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Among patients with bone marrow failure (BMF) syndrome, some are happened to have underlying Fanconi anemia (FA), a genetically heterogeneous disease, which is characterized by progressive pancytopenia and cancer susceptibility. Due to heterogeneous nature of the disease, a single genetic test, as in vitro response to DNA cross-linking agents, usually is not enough to make correct diagnosis. The aim of this study was to evaluate whether measuring repair kinetics of radiation-induced DNA double-strand breaks (DSBs) can distinguish Fanconi anemia from other BMF patients. An early step in repair of DSBs is phosphorylation of the histone H2AX, generating gamma-H2AX histone, which extends over mega base-pair regions of DNA from the break site and is visualised as foci (gamma-H2AX foci) with specific antibodies. The primary fibroblasts, established from FA patients, were exposed to gamma-rays, a dose of 2 Gy ((60)Co), incubated for up to 24 hours under repair-permissive conditions, and assayed for the level of gamma-H2AX foci and apoptosis at different recovery times after the treatment. Cell lines originating from FA patients displayed a significant delay in the repair of radiation-induced DNA DSBs relative to non-FA bone marrow failure (non-FA BMF) and control cell lines. The delay is especially evident at recovery time of 24 hours, and is seen as about 8-fold increase of residual gamma-H2AX foci compared to self-state before irradiation. The delay in repair kinetics of FA cells represents the unique feature of FA cellular phenotype, which should be exploited to distinguish FA cellular phenotype.

JOKSIC I., LESKOVAC, A., PETROVIC, S. and JOKSI, G. Vitamin B12 Reduces Ribavirin-Induced Genotoxicity in Phytohemaglutinin-Stimulated Human Lymphocytes. Toboku J. Exp. Med., 2006, 209 (4), 347-354 - Ribavirin, an N-glycosyl nucleoside (1-,6-D-ribofuranosyl-1, 2, 4-triazole-3 carboxamide), is a synthetic purine nucleoside analogue with a broad spectrum of antiviral activity, however, its high toxicity poses a major disadvantage of its use as a therapeutic. Various studies have shown that vitamin B 12 plays a significant role in maintaining the stability of the human genome. We therefore investigated the potential beneficial effect of vitamin B 12 in reducing ribavirin-induced genotoxicity. To test this, we used the cytokinesis-block micronucleus (CBMN) assay. Human blood cells were treated in vitro with increasing doses of ribavirin (0.05, 0.17, 0.32, 0.47 and 0.65,umol/ml) for three different periods of time (2, 4 and 17 hrs). Duplicate cultures were supplemented with 50 mu l of vitamin B 12 during the drug treatment (final concentration of 13.5 mu g/ml). Micronuclei formation and cell proliferation potential were then scored in both sets of samples and the corresponding controls. The results showed that supplementation with vitamin B 12 lowered the frequency of micronuclei (Z = 2.02, p LT 0.04) and recovered the proliferation potential of the treated cells for each treatment period, except for the conditions with the highest concentration of ribavirin and the shortest time. These observations underscore the unique beneficial effects of vitamin B12 in reducing genotoxicity, particularly by recovering the proliferation potential of treated cells, as demonstrated by the decrease in mononucleated cells and enhancement of binucleated and polynucleated cells. The mechanism by which vitamin B 12 reduces ribavirin-induced genotoxicity is related to de novo synthesis of nucleotides, and is worthy of further investigation. ribavirin; reduced genotoxicity; vitamin B 12; human lymphocytes (c) 2006 Tohoku University Medical Press.