褪黑素与卵泡刺激素对小鼠卵巢卵泡发育及其血管生成的影响

Effects of melatonin and FSH on ovarian follicle development and angiogenesis in mice

为了探讨褪黑素(Mel)与卵泡刺激素(FSH)对雌性动物卵巢卵泡发育及其血管的影响,选取40只3周龄ICR雌性小鼠随机均分为4组,即:对照组(Control组)、褪黑素组(Mel组)、卵泡刺激素组(FSH组)和卵泡刺激素+褪黑素组(FSH+Mel组),腹腔分别连续注射生理盐水、Mel、FSH和FSH+Mel后的第13天8:00采集血样及卵巢。每组采集5只小鼠的左侧卵巢进行固定,HE染色观察卵泡发育状况,免疫荧光技术观察卵泡血管状况;剩余卵巢利用Western blot测定Bax、Bcl-2的蛋白表达量。结果:与对照组相比,显著升高的有:Mel组和FSH组卵泡直径50~150μm卵泡数,FSH组和FSH+Mel组血清雌二醇、孕酮浓度,Mel组、FSH组和FSH+Mel组Bcl-2蛋白表达,Mel组>250~350μm卵泡CD34荧光强度,FSH组和FSH+Mel组>350~450μm卵泡毛细血管数和CD34荧光强度(P<0.05);显著降低的有:Mel组、FSH组和FSH+Mel组Bax蛋白和卵泡闭锁率(P<0.05);极显著升高的有:FSH组和FSH+Mel组卵巢重量,FSH+Mel组50~150μm卵泡数,FSH组和FSH+Mel组>250~350μm、>350~450μm、>450μm卵泡数,Mel组和FSH+Mel组50~150μm卵泡毛细血管数及CD34荧光强度,Mel组>250~350μm卵泡毛细血管数,FSH组和FSH+Mel组>250~350μm和>450μm卵泡毛细血管数及CD34荧光强度(P<0.01)。结论:连续注射褪黑素能促进卵巢50~150μm次级卵泡发育及其血管生成,促进>250~350μm有腔卵泡血管发育,FSH可促进卵巢50~150μm次级卵泡发育以及>250μm有腔卵泡发育及其血管生成;褪黑素和FSH均能显著升高抗凋亡蛋白Bcl-2的表达和显著降低凋亡蛋白Bax的表达,降低卵泡闭锁率。

In order to investigate the effects of melatonin(Mel)and follicle-stimulating hormone(FSH)on the ovarian follicular development and blood vessels of female animals,403-week-old female ICR mice were used and randomly divided into 4 groups,namely:the Control Group,the Mel,FSH,FSH+Mel groups.Blood samples and ovaries were collected at 8:00 a.m.on the 13th day after successive intraperitoneal injections of normal saline,Mel,FSH and FSH+Mel,respectively.The left ovaries of 5 mice in each group were fixed,the development of follicles was observed by HE staining,and the vascular conditions of follicles were observed using immunofluorescence techniques.The protein expression levels of Bax and Bcl-2 in the remaining ovaries were determined by Western blot.The results were that,compared with the control group,the 50-150μm follicle number of the Mel and FSH Groups,the serum estradiol and progesterone concentrations of the FSH and FSH+Mel Groups,the bcl-2 protein expression of the Mel,FSH and FSH+Mel Groups,the CD34 fluorescence intensity of>250-350μm follicle in the Mel Group,the follicular capillary number and CD34 fluorescence intensity of>350-450μm in the FSH and FSH+Mel Groups were significantly increased(P<0.05);that the Bax protein and follicular atresia rate were significantly decreased in the Mel,FSH and FSH+Mel Groups(P<0.05);that the ovarian weight of the FSH and FSH+Mel Groups,the 50-150μm follicle number of the FSH+Mel Group,the>250-350μm,>350-450μm,>450μm follicle numbers of the FSH and FSH+Mel groups,the follicular capillary number and CD34 fluorescence intensity of 50-150μm in the Mel and FSH+Mel groups,the follicular capillary number of>250-350μm in the Mel Group,the follicular capillary number and CD34 fluorescence intensity of>250-350μm and>450μm in the FSH and FSH+Mel Groups were extremely significantly increased(P<0.01).In conclusion,successive injections of Mel promoted the development of 50-150μm secondary follicles and their angiogenesis,and the development of>250-350μm antral follicles;and FSH promoted the development of 50-150μm secondary follicles and the development of>250μm antral follicles and their angiogenesis.Both Mel and FSH significantly increased the expression of anti-apoptotic protein Bcl-2 but significantly decreased the expression of apoptotic protein Bax,and reduced the follicular atresia rate.