菊苣酸对大鼠软骨关节炎的作用及机制

目的: 探究菊苣酸(cichoric acid)对大鼠软骨关节炎的作用及机制。方法: 分离大鼠膝关节处软骨原代细胞,利用白细胞介素1β(interleukin 1 beta, IL-1β)和肿瘤坏死因子α (tumor necrosis factor alpha, TNF-α)诱导软骨关节炎细胞模型,给予低中高剂量的菊苣酸和姜黄素干预。免疫荧光分析Type Ⅱ collagen表达水平,甲苯胺蓝染色和阿利新蓝染色观察干预后软骨关节炎细胞变化,β-半乳糖苷酶染色进行衰老检测,TUNEL染色检测细胞凋亡情况,Western blot检测FOXO4 (forkhead box O4)、p53 (tumor protein P53)和Type Ⅱ collagen蛋白表达水平,生化检测超氧化物歧化酶(Superoxide dismutase, SOD),丙二醛(Malondialdehyde, MDA)和一氧化氮(nitric oxide, NO)含量。结果: 分离得到大鼠软骨原代细胞,IL-1β和TNF-α诱导细胞炎性,甲苯胺蓝染色和阿利新蓝染色面积增加,大鼠软骨关节炎细胞构建成功。与模型组相比菊苣酸高剂量组,甲苯胺蓝染色、阿利新蓝染色、β-半乳糖苷酶染色和TUNEL染色平均光密度比值,MDA含量、NO含量,FOXO4和Type Ⅱ collagen蛋白表达水平显著降低(P＜0.05),p53蛋白表达水平,SOD含量显著增加(P＜0.05)。结论: 菊苣酸阻止软骨关节炎导致的细胞衰老,抑制软骨关节炎导致的细胞凋亡,进而阻止原代细胞中软骨关节炎的进展,其机制与菊苣酸下调FOXO4蛋白参与的衰老进程相关。

Effect and mechanism of cichoric acid on chondroarthritis in rats

OBJECTIVE To explore the effect and mechanism ofcichoric acid on chondroarthritis in rats.METHODS The primary chondrocytes were isolated from the knee joint of rats. The arthritis cell model was induced by interleukin 1 beta (IL-1β) and tumor necrosis factor alpha (TNF-α), and treated with low, middle and high doses of cichoric acid and curcumin. Immunofluorescence was used to analyze type Ⅱ collagen expression, toluidine blue staining and Alcian blue staining were applied to observing the effect of drugs on osteoarthritis, β-galactosidase staining was used to detect cellular senescence, TUNEL staining was employed to detect apoptosis, Western blot was used to detect the expression of FOXO4 (forkhead box O4), p53 (tumor protein P53) and type Ⅱ collagen protein, Biochemical testing was used to detect the contents of superoxide dismutase, malondialdehyde, and nitric oxide.RESULTS The primary chondrocytes of rats were isolated. After induced by IL-1β and TNF-α, the cells showed inflammation, and the areas of toluidine blue staining and Alcian blue staining increased. Rat chondroarthritis cells were successfully constructed. Compared with those of the model group, the mean density values of toluidine blue staining, Alcian blue staining, β-galactosidase staining and TUNEL staining, the contents of MDA and NO, the expression levels of FOXO4 and type Ⅱ collagen protein were decreased significantly (P＜0.05), the expression level of p53 protein and the content of SOD were increased significantly (P＜0.05) in cichoric acid high dose group.CONCLUSION Cichoric acid can prevent cellular senescence caused by chondroarthritis, inhibit apoptosis caused by chondroarthritis, and then prevent the progression of chondroarthritis in primary cells. The mechanism is related to down-regulating FOXO4 protein involving in senescence process by cichoric acid.