Endocrine Abstracts

Background: Pituitary adenomas usually occur as sporadic tumours, but familial cases are increasingly identified. Patients of 15–40% with familial-isolated-pituitary-adenoma (FIPA) harbour germline mutations in the aryl-hydrocarbon receptor interacting gene (AIP). AIP is thought to act as a tumour suppressor gene, with loss of heterozygosity shown in pituitary tumour samples at the 11q13 locus, where AIP is located. Previously we have shown AIP has properties consistent with a tumour suppressor role, with wild-type AIP attenuating cell proliferation, whereas mutant AIP losing this effect and siRNA knock-down of endogenous AIP resulting in increased cell proliferation. However, it is unclear whether the reduced cell proliferation is due to increased activation of the apoptosis pathway.

Aims and objectives: To study the effect of AIP on apoptosis.

Methods: GH3 cells were transiently transfected with wild-type AIP, mutant AIP and empty vector. Caspase assays specific for the common (Caspase-Glo^TM 3, 7), intrinsic (Caspase-Glo^TM 9) and extrinsic (Caspase-Glo^TM 8) caspase pathways were performed to determine the level of apoptosis. Nuclear staining with DAPI was used as a second method to confirm the effect of AIP on apoptosis. GH3 cells were treated with a caspase inhibitor to assess whether the attenuated cell proliferation induced by AIP could be reversed.

Results: Overexpression of AIP resulted in significant increase in caspase 3, 7, 8, and 9 activity compared to empty vector (P<0.0001) and mutant AIP (P<0.05). Nuclear staining confirmed an increase of apoptosis in cells transiently transfected with wild-type AIP compared to mutant AIP and empty vector. Addition of a caspase inhibitor reversed the attenuated cell proliferation.

Discussion: In a rodent somato-mammotroph pituitary cell line overexpression of AIP resulted in the activation of caspase cascade via both the intrinsic and extrinsic pathways. Our results suggest that AIP may express its tumour suppressor effect via the apoptosis pathway.