Endocrine Abstracts

Introduction: Primary aldosteronism (PA) is the most frequent form of endocrine hypertension and is commonly caused by an aldosterone producing adenoma (APA). Germline and somatic mutations in the KCNJ5 gene have been found in up to 40% of APAs and demonstrated to play a crucial role in the pathophysiology of PA.

Aim: Here we characterize and investigate the effects of the most common KCNJ5 mutations on cellular death mechanisms based on an in vitro model.

Methods: Cell lines in COS7 or HAC15 cells stably expressing KCNJ5 mutants (G151R, L168R, G151E, T158A) and a control cell line transfected with empty vector were established using a cumate-inducible PiggyBac vector system. Cell viability and cell death (necrosis and apoptosis) were determined by WST-1 assays or flow cytometry following induction with cumate.

Results: In COS7 cells, the KCNJ5 -G151E and -L161R mutants caused the highest proportion of cell death followed by -G151R and -T158A following cumate induction (0 μg/ml to100 μg/ml). Expression of KCNJ5-T158A in COS7 or HAC15 cells resulted in a similar loss of cell viability. All KCNJ5 mutants tested showed an increase in cell death by apoptosis or necrosis with necrosis causing a greater proportion of cell death than apoptosis, when induced with cumate (5 μg/ml and 25 μg/ml). The KCNJ5 mutations that induced the highest level of necrosis were G151E and L168R followed by G151R and T158A.

Conclusion: KCNJ5 mutations cause cell death mostly through necrosis, albeit to a different extent.