Luteal protein secretion during preimplantation in the ferret.
Luteal protein secretion during preimplantation in the ferret.
复制标题
雪貂植入前黄体蛋白的分泌。
DOI：
10.1095/biolreprod48.3.647
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发表时间：
1993
影响因子：
3.6
通讯作者：
Mead,RA
中科院分区：
文献类型：
--
作者：
Huang,JL;Powell,M;Mead,RA
在妊娠或假妊娠第5-11天收集雪貂CL，并在含有放射性标记氨基酸的McCoy's培养基中孵育，以确定雪貂CL在植入前期间合成和分泌蛋白质的能力。从培养基中回收的产物通过一维和二维 SDS-PAGE 分离，然后进行荧光照相，并通过光密度测定法进行定量。选定的分泌蛋白在蛋白质印迹上用特异性抗体初步鉴定。 Ferret CL合成并分泌较多的放射性标记产物。主要分泌蛋白的分子量为 16、22、28、32、47、68 和 185 kDa，并在植入前的所有阶段分泌。妊娠第 5-11 天期间，雪貂黄体蛋白质合成和分泌没有发生质的变化，绵羊催乳素 (oPRL) 和二丁酰 cAMP (dcAMP) 都不影响蛋白质分泌模式。然而，oPRL（100 和 1000 ng/ml）在 36 小时孵育期间增加了放射性标记氨基酸与黄体蛋白的结合。 185 kDa 放射性标记产物的相对迁移率与 α2-巨球蛋白 (M) 亚基的相对迁移率相同。人 2, M 抗体与雪貂黄体提取物和培养基中的产物 (185 kDa) 发生交叉反应，部分纯化的蛋白 (185 kDa) 抑制胰蛋白酶活性。主要放射性标记的分泌蛋白 (32 kDa) 与人组织金属蛋白酶抑制剂 (TIMP) 的抗体表现出微弱的交叉反应。这项研究证明了雪貂 CL 的多种蛋白质分泌产物，其中两种已被初步鉴定为蛋白酶抑制剂。
Ferret CL were collected on Days 5-11 of pregnancy or pseudopregnancy and incubated in McCoy's medium with radiolabeled amino acids to determine the ability of ferret CL to synthesize and secrete proteins during the preimplantation period. Products recovered from the medium were separated by one-and two-dimensional SDS-PAGE followed by fluorography and were quantified by densitometry. Selected secretory proteins were tentatively identified with specific antibodies on Western blots. Ferret CL synthesized and secreted a relatively large number of radiolabeled products. The predominant secretory proteins had molecular masses of 16, 22, 28, 32, 47, 68, and 185 kDa and were secreted at all stages of the preimplantation period. There were no qualitative changes in ferret luteal protein synthesis and secretion between Days 5-11 of pregnancy, and neither ovine prolactin (oPRL) nor dibutyryl cAMP (dcAMP) affected the pattern of protein secretion. However, oPRL (100 and 1000 ng/ml) increased incorporation of radiolabeled amino acids into luteal proteins during a 36-h incubation. The relative mobility of a 185-kDa radiolabeled product was identical to that of a2-macroglobulin (M) subunits. Antibody to human 2, M cross-reacted with a product (185 kDa) in ferret luteal extracts and culture medium, and the partially purified protein (185 kDa) inhibited trypsin activity. The major radiolabeled secretory protein (32 kDa) exhibited weak cross-reaction with antibody to a human tissue inhibitor of metalloproteinase (TIMP). This study demonstrates the wide range of proteinaceous secretory products of the ferret CL, two of which have been tentatively identified as protease inhibitors.