The role of mitochondria in the ischemic-reperfusion damage of the brain in operations on aortal arch in conditions of experiment
Authors: Gramatiuk S. M. , Ivanova Yu. V. , Kravchenko V. I. , Lohvinenko O. O. 
Published: October 2020

Objective. To investigate in experiment a functional state of mitochondria from the brain cells after its acute ischemic injury and processes, which take place in the brain in conditions of experimental ischemia/reperfusion; to analyze a correlation of the states investigated with the ischemic-reperfusion changes in the brain of patients, having the aortal arch aneurism, operated in environment of hypothermia and artificial cerebral perfusion.

Materials and metods. First stage of the investigation consisted of estimation of the cerebral cells mitochondria state in conditions of ischemia/reperfusion of the neurons primary culture, obtained from embryos of rats of the 17 - 18 gestation days, using mechanical and the enzyme dissociation in the brain cortex. The second stage of the investigation was conducted on 14 white male rats (Rattus norvegicus). In 7 animals ischemia/reperfusion of the brain was simulated, using temporary mechanical blockade of blood flow along vessels of aortal arch with its subsequent restoration, as following: thoracotomy was performed along lower edge of the third rib, making surgical access to aortal arch and main vessels with their mechanical occlusion during 30 min. Additionally in 7 experimental animals thoracotomy only without impact on cerebral blood flow was performed. After extraction of animals from the experiment the blood was collected from the heart cavities, the brain was extirpated from the skull in accordance to standard method. After finishing of the experiment MTT-test was accomplished for estimation of vital capacity of the brain neurons. The principal parameters of the mitochondria functioning and velocity of their respiration in States II and III were estimated in accordance to Chance and in a respiratory control - a ratio of respiration velocity in a State III towards respiration velocity in a State IV in accordance to Chance. Transmembranous mitochondrial potential in cellular suspension, consisting of cells of the brain. Activity of lactate dehydrogenase in the culture environment was determined. Immunohistochemical investigation of the neurons culture was conducted. Membranous mitochondrial potential was estimated in the neurons cultivated. Respiration of isolated mitochondria was measured. Energetic state of mitochondria was determined. The development degree of apoptotic processes was determined in accordance to the DNA fragmentation degree.

Results. There was established, that index of respiratory control of mitochondria from the ischemized brain, measured in pH 7.4, was lowered more than by 40%, comparing with corresponding index of mitochondria, obtained from a control culture. Velocity of a completely stimulated respiration of mitochondria (State III), which were extracted from the brain in 24 h after ischemia/reperfusion, was significantly lower, than velocity of respiration in mitochondria of control cultures, witnessing presence of damage in respiratory chain and transition of electrons of cerebral cellular mitochondria in ischemia/reperfusion, as well as incapacity of such mitochondria to transit electrons effectively from oxidation substrate towards oxygen due to damage of some complexes of the electron-transporting chain. Thus, in conditions of ischemia several processes, impacting mitochondrial functioning and cellular bioenergetics, occur simultaneously. Due to simultaneous action of these processes the mitochondrial functioning fails so significantly, that becomes one of the key moments in cerebral cellular dysfunction and death. Statistically meaningful difference, taking into account the results, between two groups of investigated animals was not revealed.

Conclusion. In environment of ischemia/reperfusion of the brain in its cells a significant disorders of mitochondrial function occur, which are demonstrated by raising of generation of the oxygen active forms, and by lowering of transmembranous mitochondrial potential and velocity of mitochondrial respiration by 40%, what is accompanied by development of lactate-acidosis. It is expedient to apply a mitochondria-directed antioxidants in complex of the treatment measures in the brain ischemia/reperfusion, as well as desintegrators of respiration and oxidated phosphorilation.