Wnt5a在小鼠牙骨质发育中的表达特点及其对成牙骨质细胞分化的影响  

Expression characteristics of Wnt5ain different periods of cementum development and its effect on differentiation of cementoblasts of mice

作　　者：包幸福[1] 倪宇昕[2] 李莹[1] 郭小凯[1] 王玉琢[1] 方蛟[2] 胡敏[1] BAO Xingfu;NIYuxin;L IYing;GUO Xiaokat;WANG Yuzhuo;FANG Jiao;HU Min(Department of Orthodontics,Stomatology Hospital,Jilin University,Changchun 130021,China;Department of Implantation,Stomatology Hospital,Jilin University,Changchun 130021,China)

机构地区：[1]吉林大学口腔医院正畸科,吉林长春130021 [2]吉林大学口腔医院种植科,吉林长春130021

出　　处：《吉林大学学报（医学版）》2018年第3期461-465,693,共6页Journal of Jilin University：Medicine Edition

基　　金：国家自然科学基金资助课题(81470764);吉林省卫计委青年科研基金资助课题(2015Q003;2015Q013);吉林大学白求恩计划项目资助课题(2015321);吉林省科技厅科技发展计划项目资助课题(20160520156JH)

摘　　要：目的:探讨Wnt家族5A蛋白(Wnt5a)在小鼠出生后牙骨质中的表达特点以及其对体外培养成牙骨质细胞分化的影响,阐明Wnt非经典通路调控牙根发育的机制。方法:选择出生后0.5、4.5、10.5、16.5、20.5、24.5、26.5和30.5d昆明小鼠并按以上相应时间点分组,每组3只;在相应时间点采用脱臼法处死小鼠,分离下颌第一磨牙及牙周组织。免疫组织化学染色法观察小鼠牙体和牙周组织中Wnt5a的表达特点。将小鼠成牙骨质细胞OCCM30分为实验组和对照组。实验组加入200μg·L^(-1)重组Wnt5a蛋白,对照组无添加;提取细胞总RNA。RT-PCR法检测2组OCCM30细胞中Ocn、Opn、Alp、Bsp、Wnt5a和Runx-2基因的相对表达水平。结果:小鼠出生后0.5d,第一磨牙牙胚中无Wnt5a阳性表达;出生后4.5d,成釉细胞和成牙本质细胞中Wnt5a阳性表达较强;出生后16.5d,可见成牙骨质细胞中Wnt5a呈阳性表达;出生后20.5~30.5d,成牙本质细胞和牙周膜细胞中Wnt5a呈阳性表达。与对照组比较,实验组成牙骨质细胞中Ocn和Alp基因相对表达水平降低(P<0.05)。结论:Wnt5a在牙根发育过程中的表达特点为从无到有,呈现规律性表达;Wnt5a下调分化基因的表达,在牙骨质发育中发挥重要作用。Objective：To study the expression characteristics of Wnt5a in cementum of the mice after birth and the effect of Wnt5a on the differentiation of cementoblasts in vitro,and to illuminate the mechanism of Wnt noncanonical signaling in regulating the tooth development.Methods：The postnatal Kunming mice at the days 0.5,4.5,12.5,18.5,24.5 and 30.5 after birth were selected and divided into various groups by different time points（n=3）.The mice were sacrificed and the lower first molar and peridentium were prepared.Immunohistochemical staining was used to detect the expression characteristics of Wnt5a in tooth and peridentium of the mice.The OCCM30 cells were divided into control group and experiment group.200μg·L-（-1） recombinant Wnt5a protein were added in experiment group,and nothing was added in control group;the total RNA in the cells was exrracted.The relative expression levels of Ocn,Opn,Alp,Bsp,Wnt5a ,and Runx-2 genes in the OCCM30 cells in two groups were detected by RT-PCR method.Results：At the day 0.5 after birth,the expression of Wnt5a in the lower first molar tooth germ was negative.At the day 4.5 after birth,the expression of Wnt5a in ameloblasts and odontoblasts were positive.At the day 16.5 after birth,the expression of Wnt5a in cementoblasts was positive.At the days 20.5-30.5 after birth,the expressions of Wnt5a in odontoblasts and periodontal ligament cells were positive.Compared with control group,the relative expression levels of Ocn and Alp gene in the OCCM30 cells in experiment group were decreased（P〈0.05）.Conclusion：The expression of Wnt5a is regular at different tooth developmental stages after birth.Wnt5a can reduce the expression of differentiated genes and play an important role in the growth of cementum.

关 键 词：WNT5A 牙发育 牙骨质 成牙骨质细胞 免疫组织化学 

分 类 号：R78[医药卫生—口腔医学] R332[医药卫生—临床医学]