EFFECT OF INSULIN SHOCK ON OXIDATIVE MODIFICATION OF PROTEINS IN THE NERVOUS TISSUE OF THE BRAIN OF LABORATORY RATS
Authors: Nikolskaya V. A. , Cheretaev I. V. 
Published: August 2020

The rationale for the study of the formation of products of oxidative modification of proteins (OMP) in the nervous tissue in an experimental study of the effects of insulin shock was the literature data indicating that, oxidative destruction of proteins is one of the early indicators of tissue damage. The OMP considered as a factor in the pathogenesis of nervous tissue in many diseases. In some studies, there is information about the effect of experimental GP on oxidative processes in blood serum, red blood cells, adrenal glands and some other tissues, but the effect of this pathology on the processes of free radical oxygenation in the nervous tissue not studied.

The purpose of this article was to evaluate changes in the level of OMP in the nervous tissue of laboratory rats under the influence of insulin shock.

The experiment was performed on 32 sexually mature mongrel male rats weighing 380–400 g (age 6 months), divided into 4 groups of 8 individuals: control (intact) group, group 1-laboratory animals exposed to a single insulin shock (1 time per day, 1 day); group 2 – laboratory rats exposed to a double insulin shock (1 time per day, 2 days); group 3 – laboratory rats exposed to triple exposure to insulin shock (1 time per day, 3 days). Rats of the experimental groups insulin (3.5 Units) subcutaneously were injected. The presence of hypoglycemic coma determined by the appearance of convulsions. Animals that are intact at this time remained in their cells. Experimental animals slaughtered by decapitation on a guillotine.

The material for research was a homogenate of nervous tissue obtained by the method Of Kutlubaev M. A. and co-authors. The study of the level of OMB in nervous tissue was performed using the Dubinina E. E. method, based on the reaction of oxidized amino acid residues of protein with 2,4-dinitrophenylhydrazine (2,4 – DNFG) with the formation of anhydrous 2,4 – DNFG. Measurements of supernatant extinction at wavelengths (λ) of 356, 370, 430 and 530 nm performed using a 5400-UV spectrophotometer (Ekros-Analytica, Russia).

Statistical processing of the experiment results performed using the student’s parametric t-test, since the experimental data obeyed the law of normal distribution.

It was found, that insulin shock and experimental hyperinsulinemia (HI) contributed to a decrease in free radical protein oxidation in rat nervous tissue homogenate, reducing the total level of OMP carbonyl products in all the study groups exposed to insulin shock: in the 1st group – by 37.6 % (n=8; p≤0.01), in the 2nd group – by 40.4 % (n=8; p≤0.01), in the 3rd group – by 40.8 % (n=8; p≤0.01).

Experimental HI causes significant changes in the level of OMP products in the nervous tissue of laboratory animals, which expressed in a decrease in aldehyde and ketone derivatives of both the main and neutral characters in comparison with the indicators of the intact group.

In all three groups with experimental HI, compared with the intact group, there was a significant decrease in primary and secondary carbonyl products of OMP, which indicates that free radical oxidation processes associated with both fragmentation of protein molecules and their aggregation suppressed under conditions of insulin shock. At one- and three-time exposure to HI, primary markers of oxidative stress, formed mainly due to fragmentation of protein molecules, prevailed among the carbonyl products of OMP in the nervous tissue. The content of primary and secondary carbonyl products was almost equal when the HI applied twice, which indicates that the processes of fragmentation and aggregation of protein molecules were almost the same under these conditions.

In all three experimental groups, compared with the intact group, there was a significant decrease in carbonyl products of both neutral and basic nature due to a decrease in free radical damage to neutral and basic amino acids of protein molecules. In all the study groups, including the intact group, the level of OMPo was significantly lower than the level of OMPn. The most pronounced differences between the level of carbonyl products of OMP and OMPo were observed with three-fold HI, and the least pronounced-with a single one. With increasing duration of insulin shock, the spectrum of carbonyl products of OMP shifted towards increasing the content of the main products, as the role of the main amino acids as targets for damage to protein molecules by free radicals increased.