ROLE OF ΚB AND AP-1 TRANSCRIPTION FACTORS IN CHANGES IN NITRIC OXIDE PRODUCTION AND UTILIZATION IN THE HEART OF RATS UNDER CHRONIC FLUORIDE INTOXICATION

Fluorides, being hazardous contaminants of soil and drinking water, can get in excessive amounts into human and animal bodies. This is especially true for regions where the fluoride content in soils is very high, for example, Poltava, Dnipropetrovsk, and Kirovohrad regions in Ukraine. Excessive fluoride intake can change the rate of nitric oxide production. The impact of fluorides on changes in nitric oxide production and metabolism in the heart and the role of redox-sensitive transcription factors in these changes are poorly understood. The aim of this study was to determine the effect of activation of κB transcription factors and activator protein 1 on the activity of inducible NO-synthase, constitutive isoforms of NO-synthase, nitrite and nitrate reductase, arginase, concentration of nitrites, peroxynitrite and nitrosothiols in the heart of rats during chronic fluoride intoxication. Materials and methods. The study was performed on 24 adult male Wistar rats weighing 220-260 grams. Animals were randomly divided into 4 groups of 6 animals in each (control, chronic fluoride intoxication group, κB blockade group and activator protein 1 blockade group). The experiment lasted 30 days. We determined the activity of inducible NO-synthase, constitutive isoforms of NO-synthase, the concentration of peroxynitrite alkali and alkaline earth metals, the concentration of nitrites and nitrosothiols, the activity of nitrite reductase, nitrate reductase and arginase. Results. Chronic fluoride intoxication increases the activity of inducible NO-synthase by 1.74 times, does not affect the activity of constitutive isoforms and reduces the activity of arginase by 35.68% compared with the control group of animals. The concentration of nitrites in the heart of rats increases 1.73 times, peroxynitrite 1.43 times, and the concentration of nitrosothiols doubled. The use of κB transcription factor blockers and activator protein 1 reduces nitric oxide production from NO synthases and reduces the concentrations of all nitric oxide metabolites in the heart of rats under conditions of chronic fluoride intoxication. Conclusions. Activation of κB transcription factors and activator protein 1 during chronic excessive intake of fluoride leads to hyperproduction of nitric oxide in the heart of rats due to increased activity of inducible NO-synthase and nitrite reductases. Excess production of nitric oxide under chronic fluoride intoxication leads to the accumulation of nitrites, peroxynitrite and nitrosothiols in the heart of rats.

Fukutin Protein Participates in Cell Proliferation by Enhancing Cyclin D1 Expression through Binding to the Transcription Factor Activator Protein-1: An In Vitro Study

The causative gene of Fukuyama congenital muscular dystrophy (fukutin) is involved in formation of the basement membrane through glycosylation of alpha-dystroglycan. However, there are other proposed functions that have not been fully understood. Using cultured astrocytes (1321N1), we found nuclear localization of fukutin and a positive relationship between fukutin expression and cell proliferation. Among potential proteins regulating cell proliferation, we focused on cyclin D1, by reverse-transcription polymerase chain reaction, Western blotting, immunocytochemistry, enzyme-linked immunosorbent assay (ELISA), and sandwich ELISA. Expression of cyclin D1 was significantly downregulated by fukutin knockdown and significantly upregulated by fukutin overexpression. Moreover, fukutin was proven to bind to the activator protein-1 (AP-1) binding site of cyclin D1 promoter, as well as the AP-1 component c-Jun. The c-Jun phosphorylation status was not significantly influenced by knockdown or overexpression of fukutin. The present results provide in vitro evidence for a novel function of fukutin, which participates in cell proliferation by enhancing cyclin D1 expression through forming a complex with AP-1. It is likely that fukutin is a potential cofactor of AP-1.