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ANALYSIS OF REGULATORY MECHANISM OF VIATAMIN D IN THE GROWTH AND DIFFERENTIATION OF EPIDERMAL KERATINOCYTES

维生素D对表皮角质细胞生长和分化的调控机制分析

基本信息

批准号：
04670641
负责人：
HASHIMOTO Koji
金额：
$ 1.34万
依托单位：
OSAKA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1993
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04670641/
关键词：
VITAMIN D_3 KERATINOCYTES INVOLUCRIN FLOWCYTOMETRY TGF-beta SV40 CELL CYCLE RETINOBLASTOMA GENE PRODUCT Vitamin D_3 c-Mlyc SV40ウイルス Vitamin D_3レセプター

项目摘要

WE EXAMINED THE EFFECT OF VITAMIN D_3 ON THE CELL CYCLE OF EPIDERMAL KERATINOCYTES.VITAMIN D3 INDUCED THE GROWTH ARREST MAINLY IN G1/GO PHASE ALTHOUGH IT ARRESTS THE KERATINOCYTE GROWTH PARTIALLY IN M PHASE.THE EFFECT OF VITAMIN D3 ON THE KERATINOCYTE DIFFERENTIATION WAS EXAMINED BY MEASURING THE NUMBER OF INVOLUCRIN-POSITIVE CELLS USING ANTI-INVOLUCRIN ANTIBODY AND FLOWCYTOMETRY.VITAMIN D_3 AT 10^<-6> M INCREASED THE INVOLUCRIN-POSITIVE CELLS.THIS INDICATES THAT VITAMIN D_3 INDUCED THE KERATINOCYTE DIFFERENTIATION.RETINOBLASTOMA GENE WAS IDENTIFIED AS TUMOR SUPPRESSOR GENE ORIGINALLY.LATER, IT WAS SHOWN THA RETINOBLASTOMA GENE PRODUCT (PRB) REGULATES THE CELL CYLCE BY MODULATING THE TRANSITION FROM G1/G0 TO S PHASE.NAMELY, PHOSPHORYLATION OF PRB INTIATES THE TRANSITION FROM G1/G0 TO S PHASE.THEREFORE, WE STUDIED THE INBOLBEMENT OF PRB IN THE GROWTH INHIBITORY MECHANISM OF HUAMAM KERATINOCYTES BY VITAMIN D_3. PRB WERE VISUALIZED THE DIFFERENCE OF MOLECULAR WEIGHT AND QUANTITATED BY DENSITOMETER.THE ADDITION OF 10^<-6> M VITAMIN D_3 INCREASED THE DEPHOSPHORYLATED PRB IN A TIME DEPENDENT MANNER FROM 23% AT 0 H TO 58% AT 48 H.THUS, THE INCREASE OF DEPHOSPHORYLATED PRB WAS CLOSELY RELATED TO THE GROWTH INHIBITION OF HUMAN KERATINOCYTES BY VITAMIN D_3. WE COMPARED THE GROWTH INHIBITORY EFFECT OF VITAMIN D_3 ON NORMAL AND SV40 TRANSFORMED HUMAN KERATINOCYTES.VITAMIN D_3 AT 10^<-6> M INHIBITED THE GROUTH OF NORMAL HUMAN KERATINOCYTES BY MORE THAN 95%. IN CONTRAST, THE GROWTH OF TRANSFORMED HUMAN KERATINOCYTES WAS REDUCED ONLY BY 50%. SINCE THE GROWTH INHIBITION OF HUMAN KERATINOCYTES BY TGF-beta WAS ABOLISHED IN SV40-TRANSFORMED KERATINOCYTES PROBABLY THROUGH THE BINDING OF LARGE T ANTIGEN TO PRB, THE POSSIBLE INVOLVEMENT OF TGF-beta IN THE GROWTH INHIBITORY MECHANISM BY VITAMIN D_3 WAS SUGGESTED.

本文研究了维生素D_3对表皮角朊细胞周期的影响。维生素D_3虽然能使角朊细胞部分阻滞于M期，但主要使其生长阻滞于G_1/G_0期。用抗Involucrin抗体和流式细胞术测定Involucrin阳性细胞数，以观察维生素D_3对角朊细胞分化的影响。维生素D_3在10 μ M时能增加Involucrin-1的表达<-6>。阳性细胞，这表明维生素D_3诱导角化细胞分化。视网膜母细胞瘤基因被鉴定为肿瘤提供基因或原始基因。后来发现，视网膜母细胞瘤基因产物（PRB）通过调节G1/G 0期向S期的转变来调节细胞周期。也就是说，PRB的磷酸化启动G1/G 0期向S期的转变。因此，本文研究了维生素D_3对人角质形成细胞生长抑制机制中PRB的抑制作用。10 μ <-6>M维生素D_3使脱磷酸化的PRB从0小时的23%增加到48小时的58%，并呈时间依赖性，因此，脱磷酸化的PRB的增加与维生素D_3对人角质形成细胞的生长抑制密切相关。本文比较了维生素D_3对正常人角朊细胞和SV 40转化人角朊细胞的生长抑制作用，10 μ M维生素D_3<-6>对正常人角朊细胞的生长抑制率达95%以上。最后，转化的人角质形成细胞的生长仅减少50%。由于TGF-β对人角质形成细胞的生长抑制作用在SV_（40）转化的角质形成细胞中可能通过大T抗原与PRB的结合而被解除，提示TGF-β可能参与了维生素D_3的生长抑制机制。