Daston G P, Baines D, Yonker J E, Lehman-McKeeman L D
Human and Environmental Safety Division, Miami Valley Laboratories, Procter & Gamble Company, Cincinnati, Ohio 45239.
Teratology. 1991 Mar;43(3):253-61. doi: 10.1002/tera.1420430309.
Altered lysosomal function in the visceral yolk sac can result in abnormal development. As proteolysis is an important function of the rodent visceral yolk sac during early and mid-gestation, we characterized the lysosomal proteolytic enzyme activity of this extraembryonic membrane and determined the effects of inhibitors of protein degradation on embryonic development. Constituent activities of cysteine and aspartic acid proteinases were measured in rat visceral yolk sac on gestation day 12, and the effects of the cysteine proteinase inhibitors leupeptin, E-64 [trans-epoxysuccinyl-l-leucylamido(4-guanido)butane] and N-ethylmaleimide and the aspartic acid proteinase inhibitor pepstatin were determined in Sprague-Dawley rat embryos cultured in vitro from gestation days 10-12. It was determined that only cysteine proteinases, primarily cathepsins B and L, are active in the mid-gestation visceral yolk sac. The cysteine proteinase inhibitors leupeptin and E-64 both produced a concentration-related decrease in embryonic growth, as measured by crown-rump length, somite number, and embryonic protein content, and a concentration-related increase in incidence of abnormalities. A characteristic pattern of abnormalities was produced which involved a decrease in neural tube volume and the formation of a subectodermal blister opposite the point of attachment of the vitelline vessels. At high concentrations, anophthalmia was also observed. The decreased neural tube volume was associated with increased osmolality of the exocoelomic fluid, the major extraembryonic fluid compartment. It is possible that the osmotic change decreased neural tube volume by causing water to move to the compartment with a higher solute concentration, out of the embryo.(ABSTRACT TRUNCATED AT 250 WORDS)
内脏卵黄囊中溶酶体功能的改变可导致发育异常。由于蛋白水解是啮齿动物妊娠早期和中期内脏卵黄囊的一项重要功能，我们对这种胚外膜的溶酶体蛋白水解酶活性进行了表征，并确定了蛋白质降解抑制剂对胚胎发育的影响。在妊娠第12天测定大鼠内脏卵黄囊中半胱氨酸和天冬氨酸蛋白酶的组成活性，并在妊娠第10 - 12天体外培养的斯普拉格-道利大鼠胚胎中确定半胱氨酸蛋白酶抑制剂亮抑酶肽、E - 64 [反式环氧琥珀酰-L-亮氨酰胺(4-胍基)丁烷]和N - 乙基马来酰胺以及天冬氨酸蛋白酶抑制剂胃蛋白酶抑制剂的作用。结果表明，只有半胱氨酸蛋白酶，主要是组织蛋白酶B和L，在妊娠中期的内脏卵黄囊中具有活性。半胱氨酸蛋白酶抑制剂亮抑酶肽和E - 64均导致胚胎生长呈浓度依赖性下降，通过顶臀长度、体节数和胚胎蛋白含量来衡量，同时异常发生率呈浓度依赖性增加。产生了一种特征性的异常模式，包括神经管体积减小以及在卵黄血管附着点相对处形成外胚层下水泡。在高浓度时，还观察到无眼畸形。神经管体积减小与胚外主要液体腔室——外体腔液渗透压升高有关。有可能这种渗透压变化通过使水从胚胎中转移到溶质浓度较高的腔室而降低了神经管体积。（摘要截短于250字）
