Mechanism Of Qianggubao On Articular Cartilage Of Fatigue Rats With Knee Osteoarthritis

Objective Investigate the effect of Qianggubao on the histomorphological changes of osteoarthritis cartilage in fatigue rats and the detection of MMP-3 and ADAMTs-7 expressions in the articular cartilage,in order to explore the mechanism of the prescription on cartilage tissue of knee osteoarthritis.Methods1.Establishment and identification of knee osteoarthritis model in ovariectomized rats induced by fatigue injury: twenty-four 6-month-old female SD rats were randomly divided into three groups: blank control group,ovariectomized group and treadmill group,with 8 rats in each group.Under sterile conditions,bilateral ovaries were removed for ovariectomized group and the treadmill group,while the blank control group was left untreated.The blank control group and the ovariectomized group were normally active,and the treadmill group were trained on the uphill running exercise to establish a strain model.The treadmill exercise program is set to 30m/min,30min/time,60min/d,slope of 16 °,rest for 1 day per week,and continuous training for 6 weeks.The femur of rats was then taken to observe the gross shape of the articular cartilage,and the morphological changes of femur were observed under light microscope.The blood of abdominal aorta was taken,and the serum levels of TNF-α,IL-1β and COMP were detected to identify the fatigue KOA model.2.Protective effect of Qianggubao on cartilage of knee Osteoarthritis in Rats: thirty-two 6-month-old female SD rats were randomly divided into four groups: blank control group,saline group,Jingutongxiao Pill group and Qianggubao group,with 8 rats in each group.Except for the blank control group,the other three groups were used to establish the model rats according to the method mentioned above.The other three groups were given placebo(0.9% saline),Jingutongxiao Pill and Qianggubao orally twice a day for 1 month.The blood was then collected from abdominal aorta,and the joints and articular cartilage were taken.The levels of IL-1β,TNF-α and COMP in serum were measured by Elisa.HE staining was performed after decalcification of knee joint.Morphological changes were observed under light microscope.The expressions of MMP-3 and ADAMTs-7 in cartilage were detected by Western blot and qPCR.Results1.Model identification results: After 6 weeks of modeling,the Mankin score of knee cartilage of rats showed that the ovariectomized group and the treadmill group were higher than the blank control group(P<0.01),and the score of treadmill group was also higher than that of ovariectomized group(P<0.01).The concentrations of IL-1β,TNF-α and COMP in the other two groups were significantly increased compared with those in the blank control group(P<0.01);the treadmill group also had higher concentration than the ovariectomized group(P<0.01).Therefore,both serological examination and morphological microscopic observation of the cartilage of the knee joint indicated that the 6-month-old SD female rat model of osteoarthritis was successful.2.Study on the mechanism of Qianggubao:(1)Compared with the blank control group,the Mankin scores of knee joint cartilage were higher in the other three groups(P<0.01).Compared with the saline group,the Mankin scores of the Jingutongxiao Pill group and the Qianggubao group were significantly lower(P<0.01).There was no significant difference between the Jingutongxiao Pill group and the Qianggubao group.(2)The comparison of contents of IL-1β,TNF-α and COMP in serum showed that the Jingutongxiao Pill group and the Qianggubao group were significantly higher than those of the blank control group(P<0.01).Compared with the saline group,the concentrations in the Jingutongxiao Pill group and the Qianggubao group were significantly lower(P<0.01).There was no statistical difference between the Jingutongxiao Pill group and the Qianggubao group(P>0.05).(3)The results of Western blot showed that the protein expressions of MMP-3 and ADAMTs-7 in the saline group,the Jingutongxiao Pill group and the Qianggubao group were higher than the blank control group(P<0.01);compared with the saline group,the protein expressions in the Jingutongxiao Pill group and the Qianggubao group were lower(P<0.05).There was no significant difference between the Jingutongxiao Pill group and the Qianggubao group(P>0.05).(4)The results of qPCR showed that the protein expressions of MMP-3 mRNA and ADAMTs-7 mRNA in the saline group,the Jingutongxiao Pill group and the Qianggubao group was significantly higher than those in blank control group(P<0.01);and compared with saline group,the protein expressions in the Jingutongxiao Pill group and the Qianggubao group were significantly lower(P<0.01).There was no significant difference between the Qianggubao group and the Jingutongxiao Pill group(P>0.05).Conclusion Qianggubao is beneficial to the repair of knee osteoarthritis by inhibiting the expression of MMP-3 and ADAMTs-7,reducing the degradation of COMP and stabilizing the internal environment of cartilage extracellular matrix.