Embryonic diapause, an arrest in development at the blastocyst stage, has been best studied in tammar wallaby Macropus eugenii. Diapause is maintained through a neuroendocrine reflex by the sucking of the young in the pouch. Removing the pouch young (RPY) causes reactivation. During diapause and for 2 weeks after, the embryo is surrounded by an acellular shell coat, so control of diapause and reactivation must be mediated by soluble factors inthe uterine environment. This study uses MALDI imaging mass spectrometry to address this question.
Cryosections of tammar endometrium from diapause (d0RPY), day 4, day 5, day 6 and day 8 after the removal of pouch young were analysed using MALDI imaging mass spectrometry which allows spatial correlation of protein/peptide profiles with H/E stained images. Kruskal Wallis rank-sum test was used to compare MALDI peak intensities between diapause and different days of reactivation. Principal components analysis and a heat map with Hierarchical clustering were generated from average peptide peak intensities. The first two principal components accounted for 80% of the variation between stages. Diapause was distinct from the reactivation stages.PC1 contained peptides which are highly expressed during d4 and d5 RPY which may be involved in the expression of specific proteins which initiate reactivation with an increase in metabolism. D6 and d8 which cluster well in PC2 contain another distinct set of peptides which might promote growth and initiate the expansion of blastocyst. Increase in uterine secretions by d4RPY was evident by the increase in peptide peak intensities in comparison to d0RPY. This supports the contention that changes in the uterine secretions causes embryonic reactivation.