發(fā)布時間：2018-04-20 15:44

  本文選題：Gsdma3 + 小鼠皮膚上皮細胞��； 參考：《第三軍醫(yī)大學》2014年碩士論文

【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號染色體的亞家族Gsdma基因家族成員之一，Gsdma基因家族成員同視黃酸受體α及角質(zhì)蛋白1的編碼基因緊鄰。Gsdma3基因存在的熱點突變區(qū)域能分別產(chǎn)生6種Gsdma3突變體，Bsk, Dfl, Rco2, Fgn,Rex-denuded和Rim3，這6種突變體在皮膚上皮組織中會引起毛囊周期中第一次生長期的延長，皮膚的過度增生，并使得皮膚產(chǎn)生炎癥反應(yīng)。經(jīng)典Wnt／β-catenin信號通路對皮膚上皮細胞的增殖，分化具有重要的調(diào)節(jié)作用，而我們前期研究結(jié)果提示，Gsdma3基因同經(jīng)典Wnt／β-catenin信號通路在小鼠皮膚上皮細胞中存在關(guān)聯(lián)，所以本研究的目的在于: 1、初步探明在小鼠皮膚上皮細胞中，Gsdma3基因?qū)σ种平?jīng)典Wnt/β-catenin信號通路的Wnt5a表達的影響。 2、初步探明在小鼠皮膚上皮細胞中，Gsdma3基因?qū)σ种平?jīng)典Wnt/β-catenin信號通路的E-cadherin表達的影響。 3、初步探明在小鼠皮膚上皮細胞中，Gsdma3基因?qū)δ軌蚺cβ-catenin蛋白形成激活蛋白復(fù)合體的NF-κB蛋白表達的影響。 方法： 1、本研究利用C57BL/6J小鼠，Gsdma3基因突變鼠及Gsdma3基因表達質(zhì)粒等材料，通過質(zhì)粒轉(zhuǎn)染、組織切片、免疫熒光、RT-PCR、WB等技術(shù)手段，在Gsdma3基因突變及突變挽救后，分析在體小鼠皮膚上皮細胞中，Gsdma3基因?qū)nt5a，E-cadherin，NF-κB表達變化的影響情況。 2、利用體外培養(yǎng)的小鼠皮膚上皮細胞和Gsdma3基因表達質(zhì)粒，質(zhì)粒轉(zhuǎn)染、流式細胞術(shù)、免疫熒光、RT-PCR、WB等技術(shù)手段，在離體水平上分析了培養(yǎng)的小鼠皮膚上皮細胞Gsdma3基因過表達后，Gsdma3基因?qū)nt5a，E-cadherin，NF-κB表達變化的影響情況。 結(jié)果： 1、在體水平上，免疫熒光結(jié)果顯示，Gsdma3基因突變鼠皮膚上皮細胞中Wnt5a的表達上調(diào)；Gsdma3基因突變鼠進行Gsdma3基因表達挽救后，Wnt5a的表達下調(diào)。同樣，在7周C57BL/6J小鼠毛囊周期同步化的背部皮膚上皮細胞中過表達Gsdma3基因后也會引起Wnt5a的表達下調(diào)。 在離體水平上，RT-PCR, Western blot結(jié)果顯示，在培養(yǎng)的小鼠皮膚上皮細胞中過表達Gsdma3基因后，Wnt5a的表達呈現(xiàn)下調(diào)趨勢。 2、在體水平上，免疫熒光，RT-PCR, Western blot結(jié)果顯示，Gsdma3基因突變鼠皮膚上皮細胞中E-cadherin的表達在突變鼠中具有顯著的上調(diào)現(xiàn)象；Gsdma3基因突變鼠進行Gsdma3基因表達挽救后，E-cadherin的表達下調(diào)。同樣，在7周C57BL/6J小鼠毛囊周期同步化的背部皮膚上皮細胞過表達Gsdma3基因后也會引起E-cadherin的表達下調(diào)。 在離體水平上，免疫熒光，RT-PCR, Western blot結(jié)果顯示，在培養(yǎng)的小鼠皮膚上皮細胞中過表達Gsdma3基因后E-cadherin的表達具有顯著的下調(diào)現(xiàn)象。 3、在體水平上，免疫熒光，RT-PCR, Western blot結(jié)果顯示，Gsdma3基因突變鼠皮膚上皮細胞中NF-κB的表達具有顯著的上調(diào)現(xiàn)象。 在離體水平上，免疫熒光，，RT-PCR, Western blot結(jié)果顯示，在培養(yǎng)的小鼠皮膚上皮細胞中過表達Gsdma3基因后NF-κB的表達具有顯著的上調(diào)現(xiàn)象。 結(jié)論： 1、在小鼠皮膚上皮細胞中Gsdma3基因能抑制Wnt5a的表達。 2、在小鼠皮膚上皮細胞中Gsdma3基因能抑制E-cadherin的表達。 3、在小鼠皮膚上皮細胞中Gsdma3基因能促進NF-κB的表達。
[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 偽 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 .

【學位授予單位】：第三軍醫(yī)大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R329.2

【參考文獻】

相關(guān)期刊論文 前2條

1 余裕;白秀峰;賴向東;楊恬;連小華;;GasderminA3基因在小鼠皮膚上皮中對NF-κB表達的影響[J];第三軍醫(yī)大學學報;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

本文編號：1778396