Tissue Eng Part A

Tissue Eng Part A. p-Akt levels were detected using western blotting analysis. The binding between PLC1 and Akt was assessed with co-immunoprecipitation assays PS 48 in human OA chondrocytes. These results showed that PLC inhibition guarded chondrocytes against OA, but Akt inhibition did not dramatically aggravate OA progression. There were mutual antagonism and binding between PLC1 and Akt that could be regulated by their phosphorylation levels. Consequently, the data reveal that the inhibition of PLC1 may provide an attractive therapeutic target for OA therapy, implicating its binding to Akt. 0.05, **0.01, ***0.001, ****0.0001, NS group). Nevertheless, the architecture of chondrocyte and matrix of articular cartilage in TCN-treated groups at different concentrations were similar to that in NS group along with the decreased thickness of articular cartilage and increased OARSI scores (Figure ?(Figure1).1). Like 1 month-treated groups, the protective effect of U73122 on OA articular cartilage was observed in 2 month-treated groups (Figure ?(Figure2,2, *0.05, **0.01, ***0.001, ****0.0001, NS group). Although TCN (10 g/kg) caused a significant decrease at the grade of OARSI scoring system compared with NS group (Figure ?(Figure2C,2C, **0.01), the other two concentration of TCN did not change the thickness of articular cartilage and OARSI scores compared with NS group. Taken the data together, U73122 treatment for either 1 or 2 2 month had a significant chondroprotective effect on articular cartilage in a rat OA model, but TCN had not any intensifying effect on OA cartilage degeneration. Open in a separate window Figure 1 Histopathological evaluation of OA in a rat OA model treated with different inhibitors of PLC1 and Akt for 1 month(A) Representatives images of Safranin O CFast green staining from the rats treated by different inhibitors of PLC1 and Akt after ACLT+MMx (original magnification 100). (B) Graph indicating the relative cartilage thickness of each femur condyle (from superficial zone to tidemark). (C) Graph indicating the OARSI scores (*0.05, **0.01, ***0.001, ****0.0001, NS group). Open in a separate window Figure 2 Histopathological evaluation of OA in a rat OA model treated with different inhibitors of PLC1 and Akt for 2 month(A) Representatives images of Safranin O CFast green staining from the rats treated by different inhibitors of PLC1 and Akt after ACLT+MMx (original magnification 100). (B) Graph indicating the relative cartilage thickness of each femur condyle (from superficial zone to tidemark). (C) Graph indicating the OARSI scores (*0.05, **0.01, ****0.0001, NS group). Effect of PLC1 inhibitor on the expression levels of Aggrecan and Col 2 in a rat OA model Aggrecan and Collagen Type 2(Col 2) are main biomarkers of cartilage matrix synthesis, the effect of PLC1 inhibition on them was then assessed with immunohistochemistry assay in a rat OA model. Considering the heavy cartilage degeneration in NS group, the difference between U73122-treated and normal groups was analysed. In 1 month-treated group, Figure ?Figure3A3A showed that the expression level of Aggrecan in U73122- treated groups at different concentrations was less than that in normal group (***0.001). In 2 month-treated groups, the expression level of Aggrecan in U73122-treated groups at different concentrations was close to that in normal group (Figure ?(Figure3B).3B). These results indicated that the treatment of U73122 for 2 month enhanced the expression level of Aggrecan, reaching normal level, and that the efficacy of U73122 treatment for 1 month was inferior to that for 2 month. In addition, the expression level of Col 2 in U73122 (200 g/kg)-treated group at for 1 month was close to that in normal group, whereas Col 2 expressions in the other PS 48 two concentrations of U73122 were significantly less than that in normal group, indicating that the efficacy of U73122 (200 g/kg) was superior to that of the other two groups (Figure ?(Figure4A,4A, **0.01, ****0.0001). In 2 month-treated groups, the expression level of Col 2 in U73122 (10 g/kg)-treated group was close to that in normal group, whereas Col 2 expressions in the other two groups of U73122 were less than that in normal group, indicating that the efficacy of MIF U73122 (10 g/kg) treatment was superior to that of the two other groups (Figure ?(Figure4B,4B, *0.05,**0.01). PS 48 Therefore, treatment of both U73122 (200 g/kg, 1 month) and U73122 (10 g/kg, 2 month) could enhance the expression of Col 2 reaching normal level, and that the efficacy of U73122 may partially depend on the time and concentration of its treatment. Consequently, definite concentration PLC1 inhibitors promoted matrix synthesis through increasing Aggrecan and Col 2 expression levels in a rat OA model. Open.