基于肠道益生菌的杜仲抗骨质疏松机制研究

The ongoing increase of incidence of osteoporosis (OP) has threatened global health care. Kidney is considered as the origin of congenital constitution in traditional Chinese medical science, which is able to generate marrow and dominate bone. Kidney tonifying traditional Chinese medicine (TCM) has been widely used for clinical prevention and treatment of OP. Pharmacological studies have shown that Eucommia ulmoides Oliv. induced anti-OP effects. However, pharmacokinetic studies found that its blood concentration of active compounds was low, which were not sufficient to explain the mechanism of its anti-OP action. We preliminarily revealed that E. ulmoides Oliv. could promote the growth of probiotics, while the mode of action is unclear. Therefore, we aim to study the probiotics-dependent function and mechanism of E. ulmoides Oliv. for preventing OP in SAMP6 mice. Biological detection is used for revealing the anti-OP function. High-throughput sequencing technology is performed for indicating the effects of E. ulmoides Oliv. on gut microbiota (GM). In order to explore the target effect of E. ulmoides Oliv. on probiotics, the targeted microorganisms are isolated and identified from fecal sample. Afterwards, promoting mechanism of E. ulmoides Oliv. on probiotics is investigated by microbiological and transcriptomics analysis. Finally, culture of targeted probiotic strains and targeted fecal are transplanted to SAMP6 mice and the intestinal probiotics-dependent effects of E. ulmoides Oliv. on OP are verified. Above all, our findings would provide scientific basis for the development and clinical rational administration of E. ulmoides Oliv..

骨质疏松（OP）发病率日益增高，已成为严重的全球性公众健康问题。中医认为肾为先天之本，肾主骨生髓，临床上也多用补肾中药防治OP。前期研究发现杜仲具有抗OP作用，然而药代动力学研究发现其有效成分血药浓度低，不足以解释其抗OP的临床药效。实验证实杜仲对肠道益生菌生长具有促进作用，基于此，我们提出杜仲通过调节肠道益生菌而发挥抗OP作用的工作假说。课题以杜仲为研究对象，对SAMP6骨质疏松小鼠进行抗OP机制研究。通过生物学检测明确杜仲对OP的防治作用；采用高通量测序技术阐明杜仲对肠道菌群（GM）的影响；根据杜仲对肠道中益生菌的影响，分离、鉴定靶点肠道益生菌；结合微生物学、转录组学技术分析杜仲影响靶点肠道益生菌生长及代谢的机制；通过粪菌移植技术验证杜仲基于肠道益生菌治疗OP的靶点机制。本研究为杜仲的开发及临床合理用药提供科学依据。

骨质疏松发病率日益增高，已成为严重的全球性公众健康问题。中医认为肾为先天之本，肾主骨生髓，临床上也多用补肾中药防治骨质疏松。前期研究发现杜仲具有抗骨质疏松作用，然而药代动力学研究发现其有效成分血药浓度低，不足以解释其抗骨质疏松的临床药效。本项目以杜仲为研究对象，重点探讨其对肠道益生菌的生长及对骨质疏松的影响。通过生物学检测明确杜仲对雌激素缺乏导致和老年性小鼠骨质疏松的防治作用；采用高通量测序技术阐明杜仲对肠道菌群的影响；通过粪菌移植技术验证杜仲基于肠道益生菌治疗骨质疏松的靶点作用。根据杜仲对肠道中益生菌的影响，分离、鉴定靶点肠道益生菌；结合微生物学、转录组学技术分析杜仲影响靶点肠道益生菌生长及活性机制。. 研究表明，不同提取溶剂提取杜仲叶的成分不同，研究发现杜仲叶水提物对常见益生菌，尤其是乳杆菌生长具有较强的促进作用。在动物模型上给予杜仲叶水提物干预后，小鼠骨相关性能及肠道菌群多样性均有所改善，还上调了乳杆菌丰度及短链脂肪酸含量。将乳杆菌剂给予骨质疏松小鼠后，发现其同样具有抗骨质疏松和调节肠道菌群的功效。基于此得出杜仲通过促进乳杆菌的丰度而发挥治疗骨质疏松的作用。从小鼠粪便和杜仲提取物中分离乳杆菌等益生菌株，选择具有酸性耐受性、胆盐耐受性的菌落EU01、EU02、EU03和EU04进行分子生物学鉴定。研究表明EU01菌株为嗜热链球菌，EU02菌株为食窦魏斯氏菌，EU03菌株为约氏乳杆菌，EU04菌株为鼠乳杆菌。转录组学研究表明，杜仲叶水提物对菌株EU01干预后，66个差异表达基因上调，229个下调，其中DNA修复和合成等益生活性相关功能基因显著上调，表明杜仲促进了该菌的益生活性。. 本研究通过研究杜仲影响骨质疏松小鼠肠道益生菌生长及生物活性，筛选分离得到靶点益生菌，解析杜仲基于肠道益生菌防治骨质疏松的靶点机制，为杜仲的药物开发和合理用药提供科学依据。