Four-day-old bovine corpus luteum: progesterone production and identification of matrix metalloproteinase activity in vitro.
Four-day-old bovine corpus luteum: progesterone production and identification of matrix metalloproteinase activity in vitro.
复制标题
四日龄牛黄体：体外黄体酮的产生和基质金属蛋白酶活性的鉴定。
DOI：
10.1095/biolreprod53.5.1160
复制
发表时间：
1995
影响因子：
3.6
通讯作者：
Condon,WA
中科院分区：
文献类型：
--
作者：
Tsang,PC;Poff,JP;Boulton,EP;Condon,WA
Little is known about matrix metalloproteinases (MMPs), enzymes that are required for the structural remodeling and angiogenesis that occur in the corpus luteum (CL) during the first several days postovulation. In fact, little attention has focused on early CL function including the regulation of progesterone (P4) production. Thus, the objective of the present study was 1) to investigate the effects of insulin, LH, and dibutyryl cAMP on P4production and cell numbers and 2) to identify MMPs in the 4-day-old CL, with use of a defined culture system. Cultures were seeded with either 1 × 106or 0.5 × 106cells. All cultures containing insulin had higher P4levels and cell numbers (p< 0.05) than those without. In cultures containing insulin, basal P4levels were high throughout the culture period. Furthermore, neither LH nor dibutyryl cAMP stimulated P4production (p> 0.05) at a seeding density of 1 × 106, whereas they stimulated P4production (p< 0.05) at a seeding density of 0.5 × 106on Days 6 and 8 of culture. In conditioned medium of control cultures seeded with 0.5 × 106cells, substrate gel electrophoresis (zymography) showed two intense bands that migrated atMr∼97 000 and ∼65 000–64 000, while two weaker ones migrated atMr∼88 000 and ∼64 000–63 000. The molecular weights of theMr∼97 000 and ∼88 000 species were consistent with MMP-9 family members, while the molecular weights of theMr∼65 000–64 000 and ∼64 000–63 000 species were consistent with MMP-2 family members. Incubation with 1,10 phenanthroline verified that these enzymes were bona fide MMPs. Furthermore, treatment of conditioned medium with p-aminophenylmercuric acetate (APMA) diminished the intensity of theMr∼97 000 and ∼65 000–64 000 bands and enhanced theMr∼63 000–62 000 band, indicating that theMr∼97 000 and ∼65 000–64 000 species were latent forms of MMP-9 and MMP-2, respectively. In addition, no differences in enzyme activity were observed between control and LH samples, while only theMr∼97 000 band decreased significantly in intensity over the 8-day culture period, as corroborated by densitometry. In summary, cells from 4-day-old CL secreted high levels of basal P4, and these levels and cell numbers required insulin. Furthermore, at the lower seeding density and in the presence of insulin, LH and dibutyryl cAMP significantly stimulated P4over controls on Days 6 and 8 of culture. In addition, MMP-9-like and MMP-2-like enzymes were identified in conditioned medium, and their activity was not regulated by LH. Thus, the present findings suggested that luteal cells from 4-day-old CL produce P4and are a potential source of MMP-like enzymes.
