wnt5a b  (Proteintech)

Journal: Biomolecules

Article Title: Therapeutic Effects of Wnt5a-Overexpressing Bone Marrow Mesenchymal Stem Cells on Endothelial Barrier Repair in Acute Lung Injury

doi: 10.3390/biom16091356

Figure Lengend Snippet: Isolation and characterization of rat BMSCs. ( A ) Flow cytometry analysis of BMSC surface markers, positive for CD29 and CD90, negative for CD45. ( B ) Osteogenic, adipogenic and chondrogenic differentiation assessed by Alizarin Red, Oil Red O, and Alcian Blue staining, respectively (100×). ( C , D ) Wnt5a expression in BMSCs were determined by Western blot analysis. Data are presented as means ± SEMs, n = 3. Compared with the corresponding control group. The blots were cropped and these uncropped images placed in .

Article Snippet: Membranes were blocked for 15 min at room temperature and incubated overnight at 4 °C with primary antibodies against phospho-PI3K (1:1000, Abmart, Shanghai, China, Cat# T40116F), PI3K (1:1000, Abmart, Shanghai, China, Cat# T40115F), phospho-AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4060S), AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4691S), ZO-1 (1:1000, Proteintech, Wuhan, China, Cat# 21773-1-AP), VE-cadherin (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2500), Bcl-2 (1:1000, Bioworld, Nanjing, China, Cat# BS1511), Bax (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2772), Wnt5a (1:1000, Abclonal, Wuhan, China, Cat# A2133), GAPDH (1:10000, Proteintech, Wuhan, China, Cat# 160004-1-Ig), and Tubulin (1:1000, Proteintech, Wuhan, China, Cat# 10094-1-AP).

Techniques: Isolation, Flow Cytometry, Analysis, Staining, Expressing, Western Blot, Control

Journal: Biomolecules

Article Title: Therapeutic Effects of Wnt5a-Overexpressing Bone Marrow Mesenchymal Stem Cells on Endothelial Barrier Repair in Acute Lung Injury

doi: 10.3390/biom16091356

Figure Lengend Snippet: Wnt5a-overexpressing BMSCs promoted the proliferation, migration, and tube formation of LPS-induced EC injury in vitro. ( A ) EdU assay was used to evaluate the proliferation of LPS-induced EC injury, the blue color indicating nuclei and the red color indicating the proliferation-active cells (magnification: 200×). ( B ) Quantitative analysis was conducted by calculating the percentage of proliferation-active cells. ( C ) The scratch assay was used to assess the migration of ECs, and the representative images of the scratches at 0 h and 24 h are shown above (magnification: 50×). ( D ) Quantitative analysis of changes in the scratched area was performed using ImageJ. ( E ) Tube-formation assays were used to detect the angiogenic capacity of injured ECs. ( F ) Quantitative analysis indicated that coculture with BMSC Wnt5a cells significantly promoted the tube formation of injured ECs compared with that in the BMSC Vector coculture group. Data are presented as means ± SEMs, n = 3. Compared with the control group, ** p < 0.01, *** p < 0.001, **** p < 0.0001; compared with the LPS group, # p < 0.05, ### p < 0.001, #### p < 0.0001.

Article Snippet: Membranes were blocked for 15 min at room temperature and incubated overnight at 4 °C with primary antibodies against phospho-PI3K (1:1000, Abmart, Shanghai, China, Cat# T40116F), PI3K (1:1000, Abmart, Shanghai, China, Cat# T40115F), phospho-AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4060S), AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4691S), ZO-1 (1:1000, Proteintech, Wuhan, China, Cat# 21773-1-AP), VE-cadherin (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2500), Bcl-2 (1:1000, Bioworld, Nanjing, China, Cat# BS1511), Bax (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2772), Wnt5a (1:1000, Abclonal, Wuhan, China, Cat# A2133), GAPDH (1:10000, Proteintech, Wuhan, China, Cat# 160004-1-Ig), and Tubulin (1:1000, Proteintech, Wuhan, China, Cat# 10094-1-AP).

Techniques: Migration, In Vitro, EdU Assay, Analysis, Scratch Assay, Plasmid Preparation, Control

Journal: Biomolecules

Article Title: Therapeutic Effects of Wnt5a-Overexpressing Bone Marrow Mesenchymal Stem Cells on Endothelial Barrier Repair in Acute Lung Injury

doi: 10.3390/biom16091356

Figure Lengend Snippet: Wnt5a knockdown in BMSCs attenuated the proliferation, migration, and tube formation of LPS-induced EC injury in vitro. ( A ) EdU assay was used to measure the proliferation of LPS-induced EC injury, the blue color indicating nuclei and the red color indicating the proliferation-active cells (magnification: 200×). ( B ) Quantitative analysis was conducted by calculating the percentage of proliferation-active cells. ( C ) The scratch assay was used to assess the migration of ECs, and the representative images of the scratches at 0 h and 24 h are shown above (magnification: 50×). ( D ) Quantitative analysis of changes in the scratched area was performed using ImageJ, and the results showed that the migration ability of ECs was significantly decreased in the siWnt5a-BMSC coculture group compared with the siNC-BMSC group. ( E ) Tube-formation assays were used to detect the angiogenic capacity of LPS-induced ECs. ( F ) Quantitative analysis indicated that coculture with siWnt5a-BMSCs significantly decreased the tube formation of injured ECs compared with the siNC-BMSC group. Data are presented as means ± SEMs, n = 3. Compared with the control group, *** p < 0.001, **** p < 0.0001; compared with the LPS group, # p < 0.05, ## p < 0.01, ### p < 0.001, #### p < 0.0001; ns: not significant.

Article Snippet: Membranes were blocked for 15 min at room temperature and incubated overnight at 4 °C with primary antibodies against phospho-PI3K (1:1000, Abmart, Shanghai, China, Cat# T40116F), PI3K (1:1000, Abmart, Shanghai, China, Cat# T40115F), phospho-AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4060S), AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4691S), ZO-1 (1:1000, Proteintech, Wuhan, China, Cat# 21773-1-AP), VE-cadherin (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2500), Bcl-2 (1:1000, Bioworld, Nanjing, China, Cat# BS1511), Bax (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2772), Wnt5a (1:1000, Abclonal, Wuhan, China, Cat# A2133), GAPDH (1:10000, Proteintech, Wuhan, China, Cat# 160004-1-Ig), and Tubulin (1:1000, Proteintech, Wuhan, China, Cat# 10094-1-AP).

Techniques: Knockdown, Migration, In Vitro, EdU Assay, Analysis, Scratch Assay, Control

Journal: Biomolecules

Article Title: Therapeutic Effects of Wnt5a-Overexpressing Bone Marrow Mesenchymal Stem Cells on Endothelial Barrier Repair in Acute Lung Injury

doi: 10.3390/biom16091356

Figure Lengend Snippet: Wnt5a-overexpressing BMSCs attenuated permeability and increased ZO-1 and VE-cadherin expression in ECs, accompanied by enhanced PI3K/AKT signaling. ( A ) Transwell inserts were used to assess the permeability of injured ECs. ( B ) Quantitative analysis of the changes in permeability of EC cocultures with indicated BMSCs. ( C ) Immunofluorescence staining for VE-cadherin (Green) and nuclei (Blue) (400×). ( D ) Immunofluorescence staining for ZO-1 (Red) and nuclei (Blue) (400×). ( E ) Western blot analysis of ZO-1, VE-cadherin, Bcl-2, Bax and PI3K/AKT signaling-related proteins. ( F – K ) Densitometric analysis of Western blots. GAPDH served as an internal reference. Data are presented as means ± SEMs, n = 3. Compared with the control group, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; compared with the LPS group, # p < 0.05, ## p < 0.01, ### p < 0.001, #### p < 0.0001; ns: not significant. The blots were cropped and these uncropped images placed in .

Article Snippet: Membranes were blocked for 15 min at room temperature and incubated overnight at 4 °C with primary antibodies against phospho-PI3K (1:1000, Abmart, Shanghai, China, Cat# T40116F), PI3K (1:1000, Abmart, Shanghai, China, Cat# T40115F), phospho-AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4060S), AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4691S), ZO-1 (1:1000, Proteintech, Wuhan, China, Cat# 21773-1-AP), VE-cadherin (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2500), Bcl-2 (1:1000, Bioworld, Nanjing, China, Cat# BS1511), Bax (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2772), Wnt5a (1:1000, Abclonal, Wuhan, China, Cat# A2133), GAPDH (1:10000, Proteintech, Wuhan, China, Cat# 160004-1-Ig), and Tubulin (1:1000, Proteintech, Wuhan, China, Cat# 10094-1-AP).

Techniques: Permeability, Expressing, Analysis, Immunofluorescence, Staining, Western Blot, Control

Journal: Biomolecules

Article Title: Therapeutic Effects of Wnt5a-Overexpressing Bone Marrow Mesenchymal Stem Cells on Endothelial Barrier Repair in Acute Lung Injury

doi: 10.3390/biom16091356

Figure Lengend Snippet: Wnt5a knockdown in BMSCs decreased their protective effects on LPS-induced EC injury. ( A ) Immunofluorescence staining for VE-cadherin (Green) and nuclei (Blue) (400×). ( B ) Immunofluorescence staining for ZO-1 (Red) and nuclei (Blue) (400×). ( C ) Western blot analysis of ZO-1, VE-cadherin, Bcl-2, Bax and PI3K/AKT signaling-related proteins. ( D – J ) Densitometric analysis of Western blots. GAPDH served as an internal reference. Data are expressed as means ± SEMs, n = 3. Compared with the control group, * p < 0.05, ** p < 0.01, *** p < 0.001; compared with the LPS group, # p < 0.05, ## p < 0.01, ### p < 0.001, #### p < 0.0001; ns: not significant. The blots were cropped and these uncropped images placed in .

Article Snippet: Membranes were blocked for 15 min at room temperature and incubated overnight at 4 °C with primary antibodies against phospho-PI3K (1:1000, Abmart, Shanghai, China, Cat# T40116F), PI3K (1:1000, Abmart, Shanghai, China, Cat# T40115F), phospho-AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4060S), AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4691S), ZO-1 (1:1000, Proteintech, Wuhan, China, Cat# 21773-1-AP), VE-cadherin (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2500), Bcl-2 (1:1000, Bioworld, Nanjing, China, Cat# BS1511), Bax (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2772), Wnt5a (1:1000, Abclonal, Wuhan, China, Cat# A2133), GAPDH (1:10000, Proteintech, Wuhan, China, Cat# 160004-1-Ig), and Tubulin (1:1000, Proteintech, Wuhan, China, Cat# 10094-1-AP).

Techniques: Knockdown, Immunofluorescence, Staining, Western Blot, Analysis, Control

Journal: Biomolecules

Article Title: Therapeutic Effects of Wnt5a-Overexpressing Bone Marrow Mesenchymal Stem Cells on Endothelial Barrier Repair in Acute Lung Injury

doi: 10.3390/biom16091356

Figure Lengend Snippet: Wnt5a-overexpressing BMSCs attenuated LPS-induced ALI in mice. ( A ) An overview diagram of the experimental protocol. At 4 h after intraperitoneal LPS challenge, mice in the treatment groups received BMSC Vector or BMSC Wnt5a cells suspended in 150 μL of PBS via tail-vein injection, whereas mice in the control and the LPS group received an equivalent volume of PBS. ( B ) The histopathology of mice in each group’s staining with hematoxylin–eosin 24 h after LPS or PBS intraperitoneal injection (magnification: 200×). ( C ) Quantitative analysis shows that BMSC Wnt5a dramatically reduced lung injury scores compared with the LPS group and the BMSC Vector group. ( D ) Lung wet/dry assay in each group. ( E ) Evans blue extravasation assay was used to evaluate pulmonary barrier permeability. ( F ) The detection of total cells counts in BALF. Data are presented as means ± SEMs, n = 6. Compared with the control group, ** p < 0.01, **** p < 0.0001; compared with the LPS group, ## p < 0.01, ### p < 0.001, #### p < 0.0001; ns: not significant.

Article Snippet: Membranes were blocked for 15 min at room temperature and incubated overnight at 4 °C with primary antibodies against phospho-PI3K (1:1000, Abmart, Shanghai, China, Cat# T40116F), PI3K (1:1000, Abmart, Shanghai, China, Cat# T40115F), phospho-AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4060S), AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4691S), ZO-1 (1:1000, Proteintech, Wuhan, China, Cat# 21773-1-AP), VE-cadherin (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2500), Bcl-2 (1:1000, Bioworld, Nanjing, China, Cat# BS1511), Bax (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2772), Wnt5a (1:1000, Abclonal, Wuhan, China, Cat# A2133), GAPDH (1:10000, Proteintech, Wuhan, China, Cat# 160004-1-Ig), and Tubulin (1:1000, Proteintech, Wuhan, China, Cat# 10094-1-AP).

Techniques: Plasmid Preparation, Injection, Control, Histopathology, Staining, Analysis, Permeability

Journal: Biomolecules

Article Title: Therapeutic Effects of Wnt5a-Overexpressing Bone Marrow Mesenchymal Stem Cells on Endothelial Barrier Repair in Acute Lung Injury

doi: 10.3390/biom16091356

Figure Lengend Snippet: Wnt5a-overexpressing BMSCs reduced inflammatory cytokine concentrations in LPS-induced ALI. ( A – C ) Concentrations of TNF-α, IL-1β, and CXCL1 in BALF. ( D – F ) Concentrations of TNF-α, IL-1β, and CXCL1 in serum. Data are presented as means ± SEMs, n = 6. Compared with the control group, ** p < 0.01, **** p < 0.0001; compared with the LPS group, # p < 0.05, ### p < 0.001, #### p < 0.0001; ns: not significant.

Article Snippet: Membranes were blocked for 15 min at room temperature and incubated overnight at 4 °C with primary antibodies against phospho-PI3K (1:1000, Abmart, Shanghai, China, Cat# T40116F), PI3K (1:1000, Abmart, Shanghai, China, Cat# T40115F), phospho-AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4060S), AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4691S), ZO-1 (1:1000, Proteintech, Wuhan, China, Cat# 21773-1-AP), VE-cadherin (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2500), Bcl-2 (1:1000, Bioworld, Nanjing, China, Cat# BS1511), Bax (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2772), Wnt5a (1:1000, Abclonal, Wuhan, China, Cat# A2133), GAPDH (1:10000, Proteintech, Wuhan, China, Cat# 160004-1-Ig), and Tubulin (1:1000, Proteintech, Wuhan, China, Cat# 10094-1-AP).

Techniques: Control

Journal: Biomolecules

Article Title: Therapeutic Effects of Wnt5a-Overexpressing Bone Marrow Mesenchymal Stem Cells on Endothelial Barrier Repair in Acute Lung Injury

doi: 10.3390/biom16091356

Figure Lengend Snippet: Wnt5a-overexpressing BMSCs increased ZO-1 and VE-cadherin expression in lung tissues of LPS-induced ALI mice. ( A ) Immunofluorescence staining for ZO-1 (green) and nuclei (blue) (400×). ( B ) Immunofluorescence staining for VE-cadherin (red) and nuclei (blue) (400×). ( C ) Quantitative analysis of ZO-1 expression. ( D ) Quantitative analysis of VE-cadherin. Data are presented as means ± SEMs, n = 3. Compared with the control group, *** p < 0.001, **** p < 0.0001; compared with the LPS group, # p < 0.05, ## p < 0.01, #### p < 0.0001.

Article Snippet: Membranes were blocked for 15 min at room temperature and incubated overnight at 4 °C with primary antibodies against phospho-PI3K (1:1000, Abmart, Shanghai, China, Cat# T40116F), PI3K (1:1000, Abmart, Shanghai, China, Cat# T40115F), phospho-AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4060S), AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4691S), ZO-1 (1:1000, Proteintech, Wuhan, China, Cat# 21773-1-AP), VE-cadherin (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2500), Bcl-2 (1:1000, Bioworld, Nanjing, China, Cat# BS1511), Bax (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2772), Wnt5a (1:1000, Abclonal, Wuhan, China, Cat# A2133), GAPDH (1:10000, Proteintech, Wuhan, China, Cat# 160004-1-Ig), and Tubulin (1:1000, Proteintech, Wuhan, China, Cat# 10094-1-AP).

Techniques: Expressing, Immunofluorescence, Staining, Analysis, Control

Journal: Biomolecules

Article Title: Therapeutic Effects of Wnt5a-Overexpressing Bone Marrow Mesenchymal Stem Cells on Endothelial Barrier Repair in Acute Lung Injury

doi: 10.3390/biom16091356

Figure Lengend Snippet: Wnt5a-overexpressing BMSC treatment was associated with preservation of endothelial junctional proteins, an anti-apoptotic protein profile and enhanced PI3K/AKT signaling in LPS-induced ALI. ( A ) Western blot analysis of ZO-1, VE-cadherin, Bcl-2, Bax and PI3K/AKT signaling-related proteins in lung tissues. ( B – G ) Densitometric analysis of Western blots. GAPDH served as an internal reference. Data are presented as means ± SEMs, n = 3. Compared with the control group, * p < 0.05, ** p < 0.01, *** p < 0.001; compared with the LPS group, # p < 0.05, ## p < 0.01, ### p < 0.001, #### p < 0.0001; ns: not significant. The blots were cropped and these uncropped images placed in .

Article Snippet: Membranes were blocked for 15 min at room temperature and incubated overnight at 4 °C with primary antibodies against phospho-PI3K (1:1000, Abmart, Shanghai, China, Cat# T40116F), PI3K (1:1000, Abmart, Shanghai, China, Cat# T40115F), phospho-AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4060S), AKT (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 4691S), ZO-1 (1:1000, Proteintech, Wuhan, China, Cat# 21773-1-AP), VE-cadherin (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2500), Bcl-2 (1:1000, Bioworld, Nanjing, China, Cat# BS1511), Bax (1:1000, Cell Signaling Technology, Beverly, MA, USA, Cat# 2772), Wnt5a (1:1000, Abclonal, Wuhan, China, Cat# A2133), GAPDH (1:10000, Proteintech, Wuhan, China, Cat# 160004-1-Ig), and Tubulin (1:1000, Proteintech, Wuhan, China, Cat# 10094-1-AP).

Techniques: Preserving, Western Blot, Analysis, Control

Journal: Frontiers in Immunology

Article Title: PD0325901 alleviates thrombin-inhibited osteogenic differentiation through an IL-1β-activated feedback loop between MEK-Erk1/2 and NF-κB signal pathways: insights from bioinformatics and experimental verification

doi: 10.3389/fimmu.2026.1730337

Figure Lengend Snippet: Experimental validation of bioinformatically identified inflammatory cytokines, ECM-related, and calcification-regulatory genes. (A) Heatmap illustrating genes involved in regulating osteoblast growth. (B) PPI network of calcification-regulatory genes from BP terms, highlighting hub genes such as Igf1, Tgfb3, and MMP-9. (C) PPI network of ECM-regulatory genes derived from CC terms, highlighting hub genes such as Igf1, Wnt5a, and Spp1. (D) Western blot analysis and quantitative analysis of COX-2/β-actin, MMP-9/β-actin, Wnt5a/β-actin, Spp1/β-actin, Tgfb3/β-actin and Igf1/β-actin ratios in thrombin- and PD03-treated osteoblasts. (E) The ratios of COX-2/β-actin, MMP-9/β-actin, Wnt5a/β-actin, Spp1/β-actin, Tgfb3/β-actin and Igf1/β-actin were compared among different groups. Data are presented as mean ± SD (n = 3). P-values were determined by one-way ANOVA (multi-group comparisons) (*p < 0.05; **p < 0.01; ***p < 0.001; ns, P >0.05). Scale bar: 100 μm.

Article Snippet: The following antibodies were used in this study: β-actin (AF5001, Beyotime, China), Col1α1 (ab270993, Abcam, UK), Runx2 (AF2593, Beyotime, China), OCN (AF6297, Beyotime, China), Igf1 (AF7179, Beyotime, China), Wnt5a (29793-1-AP, Proteintech, USA), Tgfb3 (AF8142, Beyotime, China), Spp1 (A5427, Bimake, USA), COX-2 (F0327, Bimake, USA), Matrix Metalloproteinase-9 (MMP-9, AF5234, Beyotime, China), Minichromosome Maintenance Complex Component 2 (MCM2, A5172, Bimake, USA), Proliferating Cell Nuclear Antigen (PCNA, SC-25280, Santa, USA), PAR-1 (AF6837, Beyotime, China), IL-1RA (AF7218, Beyotime, China), p65 (8242S, CST, USA), p-p65 (3033T, CST, USA), Erk1/2 (4695T, CST, USA), p-Erk1/2 (4370S, CST, USA), p-Stat3 (Y705) (9145S, CST, USA), p-Stat3 (S727) (9134S, CST, USA), and Stat3 (4904T, CST, USA).

Techniques: Biomarker Discovery, Derivative Assay, Western Blot