Effect of triptolide on reproductive toxicity in female rats with Ⅱ type collagen induced arthritis
In order to study the toxic effect and mechanism of triptolide(TP)on the reproductive system of female rats with Ⅱ type collagen induced arthritis(CIA),50 SD rats were randomly divided into normal control group,CIA model group,and three groups re-ceiving TP tablets at clinically equivalent doses of 0.5,1,and 2 times,respectively(with TP dosages of 3.75,7.5,and 15 μg·kg-1·d-1),each comprising 10 rats.Intragastric administration was started on the day after the first immunization,once a day,for 42 days.The results were taken on the 21st and 42nd days to calculate the uterine and ovarian organ indexes;pathological and morphological changes in uterus and ovaries were observed under a light microscope;and the levels of estradiol(E2)and cytochrome P450A1(aro-matase,CYP19A1)in ovarian homogenate were detected by ELISA.Furthermore,immunohistochemistry was employed to detect the expression levels of transforming growth factor β3(TGFβ3)pathway-related proteins,mothers against decapentaplegic homolog 3(Smad3)and steroidogenic factor-1(SF-1)in ovarian tissues.In vitro,the mouse Chinese hamster ovary(CHO)cell line was estab-lished,and after 24 hours of TP administration(30,60,120 nmol·L-1),cell proliferation was detected by the thiazolyl blue tetrazo-lium bromide(MTT)method,apoptosis by the flow cytometry,and TGFβ3,Smad3 and SF-1 protein expression in cells by the Wes-tern blot method,and the nuclear entry of SF-1 was detected by immunofluorescence.The results showed that compared with the CIA model group,all TP administration groups showed decreased number of uterine glands,total follicles,mature follicles,and corpus lu-teum on days 21 and 42 of administration,but there was no statistical difference,and only the administration of 2 times the clinically equivalent dose of TP could significantly increase the number of atretic follicles at 42 days of administration.TP at 3.75 μg·kg-1·d-1 significantly reduced the level of E2 at 21 days of administration and the expression of TGFβ3 and Smad3 factors in ovarian tissues,but had no significant effect on the rate-limiting enzyme in estrogen synthesis CYP19A1.TP at 7.5 and 15 μg·kg-1·d-1 significantly re-duced the expression of SF-1 regardless of administration for 21 days or 42 days.TP can significantly promote ovarian cell apoptosis in vitro,with apoptosis mainly concentrated in the late stage of apoptosis after 24 hours of administration.In addition,60 nmol·L-1 TP significantly reduced the protein expression of TGFβ3,Smad3 and SF-1 in a dose-dependent manner.In summary,intragastric admi-nistration of TP at less than 2 times the clinically equivalent dose for 21 days and 42 days did not cause obvious reproductive damage to the uterus and ovarian tissues of CIA rats,and the number of atretic follicles changed significantly only when the 2 times the clinically equivalent dose was administered for 42 days.TP exerted reproductive toxicity in vivo on reproductive target organs and in vitro on ova-rian cells by inhibiting the expression of TGFβ3/Smad3/SF-1 pathway.
triptolidefemale rats with collagen induced arthritisovarian functiontoxicitymechanism of toxic action
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