【作者】 刘晓丽；

【导师】 王人卫；

【作者基本信息】 上海体育学院 ， 运动人体科学， 2012， 博士

【摘要】 研究目的本研究以能量负平衡是引起运动性月经失调的关键因素为理论基础，拟在不改变运动强度的情况下，通过补糖增加能量摄入，改善大鼠体内能量状况，探讨补糖是否会延缓运动大鼠动情周期抑制出现的时间和改善运动大鼠动情周期抑制状态，同时探讨能量代谢指标与性激素指标间的相关性，为能量干预和防治运动性月经失调提供实验性依据。研究方法两月龄SD大鼠72只，随机分为8组：空白对照组(Con组，作为阴性对照组)、闭经模型组(M组，作为阳性对照组)、6周低聚糖补充运动组（6D组，在6周递增负荷运动中除正常饮食外增加30%的低聚糖补充）、6周葡萄糖补充运动组（6G组，在6周递增负荷运动中除正常饮食外增加30%的葡萄糖补充）、低聚糖干预组（3D组，在继续运动的基础上增加普食的30%低聚糖补充）、葡萄糖干预组（3G组，在继续运动的基础上增加普食的30%葡萄糖补充）、普食运动组（3U组，普食维持同等强度的运动）和安静恢复组（3Q组）。其中6D组和6G组是在6周递增负荷运动中除正常饮食外增加30%的能量补充，探讨在递增负荷运动中增加能量补充是否能延缓大鼠动情周期抑制出现的时间；3D组、3G组、3U组和3Q组是在大鼠经过6周递增负荷训练成功建立动情周期抑制模型后采用不同的恢复方式进行3周恢复，探讨各种恢复方式对动情周期抑制大鼠动情周期的改善状况。每天晚上对大鼠进行阴道脱落细胞学检查，确定大鼠动情周期状况。大鼠的训练方案参照Bedford动物运动负荷标准进行，3D组、3G组、3U组运动9周，3Q组运动6周后安静恢复；M组、6D组和6G组运动6周，在其它组运动3周后开始按训练计划运动。最后一次训练结束后加训一次，然后休息36小时后处死，其中最后8小时处于禁食期。下腔静脉取血6-8ml，分离血清后检测促性腺素释放激素(GnRH)、促卵泡激素(FSH)、黄体生成素(LH)、泌乳素(PRL)、孕激素(P)、雌二醇(E2)、睾酮(T)、皮质酮(C)、瘦素(leptin)、胃饥饿素(ghrelin)、胰岛素(insulin)和三碘甲状腺原氨酸(T3)，取下丘脑冰冻后测试瘦素长臂受体(Ob-Rb)、神经肽Y(NPY)和POMC的mRNA表达，取各大鼠右侧卵巢石蜡包埋后采用免疫组化方法测试Bax、Bcl-2、P53蛋白的表达。研究结果1.6G组和6D组与Con组相比6G组和6D组体脂率、leptin水平、IGF-1指标降低有统计学意义；6G组和6D组PRL水平、ghrelin水平升高有统计学意义；6D组的T3水平与Con组相比没有差异但6G组T3显著低于Con组，6D组和Con组间NPY mRNA表达差异没有统计学意义但6G组NPY mRNA水平显著高于Con组，6D组与Con组相比T/C降低差异有统计学意义，6D组和Con组间Bax间差异无统计学意义但6G组Bax水平显著高于C组。2.6G组和6D组与M组相比6G组和6D组体重增加、体脂率增加、leptin增加、Ob-Rb mRNA表达增加、P水平增加有统计学意义；6G组和6D组ghrelin降低有统计学意义；6D组Bax和p53蛋白的表达显著低于M组。3.6G组和6D组间的比较6G组NPY mRNA水平显著低于6D组。4.3D组和3G组与Con组相比3D组和3G组体重降低、体脂率降低、leptin降低较Con组有统计学意义；3G组P水平显著低于Con组。5.3D组和3G组与M组相比3D组和3G组体重增加、体脂率增加、leptin水平增加、T3水平增加、IGF-1水平增加、Ob-Rb mRNA表达增加、GnRH水平增加、E2水平增加、P水平增加有统计学意义；ghrelin水平降低、NPY mRNA表达降低、PRL水平降低有统计学意义。6.3D组和3G组与3Q组相比3D组和3G组大鼠体重较低与3Q组相比差异有统计学意义。7.3D组和3G组与3U组相比3D组和3G组leptin水平、T3水平、IGF-1水平、E2水平、P水平增加较3U组差异有统计学意义；3D组和3G组NPY mRNA表达降低；3G组较3U组T/C值升高有统计学意义，3D组Bax降低较3U组差异有统计学意义。8.3D组和3G组相比3G组IGF-1水平较3D组高，且差异有统计学意义。9.3U组大鼠动情周期恢复时间延长较3D组、3G组、和3Q组有统计学差异。10.外周能量代谢指标和血C水平对性激素指标的累积贡献率达到75%，贡献率较高，其中leptin、ghrelin、T3的影响较高，性激素指标中E2、LH、PRL的影响较高。研究结论1.在运动过程中增加糖的补充能降低ghrelin水平，增加leptin水平、T3水平、IGF-1水平，调节中枢能量代谢状态，并能调整体内性激素水平，减少卵巢颗粒细胞促凋亡因子的表达，从而改善生殖功能，减缓大鼠动情周期抑制的发生。并且不同种类的糖补充效果无差异。2.给予运动性动情周期抑制的运动大鼠糖的补充能及时调整体内能量状态，改善中枢能量代谢状态，并能调节体内性激素水平，减少卵巢颗粒细胞促凋亡因子的表达，改善大鼠动情周期抑制状态；通过调整训练计划也能改善大鼠动情周期抑制状态，但需要的时间延长；安静恢复能较快改善动情周期抑制状态，但能引起体重较快增加。3.能量代谢指标及血C是影响性激素变化的主要指标，在能量代谢指标中特别是leptin、ghrelin、T3的影响较明显。更多还原

【Abstract】 ObjectiveThe negative energy balance is a key factor caused exercise-induced menstrualdisorder, as is the theoretical foundation of this study. This research tried to explorethe possibility that carbohydrate supplement would delay the time of estrous cyclerestrain and improve the status of estrous cycle restrain in rats under the same exerciseintensity. In addition, the correlation between the energy metabolism index and sexhormones index is to be discussed.MethodDivided72two months old SD rats randomly into eight groups: the blank group(Congroup, the negative control group), amenorrhoea model group(M group, positivecontrol group),6D group(the oligosaccharides in which the energy were30%ofnormal diet added to rats in six weeks’ increasing load exercise),6G group(30%energy of normal diet added to rats by glucose in six weeks’ increasing load exercise),3D group(30%energy of normal diet added to rats by oligosaccharides, keeping onexercising till the end of9weeks),3G group(30%energy of normal diet added torats by glucose, keeping on exercising till the end of9weeks),3U group(normal diet,keeping on exercising, same intensity) and3Q group (normal diet, resting after sixweeks’ increasing load exercise).6D group and6G group are to explore thepossibility of energy supplement whether delay the time of estrous cycle restrain ornot;3D group, the3G group,3U group and3Q group are to explore the differenteffects of different recovery methods to improve the status of estrous cycle restrain.Vaginal exfoliative cytological examination was been done to rats every night tofigure out their estrous cycle conditions. With the reference to Bedford animalexercise load standards,3D group,3G group,3U group had done exercises for nineweeks,3Q group recovered quietly after six weeks; M group,6D group and6G haddone exercises for nine weeks,3weeks behind other groups. One more exercise wasadded to the rats’ last training. Then all rats were put to death after a36hours break,in which the last eight hours without diet; firstly took6-8ml blood from inferior venacava, after separating serum, the gonadotropin releasing hormones(GnRH),follicle-stimulating hormone (FSH), corpus luteum erythropoietin (LH), prolactin(PRL), progesterone (P), estradiol (E2), testosterone (T), cortex ketone (C), leptin,ghrelin, insulin and3,5,3’-triiodothyronine (T3) were tested; secondly leptin long armreceptor (Ob-Rb), neuropeptide Y (NPY) and the POMC mRNA expression ofhypothalamus were tested; thirdly using immunohistochemical method to test Bax,Bcl-2, P53protein expression of the right ovary.Results1.6G group and6D group compared with Con groupThe body fat, leptin level and IGF-1level of6G and6D groups decreasedsignificantly; The T3level and the ratio of T and C of6D group was decreasedsignificantly to Con group; The NPY mRNA and Bax level of6G group increasedsignificantly to Con group.2.6G group and6D group compared with M groupThe weight gain, leptin, Ob-Rb mRNA, P level of6G and6D groups increasedsignificantly; the ghrelin of6G and6D groups decreased significantly; comparedto Con group, the p53and bax of6Dgroup decreased significantly.3.6G group compared with6D groupThe NPY mRNA level of6G group was less than6D group significant. 4.3D group and3G group compared with Con groupThe weight gain, body fat rate and leptin levels decreased significantly.5.3D group and3G group compared with M groupThe body weight, body fat rate, leptin, T3, IGF-1, Ob-Rb mRNA, GnRH, E2andP level increased significantly. However, the ghrelin, NPY mRNA expression andPRL level decreased significantly.6.3D group and3G group compared with3QThe body weight of3D and3G groups decreased significantly.7.3D group and3G group compared with3U groupThe leptin, T3, IGF-1, E2and P levels of3Dand3G groups increased significantly.However, the NPY mRNA expression of3Dand3G groups decreasedsignificantly.8.3D group compared with3G groupThe IGF-1level of3D group decreased significantly.9. Compared with3D,3G and3Q groups, the recovery time of the oestrous cycle of3U rats prolonged.10. Cumulative contribution rate of Peripheral energy metabolism index and blood Clevel to sex hormones index contributed up to75%. And the leptin and ghrelin, T3effect were higher of energy metabolism index, LH E2, PRL influence were higherin sex hormones.Conclusion1. The carbohydrate supplement to rats during exercise can reduce ghrelin level,while raise the level of leptin, T3level, IGF-1level, by which center energymetabolism condition can be regulated, and the internal sex hormone level beadjusted and the ovarian granulosa cell promoting apoptosis gene expressionreduce accordingly, which improve the reproductive function and delay the timeof estrous cycle restrain in rats consequently. And it is showed that different kindsof carbohydrate supplement have no differences on effect.2. The carbohydrate supplement to rats at the time of estrous cycle restrain canadjust their energy condition in time, improve the central energy metabolismcondition, and adjust the internal sex hormone level and reduce the expression ofpromoting apoptosis gene in ovarian granulosa cell, thereby it can improve thestatus of rats’ estrous cycle restrain; the adjusting of the training programs canalso improve the status of rats’ estrous cycle restrain, although more time isneeded; Quiet Recovery could fast improve the status of rats’ estrous cyclerestrain, while it cause weight-gain rapidly.3. Energy metabolism index and blood C are the key indexes that effect the sexhormone changes, in which the effect of leptin, ghrelin and T3are especially moreobvious.更多还原