Molecular Mechanisms of Proteins - Targets for SARS-CoV-2 (Review).

The rapidly accumulating information about the new coronavirus infection and the ambiguous results obtained by various authors necessitate further research aiming at prevention and treatment of this disease. At the moment, there is convincing evidence that the pathogen affects not only the respiratory but also the central nervous system (CNS). The aim of the study is to provide an insight into the molecular mechanisms underlying the damage to the CNS caused by the new coronavirus SARS-CoV-2. Results: By analyzing the literature, we provide evidence that the brain is targeted by this virus. SARS-CoV-2 enters the body with the help of the target proteins: angiotensin-converting enzyme 2 (ACE2) and associated serine protease TMPRSS2 of the nasal epithelium. Brain damage develops before the onset of pulmonary symptoms. The virus spreads through the brain tissue into the piriform cortex, basal ganglia, midbrain, and hypothalamus. Later, the substantia nigra of the midbrain, amygdala, hippocampus, and cerebellum become affected. Massive death of neurons, astrogliosis and activation of microglia develop at the next stage of the disease. By day 4, an excessive production of proinflammatory cytokines in the brain, local neuroinflammation, breakdown of the blood-brain barrier, and impaired neuroplasticity are detected. These changes imply the involvement of a vascular component driven by excessive activity of matrix metalloproteinases, mediated by CD147. The main players in the pathogenesis of COVID-19 in the brain are products of angiotensin II (AT II) metabolism, largely angiotensin 1-7 (AT 1-7) and angiotensin IV (AT IV). There are conflicting data regarding their role in damage to the CNS in various diseases, including the coronavirus infection.The second participant in the pathogenesis of brain damage in COVID-19 is CD147 - the inducer of extracellular matrix metalloproteinases. This molecule is expressed on the endothelial cells of cerebral microvessels, as well as on leukocytes present in the brain during neuroinflammation. The CD147 molecule plays a significant role in maintaining the structural and functional integrity of the blood-brain barrier by controlling the basal membrane permeability and by mediating the astrocyte-endothelial interactions. Via the above mechanisms, an exposure to SARS-CoV-2 leads to direct damage to the neurovascular unit of the brain.

[Features of the in vitro expression profile of hippocampal neurogenic niche cells during optogenetic stimulation]. / Osobennosti ékspressionnogo profilia kletok v modeli neirogennoi nishi gippokampa in vitro pri optogeneticheskoi stimuliatsii.

In the central nervous system of mammals, there are specialized areas in which neurogenesis - neurogenic niches - is observed in the postnatal period. It is believed that astrocytes in the composition of neurogenic niches play a significant role in the regulation of neurogenesis, and therefore they are considered as a promising "target" for the possible control of neurogenesis, including the use of optogenetics. In the framework of this work, we formed an in vitro model of a neurogenic niche, consisting of cerebral endothelial cells, astrocytes and neurospheres. Astrocytes in the neurogenic niche model expressed canalorodopsin ChR2 and underwent photoactivation. The effect of photoactivated astrocytes on the expression profile of neurogenic niche cells was evaluated using immunocytochemical analysis methods. It was found that intact astrocytes in the composition of the neurogenic niche contribute to neuronal differentiation of stem cells, as well as the activation of astroglia expressing photosensitive proteins, changes the expression of molecules characterized by intercellular interactions of pools of resting and proliferating cells in the composition of the neurogenic niche with the participation of NAD+ (Cx43, CD38, CD157), lactate (MCT1). In particular, the registered changes reflect a violation of the paracrine intercellular interactions of two subpopulations of cells, one of which acts as a source of NAD+, and the second as a consumer of NAD+ to ensure the processes of intracellular signal transduction; a change in the mechanisms of lactate transport due to aberrant expression of the lactate transporter MCT1 in cells forming a pool of cells developing along the neuronal path of differentiation. In general, with photostimulation of niche astrocytes, the total proliferative activity increases mainly due to neural progenitor cells, but not neural stem cells. Thus, optogenetic activation of astrocytes can become a promising tool for controlling the activity of neurogenesis processes and the formation of a local proneurogenic microenvironment in an in vitro model of a neurogenic niche.

[Astroglia-mediated regulation of cell development in the model of neurogenic niche in vitro treated with Aß1-42]. / Astrotsit-oposredovannye mekhanizmy reguliatsii neirogeneza v modeli neirogennoi nishi in vitro pri deistviin Aß1-42.

Neurogenesis is a complex process which governs embryonic brain development and is importants for brain plasticity throughout the whole life. Postnatal neurogenesis occurs in neurogenic niches that regulate the processes of proliferation and differentiation of stem and progenitor cells under the action of stimuli that trigger the mechanisms of neuroplasticity. Cells of glial and endothelial origin are the key regulators of neurogenesis. It is known that physiological neurogeneses is crucial for memory formation, whereas reparative neurogenesis provides partial repair of altered brain structure and compensation of neurological deficits caused by brain injury. Dysregulation of neurogenesis is a characteristics of various neurodevelopmental and neurodegenerative diseases, particularly, Alzheimer's disease which is very important medical and social problem. In the in vitro model of the neurogenic niche using hippocampal neurospheres as a source of stem/progenitor cells and astrocytes, we studied effects of astrocyte activation on the expression of markers of different stages of cell proliferation and differentiation. We found that aberrant mechanisms of development of stem and progenitor cells, caused by the beta-amyloid (Aß1-42), can be partially restored by targeted activation of GFAP-expressing cells in the neurogenic niche.

TIGHT JUNCTIONS PROTEINS IN CEREBRAL ENDOTHELIAL CELLS DURING EARLY POSTNATAL DEVELOPMENT.

Formation and functional plasticity of the blood-brain barrier is associated with the molecular events that occur in the brain neurovascular unit in the embryonic and early postnatal development. To study the characteristics of barriergenesis under physiological conditions, as well as recovering from perinatal hypoxia and early life stress, we examined the expression of proteins of cerebral endothelial tight junctions (JAM, ZO1, CLDN5) in rats aged 7, 28 and 70 days of postnatal development (P7­P70). Under physiological conditions, we have found that the number of endothelial cells expressing JAM, ZO1, CLDN5 slightly increases in the cortex, hippocampus and amygdala of the brain in the period from P7 to P70. Perinatal hypoxia significantly increased the number of cells expressing proteins of tight junction proteins (JAM, CLDN5) up to the age P28­P70, whereas the number of cells expressing ZO1 was reduced in the same period of time. Early life stress led to an imbalance between the number of cells expressing ZO1 proteins and that expressing tight junctions proteins, but these changes were in opposite direction to that observed in perinatal hypoxia

[THE MODEL OF NEUROVASCULAR UNIT IN VITRO CONSISTING OF THREE CELLS TYPES].

There are many ways to model blood brain barrier and neurovascular unit in vitro. All existing models have their disadvantages, advantages and some peculiarities of preparation and usage. We obtained the three-cells neurovascular unit model in vitro using progenitor cells isolated from the rat embryos brain (Wistar, 14-16 d). After withdrawal of the progenitor cells the neurospheres were cultured with subsequent differentiation into astrocytes and neurons. Endothelial cells were isolated from embryonic brain too. During the differentiation of progenitor cells the astrocytes monolayer formation occurs after 7-9 d, neurons monolayer--after 10-14 d, endothelial cells monolayer--after 7 d. Our protocol for simultaneous isolation and cultivation of neurons, astrocytes and endothelial cells reduces the time needed to obtain neurovascular unit model in vitro, consisting of three cells types and reduce the number of animals used. It is also important to note the cerebral origin of all cell types, which is also an advantage of our model in vitro.