Freeman S J, Brown N A
J Embryol Exp Morphol. 1985 Apr;86:271-81.
The effects of two proteinase inhibitors, leupeptin and pepstatin on the development of 9.5-day rat conceptuses in vitro has been studied. All cultures were of 48 h duration and the inhibitors were present throughout the entire period. When pepstatin was added to the culture medium (5-25 micrograms/ml) conceptuses developed and grew to an extent that did not differ from untreated controls. However, leupeptin (1-4 micrograms/ml) caused severe growth retardation and abnormal development of conceptuses. The effects of the two inhibitors on the hydrolysis of 125I-labelled BSA and haemoglobin by homogenates of 10.5-day yolk sac indicated the biochemical basis for the differential toxic effects of the two inhibitors on development. Leupeptin was highly inhibitory of the degration of both substrates whereas pepstatin caused no inhibition of 125I-labelled BSA hydrolysis, and only a slight inhibition of haemoglobin hydrolysis. These observations demonstrate that cathepsin D, a lysosomal aspartic proteinase that is specifically inhibited by pepstatin is not involved in yolk-sac-mediated protein utilization by early organogenesis-phase conceptuses and that lysosomal cysteine proteinases, specifically inhibited by leupeptin, are of paramount importance in this yolk sac function.
研究了两种蛋白酶抑制剂——亮抑蛋白酶肽和胃蛋白酶抑制剂对体外培养的9.5天龄大鼠胚胎发育的影响。所有培养持续48小时，且在整个期间均存在抑制剂。当向培养基中添加胃蛋白酶抑制剂（5 - 25微克/毫升）时，胚胎发育并生长到与未处理对照无差异的程度。然而，亮抑蛋白酶肽（1 - 4微克/毫升）导致胚胎严重生长迟缓和发育异常。两种抑制剂对10.5天龄卵黄囊匀浆水解125I标记的牛血清白蛋白（BSA）和血红蛋白的影响表明了这两种抑制剂对发育产生不同毒性作用的生化基础。亮抑蛋白酶肽对两种底物的降解具有高度抑制作用，而胃蛋白酶抑制剂对125I标记的BSA水解无抑制作用，仅对血红蛋白水解有轻微抑制作用。这些观察结果表明，被胃蛋白酶抑制剂特异性抑制的溶酶体天冬氨酸蛋白酶组织蛋白酶D不参与早期器官发生阶段胚胎的卵黄囊介导的蛋白质利用，而被亮抑蛋白酶肽特异性抑制的溶酶体半胱氨酸蛋白酶在这种卵黄囊功能中至关重要。
