| Object:To observe the effect of traditional Chinese medicine combined with acupuncture onpolycystic ovary syndrome(PCOS)model rats,to study the abnormal angiogenesis in the ovary of PCOS rats,and to explore the effect of the treatment on the expression of vascular factor TSP-1 and ES,so as to elucidate the role and possible mechanism of Chinese medicine combined with acupuncture in the treatment of PCOS.Methods:Sixty female SD rats(6weeks)were selected and randomly divided into control group,model group,acupuncture group(EA group),traditional Chinese medicine group(TCM group),combination of medicine and acupuncture group(TCM+EA group),and Western medicine group,10 rats in each group.PCOS rat model was established by intragastric administration of 1 mg/(kg·d)letrozole dissolved in l%carboxymethyl cellulose for 30 days.After 20 days of modeling,the rats were subjected to vaginal smears for 10 consecutive days to judge the estrous cycle changes.After successful modeling,rats in the model group were intragastrically administered with carboxymethyl cellulose solution;rats in the EA group were treated with electroacupuncture,and the acupuncture points were selected as "ST36","SP6","RN4" and "EX-CA1".Rats in the TCM group were intragastrically administered with Bushen Huajian Recipe;Rats in the TCM+EA group were treated with electroacupuncture at the above-mentioned acupuncture points,and then they were intragastrically administered with Bushen Huajian Recipe;Rats in the Western medicine group were treated with Da Ying-35.The rats were weighed weekly during the treatment.After the rats were sacrificed,the ovaries were weighed and the blood was separated from the abdominal aorta.The levels of serum testosterone(T),follicle stimulating hormone(FSH),luteinizing hormone(LH)and anti-Muller tube hormone(AMH)were detected by ELISA.The morphological changes of ovarian tissue were observed under HE staining.The expression and localization of thrombospondin-1(TSP-1)and endostatin(ES)in ovarian tissue of each group were detected by immunohistochemistry.The expression of TSP-1 and ES protein in ovarian tissue of each group was detected by Western Blot.Results:1.Body mass gain and ovarian relative mass: After treatment,the body mass increase and ovarian relative quality were lower when compared with the model group(P<0.01).Comparing each treatment group,the body mass of the western medicine group was lower than that of the control group,and the relative quality of the ovary was also significantly reduced.The western medicine groupwas statistically different from the other treatment groups(P<0.05).2.Serum sex hormone levels: Compared with the controlgroup,the serum T,LH,AMH levels and LH/FSH ratio of the model group were significantly increased(P<0.01);compared with the model group,the levels of serum T,LH and theratio of LH/FSH in each treatmentgroups were decreased(P<0.05 or P<0.01).The EA+TCM group and the Western medicine group decreased significantly,and there was no significant difference between the two groups on the levels of serum T and theratio of LH/FSH.There was no significant difference in serum AMH level between the EA group and the model group(P>0.05).Compared with the model group,the serum AMH level of the other treatment groups was higher except the EA group.There was no significant difference in FSH level between the groups(P>0.05).3.Pathological changes of ovarian tissue: After HE staining,the follicular granule cells of control group were multi-layered,thick,well-formedand neatly arranged.There were also dominant follicles.In the model group,There were more early developping follicles,atresia follicles and cystic dilated follicles,but no mature follicles and corpus luteum.We can see increased small blood vessels in the interstitium.In the EA group,there were follicles at different developmental stages.Dominant follicles and corpus luteum were observed,and there were no cystic dilated follicles in the ovary.In the TCM group,cystic dilated follicles were still observed.There were also corpus luteum and primary follicles,of which the granule cells were irregularly arranged.In the EA+TCM group,there were no cystic dilated follicles in the ovary.Dominant follicles,primary follicles and small blood vessels in the ovarian interstitium were observed.In the Western medicine group,primary follicles and mature follicles were observed.4.The expression and localization of TSP-1 and ES: Immunohistochemistry showed positive expression of TSP-1 and ES in each group.TSP-1 is mainly expressed in granulosa cells of antral follicles and corpus luteum.Compared with the control group,the positive expression of TSP-1 in the model group was weak.After treatment,the positive expression of TSP-1 and its average optical density wereincrease.Among the treatment group,the average optical density of EA+TCM group was the most significantly incresed(P<0.01).ES is mainly expressed in follicular granulosa cells,follicular membrane cells,and follicular fluid.Compared with the control group,the positive expression of ES in the model group was increased.Compared with the model group,the positive expression and mean optical density of ES were decreased to some extent in each treatment group,and the EA+TCM group showed a significant decrease in the optical density of ES,while there was no significant difference between the western medicine group and the model group(P>0.05).5.The expression of TSP-1 and ES in ovarian tissue: Compared with the control group,the model group showed decrease in the expression of TSP-1and increase in the expression of ES,and the difference wassignificant(P<0.01).Compared with the model group,the expression of TSP-1 in each treatment group increased,while the expression of ES decreased.Among them,the expression of TSP-1 in the EA+TCM group and in the TCM group increased significantly(P<0.01),whilethe expression of ES decreasedsignificantly(P<0.01).The expression of TSP-1 in western medicine group,whichwas lower than that in other treatment groups,was significantly different from that in EA+TCM group(P<0.05).There was no statistical difference in theexpression of ES between the western medicine group and the model group(P>0.05).Conclusion:1.Letrozole can successfully induce PCOS model and can be used as a reliable and ideal animal model for research.2.Compared with the model group,the EA+TCM group showed decrease in the levels of serum T?LH?AMH and the ratio of LH/FSH.There were no cystic dilated follicles in ovarian tissue,but primary follicles,dominant follicles and corpus luteum were observed.This means acupuncture combined with medicine has a good therapeutic effect on reproductive endocrine and ovulation recovery in PCOS rats.3.Treatment of acupuncture combined with medicine affected the expression of vascular factors in ovarian tissue.The expression of TSP-1 was significantly increased,and the expression of ES was significantly decreased.It means that the treatmentplay a regulatory role in the expression of vascular factors in ovarian tissue.4.Acupuncture can improve the ovarian function by stimulating the ovary and regulating the visceral meridians,and the Chinese medicine can improce the overall condition.It is speculated that the mechanism of Acupuncture combined with medicine in the treatment of PCOS may be through regulating vascular factors.It can play a role in regulating reproductive endocrine status by improving blood flow in the ovary. |
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