小花棘豆中毒对和田羊卵巢损伤的研究

摘要

以小花棘豆(Oxytropis glabra DC)中毒和田羊母羊为研究对象,通过复制和田羊小花棘豆亚慢性中毒模型,对其卵巢的病理变化和超微结构变化进行观察,检测其氧化应激和细胞凋亡情况,Real-Time q PCR法检测繁殖基因mRNA的表达量,Western Blot检测其蛋白表达量.结果发现,小花棘豆中毒和田羊卵巢指数极显著升高(P<0.01),而且卵巢表面卵泡数极显著下降(P<0.01),镜检表明中毒和田羊卵巢中初级卵母细胞溶解、消失,间质血管扩张,原位末端标记(TUNEL)发现中毒和田羊卵巢细胞和卵泡颗粒细胞发生凋亡,中毒和田羊卵巢Bax基因mRNA表达量极显著升高(P<0.01),Bcl-2基因mRNA表达量极显著下降(P<0.01).小花棘豆中毒和田羊血清和卵巢组织中超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)和过氧化氢酶(CAT)活性极显著下降(P<0.01),丙二醛(MDA)含量极显著升高(P<0.01);血清中雌二醇(E2)和孕酮(P4)含量均极显著下降(P<0.01).卵巢中FSHR和LHR mRNA表达量均极显著下降(P<0.01),其蛋白表达量均显著下降(P<0.05).研究结果表明,小花棘豆中毒可导致和田羊卵巢病理性损伤,抑制相关繁殖基因表达,促使细胞凋亡,从而抑制和田羊机体生殖功能.%A model of Oxytropis glabra DC poisoning in Hetian sheep ewes was duplicated by adding dif-ferent levels of O.glabra DC in the ration to study its toxic effect.The changes in clinical pathology and ultra-structure of ovary were observed,the oxidative stress and apoptosis in the ovary were measured,then the mRNA expression of reproductive genes were measured by Real-time q PCR,the level of reproductive genes protein expression was measured by Western Blot.The result demonstrated that the index of ovary of the poisoned sheep was very significantly higher than that of normal sheep( P<0.01) ,the follicle number on the surface of the ova-ry in the poisoned group were very significantly smaller than that in normal sheep(P<0.01).The result of he-matoxylin-eosin staining showed that the primary oocytes in the ovary dissolved,disappear,the interstitial blood vessels expanded,TUNEL showed ovary cell and ovarian follicle granular cell apoptosis,the expression of Bax mRNA increased in the poisoned sheep,which was very significant higher than that in the control( P<0.01) , and the expression of Bcl-2 mRNA decreased,very significant lower than that in the control(P<0.01).The ac-tivities of SOD,GSH-Px and CAT in the poisoned sheep were very significantly lower than those in the normal sheep( P<0.01) ,the content of MDA in the poisoned sheep was very significantly higher than that in the nor-mal sheep(P<0.01).The contents of E2 and P4 in serum in the poisoned sheep ewes were very significantly lower than those in the normal sheep( P<0.01) ,the mRNA expression levels of FSHR and LHR in the ovary were very significantly lower than those in the normal sheep( P<0.01) ,the level of reproductive gene protein expression was significantly lower than that in the normal sheep(P<0.05).The result showed that O.glabra DC poisoning affected pathological damage of ovary in Hetian sheep ewes,interfered reproductive genes mR-NA expression,promoted ovarian follicle granular cell apoptosis,thus inhibiting the reproductive function of Hetian ewes.