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突变缝隙连接蛋白32在X连锁Charcot-Marie-Tooth病1型患者的外周血管内皮细胞异常表达
其他题名Mutant connexin 32 abnormally distributed in the vascular endothelial cells of X-linked Charcot-Marie-Tooth disease type 1 patients
栾兴华1; 洪道俊1; 乔晓会1; 吕鹤1; 王朝霞1; 袁云1
关键词夏科-马里-图斯病 连接蛋白类 血管 内皮细胞 Charcot-marie-tooth Disease Connexins Blood Vessels Endothelial Cells
刊名中华神经科杂志
2011
DOI10.3760/cma.j.issn.1006-7876.2011.10.012
44期:10页:689-693
收录类别中国科技核心期刊 ; CSCD
文章类型Journal Article
摘要目的 观察突变缝隙连接蛋白32( Cx32)在X连锁的Charcot-Marie-Tooth病1型(CMTX1)患者血管内皮细胞的表达规律。方法 对3例经Cx32基因检查证实的CMTX1患者(突变位点分别为c.379A>T、c.533A>G和c.590C>T点突变)进行腓肠神经活体组织检查,同时取非CMTXI患者的神经作为对照。以第Ⅷ因子、缝隙连接蛋白40和Cx32单克隆抗体为一抗,对患者及对照者标本中的神经滋养血管进行免疫荧光标记。同时构建携带上述3个点突变的以增强型绿色荧光蛋白(enhanced green fluorescence protein,EGFP)为报告基因的重组质粒pEGFP-N1-CX32质粒,转染至HeLa细胞,并进行Cx32和内质网标志蛋白葡萄糖调节蛋白78免疫荧光染色,以观察Cx32蛋白在HeLa细胞的分布规律。结果 Cx32蛋白呈一定间隔规律性点状表达于对照组血管内皮细胞间的缝隙连接部位,与Cx40蛋白共表达;3例Cx32基因突变患者的血管内皮细胞膜的Cx32蛋白表达显著减少。细胞转染显示c.379A>T突变的Cx32蛋白主要以团块状蓄积在HeLa细胞的胞质内,c.533A>G突变的Cx32蛋白仅有少量表达于核周,c.590C>T突变的Cx32蛋白在胞膜上和胞质内均有点状分布,较野生型显著减少。3种突变的Cx32蛋白在HeLa细胞内的分布和内质网标记不重叠。结论 不同Cx32基因突变均可导致CMTX1的血管内皮细胞膜Cx32蛋白表达下降,该蛋白主要聚集在细胞的内质网外。 Objective To investigate expression distribution of mutant connexin 32 (Cx32) protein in human endothelial cells in patients with X-linked Charcot-Marie-Tooth disease type 1 ( CMTX1 ) .Methods Nerve biopsies were performed in 3 patients with CMTX1 and in 3 non-CMTX1 controls. Cx32 mutations of c. 379A > T( I127F), c. 533A > G(D178G) and c. 590C > T(A197V) were identified in these 3 patients respectively. Immunofluorescent (IMF) staining of nerve blood vessel was processed with antibodies against Cx32, Yon Willebrand factor and Cx40. The mutant Cx32 was constructed in pEGFP-N plasmid (pEGFP-N1-Cx32) and was transfected in HeLa cells. Cx32 and GRP78, a marker of endoplasmic reticulum ( ER), were stained by IMF in HeLa cells to investigate expression of mutant Cx32. Results In 3 control cases, Cx32 was visualized by IMF staining as dots along gap junction of vascular endothelial cells,and it was coexisted with Cx40. However, immunoreactivity of Cx32 in 3 patients was predominantly decreased and was not located in endothelial gap junction. The transfection of 3 Cx32 mutants into HeLa cells demonstrated the pathogenic changes. The cells with the mutation c. 379A >T found Cx32 accumulations in the cytoplasm; the cells with mutation c. 533A >G showed few staining positive dots surrounding the nuclear and the cells with c. 590C > T showed dot-like expression of Cx32 both in the cytoplasmicand cell membrane. The mutant Cx32 was not overlapped with expression of the marker of ER.ConclusionsMutant Cx32 might cause dysfunction of endothelial gap-junctions due to the abnormal expression of Cx32 in level and location in the vascular endothelial cells of CMTX1 patients.
语种中文
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文献类型期刊论文
条目标识符http://ir.bjmu.edu.cn/handle/400002259/41159
专题北京大学第一临床医学院_神经内科
作者单位1.100034,北京大学第一医院神经内科
2.首都医科大学附属复兴医院神经内科
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栾兴华,洪道俊,乔晓会,等. 突变缝隙连接蛋白32在X连锁Charcot-Marie-Tooth病1型患者的外周血管内皮细胞异常表达[J]. 中华神经科杂志,2011,44(10):689-693.
APA 栾兴华,洪道俊,乔晓会,吕鹤,王朝霞,&袁云.(2011).突变缝隙连接蛋白32在X连锁Charcot-Marie-Tooth病1型患者的外周血管内皮细胞异常表达.中华神经科杂志,44(10),689-693.
MLA 栾兴华,et al."突变缝隙连接蛋白32在X连锁Charcot-Marie-Tooth病1型患者的外周血管内皮细胞异常表达".中华神经科杂志 44.10(2011):689-693.
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