茶多酚EGCG调控肠道厚壁菌抑制IGF-1在抗前列腺增生中的作用及机制研究

Benign prostatic hyperplasia (BPH) is one of the most common diseases in middle-aged men. With the aging of the population, the prevalence of BPH is on the rise. Risk factors for BPH include metabolic syndrome (MetS), obesity, diabetes, etc. Gut microbiota is closely related to MetS; Ghrelin is involved in the progression of MetS; Intestinal microflora is associated with Ghrelin secretion. Tea polyphenol as EGCG has been reported to regulate intestinal microbiota in patients with MetS. However, the influence and mechanism of EGCG on MetS-BPH remains unclear. In our previous study, we performed 16S rRNA sequencing analysis on the intestinal flora from MetS-BPH mice. The results showed that Firmicutes in BPH mice were significantly higher than normal mice, and Ghrelin content of plasma and prostate tissue of both BPH and normal recipient mice transplanted with BPH intestinal bacteria was significantly increased. Treatment with EGCG could inhibit Firmicutes from BPH mice significantly in vitro. The expression of Ghrelin in plasma or prostate tissues of BPH patients was higher than that from normal control. Ghrelin can significantly promote the proliferation of prostate cells of RWPE-1 and WPMY-1 while inhibit their apoptosis. Based on our previous study, we speculate that EGCG can negatively regulate BPH proliferation by inhibiting intestinal Firmicutes and reducing the gastric secretion of ghrelin. In this study, we aimed to investigate the effect of EGCG on intestinal Firmicutes and ghrelin in the development of BPH in MetS-BPH animal model and RWPE-1 &WPMY-1 cells vivo and in vitro, the mechanism investigation is also involved in our study.

良性前列腺增生（BPH）是中老年男性最常见的疾病之一，其危险因素包括代谢综合征（MetS）、肥胖、糖尿病等。肠道微生物与MetS密切相关；而IGF-1与糖尿病、肥胖及MetS相关；肠道厚壁菌影响IGF-1分泌。已有报道茶多酚EGCG能够调控MetS患者肠道微生物群，但EGCG对MetS型BPH影响尚不清楚。我们前期对MetS-BPH小鼠肠道菌群16SrRNA测序显示其肠道厚壁菌数量显著高于正常小鼠；EGCG干预显著降低BPH小鼠肠道厚壁菌数量。BPH与移植BPH肠道菌液的受体小鼠血浆、前列腺组织中IGF-1显著增高。IGF-1在BPH患者血浆及前列腺组织中高表达；体外显著促进前列腺细胞增殖。我们推测EGCG能够通过调控肠道厚壁菌，影响IGF-1从而抑制BPH。本课题拟通过体内MetS-BPH动物模型、体外前列腺细胞为对象，研究EGCG、肠道厚壁菌及IGF-1在BPH进展中的作用及机制。

已有研究表明代谢综合征、肠道微生物群在BPH的发病机理中发挥重要作用。我们前期的研究表明代谢综合征良性前列腺增生小鼠肠道菌群中厚壁菌表达量异常上调，IGF-1在BPH患者的血浆及前列腺组织中高表达，细胞功能学实验显示GF-1能够显著促进前列腺上皮细胞 RWPE-1和间质细胞 WPMY-1的增殖，抑制细胞凋亡。表没食子儿茶素-3-没食子酸酯（EGCG）是绿茶提取物茶多酚中含量最高的活性成分，EGCG能够显著降低MetS大鼠的IGFs，对睾酮诱导和MetS大鼠的BPH具有保护性作用。EGCG干预抑制离体分离的良性前列腺增生肠道微生物厚壁菌。本研究在前期研究的基础上，探索EGCG和IGF-1对前列腺增生发展的影响以及下游的作用机制，为EGCG治疗BPH提供新的治疗思路。本课题通过肠道菌群16S核糖体RNA（16S rRNA）测序分析肠道菌群的构成、分离BPH小鼠肠道菌群移植入正常小鼠证实BPH肠道菌群可以促进前列腺组织增生型改变，同时通过免疫组织化学染色，Western blot以及ELISA检测手段表明IGF-1及其受体在BPH肠道菌液移植的正常小鼠血浆和前列腺组织中表达增加；进一步通过IGF-1及其拮抗剂Linsitinib干预，CCK8、BD Matrigel 3D以及流式细胞术检测前列腺上皮细胞 RWPE-1和间质细胞 WPMY-1的增殖和凋亡水平，结果表明IGF-1促进前列腺细胞的增殖，荧光定量PCR结果显示，IGF-1促进BPH小鼠肠道厚壁菌的含量，增加前列腺指数，以上结果说明IGF-1可能通过调节肠道微生物厚壁菌含量促进BPH的发展。通过EGCG体内外干预代谢综合征前列腺增生小鼠中包括厚壁菌在内的肠道微生物的构成，结表明EGCG可以降低厚壁菌的含量，降低BPH菌液移植正常小鼠前列腺指数以及血浆和前列腺组织中IGF-1的含量，进一步通过免疫组化和Western blot 检测其下游通路关键蛋白PI3K和mTOR的表达水平，结果表明EGCG可以降低PI3K和磷酸化PI3K以及mTOR的表达水平，说明IGF-1/PI3K/mTOR可能是EGCG调控BPH疾病发展的下游通路，为其临床治疗提供新的依据。