Journal: JCI Insight

Article Title: Activation of SLIT2/ROBO1/LRP6 axis aggravates cartilage degradation via β -catenin signaling in TMJOA

doi: 10.1172/jci.insight.193632

Figure Lengend Snippet: ( A ) Western blot detection of the effects of SLIT2 overexpression in primary chondrocytes from 3-week-old C57 and SLIT2-Tg mice. Primary chondrocytes for each independent isolation batch were pooled from 3 mice, and the process was repeated 3 times. ( B ) Relative quantification of COL2A1, MMP13, and IL-1β proteins. ( C ) qRT-PCR analysis of the effect of SLIT2 overexpression in primary chondrocytes from 3-week-old C57 or SLIT2-Tg mice. ( D ) Western blot detection of COL2A1, MMP13, and MMP3 after 48 hours of treatment with rhSLIT2 at 0, 10, 20, 50, and 100 ng/mL in SW1353 cells. ( E ) Relative quantification of COL2A1, MMP3, MMP13 and SOX9 proteins. # P < 0.05, ## P < 0.01, ### P < 0.001 compared with the 0 ng/mL group. ( F ) Western blot detection of the effect of SLIT2 knockdown on human chondrocytes treated with NC or Si-SLIT2 sequence for 48 hours. ( G ) Relative quantification of MMP3, MMP13, and SOX9 proteins. ( H ) qRT-PCR detection of the effect of SLIT2 knockdown on SW1353 cells treated with NC or Si-SLIT2 sequence for 24 hours. ( I ) A heatmap illustrating differentially expressed genes related to TMJOA from RNA-seq analysis between the NC and Si-SLIT2 groups. ( J ) GO analysis of 12 key terms from the top terms. All data are shown as mean ± SD. Statistical significance was assessed by 2-tailed Student’s t test ( B , C , G , and H ) and 1-way ANOVA with Dunnett’s multiple-comparison test ( E ). * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: HEK293T cells and the human chondrocyte cell line SW1353 were purchased from Procell Life Science & Technology Co., Ltd. and cultured in DMEM containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin (P/S). rhSLIT2 (R&D Systems) or IL-1β (PeproTech) was added when the cells reached the appropriate density.

Techniques: Western Blot, Over Expression, Isolation, Quantitative Proteomics, Quantitative RT-PCR, Knockdown, Sequencing, RNA Sequencing, Comparison

Journal: JCI Insight

Article Title: Activation of SLIT2/ROBO1/LRP6 axis aggravates cartilage degradation via β -catenin signaling in TMJOA

doi: 10.1172/jci.insight.193632

Figure Lengend Snippet: ( A ) GSEA was performed using gene sets related to the Wnt/β-catenin pathway from the GO and KEGG databases to assess differences between the NC and Si-SLIT2 groups. ( B and C ) IHC images of active β-catenin in condylar cartilage, and quantitative analysis of the percentage of active β-catenin–positive cells ( n = 4). ( D and E ) IHC images of p-LRP6 in condylar cartilage, and quantitative analysis of the percentage of p-LRP6–positive cells ( n = 4). ( F and G ) Western blot detection of the effect of ROBO1 knockdown for GSK-3β and active β-catenin proteins in SW1353 chondrocytes, and relative quantification of p-GSK-3β/t-GSK-3β and active β-catenin proteins. ( H and I ) IF staining images of β-catenin proteins after being treated with rhSLIT2 and/or Si-ROBO1, and relative quantification of fluorescence intensity of β-catenin proteins. ( J and K ) Western blot analysis assessing the effect of LRP6 knockdown on rhSLIT2-induced catabolism in SW1353 chondrocytes, and relative quantification of MMP3, MMP13, and SOX9 proteins. ( L and M ) Western blot analysis showing the impact of LRP6 knockdown on rhSLIT2-induced nuclear translocation of β-catenin in SW1353 cells, and relative quantification of β-catenin proteins in the cytoplasm and nucleus. Scale bars: 50 μm. All data are shown as mean ± SD. Statistical significance was assessed by 2-tailed Student’s t test ( C , E , and G ), 2-way ANOVA with Šidák’s post hoc analysis ( I and K ), and 1-way ANOVA with Dunnett’s multiple-comparison test ( M ). * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: HEK293T cells and the human chondrocyte cell line SW1353 were purchased from Procell Life Science & Technology Co., Ltd. and cultured in DMEM containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin (P/S). rhSLIT2 (R&D Systems) or IL-1β (PeproTech) was added when the cells reached the appropriate density.

Techniques: Western Blot, Knockdown, Quantitative Proteomics, Staining, Fluorescence, Translocation Assay, Comparison

Journal: JCI Insight

Article Title: Activation of SLIT2/ROBO1/LRP6 axis aggravates cartilage degradation via β -catenin signaling in TMJOA

doi: 10.1172/jci.insight.193632

Figure Lengend Snippet: ( A and B ) Western blot analysis of p-LRP6 with the effect of rhSLIT2 and/or Si-ROBO1 in SW1353 cells, and relative quantification of p-LRP6 proteins. ( C and D ) IF staining images of the effect of rhSLIT2 and Si-ROBO1 on the expression of p-LRP6 in SW1353 cells, and relative quantification of fluorescence intensity of p-LRP6 proteins. Scale bar: 50 μm. ( E and F ) Western blot analysis examining the effect of LRP6 knockdown on ROBO1 expression in SW1353 cells, and relative quantification of ROBO1 proteins. ( G ) Visualization of representative potential binding sites between LRP6 and ROBO1 using PyMOL. ( H ) Co-IP experiments detected the interaction between ROBO1 and LRP6 in SW1353 cells. ( I and J ) Co-IP experiments using overexpression plasmids for LRP6 and ROBO1 to validate their interaction in SW1353 chondrocytes. ( K ) Co-IP analysis of the effect of rhSLIT2 on the interaction between LRP6 and ROBO1 in SW1353 chondrocytes transfected with overexpression plasmids. ( L ) IF staining showing the effect of rhSLIT2 on the colocalization of ROBO1 and LRP6 in SW1353 cells. Scale bars: 10 μm (left) and 1 μm (right). ( M ) IF staining intensity curve of ROBO1 and LRP6 in IF images. ( N ) Quantification of colocalization using Pearson’s correlation and overlap coefficients. All data are shown as mean ± SD. Statistical significance was assessed by 1-way ANOVA with Dunnett’s multiple-comparison test ( B and D ) and 2-tailed Student’s t test ( F and N ). NS, not significant. ** P < 0.01; *** P < 0.001.

Article Snippet: HEK293T cells and the human chondrocyte cell line SW1353 were purchased from Procell Life Science & Technology Co., Ltd. and cultured in DMEM containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin (P/S). rhSLIT2 (R&D Systems) or IL-1β (PeproTech) was added when the cells reached the appropriate density.

Techniques: Western Blot, Quantitative Proteomics, Staining, Expressing, Fluorescence, Knockdown, Binding Assay, Co-Immunoprecipitation Assay, Over Expression, Transfection, Comparison

Journal: bioRxiv

Article Title: Characterization of emerging Oropouche virus tropism and pathogenicity

doi: 10.64898/2026.03.25.714204

Figure Lengend Snippet: A: dPCR results of human chondrocytes (HC, left), Fibroblast-Like Synoviocytes (HFLS, middle) and muscular cells (CHQ; right) cultures supernatants infected with OROV at MOI 0.1 and collected at different times (24, 48, 72 h p.i.). The graphs show 18 points (except HFLS with 12 points) per condition, as well as the mean and geometric standard deviation. ns: not significant; *: p-value < 0.05; **: p-value < 0.01; ***: p-value < 0.001; ****: p-value < 0.0001 (Dunn-Bonferroni non-parametric test). B: HC (left), HFLS (middle), and CHQ (right) cells were infected with OROV at different MOIs. Cell lysates were collected at 48 or 72h p.i. and analysed by Western blot, using antibodies against calnexin (CNX) as internal control and against the N protein of OROV (OROV N). C: The supernatants from HC (left), HFLS (middle) and CHQ (right) cultures were collected at different times after infection with OROV at MOI 0.1. The viral titer was determined on Vero cells using xCELLigence technology from a standard range. Each condition has 6 replicates. ns: not significant; *: p-value < 0.05; **: p-value < 0.01 (Dunn-Bonferroni non-parametric test).

Article Snippet: Human chondrocytes (HC) and synoviocytes (HFLS) were obtained from Cell Applications, (Ref PB-402-05a and 408K-05a, respectively) and cultured according to manufacturer’s instructions.

Techniques: Infection, Standard Deviation, Western Blot, Control

Journal: bioRxiv

Article Title: Characterization of emerging Oropouche virus tropism and pathogenicity

doi: 10.64898/2026.03.25.714204

Figure Lengend Snippet: A: dPCR results of human chondrocytes (HC, left), Fibroblast-Like Synoviocytes (HFLS, middle) and muscular cells (CHQ; right) cultures supernatants infected with OROV at MOI 0.1 and collected at different times (24, 48, 72 h p.i.). The graphs show 18 points (except HFLS with 12 points) per condition, as well as the mean and geometric standard deviation. ns: not significant; *: p-value < 0.05; **: p-value < 0.01; ***: p-value < 0.001; ****: p-value < 0.0001 (Dunn-Bonferroni non-parametric test). B: HC (left), HFLS (middle), and CHQ (right) cells were infected with OROV at different MOIs. Cell lysates were collected at 48 or 72h p.i. and analysed by Western blot, using antibodies against calnexin (CNX) as internal control and against the N protein of OROV (OROV N). C: The supernatants from HC (left), HFLS (middle) and CHQ (right) cultures were collected at different times after infection with OROV at MOI 0.1. The viral titer was determined on Vero cells using xCELLigence technology from a standard range. Each condition has 6 replicates. ns: not significant; *: p-value < 0.05; **: p-value < 0.01 (Dunn-Bonferroni non-parametric test).

Article Snippet: Human chondrocytes (HC) and synoviocytes (HFLS) were obtained from Cell Applications, (Ref PB-402-05a and 408K-05a, respectively) and cultured according to manufacturer’s instructions.

Techniques: Infection, Standard Deviation, Western Blot, Control

Journal: bioRxiv

Article Title: Characterization of emerging Oropouche virus tropism and pathogenicity

doi: 10.64898/2026.03.25.714204

Figure Lengend Snippet: HC, HFLS and CHQ cells were infected on coverslips with OROV at MOI 0.1. Gc protein of the OROV virus was detected (green), and nuclei were also labelled (blue). The scale bar represents 200 µm.

Article Snippet: Human chondrocytes (HC) and synoviocytes (HFLS) were obtained from Cell Applications, (Ref PB-402-05a and 408K-05a, respectively) and cultured according to manufacturer’s instructions.

Techniques: Infection, Virus