Division of Histology and Embryology, Joint Laboratory for Embryonic Development & Prenatal Medicine, Medical College, Jinan University, Guangzhou 510632, China.
Department of Radiology, University of California San Diego, San Diego, CA 92103, USA.
Int J Mol Sci. 2020 Apr 4;21(7):2518. doi: 10.3390/ijms21072518.
Growing evidence suggests an adverse impact of gut microbiota dysbiosis on human health. However, it remains unclear whether embryonic osteogenesis is affected by maternal gut dysbacteriosis. In this study, we observed that elevated lipopolysaccharide (LPS) levels led to skeletal developmental retardation in an established mouse model of gut microbiota dysbiosis. Using chick embryos exposed to dysbacteriosis-derived LPS, we found restriction in the development of long bones as demonstrated by Alcian blue and alizarin red staining. Micro-CT and histological analysis exhibited decreased trabecular volume, bone mineral density, and collagen production, as well as suppressed osteoblastic gene expression (, , , and ) in chick embryonic phalanges following LPS treatment. Atomic force microscopy manifested decreased roughness of MC3T3-E1 cells and poorly developed matrix vesicles (MVs) in presence of LPS. The expression of the aforementioned osteoblastic genes was suppressed in MC3T3-E1 cells as well. High-throughput RNA sequencing indicated that retinoic acid (RA) may play an important role in LPS-induced osteopenia. The addition of RA suppressed expression in MC3T3-E1 cells, as was also seen when exposed to LPS. Quantitative PCR, Western blot, and immunofluorescent staining showed that retinoic acid receptor α (RARα) was upregulated by LPS or RA treatment, while the expression of DLX5 was downregulated. CYP1B1 expression was increased by LPS treatment in MC3T3-E1 cells, which might be attributed to the increased inflammatory factors and subsequently activated NF-κB signaling. Eventually, blocking RA signals with AGN193109 successfully restored LPS-inhibited osteoblastic gene expression. Taken together, our data reveals that maternal gut microbiota dysbiosis can interfere with bone ossification, in which expression regulated by RA signaling plays an important role.
越来越多的证据表明肠道微生物群落失调对人类健康有不良影响。然而，目前尚不清楚胚胎成骨是否受母体肠道菌群失调的影响。在本研究中，我们观察到在建立的肠道微生物群落失调小鼠模型中，脂多糖（LPS）水平升高导致骨骼发育迟缓。使用暴露于肠道菌群失调衍生 LPS 的鸡胚，我们发现长骨发育受限，通过茜素红和阿尔新蓝染色证明。微 CT 和组织学分析显示 LPS 处理后鸡胚指骨的小梁体积、骨矿物质密度和胶原蛋白产生减少，以及成骨细胞基因表达（、、、和）受到抑制。原子力显微镜显示 LPS 存在时 MC3T3-E1 细胞的粗糙度降低，基质小泡（MVs）发育不良。LPS 存在时，上述成骨细胞基因的表达也受到抑制。高通量 RNA 测序表明视黄酸（RA）可能在 LPS 诱导的骨质疏松中起重要作用。RA 的添加抑制了 MC3T3-E1 细胞中 的表达，这与 LPS 暴露时的情况相同。定量 PCR、Western blot 和免疫荧光染色显示 LPS 或 RA 处理上调了 RA 受体 α（RARα），而下调了 DLX5 的表达。LPS 处理增加了 MC3T3-E1 细胞中 CYP1B1 的表达，这可能归因于炎症因子的增加，随后激活 NF-κB 信号。最终，用 AGN193109 阻断 RA 信号成功恢复了 LPS 抑制的成骨细胞基因表达。综上所述，我们的数据表明，母体肠道微生物群落失调会干扰骨矿化，其中 RA 信号调节的 表达起着重要作用。
