Peculiarities of antioxidant system and iron metabolism in organism during development of tumor resistance to cisplatin

Aim : To study in vivo the peculiarities of changes of iron metabolism and antioxidant system in dynamics of growth of Guerin carcinoma with different sensitivity to cisplatin. Materials and Methods: In order to evaluate the content of metallothionein-1 (MT-1) in tumor homogenates and blood serum of rats with cisplatin-sensitive and cisplatin-resistant Guerin carcinoma the immunoenzyme method was used. The evaluation of ceruloplasmin activity, content of “free iron” complexes, superoxide and NO-generating acti­vity of NADPH-oxidase and iNOS activity in neutrophils, blood serum and tumor homogenates was measured by EPR-spectro­scopy. Results: Maximal accumulation of MT-1 in blood serum and tumor, more pronounced in resistant strain, at the border of latent and exponential phase of growth has been shown that is the evidence of protective role of this protein in the respect to the generation of free radical compounds. It has been determined that in animals with cisplatin-resistant strain of Guerin carcinoma, increase of “free iron” complexes is more apparent both on the level of tumor and organism on the background on increase of CP/TR ratio that is the consequence of organism antioxidant protection system disorder. Conclutions: Mentioned changes in metabolism of iron with its accumulation in tumor and further reprogramming of mitochondria metabolism and activity of NADPH-oxidase for non-transformed cells are favorable conditions for the formation of oxidative phenotype of tumor. Key Words: Guerin carcinoma, metallothionein-1, antioxidant system, transferrin, ceruloplasmin, “free iron” complexes, activity of NADPH-oxidase and iNOS neutrophils

Modifying effects of 5-azacytidine on metal-containing proteins profile in guerin carcinoma with different sensitivity to cytostatics

Aim: To assess the influence of the treatment with 5-azacytidine (5-aza) on the profile of metal-containing proteins and factors of their regulation in Guerin carcinoma cells in vivo. Materials and Methods: The study was conducted on Wistar rats transplanted with wild-type Guerin carcinoma (Guerin/WT) and its strains resistant to cisplatin (Guerin/CP) or doxorubicin (Guerin/Dox). Animals were distributed in 6 groups treated with 5-aza and control animals without treatment. 5-Aza was injected by i.v. route (1 injection in 4 days at a dose of 2 mg/kg starting from the 4th day after tumor transplantation, 4 injections in total). Ferritin levels in blood serum and tumor tissue were measured by ELISA, transferrin and free iron complexes — by low-temperature EPR, miRNA-200b, -133a and -320a levels and promoter methylation — by real-time quantitative reverse transcription polymerase chain reaction. Results: The study has shown that 5-aza treatment caused demethylation of promoter regions of fth1 and tfr1 genes in all studied Guerin carcinoma strains. 5-Aza treatment resulted in a significant decrease of ferritin levels in tumor tissue (by 32.1% in Guerin/WT strain, by 29.8% in Guerin/Dox and by 69.1% in Guerin/CP). These events were accompanied by 3.5-fold and 2-fold increase of free iron complexes levels in tumor tissue of doxorubicin and cisplatin resistant strains, respectively. Also, 5-aza treatment resulted in significantly elevated levels of miR-200b, -133a, 320a expression in tumor tissue. After 5-aza treatment, ferritin levels in blood serum of animals with Guerin/Dox were increased by 23.9%, while in Guerin/Wt and Guerin/CP they were decreased by 17 and 16%, respectively. Conclusion: Alterations of epigenetic regulation upon in vivo treatment with 5-aza change the levels of metal-containing proteins due to DNA demethylation and altered miRNA expression profiles in Guerin carcinoma cells

MODIFYING EFFECTS OF 5-AZACYTIDINE ON METAL-CONTAINING PROTEINS PROFILE IN GUERIN CARCINOMA WITH DIFFERENT SENSITIVITY TO CYTOSTATICS

Aim: To assess the influence of the treatment with 5-azacytidine (5-aza) on the profile of metal-containing proteins and factors of their regulation in Guerin carcinoma cells in vivo. Materials and Methods: The study was conducted on Wistar rats transplanted with wild-type Guerin carcinoma (Guerin/WT) and its strains resistant to cisplatin (Guerin/CP) or doxorubicin (Guerin/Dox). Animals were distributed in 6 groups treated with 5-aza and control animals without treatment. 5-Aza was injected by i.v. route (1 injection in 4 days at a dose of 2 mg/kg starting from the 4th day after tumor transplantation, 4 injections in total). Ferritin levels in blood serum and tumor tissue were measured by ELISA, transferrin and free iron complexes — by low-temperature EPR, miRNA-200b, -133a and -320a levels and promoter methylation — by real-time quantitative reverse transcription polymerase chain reaction. Results: The study has shown that 5-aza treatment caused demethylation of promoter regions of fth1 and tfr1 genes in all studied Guerin carcinoma strains. 5-Aza treatment resulted in a significant decrease of ferritin levels in tumor tissue (by 32.1% in Guerin/WT strain, by 29.8% in Guerin/Dox and by 69.1% in Guerin/CP). These events were accompanied by 3.5-fold and 2-fold increase of free iron complexes levels in tumor tissue of doxorubicin and cisplatin resistant strains, respectively. Also, 5-aza treatment resulted in significantly elevated levels of miR-200b, -133a, 320a expression in tumor tissue. After 5-aza treatment, ferritin levels in blood serum of animals with Guerin/Dox were increased by 23.9%, while in Guerin/Wt and Guerin/CP they were decreased by 17 and 16%, respectively. Conclusion: Alterations of epigenetic regulation upon in vivo treatment with 5-aza change the levels of metal-containing proteins due to DNA demethylation and altered miRNA expression profiles in Guerin carcinoma cells