Search

Human luteal cells have characteristic organelles, such as well developed smooth endoplasmic reticulum (sER), large mitochondria with tubular cristae, lysosome like granules and lipid droplets. To investigate possible roles of these characteristic organelles, 21 human active corpora lutea, obtained from women aged 32-46, were studied cytochemically on 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD), cytochrome c oxidase (CY-O), NADPH diaphorase (NP-DP) and acid phosphatase (ACP) activities, using the methods by Benkoël et al., Ogawa et al., Ago et Ogawa and Barka et Anderson. Subcellular localization of 3 beta-sterol were also examined using the 3 beta-sterol digitonin complex method by Mizuhira et al., 3 beta-HSD and NP-DP activities were demonstrated in the intercrystal and outer space of mitochondria and sER. CY-O activities were demonstrated in mitochondria. ACP activities were demonstrated in lysosomes and Golgi apparatus. Some lipid droplets contained the crystals of 3 beta-sterol digitonin complex. These data suggest that human luteal mitochondria can synthetize progesterone from cholesterol, and that mitochondrial respiratory chain activities in human luteal cells is highly elevated. Human luteal cells probably require much oxygen. Elevated ACP activities indicate that lysosomes play a role in the steroidogenesis. Some lipid droplets may contain cholesterol and other 3 beta-sterols.

The present study was undertaken to assess the ability of cultured luteal cells from human corpora lutea (CL) in the mid luteal phase and the early pregnancy to secrete steroids and prostaglandins (PGs). Luteal cells responded to hCG with a significant increase (2- to 4-fold) in progesterone (P) production. The addition of hCG to the culture media did not stimulate estradiol (E2) production. In contrast, both P and E2 secretion by luteal cells in early pregnancy were significantly lower than those found in the mid luteal phase. Exposure to hCG did not affect P production by luteal cells in early pregnancy. Arachidonic acid (AA) significantly stimulated PGE2 synthesis by luteal cells in the mid luteal phase in a dose-dependent manner. Both basal PGE2 production and the responsiveness to AA were maintained for the duration of the culture. However, hCG did not affect AA-stimulated PGE2 production. Both PGF2 alpha and 6-keto-PGF1 alpha production abruptly declined as the culture proceeded. PG synthesis by cultured luteal cells in early pregnancy was significantly lower than in the mid luteal phase. The ultrastructural characteristics of luteal cells in early pregnancy, which contained lipid droplets, granular and agranular endoplasmic reticulum and large spherical mitochondria, were maintained after 10 days in culture. The present results demonstrate that P and PGE2 production by cultured luteal cells predominate during the mid luteal phase. These data suggest that PGE2, but not PGF2 alpha, may be involved in the regulation of CL function in the menstrual cycle.

Kinetics of protein synthesis in the human granulosa lutein cell during pregnancy were investigated using the electron microscopic autoradiography of 3H-leucine. The corpus luteum of early pregnancy was chased at 10 and 30 min, 1, 3, 6, 12, 24 and 48 h after pulse labelling (10 min) with 100 microCi of 3H-leucine. Although the whole silver grains were very few at 10 min postpulse, silver grains were localized mainly over the rough endoplasmic reticulum (r-ER, 34.3%) and there were no silver grains over the smooth endoplasmic reticulum (s-ER). At 30 min postpulse, the ratio of the grain number to r-ER is decreased (18.7%) and silver grains begin to appear over the s-ER (5.4%). The total number of silver grains over the cell increases with the lapse of time, the ratio of the grain number over the r-ER slightly increases and then decreases afterwards, while that over the s-ER continues to increase gradually. From these facts, it is felt that some protein synthesized on the r-ER might be transported to the s-ER in the human granulosa lutein cell during pregnancy showing steroidogenic activity.

The purpose of the experiments covered by this report was to see whether or not cell cultures reflect conditions in vivo. We have used mainly the primary cell culture technique of human corpora lutea in TC199 containing 20% fetal calf serum. Steroid concentrations were determined by RIA. As compared with relative low production of estradiol and 17 alpha-hydroxyprogesterone, progesterone (P) production showed a peak of 27.5 +/- 8.7 ng/10(5) cells/ml/2 days on about the sixth day. When the cells were cultured in the presence of hCG, the P concentration (83.9 +/- 13.4) significantly increased and an inhibition in the growth rate of luteal cells was observed. When a drop of suspension of isolated cells was examined under a phase contrast microscope, the cells appeared spherical. In Sudan Black fat staining preparations, numerous lipid granules could be seen in the cytoplasm. Also electromicroscopic studies induced that the cells contained numerous mitochondria and well developed, smooth endoplasmic reticula. The cells exhibited strongly positive 3 beta-hydroxysteroid dehydrogenase activity and the cells on the tenth day culture showed the 46, XX karyotype. As mentioned above, we could show that this in vitro system, namely; monolayer cell culture reflected the conditions in vivo, mainly through P production by luteal cells and the effect of hCG.