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

我们研究了维生素d_3对表皮角质形成细胞细胞周期的影响。维生素d3主要在g1 / go期诱导角化细胞生长停止，但在m期部分抑制角化细胞生长。采用抗天合蛋白抗体和流式细胞术检测天合蛋白阳性细胞数，观察维生素d3对角质形成细胞分化的影响。维生素d_3在10^<-6> m时增加了involucrin阳性细胞。这表明维生素d_3诱导了角质形成细胞的分化。视网膜母细胞瘤基因最初被鉴定为肿瘤抑制基因。后来，研究表明视网膜母细胞瘤基因产物（prb）通过调节g1 / g0期向s期的转变来调节细胞周期。也就是说，PRB的磷酸化启动了g1 / g0期到s期的转变。因此，我们研究了PRB在维生素d_3对人角质形成细胞生长抑制机制中的作用。将PRB的分子量差异可视化，并用密度计定量。添加10^<-6> m维生素d_3使去磷酸化PRB从0 h时的23%增加到48 h时的58%，具有时间依赖性，因此，去磷酸化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。我们比较了维生素d_3对正常和sv40转化的人角质形成细胞的生长抑制作用。维生素d_3在10^<-6> m时对正常人角化细胞生长的抑制作用超过95%。相比之下，转化的人角质形成细胞的生长仅减少了50%。由于tgf - β对人角质形成细胞的生长抑制在sv40转化的角质形成细胞中可能通过大T抗原与PRB结合而被消除，因此tgf - β可能参与了维生素D_3的生长抑制机制。

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.