qPCR analysis for the lipogenic marker genes PLIN2 and PPARg , as well as COL1A1 in human lung fibroblasts treated with metformin or vehicle. (E, F) Staining of lipid droplets in fibroblasts using LipidTOX (red). Nuclei were counterstained with DAPI (blue). (G-H) Gating strategy for detecting LipidTOX + cells by flow cytometry. (I) Quantification of LipidTOX + cells in response to metformin treatment. (J) Heatmap representation of the top 100 differentially expressed genes in fibroblasts following metformin treatment. Scale bars: (E, F) 25 µm. (B-D) Each data point within a given group corresponds to one patient. Vehicle-treated group: n=12, Metformin-treated group: n=11. (I) n=3 per group. ** P<0.01, *** P<0.001, **** P<0.0001.

Article Snippet: Anti-collagen 1 A1 (anti-COL1A1) antibodies (Rockland, 1:200) and goat anti-rabbit antibodies (Life Technologies, 1:500) were used for immunofluorescence.

Techniques: Marker, Staining, Flow Cytometry

Journal: bioRxiv

Article Title: Metformin induces lipogenic differentiation in myofibroblasts to reverse mouse and human lung fibrosis

doi: 10.1101/401265

Figure Lengend Snippet: (A) Schematic representation of the experimental setup. (B-D) qPCR analysis for PLIN2, PPARg and COL1A1 in human lung fibroblasts treated with TGFβ1 or vehicle for 72h. (E-H) Staining of TGFβ1- and vehicle-treated cells with LipidTOX (red), anti-ACTA2 antibodies (green) and DAPI (blue). (I-K) qPCR analysis for PLIN2, PPARg and COL1A1 in human lung fibroblasts treated with TGFβ1 or vehicle for 72 h, followed by treatment with metformin or vehicle for 72 h. (L-M) Staining of TGFβ1-and vehicle-treated cells with LipidTOX (red) and DAPI (blue) at the end of treatment (t=144 h). Scale bars: (E-H) and (L-M) 25 µm. (B-D, I-K) Each data point within a given group corresponds to one patient. (B-D) n=4 per group. (I-K) n=9-10 per group. * P<0.05, **P<0.01, ****P<0.0001.

Article Snippet: Anti-collagen 1 A1 (anti-COL1A1) antibodies (Rockland, 1:200) and goat anti-rabbit antibodies (Life Technologies, 1:500) were used for immunofluorescence.

Techniques: Staining

Journal: bioRxiv

Article Title: Metformin induces lipogenic differentiation in myofibroblasts to reverse mouse and human lung fibrosis

doi: 10.1101/401265

Figure Lengend Snippet: (A) Schematic representation of the experimental setup. (B-E) Bright-field imaging of PCLS treated with metformin or vehicle for five days. (F, G) Hematoxylin and eosin staining and COL1A1 immunostaining of PCLS prepared from a non-IPF donor lung. (H-M) Hematoxylin and eosin staining, Masson’s trichrome staining and COL1A1 immunostaining of PCLS prepared from an IPF lung and treated with metformin or vehicle for five days. (N, O) 3D-reconstruction of z-stacks of metformin- and vehicle-treated PCLS stained for COL1A1 (green) and lipid droplets (red). (P) Gating strategy for flow cytomety-based quantification of LipidTOX + cells that are negative for hematopoeitic (CD45), endothelial (CD31) and epithelial (EpCAM) cell markers. (Q) Quantification of flow cytometry measurements on metformin- and vehicle-treated cells. (R) Total collagen assay for metformin- and vehicle-treated cells. Scale bars: (B-E) 2 mm, (F) 500 µm, (G, L, M) 50 µm, (H-K) 200 µm. (Q, R) Each data point within a given group corresponds to one patient. (Q) n=4 per group. ® n=3 per group. * P<0.05, **P<0.01.

Article Snippet: Anti-collagen 1 A1 (anti-COL1A1) antibodies (Rockland, 1:200) and goat anti-rabbit antibodies (Life Technologies, 1:500) were used for immunofluorescence.

Techniques: Imaging, Staining, Immunostaining, Flow Cytometry, Collagen Assay

Journal: bioRxiv

Article Title: Metformin induces lipogenic differentiation in myofibroblasts to reverse mouse and human lung fibrosis

doi: 10.1101/401265

Figure Lengend Snippet: (A) Schematic representation of the Acta2-Cre-ERT2 and tdTomato flox construct. (B) Schematic representation of the timeline of the experiment. Bleomycin was administered intratracheally at day 0. Between days 5 and 14, mice were fed tamoxifen-containing pellets and starting at day 14, metformin (1.5 mg/mL) or vehicle was administered through drinking water. Mice were sacrificed at day 28. (C-F) Hematoxylin and eosin and Masson’s trichrome staining of metformin- and vehicle-treated lungs. (G) Quantification of fibrosis in metformin- and vehicle-treated lungs. (H, I) Immunofluorescence for COL1A1 (green). Endogenous tdTomato signal (red) and DAPI (blue) are also shown. (J) LipidTOX staining (green) and tdTomato + cells (red) are shown. The box in (J) is magnified in (K). Arrowheads indicate LipidTOX + tdTomato + cells. (L-S) Gating strategy (to detect CD45 - CD31 - EpCAM - tdTomato + and/or LipidTOX + cells) and quantification of various cell populations based on tdTomato and LipidTOX detection. Scale bars: (C-F) 1 mm, (H, I) 50 µm, (J) 25 µm. (G, Q-S) Each data point within a given group corresponds to one animal. n=5 per group. * P<0.05, **P<0.01. IF: Immunofluorescence, ns: Not significant.

Article Snippet: Anti-collagen 1 A1 (anti-COL1A1) antibodies (Rockland, 1:200) and goat anti-rabbit antibodies (Life Technologies, 1:500) were used for immunofluorescence.

Techniques: Construct, Staining, Immunofluorescence

Journal: bioRxiv

Article Title: Metformin induces lipogenic differentiation in myofibroblasts to reverse mouse and human lung fibrosis

doi: 10.1101/401265

Figure Lengend Snippet: (A) Schematic representation of the gain-of-function experimental setup for AMPK signaling. (B-E) qPCR analysis of PLIN2, PPARg , COL1A1 and BMP2 in IPF fibroblasts treated with AMPK agonist GSK621 or vehicle. (F) Schematic representation of the loss-of-function experimental setup for AMPK signaling. (G-I) qPCR analysis of PLIN2, PPARg and COL1A1 in IPF fibroblasts treated with AMPK siRNA or scramble siRNA. The decrease of AMPK protein levels at the time of analysis is shown in (J, K). (L-N) Staining of GSK621- and vehicle-treated cells with LipidTOX (red) and DAPI (blue). Metformin-treated cells were used as a positive control for lipid-droplet accumulation (M). Scale bars: (L-N) 25 µm. (B-E, G-I, K) Each data point corresponds to one patient. (B-E) Vehicle-treated group: n=7-8, GSK621-treated group: n=6-8. (G-I) n=4 per group. (K) n=3 per group. * P<0.05. ns: Not significant.

Article Snippet: Anti-collagen 1 A1 (anti-COL1A1) antibodies (Rockland, 1:200) and goat anti-rabbit antibodies (Life Technologies, 1:500) were used for immunofluorescence.

Techniques: Staining, Positive Control

Journal: bioRxiv

Article Title: Metformin induces lipogenic differentiation in myofibroblasts to reverse mouse and human lung fibrosis

doi: 10.1101/401265

Figure Lengend Snippet: (A) Schematic representation of the experimental setup. (B-D) qPCR analysis of PLIN2, PPARg and COL1A1 in IPF fibroblasts treated with rhBMP2 or vehicle. (E, F) Staining of rhBMP2- and vehicle-treated cells with LipidTOX (red) and DAPI (blue). (G) Western blot showing the induction of PPARγ phosphorylation in response to rhBMP2 treatment. Lanes 1-4 and lanes 5-8 were run in parallel on different gels under the same conditions. Quantification of the immunoblot is shown in the lower panel. (H) Western blot showing the opposing effects of metformin and TGFβ1 on PPARγ phosphorylation, and the ability of metformin to partially restore PPARγ phosphorylation in TGFβ1-treated cells. Lanes 1-12 and lanes 13-18 were run in parallel on different gels under the same conditions. Quantification of the immunoblot is shown in the lower panel. (I) Model for the antifibrotic mechanism of action of metformin in human lung fibrosis. Metformin activates AMPK signaling in myofibroblasts, leading to suppression of collagen production, and induces lipogenic differentiation via an AMPK-independent mechanism involving BMP2 release and PPARγ activation. Arising lipofibroblasts are known to support type 2 alveolar epithelial stem cells in the lung. Scale bars: (E-F) 50 µm. (B-D, G, H) Each data point corresponds to one patient. (B-D) n=10-11 per group. (G) n=4 per group. (H) n=3 per group. * P<0.05, ns: Not significant.

Article Snippet: Anti-collagen 1 A1 (anti-COL1A1) antibodies (Rockland, 1:200) and goat anti-rabbit antibodies (Life Technologies, 1:500) were used for immunofluorescence.

Techniques: Staining, Western Blot, Activation Assay

Journal: Science Progress

Article Title: Low-dose urapidil mitigates renal ischemia-reperfusion injury through matrix metalloproteinase-9 inhibition and anti-inflammatory effects

doi: 10.1177/00368504261438886

Figure Lengend Snippet: Comparison of MMP-9 IHC between the kidney samples from each group. Details of MMP-9 marking were observed with IHC. MMP-9, matrix metalloproteinase-9; IHC, immunohistochemistry; SH, sham group; CR, control group; I/R, ischemia-reperfusion injury; UP, urapidil.

Article Snippet: Following the completion of the blocking procedure, the samples were treated with primary antibodies, including eNOS (Boster Bio., catalog number: A01604-2, diluted at a ratio of 1:250), caspase 3 (Thermo Scıentıfıc, catalog number: RB-1197-P0, diluted at a ratio of 1:100), TNF-α (Proteintech, catalog number:60291-1-Ig, diluted at a ratio of 1:300), IL-1β (Bioss, catalog number:bs-6319R, diluted at a ratio of 1:150), IL-6 (ST John’s, catalog number:STJ1110424, diluted at a ratio of 1:100), and MMP-9 (Boster Bio, catalog number: PA2140-2, diluted at a ratio of 1:100).

Techniques: Comparison, Immunohistochemistry, Control

Journal: Heliyon

Article Title: Modified Zhenwu Tang delays chronic renal failure progression by modulating oxidative stress and hypoxic responses in renal proximal tubular epithelial cells.

doi: 10.1016/j.heliyon.2024.e31265

Figure Lengend Snippet: Fig. 3. Adenine-induced oxidative stress, activation of the HIF pathway, and increased production of inflammatory factors and tubulointerstitial collagen fibers in rat proximal renal tubules, with MZWT mitigating these effects. (a) Immunohistochemical detection of SOD1, MDA, HIF-1α, COL1A1, IL-1β, TNF-α in rat renal tissues (IHC × 400). (b) Quantitative analysis of the positive expression areas for each marker. (N: normal group, M: model group, MZWT: Modified Zhenwu Tang group, Lotensin: benazepril hydrochloride group. ##P < 0.01, #P < 0.05 compared with the N group; **P < 0.01, *P < 0.05 compared with the M group; NS indicates no statistical significance).

Article Snippet: Zhang et al. Heliyon 10 (2024) e31265 temperature, and incubated with α-SMA and COL1A1 antibodies (1:100, boster, BM0002, bs-10423).

Techniques: Activation Assay, Immunohistochemical staining, Immunohistochemistry, Expressing, Marker, Modification

Journal: Heliyon

Article Title: Modified Zhenwu Tang delays chronic renal failure progression by modulating oxidative stress and hypoxic responses in renal proximal tubular epithelial cells.

doi: 10.1016/j.heliyon.2024.e31265

Figure Lengend Snippet: Fig. 12. Expression of fibrotic markers α-SMA and COL1A1 in HK-2 cells under hypoxic conditions and LPS treatment, showing reduction by MZWT (IF × 200). Details of expression: (a) α-SMA, (b) Relative intensity of α-SMA, (c) COL1A1, (d) Relative intensity of COL1A1. Groups: (A: Normal group, B: Model group, C: Blank serum group, D: Drug serum group, E: FM19G11 group, F: NAC group). Statistical significance: ##P < 0.01, #P < 0.05 compared with the A group; **P < 0.01, *P < 0.05 compared with the B group; NS indicates no statistical significance.

Article Snippet: Zhang et al. Heliyon 10 (2024) e31265 temperature, and incubated with α-SMA and COL1A1 antibodies (1:100, boster, BM0002, bs-10423).

Techniques: Expressing

Journal: Experimental Hematology & Oncology

Article Title: Dickkopf-1 promotes tumor progression of gefitinib- resistant non-small cell lung cancer through cancer cell-fibroblast interactions

doi: 10.1186/s40164-025-00616-9

Figure Lengend Snippet: Decreases in myofibroblast characteristics of MRC-5 by DKK1. ( A ) Volcano plot for secretory genes using RNA-sequencing data from Fig. A. Red dots represent genes satisfying p-value < 0.05 (left). GO term analysis for secreted protein genes satisfying p-value < 0.05 (right). ( B ) Myofibroblast marker genes in MRC-5 treated with HCC827-DKK1-OE culture supernatants. The data were obtained using RNA-sequencing data from Fig. A. ( C ) Col1a1 , ACTA2 mRNA expression in MRC-5 treated with HCC827-DKK1-OE culture supernatants for 6 h. ( D ) Col1a1 and α-SMA protein expression in MRC-5 treated with HCC827-DKK1-OE culture supernatants for 48 h. ( E ) TGF-β concentration in culture media and supernatant obtained from HCC827-LV con and HCC827-DKK1 OE. ( F ) Masson’s trichrome staining using lung tissue obtained from Fig. F. Representative images (left) and collagen area fraction (%) analyzed by ImageJ software (right). ( G ) Protein levels of Col1a1 and a-SMA in tumor sample obtained from Fig. I. All statistical significance of the differences was determined by unpaired two-tailed Student t-test. ns, non-significant; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001

Article Snippet: Antibodies targeting DKK1 (10170-R015, SinoBiological, Beijing, China), glyceraldehyde 3-phosphate dehydrogenase (GAPDH, CB1001, Merck Millipore, Burlington, MA, USA), JNK (3496-1, Epitomics, Burlingame, CA, USA), Col1a1 (PB9938, Boster Biological Technology, Pleasanton, CA, USA), α-smooth muscle actin (α-SMA, A5228, Sigma-Aldrich, St. Louis, MO, USA) were used.

Techniques: RNA Sequencing, Marker, Expressing, Concentration Assay, Staining, Software, Two Tailed Test