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Background: The biosafety of gene therapy products remains a major challenge to their introduction into the clinic. In particular, the problem of immunogenicity of viral vectors is the focus of attention. Large animals such as pigs, whose anatomical and physiological characteristics are similar to those of humans, have an advantage in testing vector systems., Methods: We performed a comprehensive in vitro and in vivo study to evaluate the biosafety of a chimeric adenoviral vector carrying a green fluorescent protein gene (Ad5/35F-GFP) in a mini-pig model., Results: Transcriptome and secretome analyses of mini-pig leucocytes transduced with Ad5/35F-GFP revealed changes restraining pro-inflammatory processes and cytokine production. No adverse effects were revealed through the clinical, instrumental, laboratory, and histological examinations conducted within a week after the direct or autologous leucocyte-mediated administration of Ad5/35F-GFP to mini-pigs. The decrease in cytokine levels in the blood of experimental animals is also consistent with the in vitro data and confirms the immune tolerance of mini-pigs to Ad5/35F-GFP., Conclusions: Here, we show the safety of Ad5/35F in a mini-pig model and provide evidence that Ad5/35F is a promising vector for gene therapy. These results advance our understanding of vector-host interactions and offer a solid foundation for the clinical application of this vector.

Hypertriglyceridemia, obesity, and aging are among the key risk factors for severe COVID-19 with acute respiratory distress syndrome (ARDS). One of the main prognostic biomarkers of ARDS is the level of cytokines IL-6 and TNF-α in the blood. In our study, we modeled hyperglyceridemia and hypercholesterolemia on 18-month-old Syrian hamsters ( Mesocricetus auratus ). By 18 months, the animals showed such markers of aging as weight stabilization with a tendency to reduce it, polycystic liver disease, decreased motor activity, and foci of alopecia. The high-fat diet caused an increase in triglycerides and cholesterol, as well as fatty changes in the liver. On the third day after infection with the SARS-CoV-2 virus, animals showed a decrease in weight in the groups with a high-fat diet. In the lungs of males on both diets, there was an increase in the concentration of IFN-α, as well as IL-6 in both males and females, regardless of the type of diet. At the same time, the levels of TNF-α and IFN-γ did not change in infected animals. Morphological studies of the lungs of hamsters with SARS-CoV-2 showed the presence of a pathological process characteristic of ARDS. These included bronchointerstitial pneumonia and diffuse alveolar damages. These observations suggest that in aging hamsters, the immune response to pro-inflammatory cytokines may be delayed to a later period. Hypertriglyceridemia, age, and gender affect the severity of COVID-19. These results will help to understand the pathogenesis of COVID-19 associated with age, gender, and disorders of fat metabolism in humans., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Fomin, Tabynov, Islamov, Turebekov, Yessimseit and Yerubaev.)

The effect of transtraumatic epidural electrostimulation (TEES) above (T5) and below (L2) spinal cord injury in the lower thoracic region (T8-T9) in combination with treadmill exercise in pigs was evaluated using electrophysiological examination methods and behavioral tests. Two weeks after spinal cord injury, motor evoked potentials of m. soleus were recorded during electrostimulation at the level of T5 and L2 segments, which indicated activation of spinal cord structures above and below the focus of injury. After 6 weeks of TEES in combination with physical training, restoration of the characteristics of M-response and H-reflex of the soleus muscle in response to stimulation of the sciatic nerve, improvement of joint mobility, and appearance of voluntary motor activity in the hindlimbs were observed. Neuromodulation with TEES had been proven to be an effective way to stimulate posttraumatic spinal cord regeneration and can be used in the development of a neurorehabilitation protocol for patients with spinal cord injury., (© 2023. Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Pathological changes associated with spinal cord injury (SCI) can be observed distant, rostral, or caudal to the epicenter of injury. These remote areas represent important therapeutic targets for post-traumatic spinal cord repair. The present study aimed to investigate the following in relation to SCI: distant changes in the spinal cord, peripheral nerve, and muscles., Methods: The changes in the spinal cord, the tibial nerve, and the hind limb muscles were evaluated in control SCI animals and after intravenous infusion of autologous leucoconcentrate enriched with genes encoding neuroprotective factors (VEGF, GDNF, and NCAM), which previously demonstrated a positive effect on post-traumatic restoration., Results: Two months after thoracic contusion in the treated mini pigs, a positive remodeling of the macro- and microglial cells, expression of PSD95 and Chat in the lumbar spinal cord, and preservation of the number and morphological characteristics of the myelinated fibers in the tibial nerve were observed and were aligned with hind limb motor recovery and reduced soleus muscle atrophy., Conclusion: Here, we show the positive effect of autologous genetically enriched leucoconcentrate-producing recombinant neuroprotective factors on targets distant to the primary lesion site in mini pigs with SCI. These findings open new perspectives for the therapy of SCI.

The development of hypogravitational motor syndrome is an essential negative consequence of weightlessness for humans; an important role in the pathogenesis of this syndrome is played by changes in axons of the spinal tracts. Myelinated fibers and transcriptome of the spinal cord were studied in mice exposed to hypogravity during a 30-day flight on a biosatellite. Morphometric analysis of myelinated fibers of the spinal tracts showed a decrease in the thickness of the myelin sheath. Analysis of spinal cord transcriptome revealed a decrease in the expression of genes involved in the myelination of nerve fibers. These results suggest that the processes of nerve fiber myelination are involved in the development of the hypogravitational motor syndrome under weightless conditions; the 7-day readaptation period was found to be insufficient for reversion of the negative changes in the myelinated fibers of the spinal cord., (© 2023. Springer Science+Business Media, LLC, part of Springer Nature.)

The natural limitations of regeneration in the CNS are major problems for the treatment of neurological disorders, including ischaemic brain strokes. Among the approaches being actively developed to inhibit post-ischaemic negative consequences is the delivery of therapeutic genes encoding neuroprotective molecules to the brain. Unfortunately, there are currently no proven and available medicines that contain recombinant human genes for the treatment of ischaemic cerebral stroke. Of particular interest is the development of treatments for patients at risk of ischaemic stroke. In the present study, we propose a proof of concept for the use of an autologous, genetically enriched leucoconcentrate temporally secreting recombinant vascular endothelial growth factor (VEGF), glial-cell-line-derived neurotrophic factor (GDNF) and the neural cell adhesion molecule (NCAM) for the treatment of stroke. In a mini-pig ischaemic stroke model, genetically enriched leucoconcentrate was infused 4 h after surgery (gene therapy in acute phase) or 2 days before stroke modelling (preventive gene therapy). On day 21, after the stroke modelling, the post-ischaemic brain recovery was examined by morphologic and immunofluorescence analysis. The benefits of treating a stroke with genetically enriched leucoconcentrate both for preventive purposes and in the acute phase were confirmed by an improved performance in behavioural tests, higher preservation of brain tissue and positive post-ischaemic brain remodelling in the peri-infarct area. These results suggest that the employment of autologous leucocytes enabling the temporary production of the recombinant therapeutic molecules to correct the pathological process in the CNS may be one of the breakthrough approaches in gene therapy.

The contemporary strategy for spinal cord injury (SCI) therapy aims to combine multiple approaches to control pathogenic mechanisms of neurodegeneration and stimulate neuroregeneration. In this study, a novel regenerative approach using an autologous leucoconcentrate enriched with transgenes encoding vascular endothelial growth factor (VEGF), glial cell line-derived neurotrophic factor (GDNF), and neural cell adhesion molecule (NCAM) combined with supra- and sub-lesional epidural electrical stimulation (EES) was tested on mini-pigs similar in morpho-physiological scale to humans. The complex analysis of the spinal cord recovery after a moderate contusion injury in treated mini-pigs compared to control animals revealed: better performance in behavioural and joint kinematics, restoration of electromyography characteristics, and improvement in selected immunohistology features related to cell survivability, synaptic protein expression, and glial reorganization above and below the injury. These results for the first time demonstrate the positive effect of intravenous infusion of autologous genetically-enriched leucoconcentrate producing recombinant molecules stimulating neuroregeneration combined with neuromodulation by translesional multisite EES on the restoration of the post-traumatic spinal cord in mini-pigs and suggest the high translational potential of this novel regenerative therapy for SCI patients.

Resistance to antibacterial therapy requires the discovery of new methods for the treatment of infectious diseases. Lactoferrin (LTF) is a well-known naïve first-line defense protein. In the present study, we suggested the use of an adenoviral vector (Ad5) carrying the human gene encoding LTF for direct and cell-mediated gene therapy of maxillofacial area phlegmon in rats. Abscesses were developed by injection of the purulent peritoneal exudate in the molar region of the medial surface of the mandible. At 3-4 days after phlegmon maturation, all rats received ceftriaxone and afterward were subcutaneously injected around the phlegmon with: (1) Ad5 carrying reporter gfp gene encoding green fluorescent protein (Ad5-GFP control group), (2) Ad5 carrying LTF gene (Ad5-LTF group), (3) human umbilical cord blood mononuclear cells (UCBC) transduced with Ad5-GFP (UCBC + Ad5-GFP group), and (4) UCBC transduced with Ad5-LTF (UCBC + Ad5-LTF group). Control rats developed symptoms considered to be related to systemic inflammation and were euthanized at 4-5 days from the beginning of the treatment. Rats from therapeutic groups demonstrated wound healing and recovery from the fifth to seventh day based on the type of therapy. Histological investigation of cervical lymph nodes revealed purulent lymphadenitis in control rats and activated lymphatic tissue in rats from the UCBC + Ad5-LTF group. Our results propose that both approaches of LTF gene delivery are efficient for maxillofacial area phlegmon recovery in rats. However, earlier wound healing and better outcomes in cervical lymph node remodeling in the UCBC + Ad5-LTF group, as well as the lack of direct exposure of the viral vector to the organism, which may cause toxic and immunogenic effects, suggest the benefit of cell-mediated gene therapy.

The data obtained by transcriptome analysis of lumbar spinal cord segments, sciatic nerve, and the respiratory diaphragm of the mice performed after a space flight on board Bion-M1 biosatellite were processed by bioinformatic methods aimed at elucidation of the regularities in hypogravity-induced transcriptome changes in various compartments of motor neurons. The study revealed abnormalities of axonal transport in spinal motor neurons provoked by weightlessness. These data agree with the results of electron microscopy examination of the spinal cord in experimental animals. In space group mice sacrificed on the landing day, the content of perinuclear ribosomes in lumbar motoneurons surpassed that in control mice or in the recovery group examined 1 week after the flight. The data corroborate our hypothesis on contribution of axonal transport disturbances into pathogenesis of hypogravity motor syndrome. They can be employed as a launching pad for further study of hypogravity-triggered motor disorder mechanisms in order to elaborate the preventive therapy against the development of hypogravity motor syndrome in space flights.

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 synaptic proteins. Our data demonstrate effects of combined EES-facilitated motor training and cell-mediated triple gene therapy after spinal contusion in large animals, informing a background for further animal studies and clinical translation.

This study evaluates the effect of combined epidural electrical stimulation (EES) applied above (C5) and below (L2) the spinal cord injury (SCI) at T8-9 combined with motor training on the restoration of sensorimotor function in mini pigs. The motor evoked potentials (MEP) induced by EES applied at C5 and L2 levels were recorded in soleus muscles before and two weeks after SCI. EES treatment started two weeks after SCI and continued for 6 weeks led to improvement in multiple metrics, including behavioral, electrophysiological, and joint kinematics outcomes. In control animals after SCI a multiphasic M-response was observed during M/H-response testing, while animals received EES-enable training demonstrated the restoration of the M-response and H-reflex, although at a lower amplitude. The joint kinematic and assessment with Porcine Thoracic Injury Behavior scale (PTIBS) motor recovery scale demonstrated improvement in animals that received EES-enable training compared to animals with no treatment. The positive effect of two-level (cervical and lumbar) epidural electrical stimulation on functional restoration in mini pigs following spinal cord contusion injury in mini pigs could be related with facilitation of spinal circuitry at both levels and activation of multisegmental coordination. This approach can be taken as a basis for the future development of neuromodulation and neurorehabilitation therapy for patients with spinal cord injury.

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 infarction volume accompanied with a lower number of apoptotic cells and decreased expression of Hsp70 in the peri-infarct region in gene-treated animals. The lower immunopositive areas for astrocytes and microglial cells markers, higher number of oligodendrocytes and increased expression of synaptic proteins suggest the inhibition of astrogliosis, supporting the corresponding myelination and functional recovery of neurons in animals receiving preventive gene therapy. In this study, for the first time, we provide evidence of the beneficial effects of preventive triple gene therapy by an adenoviral- or UCB-MC-mediated intrathecal simultaneous delivery combination of vegf165, gdnf, and ncam1 on the preservation and recovery of the brain in rats with subsequent modelling of stroke.

Background: Complexes of iodine (povidone-iodine and cadexomers) are among the most important antiseptics used in clinical and veterinary medicines. However, high local irritation activity and systemic toxicity limits their oral administration. The purpose of the study was to compare the effect of a new complex of iodine (PA, potentiator of anticancer antibiotics), in which iodine is coordinated by carbohydrates and polypeptides) on the organisms of rats and dogs treated orally with the drug for 30 days. Methods: Wistar rats and Beagle dogs served as experimental animal models. Effect of PA on the animal organism was examined through the measurements of hormones level changes, hematological and clinical chemistry parameters alterations, necropsy and histological examination. Results: The established maximum tolerated dose (MTD) of 2,000 mg/kg PA led to a decrease in the rate of body weight gain in male and female rats. Changes in hematological and certain biochemical parameters in rats at doses of 1,000 and 2,000 mg/kg were observed. Histological study of the thyroid gland revealed changes in the shape and size of the follicles along with colloid resorption. Administration of a half of MTD (180 mg/kg) and lower doses did not result in any change in dogs (thyroid-stimulating hormone, triiodothyronine, and thyroxine). Conclusions: The results of our study show that the pathogenetic action of PA takes place along the path of induction of an inflammatory response with the development of thyrotoxicosis, rather than hypothyroidism. The mechanism of induction of an inflammatory response is also confirmed by histological studies of lesions of the thyroid gland and testes in rats (Figure S1). The no-observed-adverse-effect level (NOAEL) of PA is estimated to be 180 mg/kg (or iodine 22.8 mg/kg) in dogs, which is equivalent to 100 mg/kg (or iodine 12.3 mg/kg) in humans., (Copyright © 2020 Islamov, Kustova, Nersesyan and Ilin.)

We revealed a decrease in the thickness of the myelin sheath and myelin delamination in the tibial nerve of C57BL/6N mice after a 30-day flight aboard the biosatellite Bion-M1. The processes of myelin degeneration continued for seven days after return of the animals to Earth and adaptation to the conditions of natural gravity. Our data add to hypothesis on the role of neurogenic component in pathogenesis of hypogravity motor syndrome.

OBJECTIVE The most actively explored therapeutic strategy for overcoming spinal cord injury (SCI) is the delivery of genes encoding molecules that stimulate regeneration. In a mouse model of amyotrophic lateral sclerosis and in preliminary clinical trials in patients with amyotrophic lateral sclerosis, the combined administration of recombinant adenoviral vectors (Ad5-VEGF+Ad5-ANG) encoding the neurotrophic/angiogenic factors vascular endothelial growth factor ( VEGF) and angiogenin ( ANG) was found to slow the development of neurological deficits. These results suggest that there may be positive effects of this combination of genes in posttraumatic spinal cord regeneration. The objective of the present study was to determine the effects of Ad5-VEGF+Ad5-ANG combination therapy on motor function recovery and reactivity of astrocytes in a rat model of SCI. METHODS Spinal cord injury was induced in adult Wistar rats by the weight-drop method. Rats (n = 51) were divided into 2 groups: the experimental group (Ad5-VEGF+Ad5-ANG) and the control group (Ad5-GFP [green fluorescent protein]). Recovery of motor function was assessed using the Basso, Beattie, and Bresnahan scale. The duration and intensity of infectivity and gene expression from the injected vectors were assessed by immunofluorescent detection of GFP. Reactivity of glial cells was assessed by changes in the number of immunopositive cells expressing glial fibrillary acidic protein (GFAP), S100β, aquaporin 4 (AQP4), oligodendrocyte transcription factor 2, and chondroitin sulfate proteoglycan 4. The level of S100β mRNA expression in the spinal cord was estimated by real-time polymerase chain reaction. RESULTS Partial recovery of motor function was observed 30 days after surgery in both groups. However, Basso, Beattie, and Bresnahan scores were 35.9% higher in the Ad5-VEGF+Ad5-ANG group compared with the control group. Specific GFP signal was observed at distances of up to 5 mm in the rostral and caudal directions from the points of injection. A 1.5 to 2.0-fold increase in the number of GFAP + , S100β + , and AQP4 + cells was observed in the white and gray matter at a distance of up to 5 mm from the center of the lesion site in the caudal and rostral directions. At 30 days after injury, a 2-fold increase in S100β transcripts was observed in the Ad5-VEGF+Ad5-ANG group compared with the control group. CONCLUSIONS Intraspinal injection of recombinant adenoviral vectors encoding VEGF and ANG stimulates functional recovery after traumatic SCI. The increased number of S100β + astrocytes induced by this approach may be a beneficial factor for maintaining the survival and function of neurons. Therefore, gene therapy with Ad5-VEGF+Ad5-ANG vectors is an effective therapeutic method for SCI treatment.

Myelinated fibers and myelin-forming cells in the spinal cord at the L3-L5 level were studied in C57BL/6N mice that had spent 30 days in space. Signs of destruction of myelin in different areas of white matter, reduction of the thickness of myelin sheath and axon diameter, decreased number of myelin-forming cells were detected in "flight" mice. The stay of mice in space during 30 days had a negative impact on the structure of myelinated fibers and caused reduced expression of the markers myelin-forming cells. These findings can complement the pathogenetic picture of the development of hypogravity motor syndrome.

Study Design: Experimental study., Objective: Several neuro-degenerative disorders such as Alzheimer's dementia, Parkinson's disease and amyotrophic lateral sclerosis (ALS) are associated with genetic mutations, and replacing or disrupting defective sequences might offer therapeutic benefits. Single gene delivery has so far failed to achieve significant clinical improvements in humans, leading to the advent of co-expression of multiple therapeutic genes. Co-transfection using two or more individual constructs might inadvertently result in disproportionate delivery of the products into the cells. To prevent this, and in order to rule out interference among the many promoters with varying strength, expressing multiple proteins in equimolar amounts can be achieved by linking open reading frames under the control of only one promoter., Setting: Kazan, Russian Federation., Methods: Here we describe a strategy for adeno-viral co-expression of vascular endothelial growth factor (VEGF) and fibroblast growth factor 2 (FGF2) interconnected through picorna-viral 2A-amino-acid sequence in transfected human umbilical cord blood mono-nuclear cells (hUCB-MCs)., Results: Presence of both growth factors, as well as absence of immune response to 2A-antigen, was demonstrated after 28-52 days. Following injection of hUCB-MCs into ALS transgenic mice, co-expression of VEGF and FGF2, as well as viable xeno-transplanted cells, were observed in the spinal cord after 1 month., Conclusion: These results suggest that recombinant adeno-virus containing 2A-sequences could serve as a promising alternative in regenerative medicine for the delivery of therapeutic molecules to treat neurodegenerative diseases, such as ALS.

Study Design: Experimental study., Objective: To evaluate the treatment of spinal cord injury with glial cell-derived neurotrophic factor (GDNF) delivered using an adenoviral vector (AdV-GDNF group) in comparison with treatment performed using human umbilical cord blood mononuclear cells (UCB-MCs)-transduced with an adenoviral vector carrying the GDNF gene (UCB-MCs+AdV-GDNF group) in rat., Setting: Kazan, Russian Federation., Methods: We examined the efficacy of AdV-GDNF and UCB-MCs+AdV-GDNF therapy by conducting behavioral tests on the animals and morphometric studies on the spinal cord, performing immunofluorescence analyses on glial cells, investigating the survival and migration potential of UCB-MCs, and evaluating the expression of the recombinant GDNF gene., Results: At the 30th postoperative day, equal positive locomotor recovery was observed after both direct and cell-based GDNF therapy. However, after UCB-MCs-mediated GDNF therapy, the area of preserved tissue and the number of spared myelinated fibers were higher than those measured after direct GDNF gene therapy. Moreover, we observed distinct changes in the populations of glial cells; expression patterns of the specific markers for astrocytes (GFAP, S100B and AQP4), oligodendrocytes (PDGFαR and Cx47) and Schwann cells (P0) differed in various areas of the spinal cord of rats treated with AdV-GDNF and UCB-MCs+AdV-GDNF., Conclusion: The differences detected in the AdV-GDNF and UCB-MCs+AdV-GDNF groups could be partially explained by the action of UCB-MCs. We discuss the insufficiency and the advantages of these two methods of GDNF gene delivery into the spinal cord after traumatic injury.

Previously, we formulated the hypothesis of compartmentalized protein synthesis in axons of motor neurons. In the axon hillock, along the entire length of the axon and in its ending, specific proteins are locally synthesized, which ensure the function of each compartment. In support of this hypothesis, in this work we studied the local protein synthesis in mouse motor nerve ending.

Exposure to microgravity has been shown to result in damaging alterations to skeletal muscle, bones, and inner organs. In this study, we investigated the effects of microgravity by using a hindlimb unloading model (HUM) in mice. The characteristics of the lumbar spinal cords of HUM mice 30 days after hindlimb unloading were examined. Morphometric analysis showed reductions of the total area, gray matter, and white matter by 17%, 20%, and 12%, respectively. Myelinated fibers in the white matter showed prominent myelin destruction. Analysis of the number of glial fibrillary acidic protein (GFAP+)/S100 calcium-binding protein B (S100B-), GFAP+/S100B+, and GFAP-/S100B+ astrocytes in the ventral horn (VH), central channel area (CC), dorsal root entry zone (DREZ), main corticospinal tract (CST), and ventral funiculi (VF) showed that the number of GFAP+/S100B- astrocytes was increased in the DREZ and CST of HUM mice. Additionally, GFAP+/S100B+ cell numbers were significantly decreased in the VH and CST but did not differ in the CC or DREZ of HUM mice, as compared with the control. The numbers of GFAP-/S100B+ cells were significantly reduced only in the VH of HUM mice. Moreover, the number of ionized calcium-binding adaptor molecule 1 (Iba1+) microglia cells was significantly increased in the CC and DREZ of HUM mice. In control mice, homeobox protein HoxB8 (HoxB8+) cells were found only in the CC; in contrast, HoxB8+ cells were observed in all studied areas in HUM mice, with the greatest number found in the CC. Genome-wide transcriptome analysis of the lumbar spinal cords of HUM mice showed decreased expression of genes encoding myelin, extracellular matrix, cytoskeleton, and cell adhesion proteins. Real-time polymerase chain reaction (PCR) confirmed reductions in the expression of mpz, pmp2, pmp22, and prx genes, which are involved in myelination, as well as decreases in the levels of genes encoding extracellular matrix molecules, including glycoproteins (matrix gla protein (MGP), osteoglycin (OGN), microfibrillar associated protein 5 (MFAP), and collagen, type IV, alpha 1 (COL4A)), proteoglycans (perlecan (heparan sulfate proteoglycan) (HSPG)), and metalloproteinases (lysyl oxidase (LOX)). Thus, our results showed that hindlimb unloading caused decreases in gray and white matter areas, changes in gene expression, alterations in myelination, and phenotypic modifications in glial cells in the lumbar spinal cords of mice., (Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.)

Studies results of gravity unloading influence on spinal control system of muscle structure and functions are summarized. It was shown that demyelization of axons due to reduction of genes expression responsible for myelin proteins synthesis, decrease in one of the key enzymes of cholinergic system--cholineacetyltransferase activity, alteration of normal kinetics of quantal and non-quantal neurotransmitter secretion, impaired autoregulation of acetylcholine secretion from motor nerve endings through presynaptic cholinergic receptors, slowing of axonal transport of substances in motor neurons that innervate postural muscles played the important role in the development of hypogravitational motor symptoms. At the same time, the evidences of neuroprotective mechanisms enclosing (increase in heat shock proteins Hsp25 and Hsp70 expression), that hinder apoptosis development in motor neurons and glial cells in the spinal cord under conditions of model hypogravity, were revealed.

Amyotrophic lateral sclerosis is a neurodegenerative disease characterized by progressive death of cerebral and spinal motorneurons. Using behavioral tests we studied the efficiency of gene-cell therapy in SOD1 G93A transgenic mice receiving xenotransplantation of human umbilical cord blood mononuclear cells genetically modified with adenoviral vectors encoding vascular endothelial growth factor (VEGF) and reporter green fluorescent protein (EGFP) genes. The cells were transplanted to mice on week 27 of life (preclinical stage of the disease). Behavioral tests (open field, grip strength test) showed that transplantation of umbilical cord blood mononuclear cells expressing VEGF significantly improved the parameters of motor and explorative activity, grip strength, and animal survival. Thus, gene-cell therapy based on genetically modified mononuclear cells expressing VEGF can be efficient for the treatment of amyotrophic lateral sclerosis.

Background: Complete following existing guidelines for management of acute coronary syndrome (ACS) is known to be associated with better outcomes. Partly this is explained by lesser adherence to recommendations in high risk patients. Aim of our study was to assess relationship between degree of following current guidelines and in hospital outcomes independently from initial assessment of risk., Methods: Each key recommendation from guidelines issued between 2008 and 2011 (13 for STE ACS, 12 for NSTE ACS) was given weight of 1. Sum of these units constituted index of guideline adherence (IGA). IGA was retrospectively calculated for 1656 patients included in Russian independent ACS registry RECORD-2 (7 hospitals, duration 04.2009 to 04.2011). The patients were divided into 2 groups according to quartiles of IGA distribution: 1) low adherence group (quartiles I-II); 2) high adherence group (quartiles III-IV)., Results: In low adherence compared with high adherence group there were significantly more patients more or equal 65 years (=0.0007), with chronic heart failure [CHF] (<0.0001), previous stroke (<0.0001), atrial fibrillation [AF] (=0.0002), Killip class more or equal II (=0.0065), high risk of death by GRACE score (=0.035). Inhospital mortality was 9.3 and 2.4% in low and high adherence group, respectively (p<0.0001). The following independent predictors of inhospital death were identified: IGA quartiles I-II (odds ratio [OR] 4.0; 95% confidence interval [CI] 2.3-7.1; <0.0001), high GRACE score (OR 3.3; 95% CI 1.8-6.0; <0.0001), admission systolic BP less or equal 100 mm Hg (OR 3.1; 95% CI 1.8-5.4; <0.0001), admission serum glucose more or equal 8 mmol/l (OR 2.9; 95% CI 1.8-4.7; <0.0001), age more or equal 65 years (OR 2.3; 95% CI 1.3-4.0; =0.005), ST elevation more or equal 1 mm on first ECG (OR 1.7; 95% CI 1.1-2.5; =0.013). From groups with low and high adherence to guidelines we selected pairs of patients (n=588) with similar (or close) age, type of ACS, GRACE score, Killip class, presence of other important risk factors (CHF, AF, previous stroke), and formed 2 equal subgroups without significant differences in important demographic, anamnestic, clinical and laboratory data. Hospital mortality was 7.8 and 2.7% in low and high adherence subgroup, respectively (p<0.0001)., Conclusions: In RECORD-2 ACS registry low adherence to guidelines was more frequent among high risk patients and was independent predictor of inhospital death. Association between degree of guidelines adherence and outcomes persisted after equalizing groups by some factors of risk of mortality.

Positive changes are declared to occur during recent years in management of hospitalized patients with acute coronary syndromes (ACS) in Russia. Most of these changes are related to availability of invasive treatment. But considerable portion of patients (pts) are still treated in hospitals without facilities for invasive myocardial revascularization (noninvasive hospitals - NIHs). Aim of this study was to compare some characteristics of management of ACS in NIHs which participated in ACS registries RECORD (2007-2008, 8 NIHs from 6 cities; n=381) and RECORD-2 (2009-2011, 3 NIHs from 3 cities, n=680). Results. Groups of pts recruited in these NIHs had similar mean age and portion of women (67.6 and 66.5 years, 51.1 and 53.1 % in RECORD-2 and RECORD, respectively, p=0.64). Time from symptoms onset to hospitalization was shorter in RECORD-2 (3.2 vs 4.1 hours for ST-elevation [STE], =0.03; 4.0 vs 6.5 hours for non ST elevation [NSTE] ACS, <0.0001). Among RECORD-2 NSTEACS pts more had ST depressions (50.6 vs 28.7%, <0.0001), high risk of death according to GRACE score (39.1 vs 20.9 %, <0.0001), but less Killip class >II (15.0 vs 21.6%, p=0.025). No such differences existed among STEACS pts. Thrombolysis was more often used in RECORD-2 (62.6 vs 34.1%, <0.0001). Both STEACS and NSTEACS RECORD-2 pts more often received clopidogrel (63.5 vs 18.8%, p<0.0001, and 41.6 vs 11.1%, <0.0001, respectively). More NSTEACS RECORD-2 pts were given parenteral anticoagulants (93.4 vs 80.4%, <0.0001), low molecular weight heparins (23.4 vs. 3.4%, <0.0001) and fondaparinux (10.4 vs 0.7%, <0.0001), but still in 20% of NSTEACS RECORD-2 pts unfractionated heparin was given subcutaneously. Twenty RECORD-2 pts (2.9%) were transferred to invasive hospital but none during first 24 hours. There were no significant differences between registers in hospital mortality (20.0 vs 21.2%, =0.84; 4.2 vs 2.7%, =0.24 in STE and NSTE ACS pts of RECORD-2 and RECORD, respectively). Conclusions. Despite some improvement in management of pts occurring in 2-3 years NIHs mortality in STEACS remained very high. Numerically higher mortality in NSTEACS could be partially attributed to higher risk of RECORD-2 pts.

When modeling effects of hypogravitation by the method of hindlimb unloading in rats the area of cross-section in lumbar part of a spinal cord was found to reduce. The analysis of spinal cord slides showed that these changes are associated with a decrease in the area of white substance of a spinal cord. Data obtained are consistent with our previous observation of a decrease in expression of the genes encoding myelin proteins. Results of our researches give the good reasons to believe that miyelinization failure in CNS is one of the factors that underlie the development of hypogravitational motor syndrome.

Due to the constant formation of the antibiotic resistance of pathogens the search for alternative and synergistic ways antibiotic prophylaxis and treatment of infectious complications of wounds becomes current. This article is devoted to various aspects of the use of bacteriophages. The historical background of these drugs, and the biological properties are given. The authors assessed the capabilities of phage preparations for treatment and prevention of infectious complications of wounds. It is noted that for these purposes most suitable preparations containing several phages active against major pathogens of surgical infections. The authors assessed the properties piobacteriofag polyvalent "Sekstafag" in vitro and its efficacy in clinical practice. Based on analysis of published data and own investigations we may conclude that the of using piobacteriofag for the prevention and treatment of infectious complications of wounds may be perspective.

The spontaneous quantum secretion of neurotransmitter and its regulation through the system of presynaptic acetylcholine receptors have been studied on a neuromuscular preparation of rat m. soleus of intact animals and animals in which the axonal transport was blocked via the application of colchicine to the sciatic nerve. It was shown that, after six days of colchicine application, the spontaneous quantum secretion, the reaction of presynaptic membrane, and the reaction of neurosecretory apparatus to the depolarization of nerve endings via increase of the content of potassium ions in the environment and to the activation of presynaptic receptors by carbachol are not disturbed. Keeping in mind a rather short half-life of proteins that take part in the exocytosis and its regulation, it may be concluded that their functioning does not depend on the state of the axonal transport. These data correspond to the hypothesis put forward earlier that the synthesis of some proteins performing their function in nerve terminals occurs directly at the site of their utilization but not in the perikaryon, as it has been traditionally assumed.

A trypsin inhibitor, isolated from whole-wheat grain (Triticum aestivum L.) by the method of bio-specific chromatography on trypsin-Sepharose, was potent in inhibiting human salivary alpha-amylase. The bi-functional alpha-amylase/trypsin inhibitor was characterized by a narrow specificity for other alpha-amylases and proteinases. The high thermostability of the inhibitor was lost in the presence of SH group-reducing agents. The inhibitor-trypsin complex retained its activity against alpha-amylase. The inhibitor-alpha-amylase complex was active against trypsin. Studies of the enzyme kinetics demonstrated that the inhibition of alpha-amylase and trypsin was noncompetitive. Our results suggest the existence of two independent active sites responsible for the interaction with the enzymes.

Until now, in the world since literature, there has been no direct evidence indicating that RNA-interference controls local protein synthesis in the mammalian motor neuron axons. In the present review we have summarized the results on intraaxonal protein synthesis, its role in the axonal transport, and mechanisms regulating local protein synthesis in the axoplasm. The new mechanisms regulating axonal transport based on RNA-interference presented in the review let us discuss the questions about pathogenesis of the neurodegenerative diseases. The estimated role of the intraaxonal protein synthesis on axonal transport suggested applying short interfering RNA for degradation of the mutant gene RNA for blocking synthesis of the aberrant protein along the whole axon.

The results of axonal regeneration in peripheral nerves and of the modulating effect of estrogens on the posttraumatic plasticity of motor neurons through the classical and alternative genomic mechanisms involving the activation of estrogen receptors ERalpha and ERbeta and increased transcription of specific neuroprotective control genes. New mechanisms of the neuroprotective effect of estrogens related to the cross interaction of estrogens with intracellular signaling cascades of many other biologically active compounds, in particular, vascular endothelial growth factor and endothelin, are proposed.

Expression of slow myosin in fast lumbrical muscle during unilateral damage to the sciatic nerve was studied in rats. Four weeks after unilateral excision of sciatic nerve segment or its crushing, the content of slow fiber in the lumbrical muscle of the contralateral leg tended to increase (detected using monoclonal antibodies against slow myosin). The extrapyramidal system seems to modulate the phenotype of muscle fibers via motoneurons of the spinal cord, while the compensatory activation of this system manifests in pronounced "deceleration" of both the fast and slow muscles in the hind legs.

According to one of the doctrines of neuroscience, protein synthesis in neuronal cells takes place only in the perikaryon and dendrites. The first indirect evidence on the possibility of protein synthesis in the axon appeared a long time ago, but convincing proof of protein synthesis in that compartment of the neuron was obtained only recently. Axoplasm contains numerous cytoskeletal and cytosolic proteins which are carried by slow axonal transport. Axonal length may vary from several micrometers to one meter or more. Thus, the transport of protein molecules along the axon may take days, weeks and even months. In urgent response of the neuron to the changes in the conditions of its functioning, the local protein synthesis may save the time and energy required for transportation of newly synthesized proteins. Moreover, intraaxonal protein synthesis is essential in order to maintain the protein contents of the axoplasm, since all the proteins possess limited half-life and undergo biological degradation during their slow transport in the axon. This paper reviews these problems and examines the significance of the local protein synthesis in the axon for realization of specific functions of the neuron.

The aim of this study was to examine the effects of LY117018, a selective estrogen receptor modulator, on peripheral nerve regeneration, using a model of sciatic nerve crush injury in mice. Sciatic functional index, an index of functional recovery, was significantly higher in LY117018 treated mice throughout regeneration. Analysis of semi-thin sections revealed a significant increase in both the total number of regenerating nerve fibers at day 7, and the mean axonal area of myelinated fibers at 7, 14, and 21 days after injury, in LY117018 treated mice. Analysis of axonal transport through retrograde labeling of motor neurons showed that LY117018 increased transport, and ICI 182,780 blocked the effects of LY117018, delineating estrogen receptors as its target. Our study suggests that LY117018 may markedly accelerate peripheral nerve regeneration and functional recovery through activation of estrogen receptors.

A physical self-consistent model is developed to explain single spots or complex current structures at the anode of elevated-pressure parallel-plate dc discharges. The model is based on a fluid description of electron and ion transport coupled with Poisson's equation and involves a pair of coupled reaction-diffusion equations of an activator-inhibitor-type. This system of one-dimensional equations containing no phenomenological (adjustable) parameters allows one to find the current-density (activator) and anode potential drop (inhibitor) distributions on the anode surface. In a certain range of supply voltage, an anode glow stratification, resulting in the formation of separate glowing regions, takes place. However, the growth of perturbations and formation of a spatially periodic current pattern are complicated by competition between the current stripes, leading to suppressing of the neighboring current stripes. The bifurcation behavior of the model with respect to the characteristic electron energy, recombination coefficient, and discharge gap has been analyzed. The properties of a single anode current structure, including the normal current density effect, have been investigated. The application of these results to available findings in experiments and two-dimensional numerical simulations is discussed.

Immunohistochemical study with monoclonal antibodies against fast myosin heavy chains showed that excision of a fragment of sciatic nerve does not change, while compression of the nerve decreased the relative content of fast muscle fibers in rat soleus muscle. The content of fast fibers in the muscles of contralateral limbs decreased in both models of denervation. Hence, contralateral limbs cannot be used as the control in experiments with disturbed neurotrophic function. Possible mechanisms underlying changes in the immunohistochemical profiles of experimental and contralateral limbs are discussed.

On the model of allotransplantation of m. lumbricalis, previously subjected to acute total ischemia at temperature 37-38 degrees it was shown that morphological integrity and characteristic immunohistochemical properties of the muscle remain only during two hours of the thermic ischemia. A week later surviving muscular fibres (MF) and newly forming muscular tubules (MT) were found in the transplants of the muscle. Processing with antibodies of the fast twitch type myosin filaments revealed both fast- and slow-twitch types of fibres among MF, while B- and C-typed MF were identified at defining the activity of succinat dehydrogenase (SDG). High SDG activity, corresponding to C-type fibres was observed in MT. Staining with antibodies revealed fast-twitch type myosin. After 3-hour ischemia of the muscles, few surviving MF were found, while after the 4-6-hours one only MT appeared to be present in the transplants. Thus, refering to our data, obtained in the previous works and to the results of the research it is ascertained that there is a regularity, allowing to predict critical terms of development of irreversal ischemic disturbances in the muscle of severed extremity, depending on the ischemia duration and temperature.

Membrane potential (MP) of the diaphragm muscle fibers of the rats was registered in the different periods of ischemia in vitro. The ability to normalization of MP up to 6 h of ischemia and an irreversible decrease of MP after 6.5 h of ischemia was revealed. It is concluded that the ability of diaphragm muscle fibers to restore their MP lost beginning from 6 h of artificial ischemia.

After transferrin injection to the slow m. soleus of guinea pig no immunohistochemical changes in myosin composition with monoclonal antibodies (AB) against fast myosin heavy chain (HCf) have been found. All muscles fibers in intact and experimental animals were identified as slow ones (type I of muscle fibers). After colchicine treatment of nerve trunk innervating slow muscle, some muscle fibers reacting with monoclonal AB against myosin HCf have been found. However, after colchicine treatment of nerve trunk and transferrin injection also some muscle fibers in slow muscle reacting with monoclonal AB against myosin HCf have been found. In appears that transferrin probably can not be the factor of neurotrophic control for myosin composition in skeletal muscle.

Polyclonal antibodies (PCAB) to smooth muscle myosin (SMM), monoclonal antibodies (MCAB) to cytokeratin 8 (clon HI, IgGI) and H4 (IgM), as well as PCAB to carbonic anhydrase III were used for identification of the cell types in the vas deferens cell culture of guinea pig. Smooth muscle cells (SMC) are identified by intensive staining of PCAB to SMM. Fibroblast-like cells (FBL) are determined by the presence of the filament finest network, apparently responding to the myosin non-muscular forms, which are present in PCAB to SMM. The epithelial cells are stained by MCAB to cytokeratins. PCAB to carbonic anhydrase III interact with all three cell types. In the majority of SMC the enzyme is detected as solitary stripes, though there are diffuse ones across the whole cytoplasms, the nucleus remains clearly visible. Carbonic anhydrase III in epithelial cells is detected only in nucleoli and along nucleus membrane while in FBL--in nucleoli and cytoplasm as focal granulation. PCAB to carbonic anhydrase III may serve as a universal marker for identification of cell type in the guinea pig vas deferens cell culture.

The immunohistochemical profile of intact and denervated soleus muscle of guinea pigs after sensibilization was studied. It is shown, that intact soleus muscle consists of slow fibers, which have low ATP-ase activity and don't react with monoclonal antibodies against fast myosin heavy chain. No changes of immunohistochemical profile were found after denervation or sensibilization. At the same time, the fibers, reacting with monoclonal antibodies against fast myosin heavy chain and having low ATP-ase activity, were found in denervated muscles after sensibilization. It is concluded, that the synthesis of fast myosin is induced after sensibilization of denervated muscles. Validity of myosin ATP-ase histochemistry for muscle fibers typing is discussed.

The influence of hypothermia on the development of the ischemic disorders was studied using allotransplantation of the rat skeletal muscle (m. lumbricalis) to the anterior chamber of the eye after different period of ischemia. The morphological and immunohistochemical (monoclonal antibodies to heavy chain of the fast myosin, PAP-method) data were found confirming that hypothermia (2-4 degrees C) prolongs the period of the ischemic disorders first appearance by 5 h (from 6 to 11 h) if compared with development of ischemia in the muscle at 21-23 degrees C.

The ability of muscle fibers to survive the ischemic condition was examined. After 0 to 11 h of ischemia of the rat m. lumbricalis was transplanted to the anterior chamber of the rats eye. Using a morphological and immunohistochemical (monoclonal antibodies to heavy chain of the fast myosin, PAP-method) methods it was revealed that population of surviving muscle fibers reduced significantly beginning from 6 h of ischemia. After 9 h of ischemia there were no muscle fibers in the transplant but one could observe the new formed muscle tubules. It is concluded that up to 6 h the muscle fibers of m. lumbricalis ischemia still preserve the ability to recover.

Carbonic anhydrase III (CAIII) is the isoenzyme purified from the human skeletal slow muscle immunohistochemically revealed in smooth muscle cells of uterus, myoepithelial cells of salivary, lactiferous and prostatic glands of man. Immunohistochemical determination of CAIII is a useful approach to correct the identification of myoepithelial cells in human salivary glands.