Abstract
Developmental competence of in vitro matured (IVM) oocytes needs to be improved and this can potentially be achieved by adding recombinant bone morphogenetic protein 15 (BMP15) or growth differentiation factor (GDF9) to IVM. The aim of this study was to determine the effect of a purified pro-mature complex form of recombinant human BMP15 versus the commercially available bioactive forms of BMP15 and GDF9 (both isolated mature regions) during IVM on bovine embryo development and metabolic activity. Bovine cumulus oocyte complexes (COCs) were matured in vitro in control medium or treated with 100 ng/ml pro-mature BMP15, mature BMP15 or mature GDF9 +/− FSH. Metabolic measures of glucose uptake and lactate production from COCs and autofluorescence of NAD(P)H, FAD and GSH were measured in oocytes after IVM. Following in vitro fertilisation and embryo culture, day 8 blastocysts were stained for cell numbers. COCs matured in medium +/− FSH containing pro-mature BMP15 displayed significantly improved blastocyst development (57.7±3.9%, 43.5±4.2%) compared to controls (43.3±2.4%, 28.9±3.7%) and to mature GDF9+FSH (36.1±3.0%). The mature form of BMP15 produced intermediate levels of blastocyst development; not significantly different to control or pro-mature BMP15 levels. Pro-mature BMP15 increased intra-oocyte NAD(P)H, and reduced glutathione (GSH) levels were increased by both forms of BMP15 in the absence of FSH. Exogenous BMP15 in its pro-mature form during IVM provides a functional source of oocyte-secreted factors to improve bovine blastocyst development. This form of BMP15 may prove useful for improving cattle and human artificial reproductive technologies.
Highlights
The oocyte and the somatic cells surrounding the oocyte communicate via a complex bidirectional signalling axis mediated via paracrine and gap junctional means [1]
We examined the effect of the mature (R&D Systems) and promature forms of human bone morphogenetic protein 15 (BMP15), as well as the mature form of mouse Growth differentiation factor 9 (GDF9) (R&D Systems), on bovine oocyte developmental competence following in vitro matured (IVM)
Bioactivity of different BMP15 and GDF9 forms All forms of the three recombinant proteins
Summary
The oocyte and the somatic cells surrounding the oocyte (cumulus cells in antral follicles) communicate via a complex bidirectional signalling axis mediated via paracrine and gap junctional means [1] This communication is important for transporting molecules such as growth factors, cyclic nucleotides, amino acids and other small regulatory molecules from cumulus cells into oocytes, and vice versa, in order to sustain cumulus cell health and oocyte development [1,2,3,4]. Oocyte expression of GDF9 is notably higher than BMP15, suggesting a minor role for BMP15 in the regulation of their reproduction Consistent with this notion, homozygous mutant GDF9 mice are sterile due to a block in follicular development beyond the primary follicle stage [16], whereas BMP15 homozygous mutant mice demonstrate only a mild reduction in fertility [21]. In humans, rare mutations and genetic variance of GDF9 and BMP15 are associated with polycystic ovary syndrome [23], dizygotic twinning [24] and premature ovarian failure [25,26,27]
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