Impact of Tetracycline Antibiotics on Skeletal Maturation

项目总结/摘要 一生中大约1/3的骨量增加是在青春期实现的。如果这一关键的 骨骼成熟对骨骼健康和骨折风险具有终身影响。全身性四环素类（即， 二甲胺四环素、强力霉素）常用于治疗青少年痤疮，但对骨的影响尚不清楚。 进行初步研究，用临床相关剂量的多西环素或其组合治疗C57 BL/6小鼠。 米诺环素在青春期/青春期后发育期间。给予强力霉素或米诺环素给特定的- 无病原体（SPF）小鼠引起肠道细菌组中的生态失调转变和骨骼成熟受损。 在无菌（GF）条件下饲养的小鼠给予米诺环素不影响骨骼表型， 这支持四环素类药物对成熟骨骼的影响取决于肠道微生物群。胆汁酸 被鉴定为有助于肠道微生物群对骨代谢的影响的新型候选调节剂。 胆汁酸在肝脏中合成并排泄到肠道中，在那里细菌代谢它们。的 肠FXR-FGF 15轴是支持胆汁酸稳态的肠-肝内分泌轴。胆汁酸活化 肠细胞-FXR诱导FGF 15的产生，FGF 15在肝细胞-FGFR 4处发出信号以抑制CYP 7A 1 - 1。 介导的胆汁酸合成。细菌具有独特的胆盐水解酶（BSH）， 胆汁酸结合状态影响胆汁酸激活FXR的潜力。肠道细菌组成的变化 改变BSH对胆汁酸的去结合，这可以破坏肠FXR-FGF 15轴。 初步研究表明，米诺环素治疗钝化了回肠FGF 15并增强了肝脏Cyp 7a 1， 这意味着四环素类药物破坏了肠FXR-FGF 15轴。米诺环素增加血清胆汁酸， 是FXR拮抗剂，并且这种改变的胆汁酸分布减弱了培养的原代成骨细胞中的骨生成。 两个目标将测试整体假设：四环素诱导的肠道生态失调破坏肠道FXR- FGF 15轴，通过血清胆汁酸失调削弱成骨细胞， FXR信号传导或降低成骨细胞-FGFR 4信号传导的血清FGF 15活化。目标1将利用宏基因组 方法和粪便微生物群从SPF转移到GF小鼠。研究将描述米诺环素-和 多西环素诱导的肠道细菌变化改变了肠道胆汁酸的转化，从而影响FXR- FGF 15轴。目的2依赖于施用脑内特异性FXR激动剂来确定FXR-FGF 15的作用。 二甲胺四环素对骨骼的影响。他莫昔芬诱导型成骨细胞缺失小鼠将用于描述 成骨细胞-FXR/成骨细胞-FGFR 4在米诺环素抑制成骨作用中的作用。拟议 这项工作将确定四环素诱导的肠道生态失调、FXR-FGF 15轴、成骨细胞- FXR/FGFR 4信号传导和骨骼成熟。青少年通常被开全身性抗生素， 痤疮，>70%接受四环素类。肠道和肝脏疾病有更高的骨质减少患病率/ 骨质疏松症，这强调了需要确定胆汁酸在肠道微生物群对骨的影响中的作用。

PROJECT SUMMARY/ABSTRACT Roughly 1/3 of bone mass accrual during life is realized during adolescence. Disruption of this critical window of skeletal maturation has lifelong implications for bone health and fracture risk. Systemic tetracyclines (i.e., minocycline, doxycycline) are commonly used to treat acne in adolescents, but the impact on bone is unclear. Preliminary studies were performed treating C57BL/6 mice with a clinically relevant dose of doxycycline or minocycline during pubertal/postpubertal development. Administering doxycycline or minocycline to specific- pathogen-free (SPF) mice caused dysbiotic shifts in the gut bacteriome and impaired skeletal maturation. Administering minocycline to mice reared under germ-free (GF) conditions did not affect the skeletal phenotype, which supports that tetracyclines’ effects on the maturing skeleton depend on the gut microbiota. Bile acids were identified as a novel candidate regulator contributing to gut microbiota effects on bone metabolism. Bile acids are synthesized in the liver and excreted into the intestine, where bacteria metabolize them. The intestinal FXR-FGF15 axis is a gut-liver endocrine axis that supports bile acid homeostasis. Bile acid activation of enterocyte-FXR induces the production of FGF15, which signals at hepatocyte-FGFR4 to inhibit CYP7A1- mediated bile acid synthesis. Bacteria have unique bile salt hydrolases (BSHs) that differentially deconjugate bile acids. Conjugation status affects bile acids’ potential to activate FXR. Shifts in intestinal bacteria composition alter BSHs' deconjugation of bile acids, which can disrupt the intestinal FXR-FGF15 axis. Preliminary studies showed that minocycline treatment blunted ileal FGF15 and enhanced hepatic Cyp7a1, which implies tetracyclines disrupt the intestinal FXR-FGF15 axis. Minocycline increased serum bile acids that are FXR antagonists, and this altered bile acid profile attenuated osteogenesis in cultured primary osteoblasts. Two aims will test the overall hypothesis: Tetracycline-induced gut dysbiosis disrupts the intestinal FXR- FGF15 axis, which impairs skeletal maturation through dysregulated serum bile acids that attenuate osteoblast- FXR signaling or reduced serum FGF15 activation of osteoblast-FGFR4 signaling. Aim 1 will utilize metagenomic approaches and fecal microbiota transfer from SPF to GF mice. Studies will delineate how minocycline- and doxycycline-induced changes in gut bacteria alter the transformation of intestinal bile acids to affect the FXR- FGF15 axis. Aim 2 relies on administering an intestinal-specific FXR agonist to define the role of the FXR-FGF15 axis in minocycline effects on the skeleton. Tamoxifen inducible osteoblast null mice will be used to delineate the role of osteoblast-FXR / osteoblast-FGFR4 in minocycline’s actions suppressing osteogenesis. The proposed work will define the relationship between tetracycline-induced gut dysbiosis, the FXR-FGF15 axis, osteoblast- FXR/FGFR4 signaling, and skeletal maturation. Adolescents are commonly prescribed systemic antibiotics for acne, and >70% receive tetracyclines. Gut and liver conditions have a higher prevalence of osteopenia / osteoporosis, which underscores the need to define the role of bile acids in gut microbiota effects on bone.