Study On The Effect Of Jiawei Yanghe Decoction On The Chondrocyte Model Of Knee Osteoarthritis Induced By IL-1β Based On Autophagy Observation

Objective:To observe the effect of Jiawei Yanghe Decoction on the IL-1β-induced chondrocyte model of knee osteoarthritis,and to explore its mechanism of treating knee osteoarthritis(KOA)by affecting the autophagy and inflammation levels of chondrocytes.Methods:(1)Rats were intragastrically administered at a dose of 1 ml/kgd for8 days to prepare serum containing Jiawei Yanghe Decoction.(2)Rat chondrocytes were isolated and cultured in vitro,passaged on time,and stained with toluidine blueserve the cell morphology.(3)CCK-8 method detects the effect of different doses of medicated serum on the proliferation activity of chondrocytes,and selects the best dose of medicated serum for intervention.(4)CCK-8 method was used to detect the proliferation activity of chondrocytes in each group after IL-1β induction.The cells were divided into blank group(20% blank serum);model group(20% blank serum);drug-containing serum group(20% Medicated serum);rapamycin group(rapamycin(10μmol/L)+ 20% blank serum).Except for the normal group,the other groups were induced with IL-1β(10ng/m L).CCK-8 was used to detect cell proliferation in each group.(5)Detect the expression of IL-6,MMP-13 and TNF-α in the cell supernatant of the blank group,model group,drug-containing serum group,and rapamycin group by ELISA method.(6)Western blot was used to detect the expression of autophagyrelated factors LC3,Beclin-1,ULK1 in cells.Results:(1)The CCK-8 test showed that the proliferation activity of chondrocytes increased under the intervention of Jiawei Yanghe Decoction-containing serum,and the difference was statistically significant(P<0.05).The enhancement effect was positively correlated with the dose,and each dose did not There was a cytotoxic reaction.(2)CCK-8 method was used to detect the proliferation activity of chondrocytes in each group after IL-1β induction.Compared with the blank group,the cell proliferation of the model group was significantly inhibited(P<0.01).Compared with the model group,the drug-containing serum group and the rapamycin group reduced the proliferation inhibition caused by IL-1β on chondrocytes to a certain extent,and the difference was statistically significant(P<0.05).(3)Enzyme-linked immunosorbent assay(ELISA)detects the expression of inflammatory factors in the cell supernatant.Compared with the blank group,the IL-6,TNF-α,and MMP-13 concentrations in the cell supernatant of the model group are significant Increase,the difference is statistically significant(P<0.01).Comparing the drug-containing serum group,rapamycin group and the model group,the concentration of IL-6,TNF-α,and MMP-13 in the cell supernatant decreased,and the difference was statistically significant(P<0.05).(4)Western Blot method was used to detect the expression of autophagy-related proteins in each group of chondrocytes.Compared with the model group,the relative expression of autophagy-related proteins LC3,ULK1,and Beclin-1 in the blank group were significantly increased,and there were differences.Statistically significant(P<0.05).Compared with the model group,the drug-containing serum group showed an increase in the relative expression of autophagy-related proteins LC3,ULK1,and Beclin-1.However,compared with the recognized autophagy agonist rapamycin,its effect on the level of autophagy is relatively limited,and this difference is statistically significant(P<0.05).Conclusion:Jiawei Yanghe Decoction down-regulates the inflammatory factors expressed in the IL-1β-induced KOA chondrocyte model: IL-6,TNF-α,MMP-13,and down-regulates the chondrocyte autophagy-related factors LC3,ULK1,and Beclin The expression of-1 has been increased.Response Jiawei Yanghe Decoction can improve chondrocyte autophagy and reduce inflammation to play a therapeutic role.