RESUMEN
The aim of this study was to examine the effect of interferons (IFNs) on the recovery of UV-damaged cells by means of measuring cell viability rates. The influence of the recombinant human interferons IFN-alpha, IFN-beta and IFN-gamma on the repair capacity of the UV-irradiated human cell lines WISH and HeLa was studied. The ability of cells to repair UV-induced damage was determined by the comet assay and both short- and long-term survival assays in proliferating cell cultures. We found that INFs negatively regulated DNA repair in cells damaged by UV light. One day after treatment, in both cell lines tested, IFN-alpha had a stronger inhibitory effect than IFN-gamma. Combined treatment with different IFNs exhibited a stronger inhibitory effect on cell recovery than treatment with each of them. The protein kinase inhibitor wortmanin further aggravated the effect of IFNs on cell survival.
Asunto(s)
Reparación del ADN/efectos de los fármacos , Reparación del ADN/efectos de la radiación , Interferones/farmacología , Rayos Ultravioleta , Supervivencia Celular , Ensayo Cometa , Células HeLa , Humanos , Óxido Nítrico/metabolismoRESUMEN
The capacity for nucleotide excision repair of a normal (WISH) and three tumour (MCF-7, HeLa, Namalva) cell lines treated with human recombinant interferons (hrIFN-alpha and hrIFN-gamma) was compared by the host cell reactivation assay. The cells were transfected with in vitro UV-damaged plasmid DNA (pEGFP-N1). The repair capacity was determined by measuring the fluorescence intensity of the expressed marker protein in total cell lysates. The correlation between the interferon-induced NO content and the suppressive effect of interferons on DNA repair was shown. The decrease of repair activity and NO induction by hrIFN-alpha were greatest in WISH, followed by MCF-7, Namalva and HeLa cells, whereas hrIFN-gamma was the best NO inducer and inhibitor for the repair of Namalva, followed by WISH, MCF-7 and HeLa cells. Our data clearly show that the two types of interferon have a strong inhibitory effect on the repair of UV-damaged DNA and this effect is cell type-dependent.
Asunto(s)
Daño del ADN/efectos de los fármacos , Reparación del ADN/efectos de los fármacos , Interferones/farmacología , Rayos Ultravioleta , Neoplasias de la Mama , Línea Celular , Daño del ADN/efectos de la radiación , Femenino , Células HeLa/efectos de la radiación , Humanos , Óxido Nítrico/análisis , Plásmidos/efectos de los fármacos , Transfección , Neoplasias del Cuello UterinoRESUMEN
A simple method for spontaneous transfection into mammalian cells (both adherent and suspension in culture) with plasmid DNA is described. This method does not require any specific DNA carrier or technical device and can be applied for obtaining both transient and stably transfected cells. The efficiency of spontaneous transfection is slightly lower in comparison with that of the conventional calcium phosphate and lipofectin transfection methods and does not depend on the type of cell culture used.