Effects Of Salvia Miltiorrhiza Injection On Anti-inflammatory And Anti-aging Effects Of Rabbit Chondrocytes In Vitro

Objective:This experiment by different concentrations of salvia miltiorrhiza injection,the intervention of rabbit chondrocytes in vitro culture,from the level of molecular biology to explore salvia miltiorrhiza injection on cartilage cell apoptosis,proliferation and aging.To study the mechanism,action target and optimal effect of danshen injection to promote the repair of OA cartilage cells.Method:The growth curves of chondrocyte 2-5 generation were detected by MTT assay.To optimize the effect of salvia miltiorrhiza injection on the activity of chondrocytes by using gradient time method;MTT assay was used to detect the changes in vitro proliferation of chondrocytes in rabbits with different concentrations of salvia miltiorrhiza injection.RT-PCR and Western blot were used to observe the effect of different concentrations of danshen injection on the expression of IL-1? and telomerase activity during the culture of rabbit chondrocytes in vitro.Results:1.2-5 generations of cartilage cell proliferation curve trends:the first day of each generation cells OD values did not differ significantly(P>0.05),the second day of proliferation is slow,gradually speed up the breeding on the third day,the fourth day the growing,fifth,six days slow proliferation,7,8 days multiplies into a plateau,9,10 days proliferation inhibition.The OD value decreased with cell transmission,and the second and third generation were higher than the fourth and fifth generation,but there was no significant difference(P<0.05);2.Optimization of salvia miltiorrhiza injection intervention cell activity limitation:there was no significant difference between two groups of intervention 24 hours OD value(P>0.05),the OD value of 48 hours intervention dose group than the control group(P<0.05),OD value intervention in 72,96 hours in the dose group significantly greater than the control group(P<0.01),the dose group intervention significantly OD value is greater than 24,48,72 hours,96 hours(P<0.01,P<0.01,P<0.05),therefore,choose cells intervention as the experimental testing time points after 72 hours.3.To optimize the dose effect of danshen injection to interfere with cell activity:to intervene in vitro culture of the third generation of chondrocytes with different concentrations of salvia miltiorrhiza injection,and there was no significant difference in OD value before intervention(P>0.05).72 hours after the intervention 0.15 mg/ml,0.3 mg/ml,0.6 mg/ml,1.25 mg/ml,2.5 mg/ml of salvia miltiorrhiza injection on cartilage cell proliferation has obvious promoting effect,compared with the control group with significant difference(P<0.01);0.3 mg/ml salvia miltiorrhiza group OD value was significantly greater than 2.5 mg/ml,1.25 mg/ml,0.15 mg/ml salvia miltiorrhiza group(P<0.01);0.6 mg/ml salvia miltiorrhiza group OD value was significantly greater than 2.5 mg/ml,1.25 mg/ml,0.15 mg/ml salvia miltiorrhiza group(P<0.01);0.6 mg/ml salvia miltiorrhiza OD value and 0.3 mg/ml salvia miltiorrhiza groups did not differ significantly(P>0.05).4.To optimize the dose effect of danshen injection to interfere with cell activity:to intervene in vitro culture of the third generation of chondrocytes with different concentrations of salvia miltiorrhiza injection,and there was no significant difference in OD value before intervention(P>0.05).72 hours after the intervention 0.15/ml,0.3 mg/ml,0.6 mg/ml,1.25 mg/ml,2.5 mg/ml of salvia miltiorrhiza injection on cartilage cell proliferation has obvious promoting effect,compared with the control group with significant difference(P<0.01);0.3 mg/ml salvia miltiorrhiza group OD value was significantly greater than 2.5 mg/ml,1.25 mg/ml,0.15 mg/ml salvia miltiorrhiza group(P<0.01);0.6 mg/ml salvia miltiorrhiza group OD value was significantly greater than 2.5 mg/ml,1.25 mg/ml,0.15 mg/ml salvia miltiorrhiza group(P<0.01);0.6 mg/ml salvia miltiorrhiza OD value and 0.3 mg/ml salvia miltiorrhiza groups did not differ significantly(P<0.05).5.Western blot method to detect the effect of rabbit chondrocytes telomerase activity:to induce the apoptosis of cartilage cells,the third generation plus salvia miltiorrhiza injection after the intervention cell telomerase expression influence in 72 hours,normal group of cartilage cells have the activity of telomerase expression,higher than that of model group;Low,medium and high concentration danshen group were significantly larger than the model group(P<0.05),and the high-dose group was significantly larger than the low-dose group(P<0.05).Conclusions:1.The second to third generation of cartilage cells in vitro are relatively stable biological characteristics,suitable for seed cell research.2.The cell activity of salvia miltiorrhiza injection was the strongest 72 hours after intervention.3.Medium and high concentrations of salvia miltiorrhiza injection can inhibit the IL-1 beta activity,different concentrations of salvia miltiorrhiza injection can enhance the activity of telomerase,a high concentration of salvia miltiorrhiza injection on the expression of telomerase activity is better than that of low concentration.4.Salvia miltiorrhiza injection of rabbit chondrocytes in vitro has anti-inflammatory,anti-aging effect,its mechanism may be through inhibiting cell inflammatory factor beta IL-1 activity,enhance the activity of telomerase,delay and control of telomere shortening,which further increases the number of cell division,extend the life of the cell.