内脏脂肪介导中年雌性大鼠黄体生成素高峰分泌异常的研究

Intra-abdominal visceral fat mediates the ovarian-steroid-induced luteinizing hormone surge dysfunction in middle-aged female rats

目的研究增龄性内脏脂肪增加可能通过中枢机制介导黄体生成素(LH)分泌异常,探索早期女性生殖衰老发生的相关因素及干预策略。方法阴道涂片法筛选性周期规律的36只中年(9~11个月)雌性Sprage-Dawley大鼠,随机分为减除内脏脂肪(visceral fat removal,VFR)组和假手术组,VFR组大鼠经腹部手术移除内脏(包括卵巢及子宫、肾周及膀胱周围)脂肪,假手术组大鼠于腹部正中切口暴露脏器后即刻缝合。3周后所有大鼠均接受去卵巢手术,术后1周给予外源性激素注射模拟雌激素正反馈。观察并比较大鼠体质量、体脂百分比及葡萄糖耐受情况;经颈静脉插管连续取血测定LH分泌水平;免疫组织化学双标记检测下丘脑促性腺激素释放激素(GnRH)神经元表达c-Fos的百分数;荧光定量PCR检测下丘脑前脑Igf1、Igf1r、Kiss1、Kiss1r、Nr1、Nr2b、Slc17a6和Slc32a1的mRNA表达。结果内脏脂肪移除术后4周,VFR组大鼠外源性激素诱导的总LH分泌量[(4 658.0±1 312.0)μg/L]及LH分泌高峰[(16.0±4.5)μg/L]明显高于假手术组[(1 906.0±241.1)μg/L;P=0.04;(6.1±0.7)μg/L,P=0.03];下丘脑GnRH神经元表达c-Fos百分数(53.9%±4.2%)较假手术组增加了1.4倍(36.2%±5.9%,P=0.01);与术前基线数值相比,大鼠的体质量、体脂百分数及葡萄糖耐受在内脏脂肪移除术后差异无统计学意义(P>0.05);内脏脂肪移除不改变下丘脑前脑Igf1、Igf1r、Kiss1、Kiss1r、Nr1、Nr2b、Slc17a6和Slc32a1的mRNA表达。结论减少内脏脂肪显著增加中年雌性大鼠外源性激素诱导LH高峰分泌及下丘脑GnRH神经元的激活,提示内脏脂肪可能通过中枢调控机制影响雌激素正反馈作用介导女性生殖衰老的发生,减少内脏脂肪可能有助于改善早期生殖衰老的特征表型。

Objective To investigate central mechanisms of age-related increased abdominal visceral fat and luteinizing hormone (LH) surge dysfunction in middle-aged females and explore possible intervention strategies for female reproductive aging. Methods Thirty-six cycling middle-aged Sprage-Dawley rats were selected by vaginal smears and randomly assigned to sham surgery or intra-abdominal/pelvic adipectomy (visceral fat removal, VFR) group. A mid-ventral abdominal incision cut was provided for VFR and sham rats, intra-abdominal visceral fat (including the gonad, perivesical and perinephric fat pads) was bilaterally excised for VFR rats but organs were exteriorized and placed back into abdominal cavity before skin closed for sham controls. Rats were allowed to recover for three weeks before they were gonadectomized (OVX). One week after OVX, estradiol and progesterone were primed to induce a LH surge. Serial blood sampling by jugular vein catheterization was used to assess the onset and magnitude of the LH surge, body weight and composition and glucose tolerance were determined as well. To quantify GnRH neuronal activation, as determined by c-Fos co-expression, rats were perfused and performed with immunohistochemistry. Real-time PCR was used to quantify hypothalamic neurotransmitters gene expression known to change with aging including Igf1, Igf1r, Kiss1, Kiss1r, Nr1, Nr2b, Slc17a6 and Slc32a1. Results Rats in VFR group exhibited a significant elevation of peak [(16±4.5)μg/L] and total LH release [(4 658.0±1 312.0)μg/L] and 1.4-fold increase of GnRH neuronal activation (53.9%±4.2%) compared with sham operated females [(6.1±0.7)μg/L, P=0.03;(1 906.0±241.1)μg/L, P=0.04;36.2±5.9, P=0.01]. However, no significant difference in glucose tolerance, body weight or body composition was found for VFR or sham females when compared with their baseline values obtained before surgery. VFR did not change neurotransmitters gene expression in the anterior hypothalamus when compared with sham females. Conclusion Intra-abdominal visceral fat removal markedly increased ovarian-steroids-induced LH surge and GnRH neuronal activation in middle-aged females, this study suggests that increased intra-abdominal visceral fat in middle-aged females may contribute to female reproductive aging through adversely affects estradiol positive feedback action on the hypothalamus.