Stimulating the process of angiogenesis in treating ischemia-related diseases is an urgent task f... more Stimulating the process of angiogenesis in treating ischemia-related diseases is an urgent task for modern medicine, which can be achieved through the use of different cell types. Umbilical cord blood (UCB) continues to be one of the attractive cell sources for transplantation. The goal of this study was to investigate the role and therapeutic potential of gene-engineered umbilical cord blood mononuclear cells (UCB-MC) as a forward-looking strategy for the activation of angiogenesis. Adenovirus constructs Ad-VEGF, Ad-FGF2, Ad-SDF1α, and Ad-EGFP were synthesized and used for cell modification. UCB-MCs were isolated from UCB and transduced with adenoviral vectors. As part of our in vitro experiments, we evaluated the efficiency of transfection, the expression of recombinant genes, and the secretome profile. Later, we applied an in vivo Matrigel plug assay to assess engineered UCB-MC’s angiogenic potential. We conclude that hUCB-MCs can be efficiently modified simultaneously with sever...
The biosafety of gene therapy remains a crucial issue for both the direct and cell-mediated deliv... more The biosafety of gene therapy remains a crucial issue for both the direct and cell-mediated delivery of recombinant cDNA encoding biologically active molecules for the pathogenetic correction of congenital or acquired disorders. The diversity of vector systems and cell carriers for the delivery of therapeutic genes revealed the difficulty of developing and implementing a safe and effective drug containing artificial genetic material for the treatment of human diseases in practical medicine. Therefore, in this study we assessed changes in the transcriptome and secretome of umbilical cord blood mononuclear cells (UCB-MCs) genetically modified using adenoviral vector (Ad5) carrying cDNA encoding human vascular endothelial growth factor (VEGF165) or reporter green fluorescent protein (GFP). A preliminary analysis of UCB-MCs transduced with Ad5-VEGF165 and Ad5-GFP with MOI of 10 showed efficient transgene expression in gene-modified UCB-MCs at mRNA and protein levels. The whole transcrip...
In order to improve the regeneration of full-layer skin defects, hydrogels were developed based o... more In order to improve the regeneration of full-layer skin defects, hydrogels were developed based on the combination of chitosan (Cs), Daba silk fibroin (DSF), and graphene oxide (GO): CS, DSF/Cs and DSF/Cs/GO. The biocompatibility of hydrogels with human dermis fibroblasts in vitro was evaluated using the MTS assay. To assess the regenerative potential of hydrogels, a model of a full-layer skin defect was reconstructed on the back of rats and closed the wound surface with CS, DSF/Cs and DSF/Cs/GO hydrogels. The morphological and morphometric characteristics of regenerate tissues were obtained by staining with hematoxylin-eosin, Heidengain azocarmine, and immunohistochemistry on days 7 and 14 of the experiment. It has been shown that the use of DSF/Cs and DSF/Cs/GO promotes enhanced healing and epithelization of a full-layer skin wound. The addition of GO to the hydrogel increased the synthetic activity of fibroblasts and improved the characteristics of the produced collagen fibers.
Multiple sclerosis (MS) is an incurable, progressive chronic autoimmune demyelinating disease. Th... more Multiple sclerosis (MS) is an incurable, progressive chronic autoimmune demyelinating disease. Therapy for MS is based on slowing down the processes of neurodegeneration and suppressing the immune system of patients. MS is accompanied by inflammation, axon-degeneration and neurogliosis in the central nervous system. One of the directions for a new effective treatment for MS is cellular, subcellular, as well as gene therapy. We investigated the therapeutic potential of adipose mesenchymal stem cell (ADMSC) derived, cytochalasin B induced artificial microvesicles (MVs) expressing nerve growth factor (NGF) on a mouse model of multiple sclerosis experimental autoimmune encephalomyelitis (EAE). These ADMSC-MVs-NGF were tested using histological, immunocytochemical and molecular genetic methods after being injected into the tail vein of animals on the 14th and 21st days post EAE induction. ADMSC-MVs-NGF contained the target protein inside the cytoplasm. Their injection into the caudal vei...
Multiple sclerosis (MS) is a debilitating chronic disease of unknown etiology. There are limited ... more Multiple sclerosis (MS) is a debilitating chronic disease of unknown etiology. There are limited treatment options due to an incomplete understanding of disease pathology. The disease is shown to have seasonal exacerbation of clinical symptoms. The mechanisms of such seasonal worsening of symptoms remains unknown. In this study, we applied targeted metabolomics analysis of serum samples using LC-MC/MC to determine seasonal changes in metabolites throughout the four seasons. We also analyzed seasonal serum cytokine alterations in patients with relapsed MS. For the first time, we can demonstrate seasonal changes in various metabolites in MS compared to the control. More metabolites were affected in MS in the fall season followed by spring, while summer MS was characterized by the smallest number of affected metabolites. Ceramides were activated in all seasons, suggesting their central role in the disease pathogenesis. Substantial changes in glucose metabolite levels were found in MS, ...
Introduction: Therapeutic application of umbilical cord blood mononuclear cells (UCB-MCs) has bee... more Introduction: Therapeutic application of umbilical cord blood mononuclear cells (UCB-MCs) has been actively studied for at least during the last 30 years. Currently, UCB-MCs are extensively being investigated in the regeneration of the central nervous system (CNS) and the treatment of human neurodegenerative disorders. Translational studies dedicated to the application of UCB-MCs have revealed their capacity for stimulation of neurogenesis in the aged brain and promising potency for being therapeutic agents for treating such diseases as Alzheimer`s disease, amyotrophic lateral sclerosis (ALS), ischemic stroke, traumatic brain injury, Parkinson`s disease etc. Still the exact mechanism providing therapeutic effect remains unclear. Several studies modulating neurodegeneration have demonstrated immunomodulating and pro-inflammatory activity of UCB-MCs. Moreover, the unique feature of UCB-MCs, which underlies in the optional concordance of HLA or immunosuppression during transplantation,...
Amyotrophic lateral sclerosis (ALS) is an incurable, chronic, fatal neuro-degenerative disease ch... more Amyotrophic lateral sclerosis (ALS) is an incurable, chronic, fatal neuro-degenerative disease characterized by progressive loss of moto-neurons and paralysis of skeletal muscles. Reactivating dysfunctional areas is under earnest investigation utilizing various approaches. Here we present an innovative gene-cell construct aimed at reviving inert structure and function. Human umbilical cord blood cells (hUCBCs) transduced with adeno-viral vectors encoding human VEGF, GDNF and/or NCAM genes were transplanted into transgenic ALS mice models. Significant improvement in be-havioral performance (open-field and grip-strength tests), as well as increased lifespan was observed in rodents treated with NCAM-VEGF or NCAM-GDNF co-transfected cells. Active trans-gene expression was found in the spinal cord of ALS mice 10 weeks after delivering genetically modified hUCBCs, and cells were detectable even 5 months following transplantation. Our gene-cell therapy model yielded prominent symptomatic c...
Currently, the main fundamental and clinical interest for stroke therapy is focused on developing... more Currently, the main fundamental and clinical interest for stroke therapy is focused on developing a neuroprotective treatment of a penumbra region within the therapeutic window. The development of treatments for ischemic stroke in at-risk patients is of particular interest. Preventive gene therapy may significantly reduce the negative consequences of ischemia-induced brain injury. In the present study, we suggest the approach of preventive gene therapy for stroke. Adenoviral vectors carrying genes encoding vascular endothelial growth factor (VEGF), glial cell-derived neurotrophic factor (GDNF) and neural cell adhesion molecule (NCAM) or gene engineered umbilical cord blood mononuclear cells (UCB-MC) overexpressing recombinant VEGF, GDNF, and NCAM were intrathecally injected before distal occlusion of the middle cerebral artery in rats. Post-ischemic brain recovery was investigated 21 days after stroke modelling. Morphometric and immunofluorescent analysis revealed a reduction of inf...
Introduction. Umbilical cord plasma (UCB-PL) was shown to be beneficial for treatment of ischemic... more Introduction. Umbilical cord plasma (UCB-PL) was shown to be beneficial for treatment of ischemic brain injury, Gerotarget/Aging and Alzheimer9s disease (Yoo et al., 2016; Castellano et al., 2017). Also, umbilical cord mononuclear cell (UCB-MC) demonstrated therapeutic effect in animal models of stroke, encephalopathy, amyotrophic lateral sclerosis and cerebral palsy (Islamov et al., 2015; Mukhamedyarov et al., 2013). Therapeutic effect of freshly isolated cells is believed t be due to transcriptional activation of numerous neurotrophic, antiapoptotic, proangiogenic pro- and anti-inflammatory factors. The main purpose of this investigation was to analyze the cytokine profile of hUCB-PL and hUCB-MC. Materials and Methods. Umbilical cord blood was collected into CPDA-1 blood bag and used for plasma (hUCB-PL) separation. Total proteins were normalized in hUCB-PL samples. UCB-MCs were isolated by density gradient sedimentation using Ficoll (1.077g/ml). Separated mononuclear cells were w...
Herein proteomic profiling of the rat hippocampus from the kindling and pilocarpine models of epi... more Herein proteomic profiling of the rat hippocampus from the kindling and pilocarpine models of epilepsy was performed to achieve new potential targets for treating epileptic seizures. A total of 144 differently expressed proteins in both left and right hippocampi by two-dimensional electrophoresis coupled to matrix-assisted laser desorption-mass spectrometry were identified across the rat models of epilepsy. Based on network analysis, the majority of differentially expressed proteins were associated with Ca2+ homeostasis. Changes in ADP-ribosyl cyclase (ADPRC), lysophosphatidic acid receptor 3 (LPAR3), calreticulin, ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1), synaptosomal nerve-associated protein 25 (SNAP 25) and transgelin 3 proteins were probed by Western blot analysis and validated using immunohistochemistry. Inhibition of calcium influx by 8-Bromo-cADP-Ribose (8-Br-cADPR) and 2-Aminoethyl diphenylborinate (2-APB) which act via the ADPRC and LPAR3, respectively, attenuated ...
Nowadays gene and cell therapy become the basic methods in regenerative medicine. However only fe... more Nowadays gene and cell therapy become the basic methods in regenerative medicine. However only few gene and cell products are currently approved for clinical usage. Biosafety problems, complexity of cell and gene technologies and high cost of manufacturing are the main reasons for the slow introduction of such approaches in practical medicine. Treatment of hereditary diseases of the immune system based on the correction of the mutant gene by delivering functional recombinant gene into WBC is the first successfully employed in the clinical practice approach of cell-mediated or ex vivo gene therapy. Earlier we have reported the strategy of the cell-mediated gene therapy based on umbilical cord blood mononuclear cells transduced with adenoviral vectors carrying recombinant genes encoding neurotrophic factors for treatment neurodegenerative diseases, neurotrauma and stroke. Significant disadvantage of this method is the usage of the umbilical cord blood mononuclear cells as a cell carri...
Vascular endothelial growth factor (VEGF) and fibroblast growth factor 2 (FGF2) are well-known gr... more Vascular endothelial growth factor (VEGF) and fibroblast growth factor 2 (FGF2) are well-known growth factors involved in the regeneration of various tissues and organs, including peripheral nerve system. In the present study, we elucidated the local and systemic effects of plasmid construct рBud-coVEGF165-coFGF2 injected into the epineurium of intact rat sciatic nerve. Results of histological examination of sciatic nerve and multiplex immunoassays of serum showed the absence of immunogenicity and biosafety of plasmid рBud-coVEGF165-coFGF2. Moreover, local administration of plasmid DNA construct resulted in significantly decreased levels of pro-inflammatory cytokines in the peripheral blood, including tumor necrosis factor α (TNFα) and interleukin-12, and significantly increased levels of cytokines and chemokines including Regulated upon Activation, Normal T Cell Expressed and Presumably Secrete (RANTES), epidermal growth factor, interleukin-2, and monocyte chemoattractant protein 1. These changes in the peripheral blood on day 7 after injection of plasmid construct рBud-coVEGF165-coFGF2 show that the plasmid construct has systemic effects and may modulate immune response. At the same time, reverse transcription-polymerase chain reaction revealed transient expression of coFGF2, coVEGF165, ratFGF2 and ratVEGFA with direct transport of transcripts from distal part to proximal part of the sciatic nerve. Immunohistochemical staining revealed prolonged presence of VEGFA in sciatic nerve till 14 days post-injection. These findings suggest that local administration of plasmid construct рBud-coVEGF165-coFGF2 at a concentration of 30 ng/µL results in the formation of pro-angiogenic stimuli and, and the plasmid construct, used as a drug for gene therapy, might potentially facilitate regeneration of the sciatic nerve. The study was approved by the Animal Ethics Committee of Kazan Federal University, procedures were approved by the Local Ethics Committee (approval No. 5) on May 27, 2014.
Tay-Sachs disease (TSD) is a progressive neurodegenerative disorder that occurs due to a deficien... more Tay-Sachs disease (TSD) is a progressive neurodegenerative disorder that occurs due to a deficiency of a β hexosaminidase A (HexA) enzyme, resulting in the accumulation of GM2 gangliosides. In this work, we analyzed the effect of umbilical cord blood cell transplantation (UCBCT) and curcumin administration on the course of the disease in a patient with adult TSD. The patient’s serum cytokine profile was determined using multiplex analysis. The level of GM2 gangliosides in plasma was determined using mass spectrometry. The enzymatic activity of HexA in the plasma of the patient was assessed using a fluorescent substrate assay. The HexA α-subunit (HexA) concentration was determined using ELISA. It was shown that both UCBCT and curcumin administration led to a change in the patient’s cytokine profile. The UCBCT resulted in an increase in the concentration of HexA in the patient’s serum and in an improvement in the patient’s neurological status. However, neither UCBCT nor curcumin were ...
The translation of new therapies for spinal cord injury to clinical trials can be facilitated wit... more The translation of new therapies for spinal cord injury to clinical trials can be facilitated with large animal models close in morpho-physiological scale to humans. Here, we report functional restoration and morphological reorganization after spinal contusion in pigs, following a combined treatment of locomotor training facilitated with epidural electrical stimulation (EES) and cell-mediated triple gene therapy with umbilical cord blood mononuclear cells overexpressing recombinant vascular endothelial growth factor, glial-derived neurotrophic factor, and neural cell adhesion molecule. Preliminary results obtained on a small sample of pigs 2 months after spinal contusion revealed the difference in post-traumatic spinal cord outcomes in control and treated animals. In treated pigs, motor performance was enabled by EES and the corresponding morpho-functional changes in hind limb skeletal muscles were accompanied by the reorganization of the glial cell, the reaction of stress cell, and...
Cell-mediated (ex-vivo) gene therapy for the treatment of adenosine deaminase (ADA)-deficient sev... more Cell-mediated (ex-vivo) gene therapy for the treatment of adenosine deaminase (ADA)-deficient severe combined immunodeficiency (SCID) had started in 1990 and nowadays it is the first marketing approval of an ex vivo gene therapy in Europe. The method based on ex-vivo transduction of peripheral blood lymphocytes with retroviral vector carrying the functional ADA gene in 2002 have been improved to use hematopoietic stem cell (HSC) for ex-vivo transduction with 100% survival and the evidence of safety and efficacy. Remarkably, umbilical cord blood mononuclear cells (UCB-MC) were successfully used for treatment of ADA deficiency in neonates as well. Meanwhile SCID is a very rare congenital disorder of the immune system although the option to use peripheral blood lymphocytes as cell carriers of the therapeutic genes for regenerative medicine is highly attractive. In our studies to overcome the neural cells death and stimulate neuroregeneration at neurodegenerative diseases (ALS), spinal ...
Background: The cytochalasin B-induced membrane vesicles (CIMVs) are suggested to be used as a ve... more Background: The cytochalasin B-induced membrane vesicles (CIMVs) are suggested to be used as a vehicle for the delivery of therapeutics. However, the angiogenic activity and therapeutic potential of human mesenchymal stem cells (MSCs) derived CIMVs (CIMVs-MSCs) remains unknown. Objectives: The objectives of this study were to analyzed the morphology, size distribution, molecular composition and angiogenic properties of CIMVs-MSCs. Methods: The morphology of CIMVs-MSC was analyzed by scanning electron microscopy. The proteomic analysis, multiplex analysis and immunostaining were used to characterize the molecular composition of the CIMVs-MSCs. The transfer of surface proteins from a donor to a recipient cell mediated by CIMVs-MSCs was demonstrated using immunostaining and confocal microscopy. The angiogenic potential of CIMVs-MSCs was evaluated using in vivo approach of subcutaneous implantation of CIMVs-MSCs in mixture with Matrigel matrix. Results: Human CIMVs-MSCs retain parental ...
Stimulating the process of angiogenesis in treating ischemia-related diseases is an urgent task f... more Stimulating the process of angiogenesis in treating ischemia-related diseases is an urgent task for modern medicine, which can be achieved through the use of different cell types. Umbilical cord blood (UCB) continues to be one of the attractive cell sources for transplantation. The goal of this study was to investigate the role and therapeutic potential of gene-engineered umbilical cord blood mononuclear cells (UCB-MC) as a forward-looking strategy for the activation of angiogenesis. Adenovirus constructs Ad-VEGF, Ad-FGF2, Ad-SDF1α, and Ad-EGFP were synthesized and used for cell modification. UCB-MCs were isolated from UCB and transduced with adenoviral vectors. As part of our in vitro experiments, we evaluated the efficiency of transfection, the expression of recombinant genes, and the secretome profile. Later, we applied an in vivo Matrigel plug assay to assess engineered UCB-MC’s angiogenic potential. We conclude that hUCB-MCs can be efficiently modified simultaneously with sever...
The biosafety of gene therapy remains a crucial issue for both the direct and cell-mediated deliv... more The biosafety of gene therapy remains a crucial issue for both the direct and cell-mediated delivery of recombinant cDNA encoding biologically active molecules for the pathogenetic correction of congenital or acquired disorders. The diversity of vector systems and cell carriers for the delivery of therapeutic genes revealed the difficulty of developing and implementing a safe and effective drug containing artificial genetic material for the treatment of human diseases in practical medicine. Therefore, in this study we assessed changes in the transcriptome and secretome of umbilical cord blood mononuclear cells (UCB-MCs) genetically modified using adenoviral vector (Ad5) carrying cDNA encoding human vascular endothelial growth factor (VEGF165) or reporter green fluorescent protein (GFP). A preliminary analysis of UCB-MCs transduced with Ad5-VEGF165 and Ad5-GFP with MOI of 10 showed efficient transgene expression in gene-modified UCB-MCs at mRNA and protein levels. The whole transcrip...
In order to improve the regeneration of full-layer skin defects, hydrogels were developed based o... more In order to improve the regeneration of full-layer skin defects, hydrogels were developed based on the combination of chitosan (Cs), Daba silk fibroin (DSF), and graphene oxide (GO): CS, DSF/Cs and DSF/Cs/GO. The biocompatibility of hydrogels with human dermis fibroblasts in vitro was evaluated using the MTS assay. To assess the regenerative potential of hydrogels, a model of a full-layer skin defect was reconstructed on the back of rats and closed the wound surface with CS, DSF/Cs and DSF/Cs/GO hydrogels. The morphological and morphometric characteristics of regenerate tissues were obtained by staining with hematoxylin-eosin, Heidengain azocarmine, and immunohistochemistry on days 7 and 14 of the experiment. It has been shown that the use of DSF/Cs and DSF/Cs/GO promotes enhanced healing and epithelization of a full-layer skin wound. The addition of GO to the hydrogel increased the synthetic activity of fibroblasts and improved the characteristics of the produced collagen fibers.
Multiple sclerosis (MS) is an incurable, progressive chronic autoimmune demyelinating disease. Th... more Multiple sclerosis (MS) is an incurable, progressive chronic autoimmune demyelinating disease. Therapy for MS is based on slowing down the processes of neurodegeneration and suppressing the immune system of patients. MS is accompanied by inflammation, axon-degeneration and neurogliosis in the central nervous system. One of the directions for a new effective treatment for MS is cellular, subcellular, as well as gene therapy. We investigated the therapeutic potential of adipose mesenchymal stem cell (ADMSC) derived, cytochalasin B induced artificial microvesicles (MVs) expressing nerve growth factor (NGF) on a mouse model of multiple sclerosis experimental autoimmune encephalomyelitis (EAE). These ADMSC-MVs-NGF were tested using histological, immunocytochemical and molecular genetic methods after being injected into the tail vein of animals on the 14th and 21st days post EAE induction. ADMSC-MVs-NGF contained the target protein inside the cytoplasm. Their injection into the caudal vei...
Multiple sclerosis (MS) is a debilitating chronic disease of unknown etiology. There are limited ... more Multiple sclerosis (MS) is a debilitating chronic disease of unknown etiology. There are limited treatment options due to an incomplete understanding of disease pathology. The disease is shown to have seasonal exacerbation of clinical symptoms. The mechanisms of such seasonal worsening of symptoms remains unknown. In this study, we applied targeted metabolomics analysis of serum samples using LC-MC/MC to determine seasonal changes in metabolites throughout the four seasons. We also analyzed seasonal serum cytokine alterations in patients with relapsed MS. For the first time, we can demonstrate seasonal changes in various metabolites in MS compared to the control. More metabolites were affected in MS in the fall season followed by spring, while summer MS was characterized by the smallest number of affected metabolites. Ceramides were activated in all seasons, suggesting their central role in the disease pathogenesis. Substantial changes in glucose metabolite levels were found in MS, ...
Introduction: Therapeutic application of umbilical cord blood mononuclear cells (UCB-MCs) has bee... more Introduction: Therapeutic application of umbilical cord blood mononuclear cells (UCB-MCs) has been actively studied for at least during the last 30 years. Currently, UCB-MCs are extensively being investigated in the regeneration of the central nervous system (CNS) and the treatment of human neurodegenerative disorders. Translational studies dedicated to the application of UCB-MCs have revealed their capacity for stimulation of neurogenesis in the aged brain and promising potency for being therapeutic agents for treating such diseases as Alzheimer`s disease, amyotrophic lateral sclerosis (ALS), ischemic stroke, traumatic brain injury, Parkinson`s disease etc. Still the exact mechanism providing therapeutic effect remains unclear. Several studies modulating neurodegeneration have demonstrated immunomodulating and pro-inflammatory activity of UCB-MCs. Moreover, the unique feature of UCB-MCs, which underlies in the optional concordance of HLA or immunosuppression during transplantation,...
Amyotrophic lateral sclerosis (ALS) is an incurable, chronic, fatal neuro-degenerative disease ch... more Amyotrophic lateral sclerosis (ALS) is an incurable, chronic, fatal neuro-degenerative disease characterized by progressive loss of moto-neurons and paralysis of skeletal muscles. Reactivating dysfunctional areas is under earnest investigation utilizing various approaches. Here we present an innovative gene-cell construct aimed at reviving inert structure and function. Human umbilical cord blood cells (hUCBCs) transduced with adeno-viral vectors encoding human VEGF, GDNF and/or NCAM genes were transplanted into transgenic ALS mice models. Significant improvement in be-havioral performance (open-field and grip-strength tests), as well as increased lifespan was observed in rodents treated with NCAM-VEGF or NCAM-GDNF co-transfected cells. Active trans-gene expression was found in the spinal cord of ALS mice 10 weeks after delivering genetically modified hUCBCs, and cells were detectable even 5 months following transplantation. Our gene-cell therapy model yielded prominent symptomatic c...
Currently, the main fundamental and clinical interest for stroke therapy is focused on developing... more Currently, the main fundamental and clinical interest for stroke therapy is focused on developing a neuroprotective treatment of a penumbra region within the therapeutic window. The development of treatments for ischemic stroke in at-risk patients is of particular interest. Preventive gene therapy may significantly reduce the negative consequences of ischemia-induced brain injury. In the present study, we suggest the approach of preventive gene therapy for stroke. Adenoviral vectors carrying genes encoding vascular endothelial growth factor (VEGF), glial cell-derived neurotrophic factor (GDNF) and neural cell adhesion molecule (NCAM) or gene engineered umbilical cord blood mononuclear cells (UCB-MC) overexpressing recombinant VEGF, GDNF, and NCAM were intrathecally injected before distal occlusion of the middle cerebral artery in rats. Post-ischemic brain recovery was investigated 21 days after stroke modelling. Morphometric and immunofluorescent analysis revealed a reduction of inf...
Introduction. Umbilical cord plasma (UCB-PL) was shown to be beneficial for treatment of ischemic... more Introduction. Umbilical cord plasma (UCB-PL) was shown to be beneficial for treatment of ischemic brain injury, Gerotarget/Aging and Alzheimer9s disease (Yoo et al., 2016; Castellano et al., 2017). Also, umbilical cord mononuclear cell (UCB-MC) demonstrated therapeutic effect in animal models of stroke, encephalopathy, amyotrophic lateral sclerosis and cerebral palsy (Islamov et al., 2015; Mukhamedyarov et al., 2013). Therapeutic effect of freshly isolated cells is believed t be due to transcriptional activation of numerous neurotrophic, antiapoptotic, proangiogenic pro- and anti-inflammatory factors. The main purpose of this investigation was to analyze the cytokine profile of hUCB-PL and hUCB-MC. Materials and Methods. Umbilical cord blood was collected into CPDA-1 blood bag and used for plasma (hUCB-PL) separation. Total proteins were normalized in hUCB-PL samples. UCB-MCs were isolated by density gradient sedimentation using Ficoll (1.077g/ml). Separated mononuclear cells were w...
Herein proteomic profiling of the rat hippocampus from the kindling and pilocarpine models of epi... more Herein proteomic profiling of the rat hippocampus from the kindling and pilocarpine models of epilepsy was performed to achieve new potential targets for treating epileptic seizures. A total of 144 differently expressed proteins in both left and right hippocampi by two-dimensional electrophoresis coupled to matrix-assisted laser desorption-mass spectrometry were identified across the rat models of epilepsy. Based on network analysis, the majority of differentially expressed proteins were associated with Ca2+ homeostasis. Changes in ADP-ribosyl cyclase (ADPRC), lysophosphatidic acid receptor 3 (LPAR3), calreticulin, ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1), synaptosomal nerve-associated protein 25 (SNAP 25) and transgelin 3 proteins were probed by Western blot analysis and validated using immunohistochemistry. Inhibition of calcium influx by 8-Bromo-cADP-Ribose (8-Br-cADPR) and 2-Aminoethyl diphenylborinate (2-APB) which act via the ADPRC and LPAR3, respectively, attenuated ...
Nowadays gene and cell therapy become the basic methods in regenerative medicine. However only fe... more Nowadays gene and cell therapy become the basic methods in regenerative medicine. However only few gene and cell products are currently approved for clinical usage. Biosafety problems, complexity of cell and gene technologies and high cost of manufacturing are the main reasons for the slow introduction of such approaches in practical medicine. Treatment of hereditary diseases of the immune system based on the correction of the mutant gene by delivering functional recombinant gene into WBC is the first successfully employed in the clinical practice approach of cell-mediated or ex vivo gene therapy. Earlier we have reported the strategy of the cell-mediated gene therapy based on umbilical cord blood mononuclear cells transduced with adenoviral vectors carrying recombinant genes encoding neurotrophic factors for treatment neurodegenerative diseases, neurotrauma and stroke. Significant disadvantage of this method is the usage of the umbilical cord blood mononuclear cells as a cell carri...
Vascular endothelial growth factor (VEGF) and fibroblast growth factor 2 (FGF2) are well-known gr... more Vascular endothelial growth factor (VEGF) and fibroblast growth factor 2 (FGF2) are well-known growth factors involved in the regeneration of various tissues and organs, including peripheral nerve system. In the present study, we elucidated the local and systemic effects of plasmid construct рBud-coVEGF165-coFGF2 injected into the epineurium of intact rat sciatic nerve. Results of histological examination of sciatic nerve and multiplex immunoassays of serum showed the absence of immunogenicity and biosafety of plasmid рBud-coVEGF165-coFGF2. Moreover, local administration of plasmid DNA construct resulted in significantly decreased levels of pro-inflammatory cytokines in the peripheral blood, including tumor necrosis factor α (TNFα) and interleukin-12, and significantly increased levels of cytokines and chemokines including Regulated upon Activation, Normal T Cell Expressed and Presumably Secrete (RANTES), epidermal growth factor, interleukin-2, and monocyte chemoattractant protein 1. These changes in the peripheral blood on day 7 after injection of plasmid construct рBud-coVEGF165-coFGF2 show that the plasmid construct has systemic effects and may modulate immune response. At the same time, reverse transcription-polymerase chain reaction revealed transient expression of coFGF2, coVEGF165, ratFGF2 and ratVEGFA with direct transport of transcripts from distal part to proximal part of the sciatic nerve. Immunohistochemical staining revealed prolonged presence of VEGFA in sciatic nerve till 14 days post-injection. These findings suggest that local administration of plasmid construct рBud-coVEGF165-coFGF2 at a concentration of 30 ng/µL results in the formation of pro-angiogenic stimuli and, and the plasmid construct, used as a drug for gene therapy, might potentially facilitate regeneration of the sciatic nerve. The study was approved by the Animal Ethics Committee of Kazan Federal University, procedures were approved by the Local Ethics Committee (approval No. 5) on May 27, 2014.
Tay-Sachs disease (TSD) is a progressive neurodegenerative disorder that occurs due to a deficien... more Tay-Sachs disease (TSD) is a progressive neurodegenerative disorder that occurs due to a deficiency of a β hexosaminidase A (HexA) enzyme, resulting in the accumulation of GM2 gangliosides. In this work, we analyzed the effect of umbilical cord blood cell transplantation (UCBCT) and curcumin administration on the course of the disease in a patient with adult TSD. The patient’s serum cytokine profile was determined using multiplex analysis. The level of GM2 gangliosides in plasma was determined using mass spectrometry. The enzymatic activity of HexA in the plasma of the patient was assessed using a fluorescent substrate assay. The HexA α-subunit (HexA) concentration was determined using ELISA. It was shown that both UCBCT and curcumin administration led to a change in the patient’s cytokine profile. The UCBCT resulted in an increase in the concentration of HexA in the patient’s serum and in an improvement in the patient’s neurological status. However, neither UCBCT nor curcumin were ...
The translation of new therapies for spinal cord injury to clinical trials can be facilitated wit... more The translation of new therapies for spinal cord injury to clinical trials can be facilitated with large animal models close in morpho-physiological scale to humans. Here, we report functional restoration and morphological reorganization after spinal contusion in pigs, following a combined treatment of locomotor training facilitated with epidural electrical stimulation (EES) and cell-mediated triple gene therapy with umbilical cord blood mononuclear cells overexpressing recombinant vascular endothelial growth factor, glial-derived neurotrophic factor, and neural cell adhesion molecule. Preliminary results obtained on a small sample of pigs 2 months after spinal contusion revealed the difference in post-traumatic spinal cord outcomes in control and treated animals. In treated pigs, motor performance was enabled by EES and the corresponding morpho-functional changes in hind limb skeletal muscles were accompanied by the reorganization of the glial cell, the reaction of stress cell, and...
Cell-mediated (ex-vivo) gene therapy for the treatment of adenosine deaminase (ADA)-deficient sev... more Cell-mediated (ex-vivo) gene therapy for the treatment of adenosine deaminase (ADA)-deficient severe combined immunodeficiency (SCID) had started in 1990 and nowadays it is the first marketing approval of an ex vivo gene therapy in Europe. The method based on ex-vivo transduction of peripheral blood lymphocytes with retroviral vector carrying the functional ADA gene in 2002 have been improved to use hematopoietic stem cell (HSC) for ex-vivo transduction with 100% survival and the evidence of safety and efficacy. Remarkably, umbilical cord blood mononuclear cells (UCB-MC) were successfully used for treatment of ADA deficiency in neonates as well. Meanwhile SCID is a very rare congenital disorder of the immune system although the option to use peripheral blood lymphocytes as cell carriers of the therapeutic genes for regenerative medicine is highly attractive. In our studies to overcome the neural cells death and stimulate neuroregeneration at neurodegenerative diseases (ALS), spinal ...
Background: The cytochalasin B-induced membrane vesicles (CIMVs) are suggested to be used as a ve... more Background: The cytochalasin B-induced membrane vesicles (CIMVs) are suggested to be used as a vehicle for the delivery of therapeutics. However, the angiogenic activity and therapeutic potential of human mesenchymal stem cells (MSCs) derived CIMVs (CIMVs-MSCs) remains unknown. Objectives: The objectives of this study were to analyzed the morphology, size distribution, molecular composition and angiogenic properties of CIMVs-MSCs. Methods: The morphology of CIMVs-MSC was analyzed by scanning electron microscopy. The proteomic analysis, multiplex analysis and immunostaining were used to characterize the molecular composition of the CIMVs-MSCs. The transfer of surface proteins from a donor to a recipient cell mediated by CIMVs-MSCs was demonstrated using immunostaining and confocal microscopy. The angiogenic potential of CIMVs-MSCs was evaluated using in vivo approach of subcutaneous implantation of CIMVs-MSCs in mixture with Matrigel matrix. Results: Human CIMVs-MSCs retain parental ...
Uploads
Papers by Ilnur Salafutdinov