REGULATORY MECHANISMS OF VASCULAR CONTRACTION AND CELL PROLIFERATION IN DIABETES MELLITUS

本研究旨在探讨糖尿病患者血管收缩和细胞增殖异常调控的机制。得到了以下结果：1。血管平滑肌细胞的增殖。我开发了在单细胞水平上测定原代培养血管平滑肌细胞周期阶段的方法。结果表明：(1)血小板衍生生长因子（platelet-derived growth factor， PDGF）刺激细胞周期由G_0期向G_1期进展不需要胞质Ca^<2+>瞬态；(2)高糖不刺激G_0细胞的细胞周期，而将pdgf预处理的G_1细胞的细胞周期刺激到S期和M期，而胞质Ca^<2+>浓度未增加（[Ca^<2+>]i）.2。内皮细胞的钙瞬态。利用前表面荧光法和原位加载fura-2的瓣膜内皮细胞，发现：(1)内皮素-1 （ET-1）升高[Ca^<2+>]和[Ca^<2+>]的内流和细胞内Ca^<2+>的释放；2) Ca^<2+>内流受百日咳毒素敏感的g蛋白调节，而细胞内Ca^<2+>的释放则不受调节。血管收缩。(1)利用RT-PCR和fura-2微荧光法发现血管紧张素II （AT-II）、蛋白激酶C和蛋白激酶A可调控AT-II受体mRNA的表达，mRNA水平的升高伴随着生理反应性的提高。(2)利用fura-2-前表面荧光法和猪冠状动脉条带，发现ET-3主要通过细胞外Ca^<2+>的内流增加[Ca^<2+>]i诱导血管收缩，并且在血管收缩剂对ET-1和ET-3的反应中，Ca^<2+>-敏感性的不同时间依赖性调节机制。(3)去甲肾上腺素增加了Ca^<2+>-的敏感性，而血清素对Ca^<2+>-的敏感性没有增强作用。少

The purpose of this research is to investigate the mechanisms of abnormal regulation of vascular contraction and cell proliferation in diabetes mellitus. The following results were obtained :1.CELL PROLIFERATION OF VASCULAR SMOOTH MUSCLE CELLS.I developed the method to determine the phase of the cell cycle of vascular smooth muscle cells in primary culture at the single cell level. It was found that : (1)cytosolic Ca^<2+> transients are not required for platelet-derived growth factor (PDGF) to stimulate the cell cycle progression from the G_0 phase to the G_1 phase ; and (2)high glucose does not stimulate the cell cycle of the G_0 cells, whereas it stimulates the cell cycle of the PDGF-pretreated G_1 cells to the S and M phases, without any increase of cytosolic Ca^<2+> concentration ([Ca^<2+>]i).2.CALCIUM TRANSIENTS OF ENDOTHELIAL CELLS.Using front-surface fluorometry and fura-2-loaded valvular endothelial cells in situ, it was found that : (1)endothelin-1 (ET-1) elevates [Ca^<2+>]i b … More y the stimulation of both Ca^<2+> influx and intracellular Ca^<2+> release ; and 2)the Ca^<2+>influx is regulated by an pertussis toxin-sensitive G-protein, while the release of intracellular Ca^<2+> is not.3.VASCULAR CONTRACTION.(1)Using RT-PCR and fura-2 microfluorometry, it was found that angiotensin II (AT-II), protein kinase C,and protein Kinase A regulate the expression of AT-II receptor mRNA,and the increase in mRNA level is accompanied by an increase in physiological responsiveness. (2) Using fura-2-front-surface fluorometry and porcine coronary arterial strips, it was found that ET-3 induced vasoconstriction by increasing [Ca^<2+>]i mainly through the influx of extracellular Ca^<2+>, and that distinct mechanisms of time-dependent modulation of the Ca^<2+>-sensitivity function in the vasoconstrictor responses to ET-1 and ET-3. (3)It was found that norepinephrine induced increase in the Ca^<2+>-sensitivity of contractile apparatus, while serotonin demonstrates little enhancement of the Ca^<2+>-sensitivity of contractile apparatus. Less