INSTITUTO DE BIOTECNOLOGIA Y BIOLOGIA MOLECULAR
Unidad Ejecutora - UE
congresos y reuniones científicas
Endogenous Humanin has antiapoptotic effect on somatolactotrope tumor cells
PIDRE MATIAS LUIS; ZUCCATO CAMILA; GOTTARDO MARÍA FLORENCIA; MORENO AYALA MARIELA A.; SEILICOVICH ADRIANA; ROMANOWSKI VÍCTOR; IMSEN MERCEDES; JAITA GABRIELA
Congreso; 98º Annual Meeting of the Endocrine Society; 2017
Prolactin secreting adenomas are the most frequent type among pituitary tumors. Although macroprolactinomas are benign, they can also be locally aggressive, and invade adjacent structures, resulting in neurological dysfunction. Pharmacological therapy with dopamine agonists remains the mainstay of treatment for pituitary tumors. However, a high percentage of patients with prolactin-secreting pituitary tumors show resistance to dopaminergic treatment. To find alternative therapeutic targets, we studied the role of Humanin in these tumors.Humanin (HN) is a 24-amino acid peptide originally isolated from a cDNA library of surviving neurons of Alzheimer´s disease. HN and Rattin (HNr), an homologous peptide of HN in rat, have cytoprotective action in several cell types such as neurons, lymphocytes and testicular germ cells (1). Previously, we showed that HNr expression in GH3 cells (a somatolactotrope tumor cell line) was higher than in normal anterior pituitary cells, and that exogenous HN inhibited TNF-α-induced apoptosis in these cells (2). In the present study, we evaluated the effect of inhibition of endogenous HNr expression on the apoptosis of GH3 cells using a plasmid encoding shRNA against HNr (p.shHNr). Transfection with p.shHNr increased apoptosis of GH3 cells (assessed by TUNEL) at 24 h (Control: 0.6 %; shRNA: 2.0 %, p< 0.05) and 48 h (Control: 3.8 %; shRNA: 9.4 %, p< 0.05). In presence of TNF-α (50 ng/ml), p.shHNr increased both basal and TNF-α-induced apoptosis of GH3 cells (Control: 23.7 %; shRNA: 47.1 %; TNF-α: 38.3 %; shRNA+TNF-α: 56.3 %, p< 0.05), suggesting that endogenous HNr has cytoprotective and antiapoptotic actions.In order to evaluate the effect of HN inhibition in animal models of pituitary adenoma, we generated a recombinant baculovirus (BV) encoding shHNr (BVshRNA). BV are enveloped viruses formed by a molecule of dsDNA that infects insect and mammalian cells. BVshRNA efficiently infected GH3 cells and increased apoptosis (Control: 1.2 %; BVshRNA: 5.8 %, p< 0.05). For in vivo experiments, female nude mice were injected s.c with 3 x 106 GH3 cells. When tumor volume reached 200 mm3, mice were injected intratumorally with BVcontrol or BVshRNA (MOI of 1x109). After 15 days post-treatment, mice injected with BVshRNA had smaller tumors than controls, indicating that endogenous HNr increases GH3 tumor growth.Our results suggest that endogenous HN exerts antiapoptotic effect in pituitary tumor cells and, thus, alterations in HN expression could play a role in the development of pituitary tumors. Targeting HN using baculovirus allows developing alternative therapies for the treatment of pituitary tumors.