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为探索卵巢内瘦素信号传导缺失对卵巢自身功能和全身糖脂代谢的影响,阐明瘦素对外周生殖系统的直接作用,进行了以下研究:①对瘦素长受体缺失(LR-)的雌性B6.Cg-m+/+Leprdb小鼠和其同窝出生的有完整LR的雌性野生型C57BL/6J小鼠(LR+),在性成熟期后的12周,进行卵巢相互移植,构建实验小鼠的LR基因型组合分别为A(躯体+,卵巢-)、B(躯体+,卵巢-)、C(躯体+,卵巢+)、D(躯体-,卵巢+)、E(躯体-,卵巢-)共5组(n=5),移植后观测2个动情周期取材;②检测各自的全身糖脂代谢指标、卵巢周期和生殖激素;③卵泡刺激素(0.75IU/g)刺激试验后,用免疫组化和蛋白质印迹(WB)方法,检测卵巢内脂代谢信号蛋白包括Janus激酶(JAK2)、磷酸化的丝裂原激活蛋白激酶(p-ERK)和低密度脂蛋白受体(LDLR)的表达。结果显示:①D、E组与A、B、C3组比较,体重相差约1倍,性腺周围脂肪垫重量相差约10倍(P<0.01);血葡萄糖、低密度脂蛋白胆固醇(LDL-C)和胰岛素水平相差2～3倍(P<0.01);A、B、C3组的全身血糖脂指标相似,D和E2组的高糖脂血症升高程度也相似。②A、B、C3组小鼠均恢复正常的动情周期,而D、E2组则始终处于动情间期;小鼠16周龄时D、E组与A、B、C3组比较:卵巢重量及E2、P、FSH、LH水平显著下降(P<0.05)。③瘦素刺激后JAK2在A、B、D和E组的卵巢中表达均呈弱阳性,而C组卵巢内呈强阳性表达;FSH刺激后p-ERK和LDLR在A、B、C3组卵巢内均呈强阳性表达,而D和E组则表达下降。进而得出结论:①单纯卵巢内瘦素信号缺失并不影响卵巢自身的生殖功能、脂质代谢和全身的糖脂代谢。②瘦素受体全身缺失小鼠的卵巢功能障碍,与卵巢外的全身因素即高糖脂血症和低促性腺激素状态有关。③高糖脂血症降调节卵巢内的瘦素信号蛋白JAK2的表达,诱导小鼠卵巢脂质化和功能障碍;FSH刺激可诱导ERK磷酸化后LDLR的表达增强,缓解小鼠卵巢的脂质化和功能障碍。
In order to explore the effect of ovarian leptin signaling loss on ovarian self-function and systemic glucose and lipid metabolism, and to elucidate the direct effect of leptin on the peripheral reproductive system, the following studies were carried out: (1) To investigate the effect of leptin long receptor deletion (LR-) Female B6.Cg-m + / + Leprdb mice and their littermate-born female wild-type C57BL / 6J mice (LR +) with LR were ovariectomized at 12 weeks after sexual maturation to construct experimental small The LR genotype combinations in mice were A (body +, ovary-), B (somatic +, ovarian-), C (somatic +, ovarian +), D (somatic-, ovarian +), E -) in 5 groups (n = 5). Two estrous cycles were observed after transplantation. ② The indexes of systemic glucose and lipid metabolism, ovarian cycle and reproductive hormones were tested. ③ After stimulating with follicle stimulating hormone (0.75 IU / g) Immunohistochemistry and Western blot (WB) were used to detect the expression of lipid metabolism signal proteins in ovary including Janus kinase (JAK2), phospho-mitogen-activated protein kinase (p-ERK) and low density lipoprotein receptor (LDLR) expression. The results showed that: (1) Compared with A, B and C3 groups, the difference of body weight between D and E group was about 1 time, and the weight of fat pad around gonad differed by about 10 times (P <0.01). Blood glucose, low density lipoprotein cholesterol And insulin levels 2 to 3 times (P <0.01); A, B, C3 systemic blood glucose and lipid indicators similar to D and E2 group hyperlipoidemia increased similarly. ②A, B and C3 mice returned to normal estrous cycle, while D and E2 groups were always in estrus; at 16 weeks of age, D and E groups compared with A, B and C3: ovarian weight and E2 , P, FSH, LH levels decreased significantly (P <0.05). ③ The expression of JAK2 in ovaries of A, B, D and E groups was weakly positive when leptin was stimulated, whereas it was strongly positive in ovary of C group; the levels of p-ERK and LDLR in ovaries of A, B and C groups after FSH stimulation Showed strong positive expression within the group D and E decreased expression. And then concluded: ① simple ovarian leptin signal loss does not affect ovarian own reproductive function, lipid metabolism and systemic glucose and lipid metabolism. Â’¡ leptin receptor body loss in mice ovarian dysfunction, and ovarian systemic factors that are hyperglycemia and hypo-gonadotropin status. ③ hyperlipidemia reduced ovarian leptin signaling protein JAK2 expression, induced mouse ovarian lipid and dysfunction; FSH stimulation can induce ERK phosphorylation of LDLR increased expression of alleviating mouse ovarian lipid And dysfunction.