Changes in the expression of apoptosis-associated proteins in the temporal cortex and hippocampus of rats during long-term kindling and their correction with minolexin

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Abstract

Epilepsy is one of the most common and serious diseases of the brain, affecting more than 70 million people worldwide. Available anticonvulsants are able to suppress seizures in two thirds of patients, and in the remaining third of patients, epilepsy is considered drug-resistant and other types of treatment are required, such as surgery, which also does not always lead to positive results. Overcoming resistance is a complex task that requires an understanding of the biochemical pathways and general pathological processes underlying epilepsy, primarily apoptosis. The purpose of this work was to study the effect of the antibiotic minolexin on the levels of apoptosis and the expression of apoptosis-associated molecules (p53, Bcl-2, caspase-3 and caspase-8) in the temporal cortex, underlying white matter and hippocampus of Krushinsky-Molodkina rats with hereditary audiogenic epilepsy with long-term kindling. Materials and methods. We used Krushinsky-Molodkina rats at the age of 11 months, which were subjected to audiogenic stimulation and administered intraperitoneally with 1 ml of saline solution or the second-generation tetracycline series minolexin at a dose of 45 mg/kg, dissolved in saline solution for 14 days. The temporal lobe cortex and underlying white matter, the hippocampus, were examined. Apoptosis levels (TUNEL) and expression of apoptosis-associated proteins (p53, Bcl-2, caspase-3 and -8) were assessed (immunohistochemistry, Western blotting). Results. In Krushinsky-Molodkina rats with hereditary audiogenic epilepsy, an increase in the apoptosis level was shown during long-term kindling. A p53-mediated, but caspase-independent mechanism of apoptosis activation has been identified. When minolexin was administered, an anti-apoptotic and neuroprotective effect was observed in the temporal lobe and hippocampus of rats.

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E. D. Bazhanova

Sechenov Institute of Evolutionary Physiology and Biochemistry of the Russian Academy of Sciences; The Federal State-Financed Institution Golikov Research Clinical Center of Toxicology under the Federal Medical Biological Agency

Author for correspondence.
Email: bazhanovae@mail.ru
Russian Federation, St. Petersburg; St. Petersburg

A. A. Kozlov

The Federal State-Financed Institution Golikov Research Clinical Center of Toxicology under the Federal Medical Biological Agency

Email: bazhanovae@mail.ru
Russian Federation, St. Petersburg

Yu. O. Sokolova

The Federal State-Financed Institution Golikov Research Clinical Center of Toxicology under the Federal Medical Biological Agency

Email: bazhanovae@mail.ru
Russian Federation, St. Petersburg

A. A. Suponin

Pavlov First St. Petersburg State Medical University of the Ministry of Healthcare of Russian Federation

Email: bazhanovae@mail.ru
Russian Federation, St. Petersburg

E. O. Demidova

The Federal State-Financed Institution Golikov Research Clinical Center of Toxicology under the Federal Medical Biological Agency

Email: bazhanovae@mail.ru
Russian Federation, St. Petersburg

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2. Fig. 1. Experimental plan.

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3. Fig. 2. Location of the studied structures of the rat brain.

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4. Fig. 3. Changes in the number of TUNEL-positive cells in the studied areas of the brain of Krushinsky – Molodkina rats. (a) – temporal lobe cortex, (b) – white matter, (c) – CA4 area of ​​the hippocampus, (d) – granular layer of the dentate gyrus of the hippocampus, (e) – hilus of the hippocampus. TUNEL-positive cells in the hippocampus of Krushinsky – Molodkina rats, x200. (f) – Control saline, DAPI, (g) – Kindling saline, DAPI, (h) – Kindling antibiotic, DAPI, (i) – Control saline, TUNEL, (j) – Kindling saline, TUNEL, (k) – Kindling antibiotic, TUNEL. p < 0.05 compared with: * with the Control saline group, # with the Kindling saline group.

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5. Fig. 4. The level of p53 expression in the studied areas of the brain of Krushinsky – Molodkina rats. (a) – temporal lobe cortex, (b) – white matter, (c) – CA4 area of ​​the hippocampus, (d) – granular layer of the dentate gyrus of the hippocampus, (e) – hilus of the hippocampus. p53-immunopositive cells in the hippocampus of Krushinsky – Molodkina rats, Immunohistochemistry, x200. (f) – Control saline, (g) – Kindling saline, (h) – Kindling antibiotic. p < 0.05 compared: * with the Control saline group, # with the Kindling saline group.

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6. Fig. 5. The level of Bcl-2 expression in the studied areas of the brain of Krushinsky – Molodkina rats. (a) – temporal lobe cortex, (b) – white matter, (c) – CA4 area of ​​the hippocampus, (d) – granular layer of the dentate gyrus of the hippocampus, (e) – hilus of the hippocampus. Bcl-2-immunopositive cells in the hippocampus of Krushinsky – Molodkina rats, Immunohistochemistry, x200. (f) – Control saline, (g) – Kindling saline, (h) – Kindling antibiotic. p < 0.05 compared: * with the Control saline group, # with the Kindling saline group.

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7. Fig. 6. Changes in the optical density of caspase-3-immunoreactive material in the studied areas of the brain of Krushinsky-Molodkina rats. (a) – temporal lobe cortex, (b) – white matter, (c) – hippocampus. # p < 0.05 compared to the Kindling saline group.

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8. Fig. 7. Changes in the optical density of caspase-8-immunoreactive material in the studied areas of the brain of Krushinsky-Molodkina rats. (a) – temporal lobe cortex, (b) – white matter, (c) – hippocampus. p < 0.05 compared with: * with the Control saline group, # with the Kindling saline group.

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