Journal: Journal of Orthopaedic Translation

Article Title: Targeting the senescence-related genes MAPK12 and FOS to alleviate osteoarthritis

doi: 10.1016/j.jot.2024.06.008

Figure Lengend Snippet: Verification of mRNA and protein levels of candidate senescence-specific genes in osteoarthritis. (A) The mRNA levels of MAPK12, FOS, CYR61, TNFSF15 were investigated in human normal control and osteoarthritis (OA) cartilage tissue by qRT-PCR, n = 6. (B) HE, Safranine O staining and protein levels of MAPK12, FOS, CYR61, TNFSF15 were investigated in human normal control and osteoarthritis (OA) cartilage tissue by immunohistochemistry, n = 6. Scale bar = 100 μm. (C) The mRNA levels of MAPK12, FOS, CYR61, TNFSF15 were investigated in human C28/I2 chondrocytes treated with 10 ng/ml IL-1β for 48h by qRT-PCR, n = 3. (D) The protein levels of MAPK12, FOS, CYR61, TNFSF15 were investigated in human C28/I2 chondrocytes treated with 10 ng/ml IL-1β for 48h by western blotting, n = 3. Statistical analysis was conducted using unpaired Student's t test. P values were compared between OA group and normal control group, between IL-1β group and PBS control group. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns, no significance.

Article Snippet: The following primary antibodies were used: MMP13 (Proteintech, 18165-1-AP), Col2 (Proteintech, 28459-1-AP), p16 (Proteintech, 10883-1-AP), p21 (Proteintech, 10355-1-AP), ACAN (Affinity, DF7561), C-Myc-Tag (Affinity, T0052), GAPDH (Affinity, AF7021), PCNA (CST, 13110T), LMNB1 (Affinity, BF8009), ADAMTS5 (Affinity, DF13268), SOX9 (Santa Cruz, sc-166505), MAPK12 (Proteintech, 20184-1-AP), FOS (Proteintech, 66590-1-Ig), CYR61 (Proteintech, 67656-1-Ig) and TNFSF15 (Proteintech, 29899-1-AP).

Techniques: Control, Quantitative RT-PCR, Staining, Immunohistochemistry, Western Blot

Journal: Journal of Orthopaedic Translation

Article Title: Targeting the senescence-related genes MAPK12 and FOS to alleviate osteoarthritis

doi: 10.1016/j.jot.2024.06.008

Figure Lengend Snippet: MAPK12 over-expression inhibits senescence and DNA damage of C28/I2 cells induced by H 2 O 2 . (A) The protein levels of LMNB1, PCNA, p16, p21, MMP13 were investigated in C28/I2 cells by western blotting, n = 3. (B) EdU staining was adopted for cell proliferation capability of C28/I2 cells, n = 3. Scale bar = 100 μm. (C) The protein level of γ-H2AX was investigated in C28/I2 cells by immunofluorescence, n = 3. Scale bar = 100 μm. (D) SA-β-gal staining was adopted for senescence of C28/I2 cells, n = 3. Scale bar = 100 μm. Statistical analysis was conducted using one-way ANOVA. P values were compared between OE-MAPK12 group or control group and H 2 O 2 group. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns, no significance.

Article Snippet: The following primary antibodies were used: MMP13 (Proteintech, 18165-1-AP), Col2 (Proteintech, 28459-1-AP), p16 (Proteintech, 10883-1-AP), p21 (Proteintech, 10355-1-AP), ACAN (Affinity, DF7561), C-Myc-Tag (Affinity, T0052), GAPDH (Affinity, AF7021), PCNA (CST, 13110T), LMNB1 (Affinity, BF8009), ADAMTS5 (Affinity, DF13268), SOX9 (Santa Cruz, sc-166505), MAPK12 (Proteintech, 20184-1-AP), FOS (Proteintech, 66590-1-Ig), CYR61 (Proteintech, 67656-1-Ig) and TNFSF15 (Proteintech, 29899-1-AP).

Techniques: Over Expression, Western Blot, Staining, Immunofluorescence, Control

Journal: Journal of Orthopaedic Translation

Article Title: Targeting the senescence-related genes MAPK12 and FOS to alleviate osteoarthritis

doi: 10.1016/j.jot.2024.06.008

Figure Lengend Snippet: Over-expression of MAPK12 promotes cartilage anabolism and proliferation, inhibits cartilage catabolism and senescence of IL-1β stimulated C28/I2 cells. (A, B) The protein levels and quantification of LMNB1, PCNA, p16, p21, COL2, ACAN, SOX9, MMP13, ADAMTS5 were investigated in C28/I2 cells by western blotting, n = 3. (C, D) Representative image and quantification of EdU staining adopted for cell proliferation ability of C28/I2 cells, n = 3. Scale bar = 100 μm. Statistical analysis was conducted using one-way ANOVA. P values were compared between OE-MAPK12 group or control group and IL-1β group. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns, no significance.

Article Snippet: The following primary antibodies were used: MMP13 (Proteintech, 18165-1-AP), Col2 (Proteintech, 28459-1-AP), p16 (Proteintech, 10883-1-AP), p21 (Proteintech, 10355-1-AP), ACAN (Affinity, DF7561), C-Myc-Tag (Affinity, T0052), GAPDH (Affinity, AF7021), PCNA (CST, 13110T), LMNB1 (Affinity, BF8009), ADAMTS5 (Affinity, DF13268), SOX9 (Santa Cruz, sc-166505), MAPK12 (Proteintech, 20184-1-AP), FOS (Proteintech, 66590-1-Ig), CYR61 (Proteintech, 67656-1-Ig) and TNFSF15 (Proteintech, 29899-1-AP).

Techniques: Over Expression, Western Blot, Staining, Control

Journal: Journal of Orthopaedic Translation

Article Title: Targeting the senescence-related genes MAPK12 and FOS to alleviate osteoarthritis

doi: 10.1016/j.jot.2024.06.008

Figure Lengend Snippet: MAPK12 over-expression and FOS deficiency relieve osteoarthritis and joint pain in OA. (A) Representative images of HE, Safranine O staining and synovium in mice knee joint with or without MAPK12 over-expression or FOS knock-down adenovirus, n = 6. (B) OARSI score, meniscus total scores and synovitis score of mice knee joint with or without MAPK12 over-expression or FOS knock-down adenovirus, n = 6. (C, D) Representative images and quantification of Col2, ACAN, MMP13, Adamts5 protein levels in the knee cartilage of mice by immunohistochemistry, n = 6. Scale bar = 100 μm. (E) Paw withdrawl threshold of mice hind paw was measured using von frey filaments at 6 and 8 weeks post-DMM surgery, n = 6. (F) Response time of mice hind paw retraction or licking was recorded using hot plate at 6 and 8 weeks post-DMM surgery, n = 6. P values were compared between OE-MAPK12 and OE-control group, between siFOS group and si-control group, between OE-control or si-control group and sham group. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns, no significance.

Article Snippet: The following primary antibodies were used: MMP13 (Proteintech, 18165-1-AP), Col2 (Proteintech, 28459-1-AP), p16 (Proteintech, 10883-1-AP), p21 (Proteintech, 10355-1-AP), ACAN (Affinity, DF7561), C-Myc-Tag (Affinity, T0052), GAPDH (Affinity, AF7021), PCNA (CST, 13110T), LMNB1 (Affinity, BF8009), ADAMTS5 (Affinity, DF13268), SOX9 (Santa Cruz, sc-166505), MAPK12 (Proteintech, 20184-1-AP), FOS (Proteintech, 66590-1-Ig), CYR61 (Proteintech, 67656-1-Ig) and TNFSF15 (Proteintech, 29899-1-AP).

Techniques: Over Expression, Staining, Knockdown, Immunohistochemistry, Control

Journal: Journal of Orthopaedic Translation

Article Title: Targeting the senescence-related genes MAPK12 and FOS to alleviate osteoarthritis

doi: 10.1016/j.jot.2024.06.008

Figure Lengend Snippet: MAPK12 over-expression and FOS deficiency inhibit chondrocyte senescence in OA. (A) p16 protein levels were investigated in knee cartilage of mice by immunofluorescence. Scale bar = 50 μm. (B) p21 protein levels were investigated in knee cartilage of mice by immunofluorescence. Scale bar = 50 μm. (C) γ-H2AX protein levels were investigated in knee cartilage of mice by immunohistochemistry. Scale bar = 100 μm. (D) IL6 protein levels were investigated in knee cartilage of mice by immunohistochemistry. Scale bar = 100 μm. (E) Quantification of p16, p21, γ-H2AX and IL6 protein levels in knee cartilage of mice, n = 6. (F) Schematic illustration of the MAPK12 and FOS signaling pathways in OA by regulating cellular senescence. Statistical analysis was conducted using one-way ANOVA. P values were compared between OE-MAPK12 and OE-control group, between siFOS group and si-control group, between OE-control or si-control group and sham group. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns, no significance.

Article Snippet: The following primary antibodies were used: MMP13 (Proteintech, 18165-1-AP), Col2 (Proteintech, 28459-1-AP), p16 (Proteintech, 10883-1-AP), p21 (Proteintech, 10355-1-AP), ACAN (Affinity, DF7561), C-Myc-Tag (Affinity, T0052), GAPDH (Affinity, AF7021), PCNA (CST, 13110T), LMNB1 (Affinity, BF8009), ADAMTS5 (Affinity, DF13268), SOX9 (Santa Cruz, sc-166505), MAPK12 (Proteintech, 20184-1-AP), FOS (Proteintech, 66590-1-Ig), CYR61 (Proteintech, 67656-1-Ig) and TNFSF15 (Proteintech, 29899-1-AP).

Techniques: Over Expression, Immunofluorescence, Immunohistochemistry, Protein-Protein interactions, Control