THE INVESTIGATION OF DNA DAMAGE IN LYMPHOCYTES BY COMET ASSAY IN CHRONIC VIRAL HEPATITIS B PATIENTS

Abstract

The investigation of the degree of the lymphocyte DNA damage in chronic viral hepatitis B (HBV) patients is of interest for several reasons. Firstly, it is possible to judge indirectly about the depth of the pathological process at the level of the whole organism, with bearing in mind features of the pathogenic replication of hepatitis B virus. Secondly, it is possible to give an estimation of the degree of genotoxic impact of the virus on blood cells that plays an essential role in the shaping of the immune response of the body. The study was executed on 50 blood samples from HBV patients, divided in 5 groups on the fibrosis grade according to METAVIR score: F0 (n = 10), F1 (N = 10), F2 (N = 10), F3 (n = 10), F4 (n = 10). The control group was consisted of 43 volunteers matched by the age and gender without concomitant diseases. From blood samples taken at the time of the admission to the hospital lymphocytes were isolated by density gradient on Ficoll-urografin. The degree of DNA damage in lymphocytes was determined by virtue of alkaline version of the DNA comet assay. There was noted the direct relationship between an increase in % DNA in the tail of comets and the grade of liver fibrosis. So in the control group, % DNA in the tail accounted for 3.75 ± 1.44. In the F0 group % of DNA in the tail was 5.07 ± 1.25, F1 - 6.79 ± 1.79, F2 - 7.65 ± 1.62, F3 - 8.05 ± 1.18, F4 - 9.84 ± 3.09. It is noteworthy that in groups F2, F3, F4 differences were statistically significant in comparison with the control group. Also there was noted the presence of apoptotic cells in F3, F4 groups: 1 and 0.88%, respectively. Identified changes are both important in the description of to molecular patterns of the pathogenesis of chronic hepatitis B associated with damage, and also can serve as an indirect indication of the stage of liver fibrosis.

Keywords

DNA comet assay / hepatitis B / fibrosis

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