Search

Pheochromocytomas and paragangliomas (PPGLs) with activated pseudohypoxic pathways are associated with an immature catecholamine phenotype and carry a higher risk for metastasis. For improved understanding of the underlying mechanisms we investigated the impact of hypoxia and pseudohypoxia on catecholamine biosynthesis in pheochromocytoma cells naturally lacking Hif2&alpha, (MPC and MTT) or expressing both Hif1&alpha, and Hif2&alpha, (PC12). Cultivation under extrinsic hypoxia or in spheroid culture (intrinsic hypoxia) increased cellular dopamine and norepinephrine contents in all cell lines. To distinguish further between Hif1&alpha, driven effects we expressed Hif2&alpha, in MTT and MPC-mCherry cells (naturally lacking Hif2&alpha, ). Presence of Hif2&alpha, resulted in similarly increased cellular dopamine and norepinephrine under hypoxia as in the control cells. Furthermore, hypoxia resulted in enhanced phosphorylation of tyrosine hydroxylase (TH). A specific knockdown of Hif1&alpha, in PC12 diminished these effects. Pseudohypoxic conditions, simulated by expression of Hif2&alpha, under normoxia resulted in increased TH phosphorylation, further stimulated by extrinsic hypoxia. Correlations with PPGL tissue data led us to conclude that catecholamine biosynthesis under hypoxia is mainly mediated through increased phosphorylation of TH, regulated as a short-term response (24&ndash, 48 h) by HIF1&alpha, Continuous activation of hypoxia-related genes under pseudohypoxia leads to a HIF2&alpha, mediated phosphorylation of TH (permanent status).

Contains fulltext : 215221.pdf (Publisher’s version ) (Open Access) IDH1(R132H) (isocitrate dehydrogenase 1) mutations play a key role in the development of low-grade gliomas. IDH1(wt) converts isocitrate to alpha-ketoglutarate while reducing nicotinamide adenine dinucleotide phosphate (NADP(+)), whereas IDH1(R132H) uses alpha-ketoglutarate and NADPH to generate the oncometabolite 2-hydroxyglutarate (2-HG). While the effects of 2-HG have been the subject of intense research, the 2-HG independent effects of IDH1(R132H) are still ambiguous. The present study demonstrates that IDH1(R132H) expression but not 2-HG alone leads to significantly decreased tricarboxylic acid (TCA) cycle metabolites, reduced proliferation, and enhanced sensitivity to irradiation in both glioblastoma cells and astrocytes in vitro. Glioblastoma cells, but not astrocytes, showed decreased NADPH and NAD(+) levels upon IDH1(R132H) transduction. However, in astrocytes IDH1(R132H) led to elevated expression of the NAD-synthesizing enzyme nicotinamide phosphoribosyltransferase (NAMPT). These effects were not 2-HG mediated. This suggests that IDH1(R132H) cells utilize NAD(+) to restore NADP pools, which only astrocytes could compensate via induction of NAMPT. We found that the expression of NAMPT is lower in patient-derived IDH1-mutant glioma cells and xenografts compared to IDH1-wildtype models. The Cancer Genome Atlas (TCGA) data analysis confirmed lower NAMPT expression in IDH1-mutant versus IDH1-wildtype gliomas. We show that the IDH1 mutation directly affects the energy homeostasis and redox state in a cell-type dependent manner. Targeting the impairments in metabolism and redox state might open up new avenues for treating IDH1-mutant gliomas.