Effects of in vitro maturation on gene expression in rhesus monkey oocytes

Abstract

In vitro oocyte maturation (IVM) holds great promise as a tool for enhancing clinical treatment of infertility, enhancing availability of nonhuman primates for development of disease models, and facilitating endangered species preservation. However, IVM outcomes have remained significantly below the success rates obtained with in vivo matured (VVM) oocytes from humans and nonhuman primates. A cDNA array-based analysis is presented, comparing the transcriptomes of VVM oocytes with IVM oocytes. We observe a small set of just 59 mRNAs that are differentially expressed between the two cell types. These mRNAs are related to cellular homeostasis, cell-cell interactions including growth factor and hormone stimulation and cell adhesion, and other functions such as mRNA stability and translation. Additionally, we observe in IVM oocytes overexpression of PLAGL1 and MEST, two maternally imprinted genes, indicating a possible interruption or loss of correct epigenetic programming. These results indicate that, under certain IVM conditions, oocytes that are molecularly highly similar to VVM oocytes can be obtained; however, the interruption of normal oocyte-somatic cell interactions during the final hours of oocyte maturation may preclude the establishment of full developmental competence.

Fig. 1.

Summary of procedures employed to obtain oocytes for array analysis. After 7 days of follicle-stimulating hormone (FSH) stimulation, oocytes were aspirated on day 8 and subjected to in vitro maturation (IVM). Alternatively, females were treated with human chorionic gonadotropin (hCG) and in vivo matured (VVM) oocytes were aspirated on day 9. After IVM or VVM, cumulus cells had expanded and were removed by hyaluronidase treatment. Second polar body (PB) extrusion was confirmed, and the oocytes were then lysed for analysis. MII, metaphase II.

Fig. 2.

Hierarchical clustering (HCL) of IVM and VVM oocytes.

Fig. 3.

Comparison of differences in mRNA expression revealed by microarray analysis and independent measurements using quantitative amplification and dot blotting (QADB) or quantitative real-time RT-PCR (qRT-PCR) assays.

Fig. 4.

Ingenuity pathway analysis gene interaction network 1. Red, genes upregulated in IVM oocytes; yellow, genes expressed in both oocytes and cumulus cells; blue, genes expressed in cumulus cells but not in oocytes. *More than one probe set yielded differential expression.

Fig. 5.

Ingenuity pathway analysis gene interaction network 2. Red, genes upregulated in IVM oocytes; green, genes upregulated in VVM oocytes; yellow, genes expressed in both oocytes and cumulus cells; blue, genes expressed in cumulus cells but not in oocytes; purple, genes expressed in oocytes but not in cumulus cells. *More than one probe set yielded differential expression.