Microarray analysis of mRNA from cumulus cells following in vivo or in vitro maturation of mouse cumulus–oocyte complexes

Abstract

The IVM of mammalian cumulus–oocyte complexes (COCs) yields reduced oocyte developmental competence compared with oocytes matured in vivo. Altered cumulus cell function during IVM is implicated as one cause for this difference. We have conducted a microarray analysis of cumulus cell mRNA following IVM or in vivo maturation (IVV). Mouse COCs were sourced from ovaries of 21-day-old CBAB6F1 mice 46 h after equine chorionic gonadotrophin (5 IU, i.p.) or from oviducts following treatment with 5 IU eCG (61 h) and 5 IU human chorionic gonadotrophin (13 h). IVM was performed in α-Minimal Essential Medium with 50 mIU FSH for 17 h. Three independent RNA samples were assessed using the Affymetrix Gene Chip Mouse Genome 430 2.0 array (Affymetrix, Santa Clara, CA, USA). In total, 1593 genes were differentially expressed, with 811 genes upregulated and 782 genes downregulated in IVM compared with IVV cumulus cells; selected genes were validated by real-time reverse transcription–polymerase chain reaction (RT-PCR). Surprisingly, haemoglobin α (Hba-a1) was highly expressed in IVV relative to IVM cumulus cells, which was verified by both RT-PCR and western blot analysis. Because haemoglobin regulates O2 and/or nitric oxide availability, we postulate that it may contribute to regulation of these gases during the ovulatory period in vivo. These data will provide a useful resource to determine differences in cumulus cell function that are possibly linked to oocyte competence.

Genomic expression profiles in cumulus cells derived from germinal vesicle and MII mouse oocytes

“…Furthermore, when comparing the transcriptome profiles of CCs from in vivo-and in vitromatured human COCs at different nuclear maturation stages, Ouandaogo et al (2012) found that the transcriptomic signature of the CCs varies according to both oocyte maturation stages and the maturation conditions, and suggested that there was a delay in the acquisition of the mature CC phenotype following in vitro maturation (IVM). Therefore, it seems likely that screening for changes in gene expression in CCs under different maturation conditions would make it possible to optimise the in vitro culture system (Kind et al 2013;Salhab et al 2013). However, only limited data are available regarding gene expression profiles of CCs in association with oocyte maturation.…”

Activation of Mouse Cumulus-Oocyte Complex Maturation In Vitro Through EGF-Like Activity of Versican1

“…The same gene expression and functional changes can also be induced in isolated COCs in vitro by treatment with EGF [14,15], any of the EGF-like ligands [7], or follicle-stimulating hormone (FSH) [16]. However, the temporal pattern of response is altered in vitro, including abbreviated gene induction in cumulus cells, loss of synchronization with oocyte meiotic progression, and consequently the developmental potential of oocytes matured in vitro is compromised [17][18][19]. A range of studies have investigated whether one or a combination of the EGF-like family of factors better recapitulate in vivo COC response patterns [7,[20][21][22].…”

“…An appropriate temporal response in the cumulus cells is critical to the acquisition of developmental potential in the oocyte, and it has been widely demonstrated that the temporal pattern of gene expression is suboptimal in COCs matured in vitro [18][19][20][21][23][24][25]. A number of recent studies have investigated the relative impact on oocyte quality of stimulating IVM with the different EGF-like factors.…”

“…A range of studies have investigated whether one or a combination of the EGF-like family of factors better recapitulate in vivo COC response patterns [7,[20][21][22]. Overwhelmingly, the conclusion has been that the gene expression response is truncated in COCs treated in vitro [18][19][20][21][23][24][25] and that this aberrant response contributes to poor oocyte competence in vitro.…”

“…It is clear that versican is an abundant hyaluronan cross-linking protein in the expanded cumulus matrix, which forms a complex ECM structure through protein-hyaluronan interactions as well as protein-protein binding of pentraxin 3 (PTX3) and tumor necrosis factor alpha-induced protein 6 (TNFAIP6) [9]. Each of these products is absent or reduced in cultured COCs [18,19,25], and as a result the in vitro COC matrix is functionally altered. We demonstrated this by showing a loss of capacity to control diffusion of small molecules and signaling factors in COCs matured in vitro [38].…”