Duhuo Jisheng Decoction-containing serum promotes proliferation of interleukin-1β-induced chondrocytes through the p16-cyclin D1/CDK4-Rb pathway

Duhuo Jisheng Decoction (DHJSD) is a traditional Chinese herbal medicine that has multiple uses, including as a treatment for osteoarthritis (OA). However, the molecular mechanisms underlying the therapeutic effects of DHJSD on OA remain unknown. In the present study, a serum pharmacological method was applied to investigate the effects of DHJSD on the proliferation of chondrocytes treated with interleukin-1β (IL-1β) in vitro. This is a cell model commonly used to reproduce the mechanisms involved in degenerative arthropathies, including OA. The most effective intervention conditions of DHJSD serum were examined by MTT assay. The degenerative chondrocyte model was established by IL-1β-culture for 24 h, and was verified by optical microscopy and immunohistochemical analyses. Following the successful establishment of the degenerative chondrocyte model, the chondrocytes were subsequently randomly divided into two groups: The blank serum group and the DHJSD treatment group. Subsequent to treatment with the corresponding serum, cell proliferation was detected by MTT assay and DNA staining followed by FACS analysis, and the mRNA and protein expression levels of cyclin D1, cyclin-dependent kinase 4 (CDK4), retinoblastoma tumor suppressor protein (Rb) and p16 were measured by reverse transcription polymerase chain reaction and western blotting, respectively. The results indicated that the most effective condition for the promotion of chondrocyte proliferation was 10% concentration of DHJSD 2-h serum, and the degenerative chondrocyte model was successfully reproduced by IL-1β-treatment for 24 h. The mRNA and protein expression levels of cyclin D1, CDK4 and Rb in the DHJSD serum-treated cells were significantly increased compared with those in the blank serum group, whereas p16 expression was significantly downregulated. These results indicate that treatment of cells with DHJSD-containing serum is able to promote IL-1β-induced chondrocyte proliferation by promoting G1/S phase transition via modulating the expressions of cyclin D1, CDK4, Rb and p16, which contribute to the clinical efficacy of DHJSD in OA.