HBV replication of the most prevalent subgenotypes in Argentina induces apoptosis in human hepatocytes

ELIZALDE, MERCEDES; SEVIC, INA; GONZÁLEZ LÓPEZ LEDESMA MM; FLICHMAN, DIEGO MARTIN; BARBINI, LUCIANA; CAMPOS, RODOLFO HECTOR

Mar Del Plata

Simposio; LXI Reunión Científica Anual de la Sociedad Argentina de Investigación Clínica (SAIC), LXIV Reunión Anual de la Sociedad Argentina de Inmunología (SAI), XLVIII Reunión Anual de la Sociedad Argentina de Farmacología Experimental (SAFE); 2016

Introduction. Hepatitis B virus (HBV) causesacute and chronic liver infections. The virus is classified in subgenotypes(sgts), being F1b and F4 the most prevalent in our country. Naturally occurringHBV mutations at the BCP (nts 1762-1764, K130M and V131I at X protein) are associatedwith mechanisms of pathogenesis. Objectives.Investigate if HBV replication induces different mechanisms of hepatocyte celldeath and determine whether this induction is characteristic of the sgts and theX gene variants. Methods. Huh-7 cellswere transfected with a mixture of full genome HBV clones, of F1b and F4 sgts withwild type (wt, KV) and mutated (mut, MI) 1762-64 positions, a system previouslydetermined to be replication efficient. Cell mortality was analyzed by trypanblue staining. Cell death by apoptosis was detected by the observation ofmorphological changes by contrast phase microscopy, acridine orange and ethidiumbromide (AO-EB) staining and flow cytometry. Results. Compared to control cells, HBV replication of wt and mut F1band F4 sgts significantly increased cell mortality, at different post-transfectiontimes. It also induced significant morphological changes (cell shrinkage,chromatin condensation and margination at the nuclear membrane, plasma membraneblebbing and apoptotic bodies) typical of apoptosis and significant increasesin the percentage of early and late apoptotic cells (AO-EB staining and flowcytometry). In addition, it was evident that HBV replication of mutant sgtsinduced higher percentages of apoptotic cells than the wt virus. Conclusions. HBV replication of themost prevalent sgts in Argentina induces human hepatocyte death by apoptosis.These results contribute to describe the molecular mechanisms of HBV pathogenesisin chronic infections.