N-homocysteiniylation alters erythropoietin functions

SCHIAPPACASSE, AGUSTINA; WETZLER, DIANA; MALTANERI, ROMINA; NESSE, ALCIRA; CHAMORRO, M. EUGENIA; VITTORI, DANIELA

Mar del Plata

Congreso; LXI Reunión Anual de la Sociedad Argentina de Investigación Clínica; 2016

Sociedad Argentina de Investigación Clínica

Erythropoietin (Epo) is a cytokine known by its hematopoietic properties. Despite the successful introduction of recombinant human Epo to overcome the anemia associated with different pathologies, a significant number of patients fails to respond. High levels of homocysteine ?a risk factor for cardiovascular disease? have been linked with altered protein structure due to N-homocysteinylation of protein lysine residues by reaction with the highly reactive homocysteine thiolactone (HTL). Therefore, it was interesting to study whether the erythropoietic and antiapoptotic properties of Epo changed after N-homocysteinylation. Epo was incubated with HTL at 37 °C for 24 h (Epo:HTL 1:2000 M) and residual HTL was removed by washing. Changes undergone by the molecule were evaluated by polyacrylamide gel electrophoresis, zone capillary electrophoresis, the Ellman reaction and fluorescence and circular dichroism measurements. Regarding erythropoietic action, Epo and Epo-HTL (8 ng/mL) were compared in 48 h-growth of the Epo-dependent UT-7 cells capable of erythroid differentiation. Cell proliferation assay (trypan blue): Epo 45.2±8.8 x104 cell/mL; *EpoHTL 29.6±2.7 x104 cell/ml; *P