Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Cumulus Cells”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 55 records · Page 3Linked to original sources
PubMed2002

Lysophosphatidic acid stimulates nuclear and cytoplasmic maturation of golden hamster immature oocytes in vitro via cumulus cells.

Lysophosphatidic acid (LPA), a member of the phospholipid autacoid family, is induced in incubated human follicular fluid by lysophospholipase D. It is well known that LPA functions as a growth factor and the hypothesis that LPA in human follicular fluid takes a part in meiosis of oocytes is quite plausible. We studied the effects of LPA on the maturation of golden hamster immature oocytes in vitro. Hamster oocytes with a germinal vesicle were cultured in Tyrode's albumin lactate pyruvate (TALP) medium with 10(-5) M LPA, 10 ng/ml epidermal growth factor (EGF), 30 ng/ml insulin-like growth factor-1, 1 ng/ml tumor growth factor-alpha or 1 ng/ml basic fibroblast growth factor. The nuclear maturation rates in the LPA and EGF groups were significantly higher than in the control group and the other growth factors did not show any stimulatory effect (LPA group; 74.3% [75/101], EGF group; 82.4% [89/108] vs. control group; 60.2% [59/98], p < 0.05, p < 0.01, respectively). When the cells of cumulus were removed, EGF and LPA did not increase the nuclear maturation rates. Cotreatment EGF and LPA did not significantly enhance the stimulatory effect observed with LPA alone on maturation in vitro. The penetration rate determined by the zona-free hamster oocyte test was significantly higher in the LPA group than in the control group (26.7% vs. 13.2%, p < 0.05) and was comparable with that of oocytes matured in vivo. In conclusion, LPA stimulates the nuclear and cytoplasmic maturation of hamster immature oocytes via cumulus cells.

Animals↗
PubMed1990

Mouse oocytes regulate hyaluronic acid synthesis and mucification by FSH-stimulated cumulus cells.

Mucification (or expansion) of the cumulus cells surrounding the oocyte is thought to depend on the direct action of gonadotropins in stimulating production and deposition of hyaluronic acid (HA) in the extracellular matrix. We now report that the oocyte is essential for this process. Either follicle-stimulating hormone (FSH) at 1 micrograms/ml or dibutyryl cAMP at 2 mM induces mucification of intact cumulus cell-oocyte complexes (COCs) in vitro, but fails to stimulate mucification of isolated cumulus cells. HA synthesis by FSH-stimulated cumulus cells is only approximately 3.5% of the value achieved by FSH-stimulated COCs. Isolated oocytes cultured with or without FSH do not synthesize detectable amounts of HA but induce isolated cumulus cells to increase HA synthesis approximately 13-fold in cocultures with FSH. Medium conditioned by isolated oocytes for 5 hr induces nearly the same level of HA synthesis by cumulus cells under the same culture conditions. FSH also stimulates cumulus cells to increase synthesis of dermatan sulfate proteoglycans (DS-PGs) approximately 3-fold, but this stimulation does not depend upon the presence of oocytes. The results indicate that oocytes produce a soluble factor(s) essential in combination with FSH to stimulate HA, but not DS-PG, synthesis by cumulus cells in vitro and that this factor(s) acts independently or downstream from the FSH-induced formation of cAMP.

Animals↗
PubMed2001

Effects of maturational stage, cumulus cells and coincubation of mature and immature cumulus-oocyte complexes on in vitro penetrability of porcine oocytes.

The in vitro penetrability of porcine oocytes is conditioned by several factors, some of which remain unclear. Knowledge of the different effects of the cellular components involved in penetrability would no doubt serve to simplify laboratory IVF methods. This study was designed to evaluate the effects of the following factors on penetrability: oocyte maturational stage, the presence of isolated or oocyte-attached cumulus cells, and coincubation of in vitro-matured and immature oocytes. Immature oocytes and oocytes matured in Waymouth medium were obtained from non atretic follicles and fertilized in TCM 199 medium. Sperm-rich fractions were collected by the gloved hand method and semen was used for IVF at a final concentration of 1 x 10(6) cells/mL in all experiments. Under the same conditions of IVF, the penetrability of the immature cumulus-oocyte complexes (COCs) was significantly lower than that of mature COCs, in terms of penetration rate and mean number of sperm per penetrated oocyte. This difference was abolished when the oocytes were denuded, leading to similar penetration rates. Coincubation of mature and immature COCs reduced the penetrability of immature COCs compared with that observed when these were incubated in isolation. However, neither the addition of isolated cumulus cells from decumulated mature oocytes nor the addition of denuded mature oocytes to immature COCs modified the penetration rate. These findings suggest that the presence of surrounding cumulus cells is mainly responsible for the differences observed in penetrability, regardless of the maturational stage of the oocyte. Moreover, when mature and immature COCs are coincubated, penetrability of immature COCs is diminished by the effects of the mature COC and not by the independent actions of the cellular components.

Animals↗
PubMed1990

Preendocytotic alterations in cumulus cell gap junctions precede meiotic resumption in the rat cumulus-oocyte complex.

Cumulus cells in the mammalian ovary are normally connected to each other and to their enclosed oocyte by an extensive network of gap junctions (GJs). We have shown that the loss of cumulus cell GJs is correlated temporally with meiotic resumption in the intact preovulatory rat follicle (Larsen et al., 1986). Here we describe morphological changes in GJ particle packing patterns (PPPs) that occur prior to GJ loss and meiotic resumption in hormonally stimulated rat cumulus-oocyte complexes (COCs). In the PMSG-primed rat, 89% of the cumulus cell GJ area detected by freeze-fracture electron microscopy consists of tightly packed junctional particles: 4% exhibit loose PPPs of randomly dispersed particles; and 7% contain a mixture of both tight and loose PPPs. One to 2 hr after stimulation with hCG, the area of GJs containing tight PPPs drops by 50%-60%, while junctions exhibiting loosely organized and mixed patterns increase concomitantly. These shifts in PPPs are accompanied by the appearance of unusual particle-free areas of puckered or ruffled nonjunctional membrane at the GJ periphery. Cumulus cell GJs from isolated COCs incubated in FSH-containing medium demonstrate a similar shift in PPPs prior to meiotic resumption. The appearance of fusing areas of particle-free nonjunctional membrane at the GJ periphery in vitro is correlated with GJ loss and is not seen in COCs treated with dihydrocytochalasin B to inhibit endocytotic removal of cumulus GJs. The structural and temporal nature of these morphological observations supports the hypothesis that interruption of junctional communication plays a role in meiotic maturation of the preovulatory oocyte.

Animals↗
PubMed2000

Development of reconstituted pig embryos by nuclear transfer of cultured cumulus cells.

This study tested the effects of oocyte collection method, activation protocol and maturational age of recipient oocytes on the in vitro development of nuclear transfer embryos reconstructed with cultured cumulus cells. Cumulus cells synchronized in G0/G1 phase by serum-starvation culture were transferred into enucleated oocytes that were collected by aspiration or dissection method and cultured for 33 or 44 h. Reconstituted embryos were activated with a combination of calcium ionophore A23187 or electric pulse and cycloheximide (CHXM), and cultured for 6 days. Oocyte collection methods, activation treatment in the presence of cytochalasin B and activation protocols did not affect the developmental rate of embryos reconstituted with 44-h-matured recipients. However, the development of embryos reconstituted with 33-h-matured recipients was significantly improved (P<0.05) by activation with the combination of electric pulse and CHXM. The present study shows that reconstituted porcine embryos derived from cultured cumulus cells can develop to the blastocyst stage, and that their development can be improved by reconstruction with young oocyte cytoplasts following activation with a combination of electric pulse and CHXM.

Animals↗
PubMed2001

Cumulus cells apoptosis as an indicator to predict the quality of oocytes and the outcome of IVF-ET.

PURPOSE: Our purpose was to establish an evaluation system for oocyte quality based on the incidence of cumulus cells apoptosis and to examine the effect of coculture, using autologous cumulus cells, on the outcome of IVF-ET according to proliferative activities of helper cells and the incidence of cumulus cells apoptosis. METHODS: Cumulus cell masses were collected from 91 mature oocytes among 330 oocytes retrieved from a total of 34 IVF-ET cycles with tubal infertility and unexplained infertility. The incidence of apoptosis in cumulus cells was assessed by apoptosis detection kit fluorescein. On ovum pick up, 2nd day embryos were cocultured with autologous cumulus cells. Prior to coculture, in vitro proliferative activity of cumulus cells was evaluated. RESULTS: Cumulus cells from patient groups over 40 years old had a significantly increased apoptosis incidence, a lower fertilization rate, and the decreased number of oocytes retrieved compared to the other age groups (P < .05). The incidence of cumulus cells apoptosis was significantly lower when the number of oocytes retrieved was 5 or less (P < .05). Cumulus cells from fertilized oocytes (0.43 +/- 0.07%) and those from patients who became pregnant (0.44 +/- 0.11%) following IVF-ET showed a significantly lower incidence of apoptosis compared to those of unfertilized oocytes (1.80 +/- 0.35%; P < .001) and the nonpregnant group (0.81 +/- 0.10%; P < .05). Embryo quality also had a negative correlation with the incidence of cumulus cells apoptosis. Coculture of fertilized oocytes with cumulus cells with high proliferative activity resulted in improved rates of implantation and pregnancy compared to that with poor active cumulus cells. No significant difference was found between the in vitro proliferative activity of cumulus cells and the incidence of cumulus cells apoptosis (P < .063). CONCLUSIONS: The age of women might influence the incidence of apoptosis in cumulus cells, and the increased incidence of apoptosis is associated with the number of oocytes retrieved, the fertilization rate, and the pregnancy outcome following IVF-ET. These results suggest that the incidence of cumulus cells apoptosis can be used in predicting oocyte quality, outcome of IVF-ET, and age-related decline in fertility.

Adult↗
PubMed2002

Effect of cumulus cell removal of in vitro matured bovine oocytes prior to in vitro fertilization on subsequent cleavage rate.

The aim of this study is to identify the effect of cumulus cells removal prior to the in vitro fertilization of matured bovine oocytes on cleavage rate. Denuded, matured oocytes were fertilized in presence or absence of loose cumulus cells, cumulus cell conditioned IVF medium (CCCM), charcoal-treated CCCM and charcoal-treated CCCM supplemented with progesterone at a final concentration of 150 ng/ml. After 18 h of incubation with sperm, the presumptive embryos were cultured on a BRL monolayer and the percentage of cleaved embryos was evaluated on Day 4. Removal of cumulus cells prior to IVF significantly reduced the cleavage rate (25% for denuded oocytes versus 56% for cumulus-oocyte complexes (COCs)). The addition of loose cumulus cells partially restored the effect of denudation (cleavage rate: 37% for denuded oocytes supplemented with loose cumulus cells versus 27% for denuded oocytes and 58% for COCs). CCCM also had a positive effect on the cleavage rate of oocytes denuded prior to IVF (36% for denuded oocytes fertilized in CCCM versus 14% for denuded oocytes). Treating the CCCM with charcoal resulted in complete loss of its effect on cleavage rate (18% for denuded oocytes fertilized in charcoal-treated CCCM versus 34% for denuded oocytes fertilized in CCCM). The addition of progesterone to charcoal-treated CCCM partially restored the reduction of the cleavage rate caused by charcoal treatment (27% for denuded oocytes fertilized in charcoal-treated CCCM supplemented with progesterone versus 14% for denuded oocytes fertilized in charcoal-treated CCCM and 36% for denuded oocytes fertilized in CCCM). In conclusion, removal of cumulus cells prior to IVF adversely affects the cleavage rate through loss of a factor secreted by these cells. This factor probably is progesterone.

Animals↗
PubMed2005

Oocytes prevent cumulus cell apoptosis by maintaining a morphogenic paracrine gradient of bone morphogenetic proteins.

Paracrine factors secreted by the oocyte regulate a broad range of cumulus cell functions. Characteristically, cumulus cells have a low incidence of apoptosis and we proposed that this is due to oocyte-secreted factors acting in an anti-apoptotic manner. Bovine cumulus-oocyte complexes (COC) were aspirated from abattoir-derived ovaries and oocytectomized (OOX) by microsurgical removal of the oocyte. OOX were treated with doses of either denuded oocytes (DO) or various growth factors for 24 hours (+/- rFSH; 0.1 IU/ml). Proportions of apoptotic cumulus cells were assessed using TUNEL and laser confocal scanning microscopy followed by image analysis. Quantification of Bcl-2 and Bax proteins in OOX was undertaken by western analysis. Oocyte removal led to a significant increase in cumulus cell apoptosis compared with COC controls (35% versus 9% TUNEL positive, respectively; P<0.001). Levels of OOX apoptosis were significantly reversed (P<0.001) in a dose-dependent manner when co-cultured with oocytes. Furthermore, the anti-apoptotic effect of oocyte-secreted factors followed a gradient from the site of the oocyte(s). Growth differentiation factor 9 (GDF9) had no significant effect on cumulus cell apoptosis. By contrast, cumulus cell apoptosis was significantly (P<0.001) reduced by bone morphogenetic proteins (BMP) 15, 6 or 7. Accordingly, levels of anti-apoptotic Bcl-2 were high in OOX+DO and OOX+BMP15 and low with OOX+GDF9 or OOX alone, whereas the reverse was observed for pro-apoptotic Bax. DO, BMP15 and BMP6 were also able to protect cumulus cells from undergoing apoptosis induced by staurosporine. FSH partially prevented apoptosis in all treatment groups (P<0.001). Follistatin and a BMP6 neutralizing antibody, which antagonized the anti-apoptotic effects of BMP15 and BMP6, respectively, whether alone or combined, blocked approximately 50% of the anti-apoptotic actions of oocytes. These results are the first to demonstrate that oocyte-secreted factors, and particularly BMP15 and BMP6, maintain the low incidence of cumulus cell apoptosis by establishing a localized gradient of bone morphogenetic proteins.

Animals↗
PubMed2000

Roles of gap junctional communication of cumulus cells in cytoplasmic maturation of porcine oocytes cultured in vitro.

Cumulus cells of the oocyte play important roles in in vitro maturation and subsequent development. One of the routes by which the factors are transmitted from cumulus cells to the oocyte is gap junctional communication (GJC). The function of cumulus cells in in vitro maturation of porcine oocytes was investigated by using a gap junction inhibitor, heptanol. Cumulus-oocyte complexes (COCs) were collected from the ovaries of slaughtered gilts by aspiration. After selection of COCs with intact cumulus cell layers and uniform cytoplasm, they were cultured in a medium with 0, 1, 5, or 10 mM of heptanol for 48 h. After culture in vitro, one group of oocytes was assessed for nuclear maturation and glutathione (GSH) content, and another group was assigned to in vitro fertilization and assessed for the penetrability of oocytes and the degree of progression to male pronuclei (MPN) of penetrated spermatozoa. At the end of in vitro maturation, the oocytes reached metaphase II at a high rate (about 80%) regardless of the presence of heptanol at various concentrations. Cumulus cell expansion and the morphology of oocytes cultured in the medium with heptanol were similar to those of control COCs matured without heptanol. The amount of GSH in cultured oocytes tended to decrease as the concentration of heptanol in the medium was increased. Although there was no difference in the rates of penetrated oocytes cultured in media with different concentrations of heptanol, the proportion of oocytes forming MPN after insemination decreased significantly (P < 0.01) at all concentrations tested. A higher rate of sperm (P < 0.01) failed to degrade their nuclear envelopes after penetration into the oocytes that were treated with heptanol. GJC between the oocyte and cumulus cells might play an important role in regulating the cytoplasmic factor(s) responsible for the removal of sperm nuclear envelopes as well as GSH inflow from cumulus cells.

Animals↗
PubMed2002

Mouse sperm lacking cell surface hyaluronidase PH-20 can pass through the layer of cumulus cells and fertilize the egg.

The function of glycosylphosphatidylinositol-anchored sperm hyaluronidase PH-20 in fertilization has long been believed to enable acrosome-intact sperm to pass through the layer of cumulus cells and reach the egg zona pellucida. In this study, we have produced mice carrying a null mutation in the PH-20 gene using homologous recombination. Despite the absence of sperm PH-20, the mutant male mice were still fertile. In vitro fertilization assays showed that mouse sperm lacking PH-20 possess a reduced ability to disperse cumulus cells from the cumulus mass, resulting in delayed fertilization solely at the early stages after insemination. Moreover, SDS-PAGE of sperm extracts and subsequent Western blot analysis revealed the presence of other hyaluronidase(s), except PH-20, presumably within the acrosome of mouse sperm. These data provide evidence that PH-20 is not essential for fertilization, at least in the mouse, suggesting that the other hyaluronidase(s) may play an important role in sperm penetration through the cumulus cell layer and/or the egg zona pellucida, possibly in cooperation with PH-20, although the importance of sperm motility cannot be neglected.

Animals↗
PubMed2004

Thickness of cumulus cell layer is a significant factor in meiotic competence of buffalo oocytes.

This study evaluated the meiotic competence of buffalo oocytes with different layers of cumulus cells. A total of 588 oocytes were collected from 775 ovaries averaging 0.78 oocytes per ovary. Oocytes with homogenous cytoplasm (n = 441) were selected for in vitro maturation (IVM) and divided into four groups based on their cumulus morphology: a) oocytes with > or = = 3 layers of cumulus cells, b) 1-2 layers of cumulus cells and oocytes with partial remnants or no cumulus cells to be cocultured c) with or d) without cumulus cells. Oocytes in all four groups were matured in 100 microL drop of TCM-199 supplemented with 10 microg/mL follicle stimulating hormone (FSH), 10 microg/mL luteinizing hormone (LH), 1.5 microg/mL estradiol, 75 microg/mL streptomycin, 100 IU/mL penicillin, 10 mM Hepes and 10% FBS at 39 degrees C and 5% CO2 for 24 hours. After IVM, cumulus cells were removed from oocytes using 3 mg/mL hyaluronidase, fixed in 3% glutaraldehyde, stained with DAPI and evaluated for meiotic competence. The oocytes with > or = 3 layers of cumulus cells showed higher maturation rates (p<0.05: 64.5%) than oocytes with partial or no cumulus cells (8.6%) and oocytes co-cultured with cumulus cells (34.5%) but did not differ from oocytes having 1-2 layers of cumulus cells (51.4%). The degeneration rates were higher (p<0.05) for oocytes with partial or no cumulus cells (51%) than rest of the groups (range: 13.8% to 17.4%). These results suggest that buffalo oocytes with intact layers of cumulus cells show better IVM rates than oocytes without cumulus cells and the co-culture of poor quality oocytes with cumulus cells improves their meiotic competence.

Animals↗
PubMed1998

Involvement of meiotic resumption in the disruption of gap junctions between cumulus cells attached to pig oocytes.

The present study was undertaken to examine the mechanisms by which cumulus cells regulate meiotic resumption in pig oocytes using microinjections of lucifer yellow into cumulus-oocyte complexes combined with a fluorescent assay. Some cumulus-oocyte complexes cultured for 0, 8, 16, 24, 32, 40 and 48 h were denuded to assess the nuclear status of oocytes; the remaining complexes were injected with lucifer yellow and monitored for the transfer from the oocyte to the surrounding cumulus cells using confocal laser scanning microscopy. The proportion of oocytes undergoing germinal vesicle breakdown at 16, 24 and 32 h of cultivation was much higher than that of cumulus-oocyte complexes in which all gap junctions within cumulus cells and between cumulus cells and oocyte were disrupted (16 h: 22.7% versus 8.8%, 24 h: 66.7% versus 40.3%, 32 h: 84.0% versus 69.4%), showing that the disruption does not trigger meiotic resumption in the pig oocyte. A significant positive correlation (r = 0.99, P < 0.01) was established between the proportion of germinal vesicle breakdown oocytes and that of cumulus-oocyte complexes exhibiting loss of gap junctions within all cumulus cell layers excluding the innermost layer. From these results, it is concluded that meiotic resumption in pig oocytes is induced by the disruption of gap junctions within cumulus cells, rather than that between the oocyte and cumulus cells, which blocks the conduction of meiosis inhibitory signals from the outer cumulus cells to the oocyte.

Animals↗
PubMed2003

LH reduces proliferative activity of cumulus cells and accelerates GVBD of porcine oocytes.

It has been reported that LH receptor (LHR) mRNA is not detected in cumulus cells of porcine cumulus-oocyte complexes (COCs) just after collection from small antral follicles. The present study showed that the formation of LHR in cumulus cells was up-regulated by the cultivation with 20 ng/ml FSH. When the newly synthesized receptors were stimulated by 1.0 microg/ml LH, significantly higher levels of cAMP and progesterone production in cumulus cells were observed as compared with those of COCs cultured with FSH. A loss of proliferative activity of cumulus cells was induced by the additional LH to FSH-containing medium; however, the inhibitory effect was overcome by progesterone receptor antagonist RU486. Furthermore, the addition of LH also accelerated ongoing GVBD in cumulus cells-enclosed oocytes. These results revealed that during in vitro meiotic maturation of porcine COCs, progesterone secreted by FSH- and LH-stimulated cumulus cells reduced proliferative activity of cumulus cells; the changes of cumulus cells might be involved in inducing meiotic resumption of porcine oocytes.

Animals↗
PubMed2004

Cloned horse pregnancies produced using adult cumulus cells.

The objectives of the present study were to: (1) clone horses using adult cumulus cells; and (2) determine whether the cumulus cell donor affected the outcome. In vivo-matured cumulus-oocyte complexes were obtained using transvaginal ultrasound-guided follicle aspiration; oocytes were used as cytoplasts, whereas cumulus cells (from one of three different mares) were used as donor cells. Immediately following nuclear transfer and activation procedures, cloned embryos were transferred surgically to the oviduct of recipient mares (n = 2-5 embryos per recipient) that had ovulated within 24 h prior to the transfer. An initial pregnancy examination was performed between Days 14 and 16 (Day 0 = surgery); subsequent examinations were then performed every 7-10 days. A total of 136 follicles were aspirated in 96 mares, from which 72 oocytes were recovered (53%). Sixty-two cloned embryos were transferred to recipient mares, which resulted in seven (11.3%) ultrasonographically detectable conceptuses between Days 14 and 16. All seven conceptuses were lost spontaneously between Days 16 and 80. Cumulus cells from Mare 160 tended (P = 0.08) to result in a higher embryo survival rate than cumulus cells from Mare 221 (4/17 v. 1/25 respectively). To our knowledge, this is the first report documenting the establishment of cloned equine pregnancies derived from adult cumulus cells.

Animals↗
PubMed1998

Effects of cumulus cell density during in vitro maturation of the developmental competence of bovine oocytes.

To determine the role of cumulus cells in oocyte maturation, we carried out an investigation on the effects of addition of cumulus cells to the maturation medium on the developmental competence of corona-enclosed oocytes and oocytes denuded from their somatic cells. The addition of cumulus cell (1.6 x 10(6) cells/mL) improved the development of bovine corona-enclosed oocytes, however, addition of a similar number of cumulus cells as cumulus-oocyte-complexes (COCs, cumulus cell density: 4.2 x 10(6) cells/mL) had no effect on the development of oocytes denuded from their somatic cells. To determine if corona-enclosed oocytes can obtain developmental competence without the addition of extra cumulus cells, the effects of cell density during in vitro maturation on the developmental competence were studied. A density of 1.6 to 3.2 x 10(6) cumulus cells/mL was the most effective for in vitro maturation of oocytes with intact gap junctions. The effects of the medium conditioned by COCs on the developmental competence of oocytes was also examined. It was demonstrated that COC-conditioned medium improved the development of bovine oocytes to the blastocyst stage. These data suggest that the developmental competence of bovine oocytes surrounded with corona cells is supported in a cell density-dependent manner in the maturation medium. In addition, the data indicate that cumulus cells benefit bovine oocyte development either by secreting soluble factors which induce developmental competence or by removing an embryo development-suppressive component from the medium.

Animals↗
PubMed1993

Effects of cumulus cells on culture of bovine embryos derived from oocytes matured and fertilized in vitro.

Bovine zygotes derived from oocytes matured and fertilized in vitro were cultured in vitro. In Exp. 1, zygotes were vortexed to remove cumulus cells and then cultured in oviduct epithelial cell-conditioned TCM-199 medium with or without added cumulus cells. Embryos cultured without cumulus cells developed to blastocysts significantly less often than those cultured with added cumulus cells, 3 vs 38% of cleaved ova (P < .05). Co-culture of embryos with cumulus cells during the first 48 h only, or for the duration of the culture period, resulted in development rates similar to those of controls that were not vortexed. In Exp. 2, denuded zygotes were centrifuged (15,850 x g, 3 min) at room temperature to facilitate visualization of pronuclei. Centrifuged zygotes were either placed directly into conditioned medium for culture (with or without added cumulus cells) or DNA was first microinjected into their cytoplasm or a pronucleus. All microinjected embryos were co-cultured with added cumulus cells. Centrifugation and microinjection did not reduce viability significantly when embryos were co-cultured with cumulus cells. Development was reduced significantly when embryos were cultured without cumulus cells. Thus, when oviduct epithelial cell-conditioned medium was used, cumulus cell co-culture was beneficial during the early cleavage stages of in vitro-derived zygotes, including those that received DNA microinjection.

Animals↗
PubMed2005

Apoptosis in cumulus cells, but not in oocytes, may influence bovine embryonic developmental competence.

Aim of our study was to clarify if the occurrence of apoptosis in oocytes and cumulus cells is correlated to bovine oocyte developmental competence. The cumulus-oocyte complexes (COCs) were selected according to cumulus status: G1 with more than five layers of compact cumulus cells, G2 with one to five layers of compact cumulus cells and G3 with expanded cumulus cells. The degree of apoptosis in cumulus cells and oocytes measured by caspase staining and TUNEL assay before and after maturation, and 24 h post-insemination was compared to the cleavage, blastocyst formation and hatching rates of each group. Highest cleavage, blastocyst and hatching rates were found in cumulus-oocyte complexes with more than five layers of compact cumulus cells, but no apoptosis was detected in immature or in vitro matured oocytes, regardless of the cumulus status. Many cumulus cells contained active caspases before maturation, but caspase activity declined dramatically after maturation. TUNEL positive cells were rarely observed in each cumulus-oocyte complex upon oocyte recovery, but a huge increase of them was seen after in vitro maturation. Significantly more TUNEL and caspase positive cells were found in G2 cumulus-oocyte complexes. Our results suggest that: (i) oocyte apoptosis does not account for the inferior oocyte quality of G2 and G3; (ii) apoptosis occurs in cumulus cells regardless of the number and compactness of cumulus cells; and (iii) the degree of apoptosis in the compact cumulus-oocyte complexes (G1 and G2) is negatively correlated to the developmental competence of oocyte.

Animals↗
PubMed2004

Gonadotropin-induced delta14-reductase and delta7-reductase gene expression in cumulus cells during meiotic resumption of porcine oocytes.

Progesterone is produced from cholesterol in cumulus cells during meiotic resumption of porcine oocytes. In follicular cells, it has been shown that exogenous lipoprotein-bound cholesterol ester can be used for steroid hormone production. However, in serum-free medium, progesterone is also secreted by FSH- and LH-stimulated cumulus-oocyte complexes, suggesting that progesterone could be produced from de novo synthesized cholesterol in cumulus cells. In the present study, we investigated the expression of Delta14-reductase and Delta7-reductase, which are the members of the superfamily that converts acetyl-CoA to cholesterol in cumulus cells. The expression of both genes was analyzed by RT-PCR. Both Delta14-reductase mRNA and Delta7-reductase mRNA in cumulus cells, cultured until 4 h, were under the level of detection limit. In response to gonadotropins, both mRNA levels were dramatically up-regulated, reaching a maximum at 20 h. To clarify the role of induced enzymes in cumulus cells, cumulus-oocyte complexes were cultured with either Delta14-reductase inhibitor, AY9944-A-7, or Delta7-reductase inhibitor, BM15.766. The results indicated that these inhibitors significantly suppressed the progesterone production in cumulus cells and meiotic progression of oocytes. The inhibitory effects reached a maximum at 1 microM AY9944-A-7 or 20 microM BM15.766. The addition of 20 ng/ml progesterone overcame the inhibitory effects of both drugs on meiotic resumption of oocytes. These results imply that gonadotropin-induced expression and function of Delta14-reductase and Delta7-reductase in cumulus cells contribute to oocyte meiotic resumption via a progesterone-dependent pathway.

Amino Acid Sequence↗