Effect of cell-plating density on the steroidogenic activity of human cumulus cells

Abstract

Expansion of the cumulus mass results in dilution of cumulus cells in accumulated hyaluronic acid. This process is associated with an increase in progesterone secretion. The present study was carried out to evaluate the relationship between these two processes in vitro. Dilution of human cumulus cells was attempted by changing the cell-plating density of cumulus cells in culture. Concurrent with an increase in cell-plating density from 0.25 \m=x\10 to 8 \ m=x\104, progesterone secretion increased by 10.5 times (P < 0.001), 9 times (P < 0.001), and 5.9 times (P < 0.001), and oestradiol secretion increased by 1.4 times (P < 0.001), 1.1 times (P > 0.05), and 2.6 times (P < 0.005) during days 0\p=n-\3,3\p=n-\5, and 5\p=n-\7 of culture, respectively. However, when steroid secretion was expressed in terms of ng per number of cells, the increase in cell-plating density from 0.25 \m=x\104 to 8 \m=x\104 coincided with a decrease in progesterone secretion of 3 times (P < 0.001), 3.5 times (P < 0.001), and 5.4 times (P < 0.005), and oestradiol secretion decreased by 23 times (P < 0.001), 30 times (P < 0.001), and 12.5 times (P < 0.001) during days 0\p=n-\3,3\p=n-\5,and 5\p=n-\7of culture, respectively. The progesterone:oestradiol ratio increased gradually with the increase in cell-plating density. However, at the higher range of cell-plating density the progesterone: oestradiol ratio decreased with extension of the culture period. Addition of human FSH (hFSH) or hCG increased progesterone secretion when the cumulus cells were plated at 2 \m=x\104, but in most cases not at 0.25 \m=x\104 or 8 \m=x\104 cells per dish. However, both hFSH and hCG decreased oestradiol secretion at 2 \m=x\104 cells per dish. In the presence of gonadotrophins, an increase in cell contact and cytoplasmic processes during days 0\p=n-\3and 3\p=n-\5was noted. This study indicates that secretion of progesterone and oestradiol per number of cells is significantly increased with a decrease in cell-plating density. Furthermore, addition of gonadotrophin increased progesterone and decreased oestradiol secretion at the middle range of cell-plating density.

Effect of androgen substrates on the steroidogenic pattern of cumulus cells: Correlation with cumulus culture morphology

“…It is possible that the difference in P4 secretion between type C/D and type A is magnified in a culture period longer than 1 day, as noted in our previous study in which cumulus masses were cultured for 3 days (7). It is also possible that the nonsignificant difference in P4 secretion that was diagnosed in the present study was due to the decrease in P4 secretion over a long culture period, as described previously (19). Thus, the initial significant difference in P4 secretion between type A and type C/D was diminished after 3 days of culture, probably due to lack of further stimulation.…”

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“…In individual follicles. In vitro, P 4 secretion can be increased in both cell types either by hormonal stimula-primates, P 4 receptors and 3␤-HSD have been codetected in luteal cells, and inhibition of 3␤-HSD resulted tion (7,32) or by adding the appropriate substrate (8,33). Addition of pregnenolone increased cumulus in luteolysis without affecting P 4 receptor density (37).…”