Wnt1-Cre和Pax2-Cre标记的小鼠第一鳃弓颅颌面部神经嵴细胞异质性研究  

Heterogeneity of Wnt1-Cre-marked and Pax2-Cre-marked first branchial arch cranial neural crest cells in mice

作　　者：徐珏 刘双 符宏高 邵美瑛 陈美玲 黄镇[2,3] Xu Jue;Liu Shuang;Fu Honggao;Shao Meiying;Chen Meiling;Huang Zhen(West China School of Public Health and West China Fourth Hospital,Sichuan University,Chengdu 610041,China;College of Life Sciences,Fujian Normal University,Fuzhou 350007,China;Innovation and Transformation Center,Fujian University of Traditional Chinese Medicine,Fuzhou 350108,China)

机构地区：[1]四川大学华西公共卫生学院/华西第四医院,成都610041 [2]福建师范大学生命科学院,福州350007 [3]福建中医药大学科学创新与转化中心,福州350108

出　　处：《华西口腔医学杂志》2024年第4期435-443,共9页West China Journal of Stomatology

基　　金：国家自然科学基金面上项目(82170918);四川省自然科学基金(2024NSFSC0549)。

摘　　要：目的利用Wnt1-Cre和Pax2-Cre小鼠特异性标记颅颌面神经嵴细胞(CNCs)迁移到第一鳃弓时的分化异质性及机制。方法分别收取胚胎期(E)8.0~E9.25Wnt1-Cre;R26R^(mTmG)及Pax2-Cre;R26R^(mTmG)小鼠胚胎进行整体荧光观察,利用石蜡切片免疫荧光对E15.5的Pax2-Cre;R26R^(Ai9)和Wnt1-Cre;R26R^(Ai9)小鼠所标记的CNCs在颅面部主要组织器官中的谱系分化情况进行比较分析,最后对E10.5的Wnt1-Cre;R26R^(mTmG)和Pax2-Cre;R26R^(mTmG)小鼠的第一鳃弓组织中CNCs进行单细胞测序分析,并对差异基因进行荧光定量聚合酶链反应(q-PCR)验证。结果Pax2-Cre和Wnt1-Cre小鼠特异性标记的CNCs均在E8.0自神经板开始迁移,但Pax2-Cre小鼠仅标记迁移到第一鳃弓的CNCs,而Wnt1-Cre同时标记了迁移到第一和第二鳃弓的CNCs;在分化谱系示踪方面,二者皆标记了CNCs分化形成的颅颌面部组织器官的间充质,但Wnt1-Cre在上腭和舌中标记CNCs更多;在第一鳃弓间充质中,Pax2-Cre所标记的CNCs特异性表达基因主要参与了成骨,而Wnt1-Cre所标记的CNCs特异性表达基因主要参与了肢体发育、细胞迁移和成骨,q-PCR结果也证实了两者高表达差异基因参与了以上功能。结论本研究结果提示Pax2-Cre小鼠可特异性用于第一鳃弓CNCs及其衍生组织成骨方面的研究。Objective This study aimed to explore the heterogeneity and gene ontology of Wnt1-Cre-marked and Pax2-Cre-marked first branchial arch cranial neural crest cells(CNCs)in mice.Methods The embryos of Wnt1-Cre;R26R^(mTmG) and Pax2-Cre;R26R^(mTmG) at embryonic day(E)8.0-E9.25 were collected for histological observation.We performed immunostaining to compare green fluorescent protein(GFP)-positive CNCs in Pax2-Cre;R26R^(Ai9) and Wnt1-Cre;R26R^(Ai9) mice at E15.5.Single-cell RNA sequencing(scRNA-seq)was used to analyze the first branchial arch GFP-positive CNCs from Wnt1-Cre;R26R^(mTmG) and Pax2-cre;R26R^(mTmG)mice at E10.5.Real time fluorescence quantitative polymerase chain reaction(q-PCR)was performed to validate the differential genes.Results Wnt1-Cre-marked and Pax2-Cre-marked CNCs migrated from the neural plateto first and second branchial arches and to the first branchial arch,respectively,at E8.0.Although Wnt1-Cre-marked and Pax2-Cre-marked CNCs were found mostly in cranial-facial tissues,the former had higher expression in palate and tongue.The results of scRNA-seq showed that Pax2-Cre-marked CNCs specifically contributed to osteoblast differentiation and ossification,while Wnt1-Cre-marked CNCs participated in limb development,cell migration,and ossification.The q-PCR data also confirmed the results of gene ontology analysis.Conclusion Pax2-Cre mice are perfect experimental animal models for research on first branchial arch CNCs and derivatives in osteoblast differentiation and ossification.

关 键 词：颅颌面部神经嵴细胞 第一鳃弓 Wnt1-Cre Pax2-Cre 单细胞测序 

分 类 号：Q257[生物学—细胞生物学]