Bone morphogenetic protein 2 and 5 are down-regulated in adrenocortical carcinoma and modulate adrenal cell proliferation and steroidogenesis
Inga Johnsen1,4, Roland Kappler2, Christoph Auernhammer3 & Felix Beuschlein1,4
Bone morphogenetic proteins (BMPs) have been demonstrated to impact tumorigenesis in a variety of tumors. As for the adrenal cortex, quantitative real time PCR analyses revealed down-regulation of various BMPs (e.g. BMP2, BMP5) in tissue samples from adrenocortical carcinoma and adrenocortical tumor cell lines in comparison to normal adrenal glands. Other members, by contrast, as seen for BMP6 remained unaltered. Integrity of BMP dependent pathways in the investigated cell lines could be demonstrated by activation of the Smad1/5/8 pathway with subsequent increase of ID protein expression upon incubation with BMP2 or 5. On a functional level, BMP incubation resulted in inhibition of cell proliferation and viability in a dose- and time-dependent manner. Notably, BMP2 (50 ng/ml) and BMP5 (100 ng/ml) treatment also reduced viability of cells, which were co-incubated with the insulin-like growth factor (IGF1, 13 nM), a crucial mitogen of the adrenal and activator of the AKT pathway, by 17 and 40%, respectively. We further analyzed potential cross-signalling of BMPs with IGFs and detected a BMP dependent reduction of AKT phosphorylation under baseline conditions and under IGF co-stimulation. Furthermore, BMPs influenced steroidogenic function, whereas BMP treatment reduced MC2-R and steroidogenic enzyme expression which was accompanied by reduced aldosterone, cortisol and DHEA-S secretion. Notable, effects were more pronounced under forskolin co-treatment. Moreover, in vitro demethylation treatment resulted in re-activation of BMP dependent pathways with concomitant modulation of steroidogenesis. Taken together, we demonstrate that loss of expression of members of the BMP family of ligands is a common finding in adrenocortical tumors and we provide evidence that BMP dependent pathways are likely to be involved in modulation of the malignant and functional phenotype of adrenocortical cancer cells.