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BOR - Papers in Press, published online ahead of print October 4, 2002.
Biol Reprod 2002, 10.1095/biolreprod.101.001503
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Submitted November 15, 2001
Returned for revision December 6, 2001
Accepted May 24, 2002

Embryo


Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) Acts Independently of the Beta Common Subunit of the GM-CSF Receptor to Prevent Inner Cell Mass Apoptosis in Human Embryos

Cecilia Sjoblom 1*, Matts Wikland 1, Sarah A. Robertson 2
1 Fertilitetscentrum AB
2 Adelaide University

* To whom correspondence should be addressed. E-mail: cecilia.sjoblom{at}celltherapeutics.se.

Abstract

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is expressed in the female reproductive tract in early pregnancy and can act to promote the growth and development of pre-implantation embryos in several species. We have demonstrated in in vitro experiments that the incidence of blastulation in human embryos is increased approximately two-fold when GM-CSF is present in the culture medium. In the current study we investigate the mechanisms underlying the embryotrophic actions of GM-CSF. Using RT-PCR and immunocytochemistry, expression of mRNA and protein of the GM-CSF receptor alpha subunit (GM-R{alpha}) was detected in embryos from first cleavage through blastocyst stages of development, but the GM-CSF receptor beta common subunit (ßc) could not be detected at any stage. When neutralizing antibodies reactive with GM-R{alpha} were added to embryo culture experiments, the development-promoting effect of GM-CSF was ablated. In contrast, GM-CSF activity in embryos was not inhibited either by antibodies to ßc, or by E21R, a synthetic GM-CSF analog that acts to antagonize ßc-mediated GM-CSF signaling. Unexpectedly, E21R was found to mimic native GM-CSF in promoting blastulation. When embryos were assessed for apoptosis and cell number by confocal microscopy after TUNEL and propidium iodine staining, it was found that blastocysts cultured in GM-CSF contained 50% fewer apoptotic nuclei and 30% more viable inner cell mass cells. Together, these data indicate that GM-CSF regulates cell viability in human embryos, and that this potentially occurs through a novel receptor mechanism that is independent of ßc.



Key words: Embryo • Apoptosis • Cytokines • Growth factors • In vitro fertilization


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