Doctrow S R, Folkman J
J Cell Biol. 1987 Mar;104(3):679-87. doi: 10.1083/jcb.104.3.679.
The intracellular events regulating endothelial cell proliferation and organization into formalized capillaries are not known. We report that the protein kinase C activator beta-phorbol 12,13-dibutyrate (PDBu) suppresses bovine capillary endothelial (BCE) cell proliferation (K50 = 6 +/- 4 nM) and DNA synthesis in response to human hepatoma-derived growth factor, an angiogenic endothelial mitogen. In contrast, PDBu has no effect on the proliferation of bovine aortic endothelial cells and is mitogenic for bovine aortic smooth muscle and BALB/c 3T3 cells. Several observations indicate that the inhibition of human hepatoma-derived growth factor-stimulated BCE cell growth by PDBu is mediated through protein kinase C. Different phorbol compounds inhibit BCE cell growth according to their potencies as protein kinase C activators (12-O-tetradecanoylphorbol 13-acetate greater than PDBu much greater than beta-phorbol 12,13-diacetate much much greater than beta-phorbol; alpha-phorbol 12,13-dibutyrate; alpha-phorbol 12,13-didecanoate). PDBu binds to a single class of specific, saturable sites on the BCE cell with an apparent Kd of 8 nM, in agreement with reported affinities of PDBu for protein kinase C in other systems. Specific binding of PDBu to BCE cells is displaced by sn-1,2-dioctanoylglycerol, a protein kinase C activator and an analog of the putative second messenger activating this kinase in vivo. The weak protein kinase C activator, sn-1,2-dibutyrylglycerol, does not affect PDBu binding. A cytosolic extract from BCE cells contains a calcium/phosphatidylserine-dependent protein kinase that is activated by sn-1,2-dioctanoylglycerol and PDBu, but not by beta-phorbol. These findings indicate that protein kinase C activation can cause capillary endothelial cells to become desensitized to angiogenic endothelial mitogens. This intracellular regulatory mechanism might be invoked during certain phases of angiogenesis, for example when proliferating endothelial cells become differentiated to organize into nongrowing tubes.
调节内皮细胞增殖并使其组织形成规则毛细血管的细胞内事件尚不清楚。我们报告蛋白激酶C激活剂β - 佛波醇12,13 - 二丁酸酯（PDBu）可抑制牛毛细血管内皮（BCE）细胞增殖（K50 = 6±4 nM）以及对人肝癌衍生生长因子（一种血管生成性内皮细胞促分裂原）的DNA合成反应。相反，PDBu对牛主动脉内皮细胞的增殖没有影响，且对牛主动脉平滑肌细胞和BALB/c 3T3细胞有促有丝分裂作用。多项观察结果表明，PDBu对人肝癌衍生生长因子刺激的BCE细胞生长的抑制作用是通过蛋白激酶C介导的。不同的佛波醇化合物根据其作为蛋白激酶C激活剂的效力抑制BCE细胞生长（12 - O - 十四酰佛波醇13 - 乙酸酯＞PDBu＞＞β - 佛波醇12,13 - 二乙酸酯＞＞β - 佛波醇；α - 佛波醇12,13 - 二丁酸酯；α - 佛波醇12,13 - 二癸酸酯）。PDBu以8 nM的表观解离常数与BCE细胞上一类单一的特异性、可饱和位点结合，这与报道的PDBu在其他系统中对蛋白激酶C的亲和力一致。PDBu与BCE细胞的特异性结合被sn - 1,2 - 二辛酰甘油取代，sn - 1,2 - 二辛酰甘油是一种蛋白激酶C激活剂，也是体内激活该激酶的假定第二信使的类似物。弱蛋白激酶C激活剂sn - 1,2 - 二丁酰甘油不影响PDBu结合。BCE细胞的胞质提取物含有一种钙/磷脂酰丝氨酸依赖性蛋白激酶，该激酶可被sn - 1,2 - 二辛酰甘油和PDBu激活，但不能被β - 佛波醇激活。这些发现表明蛋白激酶C激活可使毛细血管内皮细胞对血管生成性内皮细胞促分裂原脱敏。这种细胞内调节机制可能在血管生成的某些阶段被调用，例如当增殖的内皮细胞分化形成不生长的血管时。
