Method for correcting body condition in case of traumatic brain injury

Abstract

FIELD: medicine.SUBSTANCE: invention relates can be used in the treatment of traumatic brain injury in an experiment. For this purpose, molecular hydrogen is used in the form of bubbled water at a dose of 0.3-0.5 mL / kg per day orally. The first administration of bubbled water is carried out 30 minutes after traumatic brain injury. The bubbled water is administered using a probe for 10 days.EFFECT: method reduces the risk of post-traumatic hypoxic conditions after traumatic brain injury in an experiment by limiting damage to the microvasculature, reducing endogenous intoxication, and restoring the endothelial lining of blood vessels in the post-traumatic period of traumatic brain injury, which minimizes side effects after injury.2 cl, 5 tbl, 4 ex

Description

[0001] This invention relates to experimental medicine, specifically experimental pharmacology, and concerns a method for correcting the body's condition following traumatic brain injury. The development of methods for correcting the body's condition after a traumatic brain injury is aimed at improving quality of life and health, and also significantly reducing the economic costs associated with patient rehabilitation in the post-traumatic period.

[0002] TBI is a complex multidisciplinary problem, one of the most significant in healthcare. The human brain is extremely vulnerable to injuries, as they can impair quality of life due to profound, developing cognitive and neurobehavioral dysfunction due to impaired tissue perfusion and oxygenation. According to statistics, this "silent epidemic" leads to death and disability in victims more often than any other traumatic injury (Sabirov D.M., Rosstalnaya A.L., Makhmudov M.A. Epidemiological Features of Traumatic Brain Injury / / Bulletin of Emergency Medicine, 2019. 12 (2). pp. 61-66).

[0003] In TBI, brain damage is determined by the degree of tissue blood flow shutdown at the time of ischemia and the duration of the ischemic period itself. Microthrombi form in the vessels of the microcirculatory bed, which, by disrupting vascular patency and reducing blood flow velocity, lead to the formation of ischemic foci and secondary brain damage. Ischemic brain damage is aggravated by factors such as hypotension, hypoxia, increased intracranial pressure, local tissue compression, and, in the later stages, vasospasm in the remaining vessels. Vasospasm is accompanied by venous congestion (Modi NJ, Agrawal M, Sinha VD Post-traumatic subarachnoid hemorrhage: A review / / Neurology India, 2016. 64 (1). P. S8-S13). As a result of excessive waterlogging of brain cells and intercellular space, edema and swelling of the brain develop, which causes further deterioration of blood circulation, metabolism and functional activity of the brain (Boyko AN, Batysheva T.T., Kostenko EV, Zaitsev KA Traumatic brain injury / / Consilium Medicum, 2007. 9 (8). P. 5-10).

[0004] Massive microcirculation disorders accompanied by increased formation of free radicals, damage to the endothelium of the vascular walls with the development of perivascular edema cause the death of neurons, impairment of cognitive function and neurological status of the patient (Deryugina A.V., Polozova A.V., Nikolsky V.O., Boyarinov G.A. Functional indices of erythrocytes and cerebral microcirculation against the background of cytoflavin action after traumatic brain injury / / Experimental and clinical pharmacology, 2020. 83 (1). P. 13 - 18; Karchevskaya A.E., Payushina O.V., Sharova E.V., Oknina L.B., Titov O.Yu. Neuroinflammation as a process of secondary damage in traumatic brain injury / / Annals of clinical and experimental neurology, 2023. 17 (1). pp. 55-68; Ali HT, Sula I., AbuHamdia A., Elejla SA, Elrefaey A., Hamdar H., Elfil M. Nervous System Response to Neurotrauma: A Narrative Review of Cerebrovascular and Cellular Changes After Neurotrauma / / Journal of Molecular Neuroscience, 2024.74 (1). R. 22).

[0005] TBI results in an inflammatory response that is not limited to the brain and often causes multiple organ dysfunction syndrome, which is accompanied by an increase in medium-weight molecules, toxins, and cytokines.

[0006] In this regard, the development and implementation of new effective methods and means for correcting the body's condition after injury is of utmost importance. These methods improve cerebral blood flow, limit the development of endothelial dysfunction, reduce cerebral ischemia, and decrease the level of free radicals and toxins. Moreover, these drugs and methods should have minimal side effects and improved bioavailability.

[0007] A patent search revealed various pharmacological drugs and correction methods aimed at restoring hemodynamics, stopping systemic inflammation and reducing secondary damage to eliminate the phenomena of brain hypoxia after TBI.

[0008] The patent (RU 2498826 C1, cl. A61M 37 / 00, A61K 31 / 573, A61K 31 / 522, A61P 43 / 00, published 20.11.2013) describes a method for treating patients with traumatic brain injury. For this purpose, in addition to standard drug therapy, 0.3 ml of dexamethasone solution and 0.4 ml of trental solution are additionally administered into the area of ​​the inferolateral angle of the orbit, located between the eyeball and the bony orbit, the external and inferior rectus muscles of the eye. The administration is carried out daily once a day for 5-7 days. The method ensures the achievement of the maximum concentration of drugs in the brain tissue, administered into the retrobulbar space, in a shorter time than with other routes of administration, which, in turn, leads to a more rapid removal of brain tissue edema, as well as peripapillary edema of the optic nerve and, as a consequence, an accelerated recovery from the comatose state of a patient with TBI.This invention addresses the recovery of a patient from a coma, but does not describe the drug's effect on the patient's overall health. Dexamethasone is a steroid drug. Steroids are used with good efficacy to treat cerebral edema associated with brain tumors and, as laboratory studies have shown, reduce the production of free radicals and have a protective effect on the brain. However, several clinical studies of TBI have not shown a clear positive effect on outcome or intracranial pressure (Perel P., Edwards P., Wentz R., Roberts I. Systematic review of prognostic models in traumatic brain injury / / BMC medical informatics and decision making, 2006. 6. P. 38). In addition, it is known that the main disadvantage of dexamethasone is the large number of side effects.

[0009] The patent (RU 247676 C2, cl. A61M 1 / 36, A61K 31 / 4015, A61K 35 / 18, A61P 25 / 28, published 10.03.2013) proposes a method for treating patients with traumatic brain injury in the acute period also using dexamethasone, by administering it using the patient's red blood cells. For this, the patient's blood is collected, followed by the isolation of 50 ml of red blood cells. Then, the obtained red blood cells are saturated with 3 ml of dexamethasone and returned to the patient's circulatory system. The procedure is carried out once a day with an interval of 2-3 days. The course of treatment is 5 procedures. Moreover, the effectiveness of this method has been determined for patients with a history of developing hypertension syndrome. The method allows to increase the effectiveness of treatment for this category of patients by ensuring the most complete delivery of the drug. The disadvantages of this method are the same as in the previous method, since regardless of the method of administration, the same drug is used.

[0010] A similar administration method is used in the patent "Method for treating patients with traumatic brain injury associated with hypertensive syndrome during the rehabilitation period" (RU 2470676 C2, cl. A61M 1 / 36, A61K 31 / 4015, A61K 35 / 18, A61P 25 / 28, published 27.12.2012). The difference lies in the introduction of nootropil into the body. The use of the proposed method allows for increased effectiveness and reduced treatment duration. The main disadvantage of nootropil is its side effects, such as headache, drowsiness, imbalance, stomach pain, diarrhea, weakness, nausea, and anxiety.

[0011] The patent (RU 2406506 C1, cl. A61K 31 / 685, A61K 47 / 32, A61K 9 / 08, A61P 25 / 28, published on 20.12.2010) describes a pharmaceutical composition of choline alfoscerate in the form of an injection solution (cholitilin ®), which has nootropic activity and cholinomimetic action, as well as a method for the treatment / prevention of central nervous system diseases and the consequences of traumatic brain injury. The pharmaceutical composition contains choline alfoscerate as an active ingredient in a therapeutically effective amount and povidone (plasdone or kollidon) as a pharmaceutically acceptable carrier in an amount of 0.2-0.8 wt.% based on 100% of the total composition. The pharmaceutical composition maintains its stability during long-term storage, has minimal side effects, and improved bioavailability. The main drawback of the pharmaceutical composition is the side effects associated with the cholinergic action of cholelithin, such as nausea, abdominal pain, and permanent confusion.

[0012] The patent (RU 2464976 C1, cl. A61K 31 / 132, A61K 31 / 44, A61K 31 / 7068, A61K 31 / 727 and A61P 25 / 28, published on 27.10.2012) proposes a method for intensive care of patients in a vegetative state. For this purpose, a microcatheter is inserted into the carotid basin to the level of the cavernous region of the most affected cerebral hemisphere or into the vertebrobasilar basin to the V4 segment in the presence of signs of post-dislocation damage to the brainstem in patients with the consequences of traumatic brain injury, as well as in the presence of a diffuse decrease in metabolism of more than 60% in patients with the consequences of hypoxic brain damage. Then, a continuous infusion of the following medications is administered over seven days: sequentially administering Neoton 14-20 mg / kg / day, Nimotop 0.1-0.2 mg / kg / day, and Ceraxon 85-100 mg / kg / day, with the daily dose divided into two administrations. Concurrently, administer heparin 10,000-12,000 units per day.This method provides effective treatment by creating high drug concentrations in the affected brain area in patients with various etiologies of vegetative states. The main drawback of pharmaceutical combination therapy is the side effects associated with Nimotop, including decreased blood pressure, bradycardia, hot flashes, arrhythmias, nausea, diarrhea, gastrointestinal discomfort, dizziness, headache, asthenia, depression, fatigue, drowsiness, difficulty breathing, pulmonary edema, and thrombocytopenia.

[0013] Another group of patents includes the use of physiotherapeutic interventions rather than medications in the correction of TBI.

[0014] The patent (RU 2666121 C1, class A61N 5 / 02, published 05.09.2018) describes a method for treating patients with post-traumatic brain injury in the early stages of the disease. Physiotherapeutic intervention is carried out from the moment the hemodynamic state is stabilized. Impact is carried out against the background of intensive therapy aimed at maintaining vital functions of the body. As a physiotherapeutic intervention, exposure to the projection zones of focal brain lesions with broadband infrared radiation with a wavelength range of 1-56 μm, modulated by terahertz radiation across the entire radiation spectrum, is used. The IR-Dipole device is used. The maximum radiation power is 30 mW at the base of the emitter with a reflector diameter of 9 cm. The radiation intensity density is 2.4 mW / cm 2The exposure time is 22.5 minutes. The emitter and reflector are positioned 1 cm from the surface of the focal brain lesion projection area. One procedure is performed daily. The treatment course consists of 10 procedures. This method improves the effectiveness of treatment for patients with post-traumatic brain injury in the early stages of the disease by improving tissue blood flow and nerve tissue regeneration. The main disadvantage of this method is that it can only be used after hemodynamic stabilization, which limits its use in the acute phase of TBI.

[0015] The patent (RU 2441681 C1, class A61N 5 / 06, published 10.02.2012) presents a method for treating patients in the acute period of severe traumatic brain injury. For this purpose, narrow-band LED radiation with a wavelength of 540 ± 20 nm is applied in a continuous mode to the orbital areas of both eyes with motionless contact for 5 minutes and to the “collar” zone with labile contact for 10 minutes. In this case, the radiation intensity is 100%, the radiation power is 3.2 mW, the power density is 0.5 mW / cm 2The total exposure time is 20 minutes. The treatment course consists of 10 procedures. This method restores impaired compensatory and regenerative mechanisms in the brain, improving microcirculation, normalizing vascular tone, autonomic dysfunction, and glucose levels in the peripheral blood. A disadvantage of this method is that it is used in conjunction with drug therapy, and there are no comparable examples of this therapy without electromagnetic stimulation, making it impossible to determine the effectiveness of narrowband LED radiation. Furthermore, the stimulation is performed in the orbital area of ​​both eyes, with motionless contact for 5 minutes at a time, which may negatively impact vision.

[0016] The patent (RU 2645968 C1, class A61N 2 / 00, published February 28, 2018) describes a method for treating patients with post-traumatic disorders of the bioelectrical activity of the brain using a low-frequency pulsed magnetic field using the Neuro-MS / D device. The Fc3, Fcz, and Fc4 zones of the cerebral cortex are affected with a strength of 0.4 T, using a small ring-shaped inductor with an external diameter of 100 mm and a frequency of 0.5-1.0 Hz. The effect is carried out by contact, steadily for 2 minutes on each zone. The total transcranial exposure time is 6 minutes, the treatment course is 10 procedures, one procedure performed daily. It has been shown that the use of this method causes a decrease in existing paroxysmal activity, as well as the formation of a normal alpha rhythm in case of insufficient functional activity of the cerebral cortex, that is, the bioelectrical activity of the brain is normalized.Furthermore, it improves mood and increases motivation for restorative treatment and recovery. However, this method does not improve microcirculation, reduce oxidative stress, or reduce endothelial dysfunction.

[0017] A method for using a drug containing a functionally significant fragment of human apolipoprotein A-I as a treatment for traumatic brain injury (RU 2826364 C1, cl. A61K 38 / 17, A61P 25 / 00, published 09.09.2024) is known. Apolipoproteins are natural origin, are structural components of serum lipoproteins, and are able to cross the blood-brain barrier, including through cholesterol-mediated endocytosis into transformed endothelial cells of human brain microvessels and transcytosis through the endothelium. This allows them to bind to the endothelium, initiate reparative processes, and reduce secondary inflammation and edema. The use of this nanocomplex improves locomotor activity and cognitive functions. The disadvantage of the claimed invention is the complexity of mass production of the nanocomplex and its inaccessibility for widespread use in medical institutions.

[0018] Currently, the use of molecular hydrogen as a therapeutic agent in cardiovascular diseases is known (George AK, Behera J., Homme RP, Tyagi N., Tyagi SC, Singh M. Rebuilding Microbiome for Mitigating Traumatic Brain Injury: Importance of Restructuring the Gut-Microbiome-Brain Axis / / Molecular neurobiology, 2021. 58 (8). P. 3614-3627), neurodegenerative diseases (Bai X., Liu S., Yuan L., Xie Y., Li T., Wang L., Wang X., Zhang T., Qin S., Song G., et al: Hydrogen-rich saline mediates neuroprotection through the regulation of endoplasmic reticulum stress and autophagy under hypoxia-ischemia neonatal brain injury in mice / / Brain research, 2016. 1646. P. 410-417.), acute and chronic lung lesions (Yang L., Liu S., Liu J. Zhang Z., Wan X., Huang B., Chen Y., & Zhang Y. COVID-19: immunopathogenesis and Immunotherapeutics / / Signal transduction and targeted therapy, 2020. 5 (1). P. 128), neuromuscular disorders (Hasegawa S., Ito M., Fukami M., Hashimoto M., Hirayama M., Ohno K. Molecular hydrogen alleviates motor deficits and muscle degeneration in mdx mice / / Redox report: communications in free radical research, 2017. 22(1). P. 26-34), metabolic syndrome (LeBaron TW, Singh RB, Fatima G, et al. The Effects of 24-Week, High-Concentration Hydrogen-Rich Water on Body Composition, Blood Lipid Profiles and Inflammation Biomarkers in Men and Women with Metabolic Syndrome: A Randomized Controlled Trial / / Diabetes, metabolic syndrome and obesity: targets and therapy, 2020. No. 13. P. 889-896). The high therapeutic effect is based on the antioxidant, anti-apoptotic, cytoprotective and anti-inflammatory effects of hydrogen (Chuchalin A.G. Molecular hydrogen as a biological marker and drug / / Pulmonology, 2024. 34 (5). P. 634-642).

[0019] Molecular hydrogen is a natural antioxidant with a high selective ability to neutralize cytotoxic hydroxyl radicals (OH) and peroxynitrites (ONOO - ), which are strong oxidants (Fang W., Tang L., Wang G., Lin J., Liao W., Pan W., Xu, J. Molecular Hydrogen Protects Human Melanocytes from Oxidative Stress by Activating Nrf2 Signaling / / The Journal of investigative dermatology, 2020. 140 (11). P. 2230-2241). Molecular hydrogen has synergistic antioxidant activity associated with the activation of the body's own endogenous antioxidants in various pathological disorders (Deryugina A.V., Danilova D.A., Brichkin Yu.D., Nazarov E.I., Medvedev A.P., Pichugin V.V., Makarov E.V., Taranov E.V., Fedorov S.A., Ivaschenko M.N. Possibilities of correcting the oxygen transport function of erythrocytes in cardiac surgery patients during operations with artificial circulation / / Bioradicals and antioxidants. 2020. Vol. 7, No. 3. Pp. 142-149).

[0020] Molecular hydrogen, having a low molecular weight, can penetrate the blood-brain barrier. At the same time, it has been shown that it does not have a toxic effect and does not accumulate in the body (Ohta S. Molecular hydrogen as a novel antioxidant: overview of the advantages of hydrogen for medical applications / / Methods in enzymology, 2015. No. 555. P. 289-317; Russell G., Nenov A., Kisher H., Hancock JT Molecular Hydrogen as Medicine: An Assessment of Administration Methods / / Hydrogen, 2021. 2(4). P. 444-460).

[0021] A known method for preventing organ damage during artificial circulation during cardiac surgery (RU 2822242 C2, published July 3, 2024) involves supplying a gas-air mixture to the extracorporeal circulation circuit, with hydrogen inhalation at a dose of 1.2 ppm administered before and after artificial circulation. During artificial circulation, after reaching the calculated volumetric perfusion rate, nitric oxide at a dose of 40 ppm and hydrogen at a dose of 1.2 ppm are additionally supplied to the gas-air mixture supply line, and this gas supply protocol is maintained throughout the entire period of artificial circulation. The supply of nitric oxide and hydrogen is stopped 5 minutes before the end of artificial circulation.The method allows for an increase in antioxidant activity, manifested in the inhibition of oxidative reactions and an increase in the activity of antioxidant enzymes, thereby reducing the manifestations of organ damage during artificial circulation.

[0022] This method of using molecular hydrogen is carried out in combination with the use of nitric oxide, which does not allow us to evaluate the mono-effect of molecular hydrogen on the state of the body.

[0023] The patent "Method for Protecting Red Blood Cells in Cardiac Surgery Patients Under Cardiac Bypass" (RU 2729026 C1 IPC A61H31 / 00, published August 3, 2020) demonstrates the use of molecular hydrogen in cardiovascular surgery. Molecular hydrogen is inhaled into the breathing circuit of a ventilator at a concentration of 1.5-2% immediately after tracheal intubation and throughout the surgery. This method improves the safety of red blood cells in patients. Improvements in clinical and functional parameters have been demonstrated in these patients.

[0024] Both methods of using molecular hydrogen are associated with cardiac surgery patients undergoing artificial circulation. A disadvantage of these methods is their inability to correct the body's condition in the post-traumatic period of TBI.

[0025] The objective of the invention is to develop a new method for correcting the body's condition in case of traumatic brain injury using molecular hydrogen.

[0026] The technical result of the proposed invention is to increase the effectiveness of correcting the body's condition by limiting damage to the microcirculatory bed, reducing endogenous intoxication, restoring the endothelial lining of blood vessels in the post-traumatic period of traumatic brain injury, expanding the arsenal of means that limit damage to the body in the post-traumatic period of TBI, which contributes to improving the condition of the body and minimizing side effects after injury.

[0027] This is achieved by the fact that the method for correcting the body's condition in traumatic brain injury includes the use of molecular hydrogen in the form of bubbled water at a dose of 0.3-0.5 ml / kg per day orally; the bubbled water is administered using a probe for 10 days; the first use of bubbled water is carried out 30 minutes after the traumatic brain injury; saturation of the water with molecular hydrogen is carried out using the Sputnik-3 hydrogen generator.

[0028] The proposed method for correcting the body's condition in case of traumatic brain injury is carried out as follows.

[0029] Molecular hydrogen in the form of bubbled water, which is administered orally (drinking), is used as a means of correcting the body's condition in case of TBI.

[0030] The proposed method was tested in an experiment in modeling TBI in Wistar rats weighing 180-200 g. The injury was inflicted without anesthesia on an intact skull, without preliminary manipulations. The animals were fixed on a tablet, from a cylindrical tube (∅20 mm), rigidly fixed on a tripod with two holders and centered over the parietal-occipital region of the skull, a load weighing 100 g fell from a height of 80 cm. This model allows you to reproduce a TBI model as close as possible to a similar one in humans (Rybakina E.G., Shanin S.N., Fomicheva E.E., Filatenkova T.A., Dmitrienko E.V. Cellular and molecular mechanisms of changes in the protective functions of the body in traumatic brain injury and an attempt at treatment / / Medical Academic Journal, 2014. Vol. 14, No. 4. Pp. 55-62). The application of such mechanical energy resulted in focal injuries and brain contusions, accompanied by the formation of epidural and subdural hematomas. The mortality rate from the falling load was 0-10%.Immediately after the injury, the rats exhibited asphyxia, convulsions, bleeding, and other symptoms. After 30-40 minutes, they returned to normal functioning and feeding. Animals that died during the experiments were excluded from the study.

[0031] The rats were divided into two equal experimental groups. The first group (n=24, comparison group) included animals that received molecular hydrogen-sparged water daily at a dose of 0.3-0.5 ml / kg per day for 10 days. The control group (n=24) included animals that received water in an equivalent volume for 10 days after the modeling. A Sputnik-3 hydrogen generator (China) was used. The physiological norm was determined for the group of intact animals. Blood was collected from the sublingual vein in an amount of 2.0 ml on days 1, 3, 7, and 14 after TBI.

[0032] Molecular hydrogen in the form of bubbled water was administered orally to rats using a probe (Ishibashi, 2013).

[0033] The first administration was carried out 30 minutes after the injury simulation.

[0034] Water was saturated with molecular hydrogen using the Sputnik-3 hydrogen generator (China).

[0035] The claimed invention limited free radical processes by limiting the formation of the most reactive radicals—hydroxyl radicals. This, in turn, helped limit damage to the microcirculatory bed, reduce endogenous intoxication while restoring tissue oxidative stress, maintain the integrity of the vascular endothelial lining, and normalize microcirculation in the post-traumatic period of TBI. This also allowed for the restoration of motor function impaired by brain injury and prevented the development of secondary damage in the body. The identified processes indicate a limitation of post-traumatic reactions that develop with TBI. The reduction of these processes leads to an improvement in the body's condition and minimizes side effects after injury.

[0036] In addition, the use of the proposed invention helps to expand the arsenal of means for limiting damage to the body in the post-traumatic period of TBI, which helps to improve the condition of the body and minimizes side effects after injury.

[0037] Example 1

[0038] During the study, endogenous intoxication of the rat body in the post-traumatic period of TBI was assessed based on the content of low- and medium-molecular-weight substances (LLMSM) in erythrocytes and blood plasma. LLMSM was considered as an indicator of the intensity of endogenous intoxication, since LLMSM are mostly products of free-radical oxidation and proteolysis, which are determined by the intensity of tissue hypoxia (Edelev I.S. Improvement of forensic diagnostics of the features of the premortem period: dis. ... Cand. of Medicine: 14.03.05 / Edelev Ivan Sergeevich. - N.N., 2019. - 130 p.). It has been shown that the accumulation of medium-weight molecules is not only a marker of endotoxicosis of the body, but also a factor aggravating the course of the pathological process (Kuzminova E.V., Sampiev A.M., Semenenko M.P., Abramov A.A., Kurtsevich L.V., Semenenko K.A.Medium-weight molecules as a diagnostic criterion for the development of endotoxicosis in animals / / Gestionarea fondului genetic animalier - probleme, soluţii, perspective, 20-23 septembrie 2023, Maximovca. Maximovca: "Print-Caro" SRL, 2023. P. 454-462).

[0039] The level of VNiSMM content was recorded spectrophotometrically at 238, 254, 266, and 282 nm wavelengths. The final result was calculated using the formula and expressed in arbitrary units.

[0040] VNSMM=1.013x (8xE238 + 16xE254 + 44xE266 / 3+ 64xE282 / 3), where E 238 , E 254 , E 266 , E 282 - the absorption values ​​of the sample at the corresponding wavelengths.

[0041] A study of the level of endogenous intoxication in rats during the post-traumatic period revealed high levels of VNiSMM in the erythrocytes and blood plasma of rats throughout the entire study period compared to animals in the intact group. Oral administration of bubbled water with molecular hydrogen contributed to a decrease in the VNiSMM in erythrocytes by day 3, and in blood plasma by day 7 of the study to the values ​​of the intact group, while in the control the studied indicators were significantly higher relative to the normal values ​​(Table 1).

[0042] Table 1

[0043] The content of VNiSMM in the blood of rats in the post-traumatic period of TBI in the control group and in the comparison group

[0044] Group Time after TBI, days 1 3 7 14 Substances of low and medium molecular weight in erythrocytes, conventional units Intact 4,6±0,7 TBI 9,8±0,4* 7,1±0,6* 5,1±0,9 3,9±0,3 TBI + H2 6,6±0,1*◊ 5,3±0,6◊ 4,5±0,8 3,5±0,8* Substances of low and medium molecular weight in blood plasma, conventional units Intact 6.3±0.3 TBI 13.5±0.9* 11.7±0.8* 10.7±0.7* 9.8±0.8* TBI + H2 8.2±0.7*◊ 7.6±0.9 5.1±0.6*◊ 4.8±0.6*◊ Note: “*” - statistically significant differences compared to the intact group, p≤0.05; “◊” - statistically significant differences compared to the control group, p≤0.05. Abbreviations: H2 - oral administration of water barbated with molecular hydrogen

[0045] Thus, in the post-traumatic period of TBI, an increase in endogenous intoxication and intensive formation of VNiSMM in erythrocytes and blood plasma were recorded, which indicated decompensation of protective and regulatory systems with failure of the detoxification function of the liver in animals of the control group.

[0046] Oral administration of water bubbled with molecular hydrogen 30 minutes after the injury modeling weakened the intoxication phenomena, as evidenced by the dynamics of the restoration of VNiSMM in the blood of rats to the values ​​of the intact group from the 3rd day of the study.

[0047] Example 2

[0048] To analyze the effectiveness of using molecular hydrogen as a post-traumatic stress corrector in rats with TBI, we studied tissue oxidative metabolism (TOM) and morphological identification of circulating endothelial cells (CEC). Oxidative metabolism (TOM) was considered an indicator reflecting the degree of oxidative damage in the body, with an increase in OM indicating increased oxidative stress. The level of CEC was considered as a diagnostic biomarker of endothelial dysfunction, since the degree of endothelial dysfunction is directly dependent on the severity of neurological deficit (Topuzova M.P., Alekseeva T.M., Vavilova T.V., Sirotkina O.V., and Klocheva E.G. Circulating endothelial cells and their precursors as a marker of endothelial dysfunction in patients with arterial hypertension who have suffered ischemic stroke (review) / / Arterial hypertension, 2018. 24 (1), pp. 57-64).It has been shown that a high level of CEC in the blood of patients with pathological conditions is considered an unfavorable factor (Farinacci M., Krahn T., Dinh W., Volk HD, Düngen HD, Wagner J., Konen T., von Ahsen O. Circulating endothelial cells as a biomarker for cardiovascular diseases / / Research and practice in thrombosis and haemostasis, 2018.3(1). P. 49-58).

[0049] Oxidative metabolism was measured using laser fluorescence spectroscopy. The number of CECs was determined using a light microscope after pre-staining the cells with methylene blue.

[0050] The results of the study showed that the effect of mechanical TBI was manifested by a significant increase in the POM in animals of the control group. Oral administration of bubbled water containing molecular hydrogen inhibited the increase in POM, maintaining the indicator at physiologically normal levels throughout the study period (Table 2).

[0051] A study of endothelial dysfunction revealed a significant increase in CECs in rat plasma during the post-traumatic period of TBI (Table 2). Oral administration of water bubbled with molecular hydrogen resulted in a 68% decrease in CECs on day 3 and a 60% decrease on day 7, relative to the control group. By day 14, CEC concentrations in the molecular hydrogen-bubbled water group had reached normal values. In the control group, however, elevated CECs persisted throughout the study period.

[0052] Table 2

[0053] Oxidative metabolism and the number of circulating endothelial cells in rat blood plasma during the post-traumatic period of TBI

[0054] in the control group and in the comparison group

[0055] Group Time after TBI, days 1 3 7 14 Oxidative metabolism index (OMI), relative units Intact 1,02±0,15 TBI 1,94±0,17* 1,39±0,09* 1,21±0,08 1,25±0,18 TBI + H2 1,06±0,24◊ 1,02±0,23 0,82±0,17 0,94±0,11 Circulating endothelial cells (CEC), cells / 100 µl Intact 16.7±7.1 TBI 65.3±7.5* 31.9±7.9* 30.5±5.9* 27.8±4.9* TBI + H2 52.7±9.3*◊ 25.8±4.6*◊ 20.4±4.3* 11.1±3.8*◊ Note: “*” - statistically significant differences compared to the intact group, p≤0.05; “◊” - statistically significant differences compared to the control group, p≤0.05. Abbreviations: H2 - oral administration of water bubbled with molecular hydrogen

[0056] Thus, the use of molecular hydrogen in the form of bubbled water helps limit the development of oxidative processes and endothelial damage in the post-traumatic period of TBI. Restoring the redox balance in tissues and the endothelial lining reduces the likelihood of triggering a systemic inflammatory response, progressive cerebral edema, and irreversible functional damage to the brain.

[0057] Example 3

[0058] To confirm the effectiveness of using molecular hydrogen in the form of bubbled water in the post-traumatic period of TBI in rats, the preservation of microcirculation and regulation of peripheral blood flow were assessed. Microcirculation was assessed using a LAZMA-ST laser blood microcirculation analyzer (manufactured by NPP Lazma, Russia) and LDF 2.20.0.507WL software (manufactured by NPP Lazma, Russia). During the study, the level of basal perfusion and the contribution of active and passive components to the regulation of tissue blood flow were recorded. The perfusion index (M, perf. units) was considered as the value of the average blood flow over the recording time interval in the studied tissue volume of approximately 1 mm, which reflects the dynamic characteristics of microcirculation. Amplitude-frequency analysis was used to assess the contribution of active factors controlling microcirculation - myogenic (Am, perf. units), neurogenic (An, perf. units) and endothelial vasomotions (Ae, perf. units).); and passive factors - respiratory (Ad, perf. units) and cardiac activity (Ac, perf. units).

[0059] The obtained results showed a decrease in tissue perfusion and microcirculatory blood flow oscillation amplitudes across all frequency spectra on days 1–7 of the study in control animals during the post-traumatic period of TBI (Table 3). Oral administration of molecular hydrogen in the form of bubbled water contributed to the early (from day 3 of the study) restoration of microcirculatory transport systems to intact group values, as evidenced by a significant increase in the activity of neurogenic, myogenic, and endothelial oscillations (an average of 2-fold increase) on day 7 post-injury and maintenance of perfusion levels at normal levels.

[0060] Table 3

[0061] Changes in perfusion parameters and amplitude-frequency oscillations in the microcirculatory bed of rats in the post-traumatic period in the control group and in the comparison group

[0062] Indicators Norm Values ​​of the indicators 1 day Day 3 Day 7 TBI TBI + H 2 TBI TBI + H 2 TBI TBI + H 2 Perfusion index, perf. units 6,44 ±0,29 12,51 ±1,36* 5,62 ±0,42◊ 10,83 ±1,03* 7,89 ±0,92*◊ 5,84 ±0,51 6,46 ±1,15 Amplitude of respiratory oscillations, perf. units 1,36 ±0,16 1,20 ±0,21 1,22 ±0,22 1,53 ±0,11 1,41 ±0,28 1,66 ±0,05* 1,39 ±0,11 Amplitude of cardiac oscillations, perf. units 1,31 ±0,13 0,92 ±0,07* 0,98 ±0,10* 1,26 ±0,10 1,48 ±0,18◊ 1,15 ±0,13 1,33 ±0,06 Amplitude of neurogenic oscillations, perf. units 1,58 ±0,22 0,89 ±0,26* 1,26 ±0,26 0,94 ±0,29* 1,74 ±0,24◊ 0,72 ±0,15* 1,86 ±0,21◊ Amplitude of myogenic oscillations, perf. units 1,28 ±0,21 0,81 ±0,24 1,39 ±0,22 0,75 ±0,16* 1,64 ±0,53◊ 0,89 ±0,14* 1,89 ±0,27*◊ Amplitude of endothelial oscillations, perf. units 0,98 ±0,12 0,49 ±0,17* 0,97 ±0,12 0,64 ±0,23 1,91 ±0,37*◊ 0,88 ±0,31 1,86 ±0,21*◊ Note: “*” - statistically significant differences compared to the intact group, p≤0.05; “◊” - statistically significant differences compared to the control group, p≤0.05. Abbreviations: H2 - oral administration of water bubbled with molecular hydrogen

[0063] Thus, a decrease in perfusion, “active” vasomotor mechanisms of microcirculation regulation, an increase in the amplitude of the respiratory wave indicate a deterioration in venous outflow, the formation of congestive phenomena in the venular link of the microcirculatory bed and gives reason to assume that the rats of the control group experienced the development of a congestive-ischemic form of blood circulation in the post-traumatic period of TBI.

[0064] The use of water bubbled with molecular hydrogen resulted in a significant increase in the amplitudes of tone-forming "active" factors controlling microcirculation, particularly endothelial amplitudes, indicating an increase in endothelium-dependent vasodilation and the absence of signs of arteriolar constriction. This focus on blood flow regulation mechanisms was accompanied by an increase in blood flow into the microcirculation. This suggests that the use of water bubbled with molecular hydrogen helps limit the development of ischemia and normalize blood flow in the early period of TBI, thereby reducing the risk of post-traumatic hypoxic conditions.

[0065] Example 4

[0066] The study analyzed the animals' motor activity. In the experiment, two days before the modeled TBI, rats were trained to move along a narrow wooden block 2 cm wide and 50 cm long. Movement was stimulated by a bright light at one end of the block and then moving toward the other, where a dark chamber was located, in which the animal was protected from the light. During the post-traumatic period, the ability to balance and remain on the block was assessed, i.e., the time spent moving along the block from the bright light source to the dark chamber, and the frequency of paw slippage (as points). Overall motor activity was assessed on a 10-point scale (Table 4). The maximum possible score was 10 (inability to complete the task), the minimum possible was 2 (successful completion of the task). Measurements were taken three times, and the average result was taken.

[0067] Table 4

[0068] Motor Activity Assessment System

[0069] Rating, points Time, s Frequency of paw slips 5 17-20 >4 4 13-16 3 3 9-12 2 2 5-8 1 1 4-1 0

[0070] The results of the study showed that rats with TBI exhibited a decrease in motor activity, manifested by an increase in the time spent walking along the block and the frequency of paw slippage relative to the intact group. Oral administration of molecular hydrogen, beginning three days after injury, promoted restoration of walking speed along the block and overall motor activity, and, beginning seven days after injury, a decrease in the frequency of paw slippage in the post-traumatic period. In animals of the control group, these parameters remained higher than those of the intact group until the end of the study period (Table 5).

[0071] Table 5

[0072] Motor activity of rats in the post-traumatic period of TBI and its correction with molecular hydrogen

[0073] Group Time after TBI, days 1 3 7 14 Time, s Intact 5.17±0.71 TBI 10.47±0.55* 8.81±0.84* 6.97±1.17 6.11±0.89* TBI + H2 8.82±0.84*▲ 5.19±0.72▲ 5.61±0.54 4.26±0.83▲ Frequency of paw slips Intact 0.67±0.52 TBI 3.5±0.48* 2±0.34* 1.7±0.48* 1.6±0.48* TBI + H2 3.2±0.51* 1.7±0.37*▲ 1±0.19▲ 0.8±0.15 Overall assessment of motor activity, points Intact 0.7±0.5 TBI 2.2±0.3* 1.6±0.2* 1.6±0.2* 1.1±0.1 TBI + H2 1.8±0.3* 1.2±0.3 1±0.2 0.4±0.1 Note: “*” - statistically significant differences relative to the values ​​of the intact group, p ≤ 0.05; “▲” - statistically significant differences between the experiment and the control, p ≤ 0.05.

[0074] Thus, we have established that the use of water bubbled with molecular hydrogen significantly improves the motor activity of rats after TBI, which indicates a reduction in defects in motor function and coordination of movements in the early stages of the post-traumatic period of rats.

Claims

1. A method for reducing the risk of developing post-traumatic hypoxic conditions after a traumatic brain injury in an experiment, characterized in that molecular hydrogen is used in the form of bubbled water at a dose of 0.3-0.5 ml / kg per day orally, with the first administration of bubbled water being carried out 30 minutes after the traumatic brain injury, and the administration of bubbled water is carried out using a probe for 10 days.

2. The method according to paragraph 1, characterized in that the saturation of water with molecular hydrogen is carried out using the Sputnik-3 hydrogen generator unit.