Present Ph.D. thesis is devoted to the study of MSCs therapeutic effect in the treatment of multiorgan dysfunction syndrome and to the elucidation of possible molecular mechanisms of such action. The studies were performed on laboratory ICR mice and a reproduced model of systemic organism lesions with multiorgan dysfunction syndrome.
To study the therapeutic effect of mesenchymal stem cells (MSCs), an animal model was developed and characterized, which corresponds to the severe condition of multiorgan pathology. The important feature of the created model is the genetically determined predisposition of experimental animals to tumor formation, which reproduces the growing predisposition to human carcinogenesis. So, such model provides a manifestation of possible carcinogenic properties of the studied MSCs.
Tetrachloromethane (CCl4), whose metabolism in the cell causes the appearance of great number of reactive molecules and affects the permeability of membranes, was chosen as the factor that triggers the chain of destructive reactions in the created experimental animal model. With prolonged loading, the existing mechanisms of cell detoxification are depleted, which consistently leads to their death, the development of chronic inflammatory reactions and organ dysfunction. The severity of the model is confirmed by pathomorphological changes in vital organs and the mortality index at the stage of formation of the required level of pathology, which was 18-22% of the total amount of experimental animals.
Therapeutic effects of allogeneic and xenogeneic MSCs in different concentrations, schemes of introduction and terms of therapeutic effect development are studied. This was performed against the background of the developed model by means of pathomorphological analysis and analysis of blood parameters.
The study involved mouse embryonic fibroblast-like cells of mice ICR, BALB, FVB-Cg-Tg (GFPU) 5Nagy / J lines (FM), and mesenchymal stem cells from human Wharton jelly, which were positive for markers CD73, CD90, CD105.
It has been shown that in general the therapeutic effects of allogeneic MSCs derived from different lines of mice are similar in many aspects. Differences in therapeutic effects related to the degree of induced proliferative activation in the regenerating organs. Thus, 3 weeks after MSCs transplantation from FVB mice, the proliferative activity of the liver cells of the recipient mouse was lower compared to the use of MSCs of BALB mice. Xenogeneic MSCs caused more pronounced inflammatory response than allogeneic, although the positive physiological effects and survival index were at the level of allogeneic MSCs of higher concentration use. That is, a single transplant of xenogeneic MSCs caused the same level of response as a triple transplant of allogeneic MSCs.
The dependence of therapeutic effects on the concentrations of transplanted MSCs was established. Following doses and amount of exogenous cell transplants were studied: 1x10^4, 1x10^5 and 3x10^5 cells per mouse. Moreover, the "dose" 3x10^5 was divided into 3 transplants with 7-days break between them. The dose of 1x10^5 was calculated based on therapeutic doses of MSCs long-lasting clinical trials of MSCs in humans at that time. For this dose, partial restoration of pulmonary histoarchitectonics, partial restoration of white and red spleen pulp with signs of immune response activation, and regeneration of hepatocytes near vessels in the liver were shown. A lower dose of MSC - 1x10^4 cells per mouse also led to compensatory-adaptive responses and to the absence of alternative-necrotic changes, but the number of regenerating hepatocytes in the liver was smaller. In the more severe initial state, it was low to increase the survival index of experimental animals.
It is shown that transplanted MSCs, in the conditions of systemic lesion of the organism, under the terms of exhausted detoxification mechanisms do not additionally activate and accelerate the development of tumors in animals with a hereditary predisposition to carcinogenesis. Systemic lesions realize a genetic predisposition to carcinogenesis, but even under these conditions, MSC transplantation did not lead to an intensification of this process.
Based on the results of the analysis of the results obtained, a hypothesis of the possible mechanism of action of therapeutic MSCs was formulated. According to the hypothesis, resident ("watchdog") mesenchymal cells in chronic lesions / inflammation are in a depressed state, due to the cellular analogue of the protective physiological response – a decrease in excitability with prolonged exposure to stimuli.
Thus, it is shown that transplanted MSCs under conditions of severe systemic damage of the body do not lose their restorative regulatory properties and can be proposed for detailed development of protocols for therapeutic use in these conditions.
