Изменение экспрессии генов в тканях крыс при токсоплазмозе

Abstract

The purpose of the study is to study changes in gene expression in rat tissues during toxoplasmosis. Materials and methods. The experiment was conducted on 70 Wistar females weighing 170-220 grams. To achieve this goal, the expression of the proto-oncogenes survivin (BIRC5), epidermal growth factor (ErbB-2/HER2-Neu), GLI, vascular endothelial growth factor (VEGF) and anti-oncogene TP53 was determined in comparison with the reference genes β-actin (ACTB) and GAPDH by PCR analysis in the tissues of 10 healthy female rats and 60 infected with toxoplasma. RNA isolation was performed by the column method using the ReliaPrep RNA Cell Miniprep System (Promega Corporation, USA). The quality of the isolated RNA was evaluated spectrophotometrically. Reverse transcription was performed using M-MuLV RT (New England BioLabs Inc, USA). Primers specific to the genes were prepared using Primer3 and the NCBI Nucleotide database. Amplification was performed on a Real-Time PCR Detection System CFX96 thermal cycler (Bio-Rad, USA), using a qPCRmix-HS SYBR PCR mixture (Eurogen, Russia). Comparative expression of the studied genes was carried out after normalization of each of the samples to the level of the control genes GAPDH and ACTIN-β. Expression analysis was performed by qbase+ and CFX Maestro. Statistical processing of the obtained data was carried out using the program Statistica 10.0. Results and discussion. Toxoplasma increases the expression of survivin (BIRC5) in lung tissue to 0.013 relative units, in liver – to 0.038 relative units, in spleen – to 0.061 relative units, and in brain – to 0.050 relative units. VEGF expression in lungs increased to 0.034 relative units, in liver – to 0.041 relative units, in spleen – to 0.063 relative units, in brain tissues – to 0.080 relative units. There was an increase in the expression of ErbB-2/HER2-Neu in lung tissue to 0.436 relative units, in liver – to 0.259 relative units, in spleen – to 0.271 relative units, and in brain – to 0.131 relative units. GLI expression in lung tissues after toxoplasma infection increased to 0.113 relative units, in liver – to 0.188 relative units, in spleen – to 0.388 relative units, and in brain tissues – to 0.459 relative units. An increase in the expression of the anti-oncogene TP53 in the tissues of the lungs to 0.171 relative units, liver – to 0.295, spleen – to 0.408, and brain – to 0.259 relative units was revealed. Conclusion. It has been shown that toxoplasma can cause an increase in the expression of the proto-oncogenes survivin (BIRC5), epidermal growth factor (ErbB-2/HER2-Neu), GLI and vascular endothelial growth factor (VEGF) with simultaneous enhancement of the anti-oncogene TP53