兔BMP4基因对毛囊发育相关基因表达的影响  

Effect of rabbit BMP4 gene on expression of genes related to hair follicle development in dermal papilla cells and its functional analysis

作　　者：张琛 李佳丽[1] 靳荣帅 白少成 姚凡 赵博昊 陈阳[1] 吴信生[1] ZHANG Chen;LI Jia-li;JIN Rong-shuai;BAI Shao-cheng;YAO Fan;ZHAO Bo-hao;CHEN Yang;WU Xin-sheng(College of Animal Science and Technology,Yangzhou University,Yangzhou,Jiangsu 225009,China)

机构地区：[1]扬州大学动物科学与技术学院,江苏扬州225009

出　　处：《西南农业学报》2022年第6期1448-1454,共7页Southwest China Journal of Agricultural Sciences

基　　金：财政部和农业农村部国家现代农业产业技术体系资助(CARS-43-A-1);国家自然科学基金(32072724)。

摘　　要：【目的】为了研究骨形态发生蛋白4(BMP4)对兔毛囊发育相关基因的影响。【方法】利用显微切割和二步酶消化相结合的方法分离了兔毛乳头细胞,并通过免疫荧光染色检测毛乳头细胞的特异性标记物α-SMA和VIM的表达情况。构建pcDNA3.1-BMP4过表达载体,转染至毛乳头细胞中,利用荧光定量PCR技术分析BMP4基因对毛囊发育相关基因(ID1、ID2、SAMD7、FGF1、TGF-β)mRNA表达的影响,通过CCK-8细胞增殖实验验证BMP4基因对兔毛乳头细胞生长的作用。【结果】免疫荧光结果显示,分离培养的细胞为毛囊真皮源性的毛乳头细胞。进一步成功克隆了家兔BMP4基因,发现其CDS序列全长为1230 bp,编码409个氨基酸,预测为亲水性外分泌蛋白,含有1个信号肽序列,无跨膜结构域。荧光定量PCR结果显示,过表达BMP4能显著升高ID1、ID2、SAMD7的mRNA表达水平(P<0.01),但显著抑制FGF1的mRNA表达水平(P<0.05)。通过CCK-8细胞增殖实验发现,过表达BMP4基因极显著抑制毛乳头细胞增殖(P<0.01)。【结论】BMP4基因可能通过影响毛乳头细胞中毛囊发育相关基因的表达,抑制毛乳头细胞增殖,参与家兔毛囊发育过程。【Objective】The experiment aimed to study the effects of bone morphogenetic protein 4(BMP4)on the genes related to hair follicle development in rabbits.【Method】In this study,rabbit hair papilla cells were isolated by microdissection and two-step enzyme digestion,and the expression of specific markersα-SMA and VIM were detected by immunofluorescence staining.The pcDNA3.1-BMP4 overexpression vector was constructed and transfected into dermopapilla cells,and the effects of BMP4 gene on the mRNA expression of hair follicle developing-related genes(ID1,ID2,SAMD7,FGF1,TGF-β)were analyzed by fluorescence quantitative PCR.The effect of BMP4 gene on the growth of rabbit hair papilla cells was verified by CCK-8 cell proliferation assay.【Result】Immunofluorescence showed that the cultured cells were dermal papilla cells derived from hair follicle dermal skin.BMP4 gene was cloned successfully.The CDS of BMP4 gene was 1230 bp and encoded 409 amino acids.This protein was predicted to be a hydrophilic exsecretory protein with 1 signal peptide sequence and no transmembrane domain.Fluorescence quantitative PCR showed that BMP4 overexpression significantly increased the mRNA expression levels of ID1,ID2 and SAMD7(P<0.01),but significantly inhibited that of FGF1(P<0.05).In CCK-8 cell proliferation assay,BMP4 overexpression significantly inhibited the proliferation of dermal papilla cells(P<0.01).【Conclusion】In conclusion,BMP4 gene may inhibit the proliferation of dermal papilla cells and participate in the development of rabbit hair follicles by affecting the expression of genes related to hair follicle development in dermal papilla cells.

关 键 词：兔 骨形态发生蛋白4 毛囊发育 毛乳头细胞 基因表达 

分 类 号：S829.9[农业科学—畜牧学]