The Role of JNK and p53 in the Regulation of Secretory Function of Neuroglial Cells of Various Types in ß-Amyloid-Induced Neurodegeneration.

We studied the effect of JNK and p53 inhibitors on the production of neurotrophic factors stimulating the realization of the growth potential of neural stem cells by neuroglial cells of various types under conditions of simulation of induced ß-amyloid neurodegeneration in vitro. It was shown that ß-amyloid stimulates the production of neurotrophins by astrocytes and microglial cells, but does not affect the functioning of oligodendrocytes. JNK and p53 were not involved in the secretion of neurotrophins by intact astrocytes. The stimulating role of p53 on the implementation of their secretory activity under the influence of a neurotoxic agent was revealed. At the same time, the inhibitory role of JNK and p53 in the production of neurotrophic growth factors by oligodendrocytes and microglial cells was revealed both under conditions of their optimal vital activity and when ß-amyloid was added to the cell culture.

The Role of cAMP-Dependent Intracellular Signaling Pathways in the Regulation of the Functions of Neural Stem Cells and Neuroglial Cells in Amyloid-ß-Induced Neurodegeneration.

Under conditions of neurodegeneration modeled in vitro by the ß-amyloid peptide-(25-35) fragment (Aß25-35), we studied the role of individual links of cAMP-dependent intracellular signaling pathways in determining the proliferation and differentiation status of neural stem cells (NSCs) and colony-stimulating activity of supernatants from neuroglial cells. The important role of intracellular cAMP and PKA in the inhibition of the progression of the NSC cell cycle and stimulation of the process of their specialization induced by Aß25-35 was found. The selective ability of PKA to block the production of factors constituting colony-stimulating activity by neuroglial cells under conditions of their cultivation in vitro with a neurotoxic agent was revealed. Our results suggests that inhibitors of adenylate cyclase and PKA can increase the degree of implementation of the growth potential of NSCs and conjugation of the processes of their proliferation and differentiation in Alzheimer's disease. At the same time, selective PKA blockers can also induce the production of NSC-stimulating factors by neuroglial cells.

Participation of cAMP-Mediated Signaling Transduction in the Regulation of the Secretory Function of Neuroglia during Ethanol-Induced Neurodegeneration.

We studied the involvement of cAMP and PKA in the regulation of the secretion of neurotrophic growth factors by macro-and microglial cells in the model of ethanol-induced neurodegeneration in vitro and in vivo. The stimulating role of cAMP in the secretion of neurotrophins by intact astrocytes and oligodendrocytes was shown, while PKA does not participate in this process. On the contrary, the inhibitory role of cAMP (implemented via PKA activation) in the production of neurogenesis stimulators by microglial cells under conditions of optimal vital activity was found. Under the influence of ethanol, the role of cAMP and PKA in the production of growth factors by macroglial cells was considerably changed. The involvement of PKA in the cAMP-dependent signaling pathways and inversion of the role of this signaling pathway in the implementation of the neurotrophic secretory function of astrocytes and oligodendrocytes, respectively, directly exposed to ethanol in vitro were noted. Long-term exposure of the nervous tissue to ethanol in vivo led to the loss of the stimulating role of cAMP/PKA signaling on neurotrophin secretion by macroglial cells without affecting its inhibitory role in the regulation of this function in microglial cells.

Functional State of Various Types of Regenerative-Competent Neural Tissue Cells in ß-Amyloid-Induced Neurodegeneration.

We analyzed the content and functional activity of various types of regenerative-competent cells of the subventricular zone of the cerebral hemispheres in experimental mice under conditions of in vitro modeling of ß-amyloid-induced neurodegeneration. Fundamentally different (opposite) effects of ß-amyloid on functional activity of multipotent neural stem cells (NSC) and neuronal committed progenitors (NCP) were revealed. ß-Amyloid suppressed proliferation of NSC and stimulated their differentiation. At the same time, an increase in the mitotic activity of NCP was observed with a decrease in the intensity of their specialization. These changes in the functioning of progenitor cells developed against the background of a significant drop in the production of neurotrophic growth factors by neuroglia. These phenomena indicate marked discoordination of the activity of regenerative-competent cells of various types.

Psychopharmacological Effects of Stimulation of the Functions of Neural Stem Cells by STAT3 Inhibitor under Conditions of Modeled Ethanol-Induced Encephalopathy.

The psychopharmacological effects of a stimulator of functions of progenitor cells of the nervous tissue STAT3 inhibitor (STAT3 Inhibitor XIV, LLL12) were studied under conditions of modeled alcoholic encephalopathy in C57BL/6 mice. The pharmacological agent corrected the parameters of exploratory behavior (characterizing predominantly cognitive activity) in the experimental animals at the late terms of observation. At the same time, the reproducibility of the conditioned passive avoidance response developed at the beginning of the course STAT3 inhibitor administration decreased. These effects developed against the background of a significant increase in the content of neural stem cells and their proliferative activity in the paraventricular zone of the brain.

The Role of JAKs and STAT3 in Regulation of Regenerative-Competent Cells of the Nervous Tissue in ß-Amyloid-Induced Neurodegeneration.

We studied the role of JAKs and STAT3 in the growth potential of neural stem cells and the humoral neurotrophic function of neuroglia in modeling ß-amyloid-induced neurodegeneration in vitro. It was found that these signaling molecules do not participate in the neural stem cell functioning, and JAKs plays an inhibitory role (realized, however, without STAT3) in the secretion of neurotrophins by glial cells under conditions of their optimal vital activity. The effect of ß-amyloid on progenitor cells is accompanied by the appearance of a "negative" effect of STAT3 signaling pathway on their proliferative activity. At the same time, JAKs and STAT3 during neurodegeneration stimulate specialization/differentiation of neural stem cells and production of growth factors by neuroglial cells. These results indicate the possibility of stimulating proliferation of neural stem cells coupled with their differentiation by using selective STAT3 inhibitors.

The Role of MARK ERK1/2 and p38 in Regulation of Functions of Neural Stem Cells and Neuroglia under Conditions of ß-Amyloid-Induced Neurodegeneration.

The role of ERK1/2 and p38 in the realization of the growth potential of neural stem cells and secretion of neurotrophic growth factors by glial cells was studied using in vitro model of ß-amyloid-induced neurodegeneration. It was shown that amyloid-ß fragment 25-35 significantly inhibits the cell cycle progression of neural stem cells against the background of stimulation of their differentiation and reduced production of growth factors by neuroglia. The inhibitory role of ERK1/2 and p38 in relation to the proliferative activity of neural stem cells and the secretory activity of glial elements was revealed. ERK1/2 and p38 inhibitors increased proliferation of progenitor cells of the nervous tissue and reduced the intensity of their specialization, as well as stimulated production of growth factors by neuroglial cells under conditions of simulated ß-amyloid-induced neurodegeneration.

The Role of JAK and STAT3 in Regulation of Secretory Function of Neuroglial Cells of Different Types in Ethanol-Induced Neurodegenerationt.

The effects of JAK and STAT3 inhibitors on the production of neurotrophic growth factors by different types of neuroglial cells were studied under conditions of in vitro and in vivo models of ethanol-induced neurodegeneration. It was shown that these signaling molecules do not participate in the secretion of neurotrophins by intact astrocytes and oligodendrocytes. The inhibitory role of JAK in the regulation of this function of microglial cells was revealed. We also revealed significant changes in the role of JAK and the presence of STAT3 specifics within the framework of JAK/STAT signaling in the production of growth factors by various glial elements under the influence of ethanol. Neurodegeneration modeled in vitro led to the appearance of a "negative" effect of STAT3 on the production of neurogenesis stimulants by all types of glial cells. Moreover, the role of STAT3 in oligodendrocytes and microglial cells generally corresponded to that of JAK/STAT signaling. In astrocytes, only selective blockade of STAT3 (but not JAK) led to stimulation of their function. In mice subjected to prolonged peroral alcoholization, the neuroglial responses to the pharmacological regulation of JAK/STAT signaling were different. An inversion of the role of JAK and STAT3 in the production of neurotrophins by oligodendrocytes was noted. In addition, JAK inhibitor did not stimulate secretory function of microglial cells under conditions of prolonged exposure to ethanol in vivo.

Role of MAPK ERK1/2 and p38 in the Regulation of Secretory Functions of Different Populations of Neuroglia in Ethanol-Induced Neurodegeneration.

We studied the participation of ERK1/2 and p38 in secretion of neurotrophic growth factors by various types of neuroglia under conditions of in vitro and in vivo modeled ethanol-induced neurodegeneration. The inhibitory role of these protein kinases in the production of neurotrophins by intact astrocytes and the absence of their participation in the regulation of functions of oligodendrocytes and microglial cells were shown. Under conditions of ethanol neurotoxicity, the role of ERK1/2 and p38 in the production of growth factors by glial elements was significantly changed. Neurodegeneration modeled in vitro led to inversion of the role of both protein kinases in the secretion of neurotrophins by astroglia and inhibition of the cytokine-synthesizing function of oligodendrocytes and microglial cells by ERK1/2 and p38. In mice receiving ethanol per os for a long time (as well as in cells in vitro exposed to ethanol), mitogen-activated kinases stimulated the function of astrocytes and inhibited the production of growth factors by microglial cells. At the same time, chronic alcoholization was accompanied by the appearance of the stimulating role of ERK1/2 and p38 in the implementation of the secretory function by oligodendrocytes.

The Role of Intracellular Signaling Molecules in the Production of Granulocytic CSF Mononuclear Phagocytes in Stress and Cytostatic Influence.

Under conditions of steady-state hemopoiesis, nuclear factor NF-κB, in contrast to MAP kinase p38, plays an important role in the maintenance of the initial level of secretory activity of monocytes. The increase in the production of G-CSF under stress conditions (10-h immobilization) is mainly regulated by the alternative p38MARK signaling pathway via activation of p38 synthesis. It was shown that under conditions of cytostatic-induced myelosuppression, the production of protein kinase p38 in cells decreases, and it, like NF-κB, is not the main one in the production of hemopoietin by mononuclear phagocytes.

Role of JNK and p53 in Implementation of Functions of Various Types of Regeneration-Competent Cells of the Nervous Tissue.

We studied the participation of JNK and p53 in the realization of the growth potential of different types of progenitors of the subventricular zone of mouse brain and secretion of neurotrophins by glial cells. The stimulating role of these signaling molecules in mitotic activity and specialization of multipotent neural stem cells was shown. It was found that JNK and p53 do not participate in the regulation of committed neuronal progenitor cells (clonogenic PSA-NCAM+ cells). A dependence of neurotrophic growth factors in individual populations of neuroglia on activity of these protein kinase and transcription factor was revealed. The role of JNK and p53 in astrocytes consists in stimulation of their secretion, and in microglial cells, on the contrary, in its inhibition. The secretory neurotrophic function of oligodendrogliocytes is not associated with JNK and p53 activity.

Specific Features of Intracellular Signal Transduction in the Regulation of Functions of Neural Stem Cells and Committed Neuronal Progenitors.

We studied the role of NF-κB-, cAMP/PKA-, JAKs/STAT3-, ERK1/2-, p38-, JNK- and p53- mediated signaling pathways in the realization of the growth potential of neural stem cells and committed neuronal progenitors under in vitro conditions. The method of pharmacological blockade with selective inhibitors of individual signaling molecules revealed some principal differences in their role in the determination of the proliferation and differentiation status of progenitor cells of different classes. Analysis of the peculiarities of intracellular signaling in cells and comparison of the role of its individual elements attest to the prospects of developing new drugs with neuroregenerative activity based on STAT3 inhibitors or JNK activators. These modulations of activity of signaling molecules can stimulate the realization of the growth potential of committed neuronal progenitors and neutral stem cells, respectively. The blockade of STAT3 and an increase in the content of phosphorylated forms of JNK had no "negative" effects on the functioning of multipotent neural stem cells and committed neuronal progenitors, respectively.

The Role of NF-κВ in the Realization of Functions of Various Types of Regeneration-Competent Cells of the Nervous Tissue in Ethanol-Induced Neurodegeneration.

The role of NF-κВ in the realization of the growth potential of neural progenitor cells from the subventricular area of cerebral hemispheres and secretion of neurotrophins by glial elements was studied under conditions of in vitro and in vivo modeled ethanol-induced neurodegeneration. It was found that this transcription factor does not participate in the regulation of mitotic activity of neural stem cells and neuronal-committed progenitors under optimal conditions and under the influence of ethanol in vitro. At the same time, NF-κВ suppresses differentiation/maturation of neural progenitor cells. Long-term peroral administration of ethanol to mice was accompanied by the inhibitory influence of NF-κВ on proliferation of progenitor cells. Blockade of NF-κВ in neural stem cells and committed neuronal precursors in animals with neurodegeneration induced cell cycle progression in these elements. The involvement of NF-κВ in the secretory function of astrocytes and oligodendrogliocytes was established. Inactivation of the nuclear transcription factor reduced the production of neurotrophins, in particular, in the case of ethanol exposure. At the same time, no changes in the function of microglia were noted.

Peculiarities of the Involvement of MAPKS ERK1/2 and р38 in the Implementation of the Functions of Neural Stem Cells and Neuronal Committed Precursors in Ethanol-Induced Neurodegeneration.

We studied the peculiarities of the participation of ERK1/2 and р38 in regulation of various types of progenitor cells of the nervous tissue under conditions of ethanol-induced neurodegeneration modeled in vitro and in vivo. The stimulating role of these signaling molecules in the realization of the growth potential of intact multipotent neural stem cells and committed neuronal precursors (clonogenic PSA-NCAM+ cells) was demonstrated. In vitro exposure to neurotoxic doses of ethanol led to the loss of the specified role of ERK1/2 and p38 in the cell cycle regulation. Inversion of the role of both studied MAP-kinases in determining the proliferation status of neural stem cells after long-term administration of ethanol to experimental animals was revealed. In committed neuronal precursors, this inversion (inhibition of mitotic activity instead of activation) was revealed only for ERK1/2. In mice exposed to chronic alcoholization, ERK1/2 no longer participated in the process of specialization of both types of regeneration-competent cells of the nerve tissue. The revealed fundamental difference between the functions of ERK1/2 and p38 in the cell cycle regulation in neural stem cells and committed neuronal precursors under optimal conditions and during ethanol-induced neurodegeneration does not allow drawing definite conclusions about the prospect of using modifiers of their activity for the therapy for alcohol-related CNS pathologies.

Involvement of Signaling Cascades in Granulocytopoiesis Regulation under Conditions of Cytostatic Treatment.

Suppression of the production of granulocytic CSF under the effect of 5-fluorouracyl is related to disorders in the NF-κB-, cAMP-dependent signaling pathways and MAPK cascade. These secondary messengers are involved in the regulation of functional activity of nonadherent myelokaryocytes starting from day 10 of the experiment (initial period of the hemopoietic granulocytic stem regeneration after antimetabolite challenge). Granulocytic CSF does not play essential role in the formation of colony-stimulating activity of cells of the adherent and nonadherent fractions of the bone marrow. Only cAMP-dependent pathway is involved in the regulation of the realization of the granulocytic precursor growth potential in response to the challenge.

Hemostimulating Effects of c-Jun N-Terminal Kinase (JNK) Inhibitor during Cytostatic Myelosuppression and Mechanisms of Their Development.

The hemostimulating effects of c-Jun N-terminal kinase (JNK) inhibitor were examined on the mouse model of myelosuppression provoked by 5-fluorouracil. Blockade of JNK during postcytostatic period accelerated recovery of granulomonocytopoiesis and erythropoiesis. It also increased the content of neutrophilic granulocytes and erythroid cells in the hematopoietic tissue and elevated the counts of neutrophils and reticulocytes in the peripheral blood. The development of these phenomena resulted from elevated content and up-regulated functional activity of bone marrow hematopoietic progenitors associated with the direct action of JNK inhibitor on these progenitors and enhanced secretion of hemopoietins by stromal elements of the hematopoiesis-inducing microenvironment.

Specific Roles of JAKs and STAT3 in Functions of Neural Stem Cells and Committed Neuronal Progenitors during Ethanol-Induced Neurodegeneration.

Peculiar roles of JAKs and STAT3 in realization of growth potential of various types of progenitor cells in neural tissue were examined during ethanol-induced neurodegeneration modeled both in vitro and in vivo. During in vitro action of C2H5OH, these signal molecules exerted the opposite effects on mitotic activity of multipotent neural stem cells and committed neural progenitors (the clonogenic PSA-NCAM+ cells). The JAKs and STAT3 inhibitors down-regulated the rate of neural stem cell division (proliferative activity) but up-regulated such activity of the committed neural progenitors. A long-term in vivo exposure of mice to ethanol inversed the roles of JAKs and STAT3 in determination of proliferative status of neural stem cells and eliminated involvement of JAKs in functional control over the committed progenitors of neurons. The data attest to much promise of STAT3 inhibitors in treatment of ethanol-induced CNS diseases as the remedies that stimulate realization of growth potential in multipotent neural stem cells and committed neural progenitors.

Participation of cAMP/PKA-Mediated Signaling Pathways in Functional Activity of Regeneration-Competent Cells in the Nervous Tissue under Conditions of Ethanol-Induced Neurodegeneration.

We studied the involvement of cAMP/PKA signaling in the realization of the growth potential of neural progenitors and secretion of neurotrophic growth factors by glial elements under conditions of ethanol-induced neurodegeneration in vitro and in vivo. The stimulating role of cAMP and PKA in cell cycle progression of the neural progenitor cells and in production of neurotrophins by the cells in nervous tissue under the optimal conditions to vital activity was demonstrated. Ethanol inverted the role of cAMP/PKA signaling pathways in determination of the proliferation-differentiation status of neural stem cells. Selective blockade of adenylate cyclase or PKA in neural stem cells increased the rate of their division against the background of relative decrease in differentiation rate. In addition, cAMP/PKA signaling does not longer participate in neurotrophin production by glial cells in neurodegeneration. These findings suggest that inhibitors of activity/expression of adenylate cyclase and PKA can be considered as possible drugs with regenerative activity for the treatment of nervous system pathologies provoked by alcohol.

Role of MAPK ERK1/2 and p38 in the Realization of Growth Potential of Various Types of Regeneration-Competent Cells in Mouse Neural Tissue during Ethanol-Induced Neurodegeneration In Vitro.

Ethanol-induced neurodegeneration was modeled in vitro to study the roles of ERK1/2 and p38 in realization of the growth potential of neural progenitor cells and secretion of neurotrophic growth factors by glial elements. Addition of the neurotoxic dose of C2H5OH (65 mM) to the culture medium abolished the effects of specific ERK1/2 and p38 inhibitors on the formation of colonies (neurospheres) and proliferative activity of neural CFU in cultured cells derived from paraventricular region of the mouse brain. The study established that these protein kinases are not implicated in ethanol-induced stimulation of the formation of neural CFU, differentiation of neural progenitors, and synthesis of humoral functional regulators of neural CFU by glial cells.

Peculiarities of Intracellular Signal Transduction in the Regulation of Functions of Mesenchymal, Neural, and Hematopoietic Progenitor Cells.

The role of NF-κB, cAMP/PKA, JAKs/STAT3, ERK1/2, p38, JNK, and p53 signaling pathways in the realization of growth potential of mesenchymal, neural, erythroid, and granulomonocytic progenitor cells were examined in vitro. Using selective blockers of signaling molecules, we revealed some principal distinctions of their involvement in determination of proliferation-differentiation status of the progenitor cells of different functional classes. The most salient peculiarities were observed in the roles of cAMP/PKA, JNK, and JAKs/STAT3 signaling pathways in the control of functions of various types of the regeneration-competent elements. The specific features of intracellular signaling revealed in histogenetically and functionally different progenitor cells attest to visibility of differentiated pharmacological stimulation of regeneration in individual tissues and prospectiveness in the development of targeted remedies for regenerative medicine based on modifiers of activity of the intracellular signaling molecules.