Gsdma3基因在小鼠皮肤上皮细胞中对Wnt5a，E-cadherin，NF-κB表达影响的研究

发布时间：2018-04-20 15:44

  本文选题：Gsdma3 + 小鼠皮肤上皮细胞　； 参考：《第三军医大学》2014年硕士论文

【摘要】：研究背景和目的： Gsdma3基因是Gsdm基因家族中位于小鼠第11号染色体的亚家族Gsdma基因家族成员之一，Gsdma基因家族成员同视黄酸受体α及角质蛋白1的编码基因紧邻。Gsdma3基因存在的热点突变区域能分别产生6种Gsdma3突变体，Bsk, Dfl, Rco2, Fgn,Rex-denuded和Rim3，这6种突变体在皮肤上皮组织中会引起毛囊周期中第一次生长期的延长，皮肤的过度增生，并使得皮肤产生炎症反应。经典Wnt／β-catenin信号通路对皮肤上皮细胞的增殖，分化具有重要的调节作用，而我们前期研究结果提示，Gsdma3基因同经典Wnt／β-catenin信号通路在小鼠皮肤上皮细胞中存在关联，所以本研究的目的在于: 1、初步探明在小鼠皮肤上皮细胞中，Gsdma3基因对抑制经典Wnt/β-catenin信号通路的Wnt5a表达的影响。 2、初步探明在小鼠皮肤上皮细胞中，Gsdma3基因对抑制经典Wnt/β-catenin信号通路的E-cadherin表达的影响。 3、初步探明在小鼠皮肤上皮细胞中，Gsdma3基因对能够与β-catenin蛋白形成激活蛋白复合体的NF-κB蛋白表达的影响。 方法： 1、本研究利用C57BL/6J小鼠，Gsdma3基因突变鼠及Gsdma3基因表达质粒等材料，通过质粒转染、组织切片、免疫荧光、RT-PCR、WB等技术手段，在Gsdma3基因突变及突变挽救后，分析在体小鼠皮肤上皮细胞中，Gsdma3基因对Wnt5a，E-cadherin，NF-κB表达变化的影响情况。 2、利用体外培养的小鼠皮肤上皮细胞和Gsdma3基因表达质粒，质粒转染、流式细胞术、免疫荧光、RT-PCR、WB等技术手段，在离体水平上分析了培养的小鼠皮肤上皮细胞Gsdma3基因过表达后，Gsdma3基因对Wnt5a，E-cadherin，NF-κB表达变化的影响情况。 结果： 1、在体水平上，免疫荧光结果显示，Gsdma3基因突变鼠皮肤上皮细胞中Wnt5a的表达上调；Gsdma3基因突变鼠进行Gsdma3基因表达挽救后，Wnt5a的表达下调。同样，在7周C57BL/6J小鼠毛囊周期同步化的背部皮肤上皮细胞中过表达Gsdma3基因后也会引起Wnt5a的表达下调。 在离体水平上，RT-PCR, Western blot结果显示，在培养的小鼠皮肤上皮细胞中过表达Gsdma3基因后，Wnt5a的表达呈现下调趋势。 2、在体水平上，免疫荧光，RT-PCR, Western blot结果显示，Gsdma3基因突变鼠皮肤上皮细胞中E-cadherin的表达在突变鼠中具有显著的上调现象；Gsdma3基因突变鼠进行Gsdma3基因表达挽救后，E-cadherin的表达下调。同样，在7周C57BL/6J小鼠毛囊周期同步化的背部皮肤上皮细胞过表达Gsdma3基因后也会引起E-cadherin的表达下调。 在离体水平上，免疫荧光，RT-PCR, Western blot结果显示，在培养的小鼠皮肤上皮细胞中过表达Gsdma3基因后E-cadherin的表达具有显著的下调现象。 3、在体水平上，免疫荧光，RT-PCR, Western blot结果显示，Gsdma3基因突变鼠皮肤上皮细胞中NF-κB的表达具有显著的上调现象。 在离体水平上，免疫荧光，，RT-PCR, Western blot结果显示，在培养的小鼠皮肤上皮细胞中过表达Gsdma3基因后NF-κB的表达具有显著的上调现象。 结论： 1、在小鼠皮肤上皮细胞中Gsdma3基因能抑制Wnt5a的表达。 2、在小鼠皮肤上皮细胞中Gsdma3基因能抑制E-cadherin的表达。 3、在小鼠皮肤上皮细胞中Gsdma3基因能促进NF-κB的表达。
[Abstract]:Background and purpose of the study :

The Gsdma3 gene is one of the members of the Gsdma gene family in the Gsdm gene family . The Gsdma gene family members are closely related to the coding genes of retinoic acid receptor 伪 and keratin 1 . The Gsdma3 gene can produce six Gsdma3 mutants , Bsk , Dfl , Rco2 , Fgn , Rex - ATPase and Rim3 respectively . The classical Wnt / 尾 - catenin signaling pathway plays an important role in the proliferation and differentiation of skin epithelial cells . The classical Wnt / 尾 - catenin signaling pathway is associated with the classical Wnt / 尾 - catenin signaling pathway in mouse skin epithelial cells .

1 . The effect of Gsdma3 gene on the expression of Wnt5a in the classical Wnt / 尾 - catenin signaling pathway was preliminarily explored in mouse skin epithelial cells .

2 . The effect of Gsdma3 gene on the expression of E - cadherin in the classical Wnt / 尾 - catenin signaling pathway was preliminarily explored in mouse skin epithelial cells .

3 . The effect of Gsdma3 gene on the expression of NF - 魏B in mouse skin epithelial cells was preliminarily explored .

Method :

1 . The effects of Gsdma3 gene on the expression of Wnt5a , E - cadherin and NF - 魏B were analyzed by means of plasmid transfection , tissue section , immunofluorescence , RT - PCR , WB and so on .

2 . The effect of Gsdma3 gene on the expression of Wnt5a , E - cadherin and NF - 魏B in cultured mouse skin epithelial cells was analyzed by means of plasmid , plasmid transfection , flow cytometry , immunofluorescence , RT - PCR , WB and so on .

Results :

1 . At the body level , the expression of Wnt5a in skin epithelial cells of Gsdma3 gene mutation was up - regulated .
The expression of the Gsdma3 gene was down regulated by Gsdma3 gene mutation . Similarly , the expression of the Gsdma3 gene in the dorsal skin epithelial cells synchronized with the hair follicle cycle of C57BL / 6J mice in 7 weeks could also result in downregulation of Wnt5a .

RT - PCR and Western blot showed that the expression of Wnt5a was down - regulated after overexpression of Gsdma3 gene in cultured mouse skin epithelial cells .

2 . In vivo , the expression of E - cadherin in skin epithelial cells of Gsdma3 gene mutation was significantly up - regulated in mouse skin epithelial cells by immunofluorescence , RT - PCR and Western blot .
The expression of E - cadherin was downregulated after the expression of Gsdma3 gene in Gsdma3 gene mutant mice .

Western blot showed that the expression of E - cadherin was down - regulated in cultured mouse skin epithelial cells after overexpression of Gsdma3 gene .

3 . The expression of NF - 魏B in skin epithelial cells of Gsdma3 gene mutation was significantly up - regulated at body level , immunofluorescence , RT - PCR and Western blot .

Western blot showed that the expression of NF - 魏B in cultured mouse skin epithelial cells was significantly up - regulated after overexpression of Gsdma3 gene in cultured mouse skin epithelial cells .

Conclusion :

1 . Gsdma3 gene can inhibit the expression of Wnt5a in mouse skin epithelial cells .

2 . The expression of E - cadherin was inhibited by Gsdma3 gene in mouse skin epithelial cells .

3 . Gsdma3 gene can promote the expression of NF - 魏B in mouse skin epithelial cells .

【学位授予单位】：第三军医大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R329.2

【参考文献】

相关期刊论文 前2条

1 余裕;白秀峰;赖向东;杨恬;连小华;;GasderminA3基因在小鼠皮肤上皮中对NF-κB表达的影响[J];第三军医大学学报;2014年04期

2 De-Ren Fang;Zhong-Fa Lv;Gang Qiao;;Dynamic Wnt5a expression in murine hair follicle cycle and its inhibitory effects on follicular in vivo[J];Asian Pacific Journal of Tropical Medicine;2014年04期

本文编号：1778396