Division of Histology and Embryology, International Joint Laboratory for Embryonic Development & Prenatal Medicine, Medical College, Jinan University, Guangzhou, China.
Medical Records Department, Quality and Safety Management Office, The First Affiliated Hospital of Jinan University, Guangzhou, China.
FASEB J. 2022 Jan;36(1):e22113. doi: 10.1096/fj.202101343R.
Intrauterine infection would harm a developing embryo/fetus, thereby increasing the risk of developmental malformation. But, whether or not the infection-induced inflammation affects neural crest development still remains obscure. In this study, we employed meta-analysis to demonstrate the potential correlation between infection-induced inflammation and craniofacial anomalies, which was usually derived from the problems in neural crest cell development. The correlation was further verified by inflammatory cytokine release and the activation of nuclear factor kappa-light-chain enhancer of activated B cells signaling in lipopolysaccharide-treated HH10 chicken embryos. In such an inflammatory condition, AP-2α- and Pax7-labeled pre-migratory and migratory neural crest cells in HH10 chicken embryos were significantly less than the ones in control. The bioinformatics analysis of RNA-seq data demonstrated that the principal differential gene expression occurred in transforming growth factor-beta (TGF-β) signaling pathway, which was confirmed by the subsequent experimental results of quantitative PCR and immunofluorescent staining. Under this inflammatory circumstance, whole-mount in situ hybridization, immunofluorescence, and quantitative PCR showed the gene expression changes of key EMT-related transcription factors including upregulated Msx1, downregulated Slug, and FoxD3, as well as adhesion molecules and extracellular matrix protein including upregulated Cadherrin6B, E-cadherin, N-cadherin, and Laminin at the dorsal portion of neural tube of HH10 chicken embryos. Meanwhile, the bioinformatics analysis of RNA-seq data also manifested the differential gene expressions relevant to cell proliferation, which was confirmed by proliferating cell nuclear antigen Western blot data and co-immunofluorescence staining of human natural killer-1 and phosphorylated histone H3. In brief, this study revealed for the first time that the double-edged sword role of TGF-β signaling pathway between intrauterine inflammation (protective role) and cranial neural crest development (harmful role).
宫内感染会损害发育中的胚胎/胎儿，从而增加发育畸形的风险。但是，感染引起的炎症是否影响神经嵴发育仍然不清楚。在这项研究中，我们采用荟萃分析来证明感染引起的炎症与颅面异常之间的潜在相关性，这种相关性通常源自神经嵴细胞发育的问题。通过在脂多糖处理的 HH10 鸡胚中释放炎性细胞因子和激活核因子κ轻链增强子的 B 细胞信号转导，进一步验证了这种相关性。在这种炎症状态下，AP-2α和 Pax7 标记的前迁移和迁移性神经嵴细胞在 HH10 鸡胚中的数量明显少于对照组。RNA-seq 数据的生物信息学分析表明，主要的差异基因表达发生在转化生长因子-β（TGF-β）信号通路中，随后的定量 PCR 和免疫荧光染色实验结果证实了这一点。在这种炎症环境下，整体原位杂交、免疫荧光和定量 PCR 显示了关键 EMT 相关转录因子的基因表达变化，包括上调的 Msx1、下调的 Slug 和 FoxD3，以及粘附分子和细胞外基质蛋白，包括上调的 Cadherrin6B、E-钙粘蛋白、N-钙粘蛋白和 Laminin 在 HH10 鸡胚神经管的背侧。同时，RNA-seq 数据的生物信息学分析也显示了与细胞增殖相关的差异基因表达，这通过增殖细胞核抗原 Western blot 数据和人自然杀伤-1 和磷酸化组蛋白 H3 的共免疫荧光染色得到证实。总之，这项研究首次揭示了 TGF-β信号通路在宫内炎症（保护作用）和颅神经嵴发育（有害作用）之间的双刃剑作用。
