Spatio-temporal expression of the ACTH receptor during mouse adrenal development: Implication for adrenal growth and differentiation

Abstract

The ACTH receptor or melanocortin 2 receptor (MC2-R) is the key receptor regulating steroid biosynthesis in the adrenal cortex. Moreover, its expression has been implicated as a marker of adrenal tumors with higher differentiation and better clinical prognosis. Thus, ACTH as the ligand of the MC2-R might serve as a differentiation factor in the context of adrenal tumorigenesis. However, the role of ACTH during adrenal development is unknown and can not be determined without detailed knowledge about the expression of its receptor in the developing adrenal gland. Thus, timed pregnancies of 129Sv mice were performed and embryos harvested at day E9.5, E11.5, E13.5, E15.5, and E17.5 and 2 weeks post partum. MC2-R expression was assessed by means of RT-PCR and in situ hybridization (ISH) and correlated with the expression of a differentiation marker (side chain cleavage enzyme, SCC) as well as a proliferation marker (BrdU incorporation) at the various time points. Interestingly, RT-PCR of whole embryo cDNA demonstrated the expression of MC2-R mRNA as early as E9.5, at a time point when the urogenital ridge forms. In contrast, expression of steroidogenic factor 1 (SF-1) was detected at the earliest at E11.5, indicating a SF-1 independent expression of MC2-R in vivo. Comparable expression patterns were found between MC2-R mRNA and SCC mRNA. BrdU, which incorporates into cells undergoing mitosis was detected in a scattered distribution throughout the adrenal glands at each time point. Double staining experiments using BrdU immunohistochemistry and MC2-R ISH and SCC ISH, respectively, revealed the majority of cells expressing SCC or MC2-R to be BrdU negative. However, some BrdU/SCC positive cells could also be detected. These preliminary results suggest that the majority of adrenal cells with a differentiated phenotype resemble non-proliferating cells during adrenal development, whereas undifferentiated cells might represent a pool of proliferating stem cells that further differentiate in cells capable of steroidogenesis.