Differentiation of trophoblast cells from mononuclear cytotrophoblast to a multinucleated syncytiotrophoblast is paralleled by alterations in the organelles which reside within these cell lineages. The most drastic alteration is seen in mitochondria, cytotrophoblast mitochondria remain stereotypical with large well-defined cristae while syncytiotrophoblast mitochondria are much smaller, spherical, with a less defined cristae structure. However, the molecular mechanisms which drive this process remains unknown. This study aimed to identify key protein differences between trophoblast mitochondrial subpopulation and associate these to functional and morphological characteristics.
Mitochondria were isolated from the cytotrophoblast and syncytiotrophoblast of healthy term placenta with LC-MS to generate proteomic profiles. To evaluate and confirm the finding western blotting and plate based assays were used in conjunction with O2k Oroboros respirometry to assess the metabolic and bioenergetic capacity of the subpopulations.
Proteomics identified several proteins involved in complexes of the electron transport chain that were decreased in the syncytiotrophoblast mitochondria including NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 12 (p<0.05) (Complex I), Succinate dehydrogenase (p<0.05) (complex II) and ATP synthase subunit A1 (p<0.01) and B (P=<0.0001) (complex V). Furthermore, decreases in proteins involved with carbohydrate metabolism, fatty acid metabolism and amino acid utilisation were observed in syncytiotrophoblast mitochondria including: pyruvate dehydrogenase phosphatase (p<0.05), phosphoenolpyruvate carboxykinase (p<0.002), very long chain acyl-CoA dehydrogenase (p<0.007), Mitochondrial 2-oxoglutarate/malate carrier protein (p<0.05). Protein expression was validated using western blotting, respirometry and enzyme assays confirming pyruvate dehydrogenase activity decreased significantly (p<0.05), there was a decreased maximum respiratory capacity (p<0.001) decreased ATP production in syncytiotrophoblast mitochondria.
This research highlights multiple metabolic pathways which appear altered between the two mitochondrial subpopulations following trophoblast differentiation. These changes appear to have a direct effect on the bioenergetic capacity of the mitochondria present in syncytiotrophoblast with many of the identified proteins linked to the cristae structure and morphology.