Oxidative DNA Damage of Peripheral Blood Cells and Blood Plasma Сell-Free DNA as an Indicator of the Oxidative Stress Level in Children with Autism Spectrum Disorders and Schizophrenia

Abstract

Background: pathogen heterogeneity and complexity are the main obstacles for schizophrenia and autism spectrum disorders (ASD) differential diagnosis in children. The role of oxidative stress in the molecular mechanisms of schizophrenia and autism pathogenesis is beyond doubt. Free radicals that accumulate during stress can cause oxidative modifications and the formation of breaks in the сell-free DNA (cfDNA) and nuclear DNA of blood cells. To date, it has been proven that 8-hydroxy-2’- deoxyguanosine (8-OHdG) can be considered as an oxidative stress biomarker. However, it is still unclear how pronounced the genotoxic consequences of oxidative stress are in ASD of varying severity and in childhood onset schizophrenia (COS). Objective: to study the relationship between the oxidative DNA damage level in peripheral blood cells and the circulating cell-free DNA characteristics with the severity of COS and the course of ASD in children. Patients and methods: blood samples of 96 patients with childhood autism (CA — F84.0 according to ICD-10), atypical autism (AA — F84.1 according to ICD-10) and with childhood onset schizophrenia (COS — F20.8 according to ICD-10) were obtained from the Child Psychiatry Department of the Mental health research center. Blood samples of the control group (34 people) — from the collection of samples of the Research Centre for medical Genetics. The selection of patients was carried out using the clinical and psychopathological method. Cell-free DNA was isolated by extraction with organic solvents. The concentration of cfDNA was determined fluorimetrically. The level of 8-OHdG in cell-free DNA was determined by binding of the corresponding antibodies on membrane filters, endonuclease activity was determined by radial diffusion in a gel. G0-peripheral blood lymphocytes were isolated by gradient centrifugation. The level of 8-OHdG and the level of the phosphorylated form of histone H2AX (yH2AX) in G0-peripheral blood lymphocytes were analyzed in fixed cells by flow cytofluorometry using appropriate antibodies. Statistical processing was carried out using Microsoft Office Excel, Statistica 6.0, StatGraph. Results and conclusions: oxidative stress has different severity in ASD, occurring in severe form (AA) and mild/moderate form (CA). In CA, the level of oxidative damage to the DNA of lymphocytes tends to increase, but does not reach statistically significant level; the level of oxidative damage to cfDNA does not differ from the control. In AA and, to an even stronger extent, in COS, the level of oxidative damage to the DNA of cells and cfDNA is significantly increased, which indicates the development of systemic oxidative stress, which is not compensated by the body’s antioxidant system. The level of 8-OHdG in the composition of the cfDNA and DNA of the nuclei of peripheral blood cells can be a marker of oxidative stress, which is important not only for diagnosing the severity of the pathological process, but also for treatment regimens development for COS and ASD in children.