| Objective To study the effects of combination of Leonurus and Tripterygium wilfordii on immune activity,liver and kidney and reproductive toxicity of Tripterygium wilfordii.Method MTT method was used to observe the effects of different concentrations of Tripterygium wilfordii extract on the proliferation of COS-7 green monkey kidney cells and L-02 liver cells in vitro;The MTT method was used to observe the effects of different concentrations of Tripterygium wilfordii and Tripterygium wilfordii extract combined with Leonurus on cell proliferation in vitro;A female rat model of rheumatoid arthritis induced by Freund’s complete adjuvant(FCA)was established,and the rats were randomly divided into normal group,model group,Tripterygium wilfordii group and Tripterygium wilfordii Leonurus group.Tripterygium wilfordii group and Tripterygium wilfordii Leonurus group were given intragastrically according to the experimentally designed dose.The normal group and model group were given intragastrically with the same volume of purified water for 21 days.The general condition of the rats was observed and recorded every 4 days.The volume of the plantar of the foot of the rat.After 21 days,the organ index and serum sex hormone levels(E2,FSH and LH)in the peripheral blood of the rat were measured.HE staining was used to observe the pathological sections of the ovary,ankle and uterus of the rat.The value of apoptotic cells in mouse ovarian tissue and Western-blot method were used to determine the expression of p450arom protein in rat uterine tissue.Results In the experiment of liver and kidney cell proliferation in vitro,at a concentration of 3 mg/mL,the inhibition rate of liver cell proliferation in vitro was 7.158±3.003,and the proliferation rate of kidney cell in vitro was-11.942±8.997;The inhibition rate of 15mg/mL liver cell proliferation in vitro was 6.914± 6.037,the value-added rate of kidney cells in vitro was 40.698±6.375;The higher the concentration of Tripterygium wilfordii,the more obvious the effect of inhibiting the value-added of liver and kidney cells,showing the concentration-toxicity relationship;In the in vitro lymphocyte proliferation experiment,the proliferation inhibition rate of Tripterygium wilfordii group was 1.192±0.012 at the concentration of 6mg/mL,and the proliferation inhibition rate of Tripterygium wilfordii Leonurus group was 1.112±0.069,the difference was statistically significant(p<0.05);12mg/mL concentration of Tripterygium wilfordii group proliferation inhibition rate was 1.054 ± 0.043,Tripterygium wilfordii Leonurus group proliferation inhibition rate was 1.059 ± 0.030,there was no statistically significant difference between the two(p>0.05);Tripterygium wilfordii mixed with Leonurus at 6mg/mL and 12mg/mL concentration,the same concentration of Tripterygium wilfordii inhibits mouse spleen lymphocytes.Tripterygium wilfordii compatibility experiment results show Weight growth rate compared with the model group,the weight growth rate of Tripterygium wilfordii and Tripterygium wilfordii Leonurus group increased significantly(p<0.05);Plantar swelling rate Compared with the model group,the foot and sole swelling rate of Tripterygium wilfordii group showed significant changes(p<0.05);Compared with the tripterygium wilfordii group,the plantar swelling rate of Tripterygium wilfordii Leonurus group did not change significantly(p>0.05);Sex hormone levels compared with the Tripterygium wilfordii group,the Tripterygium wilfordii Leonurus group had no significant changes in liver organ index(p>0.05),and the thymus,spleen,kidney,and uterine organ indexes were significantly different(p<0.05),E2,LH value increased significantly(p<0.05),FSH did not change significantly(p>0.05);The results showed that a small amount of lymphocyte infiltration was observed in the ovarian tissue Tripterygium wilfordii group,no obvious lymphocytic infiltration in the Tripterygium wilfordii Leonurus group,ankle joint Tripterygium wilfordii group and Tripterygium wilfordii Leonurus group synovial hyperplasia were not obvious,and occasionally inflammatory cell infiltration was observed.The local epithelial cells in the group were disordered and the intimal lymphocytes were infiltrated.The Tripterygium wilfordii Leonurus group had local epithelial cell hyperplasia and no lymphocyte infiltration in the intimal layer;protein expression compared with the tripterygium wilfordii group,the tripterygium wilfordii Leonurus group ovarian tissue apoptosis cells were significantly reduced(p<0.05),and the expression of p450arom protein was significantly enhanced.Conclusions 1.In the experiment of hepato-renal cytotoxicity,Tripterygium wilfordii has toxic effects on hepatorenal cells and renal cell,and it should not be applied in large doses clinically.2.In the experiment of lymphocyte proliferation in vitro,the immunosuppressive effect of Tripterygium wilfordii after the combination of Tripterygium wilfordii and Leonurus sibiricus was not affected.3.Tripterygium wilfordii mixed with Leonurus has a therapeutic effect on rheumatoid arthritis.It exhibits attenuating effect on organ indexes such as thymus,spleen,and kidney.It does not increase liver and kidney toxicity caused by tripterygium wilfordii.The compatibility is safe and effective.According to the increase of sex hormone level,the increase of uterine organ index,the improvement of pathological status of ovarian tissue and uterine pathological sections,the reduction of ovarian tissue cell apoptosis and the increase of p450arom protein expression,it suggests that the combination has attenuating effects on reproductive toxicity.4.Through the animal experiment research of Tripterygium wilfordii,the attenuating mechanism of Tripterygium wilfordii compatible with Leonurus may be related to the estrogen-like effect of Leonurus.By affecting the synthesis of endogenous hormones,E2 levels are increased to promote the repair of uterus and ovarian function Recovery;second is to reduce ovarian tissue damage by reducing apoptosis of ovarian tissue cells and enhancing uterine p450arom protein expression,which plays a role in reducing reproductive toxicity caused by Tripterygium wilfordii. |
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