Erythropoietin action upon activation of microglial cells

WENKER SHIRLEY; CHAMORRO MARÍA EUGENIA; VITTORI DANIELA; NESSE, ALCIRA

Florencia

Congreso; 8th World Congress of IBRO (International Brain Research Organization); 2011

International Brain Research Organization

Erythropoietin (Epo) is well known as a growth factor that maintains the number of circulating erythrocytes primarily by preventing apoptosis of erythroid progenitors. However, its biological role has been expanded by the finding of specific receptors in non-hematopoietic tissues, such as neuronal cells. Previously, we observed that Epo has a neuroprotective action on neuronal SH-SY5Y cells against cytotoxicity induced by staurosporine, TNF-alfa or hypoxia. During cerebral ischemia, hypoxia may not only cause neuronal cell injury in a direct way, since neuron-microglia interactions are important for neuronal viability. Because of the role of hypoxia in proinflammatory mechanisms in the nervous system, and the protective effect of Epo against proinflammatory conditions, we focused our investigation on the role of this hormone in microglial activation. Activation was characterized in murine microglial cells (EOC-2) exposed to TNF-alfa, LPS or CoCl2-induced chemical hypoxia, by measurement of cell proliferation and proinflammatory mediators (nitrites and TNF-alfa). CoCl2 exposure increased nitrites (Griess: C 5.8±2.0, CoCl2 32.0±3.1 nmol/ml, P< 0.001, N=6) and TNF-alfa production (ELISA: C 211.8±33.2; CoCl2 463.7±37.7 pg/ml, P< 0.01, N=3) as well as iNOS expression. MTT data, microscopy cell counting, and PCNA staining suggest that CoCl2 induces cell proliferation (MTT: C 0.276±0.024; CoCl2 0.580±0.071, P< 0.01, N=7). Pretreatment with Epo prevented the CoCl2 effect on cell proliferation (MTT: CoCl2 0.580±0.071, Epo-CoCl2 0.356±0.034, P< 0.05, N=7; Cell count: CoCl2 (1.23±0.18)105, Epo-CoCl2 (0.58±0.09)105 cell/ml, P< 0.05, N=7) without altering the iNOS protein expression, nitrite production (CoCl2 30.8±5.8, Epo-CoCl2 32.6±6.0 nmol/ml, NS, N=5), or TNF-alfa secretion (CoCl2 463.7±37.7, Epo-CoCl2 545.4±175.4 pg/ml, NS, N=3). Moreover, Epo prevented the intracellular ROS levels induced by CoCl2 (DCF fluorescence microscopy). Results suggest that although Epo is unable to prevent nitrite and TNF-alfa production during activation of microglia, it has an antioxidant and antiproliferative effect which could ameliorate a long-term inflammation condition.