不含咖啡因的绿茶多酚对单纯性肥胖女童性早熟的预防作用及其机制研究

Precocious puberty, with higher incidence among obese girls， has become one of the health problems over the world. Leptin is considered as the ‘permissive factor’ to initiate the puberty in the obese. Our previous experiment found that green tea polyphenols had a preventive effect on estrogen induced precocious puberty in female rats. In this study, 7-10 years old obese girls will be recruited to test the preventive effect of decaffeinated green tea polyphenols on the risk of precocious puberty by the random, placebo-controll and double blind design. The intervention group will be given decaffeinated green tea polyphenols capsule (200mg, Bid) and the control group will be given placebo. The oral treatment will be lasted for 12 weeks.Then all the subjects will be followed up every 3 months until three months after menstruation. At the baseline and after the 12 week intervention, the clinical manifestations of secondary sexual characters, the serum levels of sex hormones will be determined as the outcome variables. After controlling confounding factors, the preventive effect of the green tea polyphenols on precocious puberty or early puberty among obese girls will be analyzed. After that, using our previous established diet-induced obese rat model, the estrogen-induced precocious puberty rat model and co-culture system of fat cells and hypothalamus, together with the above human level study, we will explore the mechanism of green tea polyphenols on preventing precocious puberty by the following hypothesis: green tea polyphenols can decrease the letpin level produced by the white adipose tissue through ameliorating the status of oxidative stress and inflammation, and further affect the expression of kiss-1/GPR54 in hypothalamus,finnally delaying sexual development. The purpose of this study is to find an preventative method to precocious puberty for obese girls, and to provide a theoretical basis for application of green tea polyphenols on the prevention of precocious puberty among obese girls.

性早熟日趋严重，肥胖女童尤甚，瘦素是肥胖导致性早熟的关键因子。有效预防措施是患儿及家长迫切需要，而相关研究极少。课题组前期发现绿茶多酚对雌性大鼠性早熟具预防作用，且已有证据证明不含咖啡因的绿茶多酚对儿童安全无害。本课题以7-10岁单纯性肥胖女童为研究对象，采用双盲随机对照试验，用不含咖啡因的绿茶多酚胶囊进行口服干预12周，之后每隔3个月随访一次直至其月经初潮后三个月。以青春发育指标为结局变量，分析绿茶多酚对单纯性肥胖女童性早熟预防作用。检测受试儿童血清瘦素、kisspeptin、氧应激和炎症指标，并建立茶多酚干预肥胖性早熟大鼠模型以及脂肪细胞与下丘脑共培养体系，深入研究绿茶多酚是否通过改善肥胖者氧应激与炎症影响瘦素生成，进而影响下丘脑kiss-1/GPR54等性发育关键分子表达，最终影响青春发育。本研究为饮食预防肥胖女童性早熟寻找有效干预手段，为食用茶多酚预防肥胖女童性早熟提供理论依据。

性早熟日趋严重，肥胖女童尤甚，有效预防措施是患儿及家长迫切需要，而相关研究极少。本课题招募90名6-10岁单纯性肥胖女童，采用双盲随机对照试验，用不含咖啡因的绿茶多酚胶囊进行口服干预12周（每日给予400mg茶多酚胶囊），通过体格检查，身体成分分析，B超检测子宫、卵巢、乳腺组织，骨龄检测，生化指标检测（包括性激素、炎症因子、性发育关键因子等）来收集研究对象干预前与12周干预后的肥胖相关指标、性发育指标；结果发现不含咖啡因的绿茶多酚对肥胖女童肥胖指标和性发育指标均有改善作用，且女童补充12周400mg/d的绿茶多酚安全且无不良反应。在此基础上，利用动物实验探究表没食子儿茶素没食子酸酯(EGCG，茶叶中生理活性最强、含量最高、研究最广泛的儿茶素成分)对高脂饮食雌性大鼠性发育的影响及其分子机制。60只21日龄离乳雌性SD大鼠随机分成4组,分别为正常对照组、普通饲料EGCG干预组、高脂饮食(HFD)组和高脂饮食EGCG干预组。选取27日龄(青春期启动前)、33日龄(青春期启动时), 36日龄(青春期启动后)为取材点，各组分别处死5只大鼠。Elisa法检测血清中性发育相关的关键因子浓度(Leptin、Kisspeptin、NKB、Ghrelin、IGF-1和GH)；RT-PCR法和Western blot 法检测大鼠下丘脑Kisspeptin及其受体Kiss1R、NKB及其受体NK3R mRNA和蛋白的表达水平；免疫组化法检测大鼠卵巢NKB蛋白的表达水平。结果，EGCG干预可显着延迟HFD喂养大鼠的阴道开放时间。 在33日龄，EGCG干预可显着降低HFD喂养大鼠血清NKB、LH水平、卵巢NKB蛋白表达和子宫内膜厚度，显著增加NKB/NK3R的mRNA和蛋白表达。各组大鼠下丘脑Kisspeptin及其受体Kiss1 R mRNA水平均无统计学差异。动物实验进一步验证了绿茶多酚对肥胖相关性早熟的预防作用，其作用机制并未通过预期的对Kisspeptin/Kiss1R信号通路的调节作用，而是可能通过上调下丘脑NKB/NK3R的表达，从而影响血清性激素水平有关。本项目首次通过临床研究明确不含咖啡因的绿茶多酚可以预防肥胖女童性早熟并初步探明其作用途径，为绿茶多酚预防肥胖女童性早熟提供理论依据。