Porcine Cumulus Cells Research Articles

Insulin-like growth factor-1, a component of serum that enables porcine cumulus cells to expand in response to follicle-stimulating hormone in vitro.

The present study was aimed at determining whether insulin-like growth factor-1 (IGF-1) is a component of fetal bovine serum (FBS) that enables porcine cumulus cells to expand in response to FSH in vitro. Cumulus-oocyte complexes (COC) obtained from 4- to 6-mm follicles of prepubertal gilts were cultured at 39 degrees C for 24 h in media that contained human recombinant IGF-1 (50 ng/ml), FBS (15% v:v), or their combination, with or without FSH (1.5 microg/ml), and cumulus expansion was scored microscopically. Expansion was FSH dependent and was observed only when IGF-1, FBS, or both were present. The proportion of FSH-stimulated COC exhibiting full expansion in response to IGF-1 alone did not differ significantly (p > 0.05) from the proportion in those cultured with FBS or IGF-1+FBS (79 +/- 2.8% vs. 84 +/- 2.2% or 76 +/- 6.2%, respectively). In a concentration-response study, FSH-stimulated expansion was observed in a significant proportion of COC (32 +/- 2.8% vs. 0% control) at 1 ng/ml IGF-1, with the proportions increasing dose-dependently to maximal values between 10 and 75 ng/ml IGF-1, and decreasing at higher IGF-1 concentrations. Exposure of COC to an IGF-1 receptor (IGF-1R)-neutralizing antibody (Ab) for 90 min before addition of FSH and FBS dose-dependently inhibited cumulus expansion, with maximal inhibition at 10 microg/ml (1 +/- 1.0% vs. 68 +/- 1.1% control). In the absence of FBS or IGF-1, some COC had a tendency toward slight expansion when cultured with FSH, and the Ab completely inhibited that effect, suggesting that this may be due to endogenous IGF-1 production by the COC. The Ab effect was reversible and was eliminated by washing twice with fresh medium followed by culture for an additional 24 h in the presence of FSH and FBS. Expression of IGF-1R mRNA in the isolated oocyte and cumulus cells was determined by reverse tramscriptase polymerase chain reaction using sequence-specific primers. The IGF-1R message was detected in both the oocyte and cumulus cells. Collectively, these observations suggest that IGF-1 is a component of serum that enables cumulus cells to expand in response to FSH in vitro, and that the effect is receptor mediated. Since IGF-1 is present in the follicle in vivo, it may have a physiological role during gonadotropin-induced cumulus expansion.

Influence of dbcAMP on the inhibitory effect of cumulus cell factor(s).

The factor(s) produced by porcine cumulus cells (cumulus cell factor (s): CCF) was described as quantitatively inhibiting the maturation of oocytes in vitro (Petr et al, 1989). When 1, 10, 20 or 40 cumulus oocyte complexes (COCs) were cultured in a droplet of medium (vol 10 microliters), germinal vesicle breakdown (GVBD) was observed in 85, 78, 57 or 19% of the oocytes, respectively. GVBD was observed in 82, 84, 80 or 90% of cumulus-free oocytes, respectively, when they were cultured at the same numbers per 10-microliters droplet. When 1, 10, 20 or 40 cumulus-free oocytes were cultured under the same conditions in a medium containing 140 dbcAMP per ml, 61, 63, 60 or 58% of them were observed at GVBD. However, when COCs were cultured in a 10 microliter droplet of medium with 140 micrograms of dbcAMP per ml, GVBD occurred in 64, 42, 9 or 0% respectively. Based on these results, we can conclude that dbcAMP exerted a further inhibitory effect on GVBD in pig oocytes cultured under the influence of inhibitory factor(s) from cumulus cells. On the other hand, dbcAMP was shown to partly overcome the effect of CCF on GVBD in porcine oocytes. This suggestion was based on the finding that a 6-h pre-culture of COCs in a medium with 1,000 micrograms of dbcAMP significantly decreased the subsequent effect of CCF (GVBD: 44%) compared with those pre-cultured in a medium with 140 micrograms of dbcAMP/ml (GVBD:5%) or without dbcAMP (GVBD: 15%).(ABSTRACT TRUNCATED AT 250 WORDS)

Quantitative inhibitory influence of porcine cumulus cells upon the maturation of pig and cattle oocytes in vitro.

Porcine cumulus oocyte complexes (COCs) were cultured together in 10-microliters droplets of culture medium. When 10 COCs were cultured for 24 h, germinal vesicle breakdown (GVBD) occurred in 81% of them. When more COCs (20 or 40) were put into the same volume of medium the frequency of GVBD gradually decreased. This inhibition was not observed in denuded oocytes. The process of GVBD was adversely influenced when 10 COCs were cultured in cumulus-preconditioned medium. It is concluded that porcine cumulus cells produced a factor inhibiting GVBD. After removing the inhibitory block and extensive washing, GVBD of arrested oocytes was significantly accelerated. The addition of LH or heparin only partially overcame the inhibitory action. This factor produced by porcine cumulus cells negatively influenced maturation of bovine oocytes; however, a similar effect was not demonstrated in the mouse. Our results suggest that a high concentration of porcine cumulus cells exerts a quantitative inhibitory effect upon GVBD of porcine and cattle oocytes cultured in vitro.