Combined treatment with ribavirin and tiazofurin attenuates response of glial cells in experimental autoimmune encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) is an animal model of multiple sclerosis (MS), a human inflammatory and demyelinating disease. Microglia and astrocytes are glial cells of the central nervous system (CNS) that play a dual role in MS and EAE pathology. The aim of this study was to examine the effect of combined treatment with two nucleoside analogues, ribavirin and tiazofurin, on microglia and astrocytes in actively induced EAE. Therapeutic treatment with a combination of these two nucleoside analogues reduced disease severity, mononuclear cell infiltration and demyelination. The obtained histological results indicate that ribavirin and tiazofurin changed activated microglia into an inactive type and attenuated astrocyte reactivity at the end of the treatment period. Since reduction of reactive microgliosis and astrogliosis correlated with EAE suppression, the present study also suggests that the obtained beneficial effect of ribavirin and tiazofurin could be a consequence of their action inside as well as outside the CNS. [Acknowledgments. This work was supported by the Serbian Ministry of Education and Science, Project No: III41014.

Pattern of chondroitin sulfate proteoglycan expression after ablation of the sensorimotor cortex of the neonatal and adult rat brain

The central nervous system has a limited capacity for self-repair after damage. However, the neonatal brain has agreater capacity for recovery than the adult brain. These differences in the regenerative capability depend on local environmental factors and the maturational stage of growing axons. Among molecules which have both growth-promoting and growth-inhibiting activities is the heterogeneous class of chondroitin sulfate proteoglycans (CSPGs). In this paper, we investigated the chondroitin-4 and chondroitin-6 sulfate proteoglycan expression profile after left sensorimotor cortex ablation of the neonatal and adult rat brain. Immunohistochemical analysis revealed that compared to the normal uninjured cortex, lesion provoked up regulation of CSPGs showing a different pattern of expression in the neonatal vs. the adult brain. Punctuate and membrane-bound labeling was predominate after neonatal lesion, where as heavy deposition of staining in the extracellular matrix was observed after adult lesion. Heavy deposition of CSPG immunoreactivity around the lesionsite in adult rats, in contrast to a less CSPG-rich environment in neonatal rats, indicated that enhancement of the recovery process after neonatal injury is due to amore permissive environment

Combined treatment with ribavirin and tiazofurin attenuates response of glial cells in experimental autoimmune encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) is an animal model of multiple sclerosis (MS), a human inflammatory and demyelinating disease. Microglia and astrocytes are glial cells of the central nervous system (CNS) that play a dual role in MS and EAE pathology. The aim of this study was to examine the effect of combined treatment with two nucleoside analogues, ribavirin and tiazofurin, on microglia and astrocytes in actively induced EAE. Therapeutic treatment with a combination of these two nucleoside analogues reduced disease severity, mononuclear cell infiltration and demyelination. The obtained histological results indicate that ribavirin and tiazofurin changed activated microglia into an inactive type and attenuated astrocyte reactivity at the end of the treatment period. Since reduction of reactive microgliosis and astrogliosis correlated with EAE suppression, the present study also suggests that the obtained beneficial effect of ribavirin and tiazofurin could be a consequence of their action inside as well as outside the CNS.Projekat ministarstva br. III4101

Animal models of multiple sclerosis: Focus on experimental autoimmune encephalomyelitis.

Multiple sclerosis (MS) is a chronic, progressive disorder of the central nervous system (CNS) that affects more than two million people worldwide. Several animal models resemble MS pathology; the most employed are experimental autoimmune encephalomyelitis (EAE) and toxin- and/or virus-induced demyelination. In this review we will summarize our knowledge on the utility of different animal models in MS research. Although animal models cannot replicate the complexity and heterogeneity of the MS pathology, they have proved to be useful for the development of several drugs approved for treatment of MS patients. This review focuses on EAE because it represents both clinical and pathological features of MS. During the past decades, EAE has been effective in illuminating various pathological processes that occur during MS, including inflammation, CNS penetration, demyelination, axonopathy, and neuron loss mediated by immune cells."This is the peer reviewed version of the following article: Bjelobaba I, Begovic-Kupresanin V, Pekovic S, Lavrnja I. Animal models of multiple sclerosis: Focus on experimental autoimmune encephalomyelitis. J Neurosci Res. 2018, which has been published in final form at [https://doi.org/10.1002/jnr.24224]. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.

Pattern of chondroitin sulfate proteoglycan expression after ablation of the sensorimotor cortex of the neonatal and adult rat brain

The central nervous system has a limited capacity for self-repair after damage. However, the neonatal brain has agreater capacity for recovery than the adult brain. These differences in the regenerative capability depend on local environmental factors and the maturational stage of growing axons. Among molecules which have both growth-promoting and growth-inhibiting activities is the heterogeneous class of chondroitin sulfate proteoglycans (CSPGs). In this paper, we investigated the chondroitin-4 and chondroitin-6 sulfate proteoglycan expression profile after left sensorimotor cortex ablation of the neonatal and adult rat brain. Immunohistochemical analysis revealed that compared to the normal uninjured cortex, lesion provoked up regulation of CSPGs showing a different pattern of expression in the neonatal vs. the adult brain. Punctuate and membrane-bound labeling was predominate after neonatal lesion, where as heavy deposition of staining in the extracellular matrix was observed after adult lesion. Heavy deposition of CSPG immunoreactivity around the lesionsite in adult rats, in contrast to a less CSPG-rich environment in neonatal rats, indicated that enhancement of the recovery process after neonatal injury is due to amore permissive environment.Centralni nervni sistem ima ograničen kapacitet za oporavak nakon povrede. Međutim, neonatalni mozak pokazuje veću sposobnost oporavka u odnosu na odrasle. Ove razlike zavise od lokalnih sredinskih faktora i stepena zrelosti aksona tokom izrastanja. U grupu molekula koji mogu da stimulišu ili inhibiraju rast aksona spada i heterogena klasa molekula označena kao hondroitin sulfatni proteoglikani (CSPG). U ovom radu ispitivanje profil ekspresije hondroitin-4 i hondroitin-6 sulfatnih proteoglikana nakonlezije leve senzomotorne kore neonatalnog i adultnog mozga pacova. Imunohistohemijska analiza pokazuje da u odnosu na normalni,nepovređeni korteks, lezija dovodi do povećanja ekspresije CSPG koji ima različiti obrazac promena u neonatalnom u odnosu na adultni mozak. Nakonlezije kod mladih, predominiraju tačkasta i membranski-vezana forma, dok kod odraslih lezija dovodi do nagomilavanja CSPG u ekstra ćelijskom matriksu. Prohodnija sredina u mozgu neonatalnih pacova koja je siromašnija CSPG, preduslov je boljeg procesa oporavka u odnosu na adulte, kod kojih se nakon povrede CSPG nagomilavaju oko mesta lezije.Projekat ministarstva br. 143005

Expression profiles of cholesterol metabolism-related genes are altered during development of experimental autoimmune encephalomyelitis in the rat spinal cord

Increased evidence suggests that dysregulation of cholesterol metabolism may be a key event contributing to progression of multiple sclerosis (MS). Using an experimental autoimmune encephalomyelitis (EAE) model of MS we revealed specific changes in the mRNA and protein expression of key molecules involved in the maintaining of cholesterol homeostasis in the rat spinal cord: 3-hydroxy-3-methylglutaryl-coenzyme-A reductase (HMGCR), apolipoprotein E (ApoE) and cholesterol 24-hydroxylase (CYP46A1) during the course of disease. The presence of myelin lipid debris was seen only at the peak of EAE in demyelination loci being efficiently removed during the recovery period. Since CYP46A1 is responsible for removal of cholesterol excess, we performed a detailed profiling of CYP46A1 expression and revealed regional and temporal specificities in its distribution. Double immunofluorescence staining demonstrated CYP46A1 localization with neurons, infiltrated macrophages, microglia and astrocytes in the areas of demyelination, suggesting that these cells play a role in cholesterol turnover in EAE. We propose that alterations in the regulation of cholesterol metabolism at the onset and peak of EAE may add to the progression of disease, while during the recovery period may have beneficial effects contributing to the regeneration of myelin sheath and restoration of neuronal function.Scientific Reports (2017), 7(1): 270

Theta burst stimulation influence the expression of BDNF in the spinal cord on the experimental autoimmune encephalomyelitis.

Repetitive transcranial magnetic stimulation (rTMS) induces changes in expression of proteins engaged in the activity of excitatory and inhibitory systems, restores these functions and suppresses the progression of disability in experimental autoimmune encephalitis (EAE). The structural type of TMS, the arrangement as theta burst stimulation (TBS) has been applied as intermittent TBS (iTBS) and continuous TBS (cTBS) protocols to female adult DA rats. The animals were randomly divided into experimental groups: control group (C), group treated with complete Freund's adjuvant (CFA), experimental autoimmune encephalomyelitis (EAE) group, group treated with iTBS post EAE immunization (EAE + iTBS), group treated with cTBS post EAE immunization (EAE + cTBS), group of healthy animals treated with iTBS or cTBS. Therapeutic protocols of iTBS or cTBS in all EAE groups of animals were performed starting from 14 days post immunization (dpi), for 10 days with time point decapitation at 24 dpi. After decapitation, spinal cords were analysed for BDNF and Ki67 expression. The results revealed reduced BDNF expression in the rat's spinal cord of EAE animals in the stage of remission, which was associated with increased Ki67 and GFAP expressions. Decreased Iba 1 and BDNF expression, contrary to increased Iba 1 and Ki67 expression, suggests clustered microglia in the resolution phase of EAE. Enhanced GABA expression in spinal cord sections indicates higher GABA metabolic turnover, and also GAD activity in astrocytes, or prominent activity of GABAergic neurons. Both TBS protocols induced advance BDNF expression; amongst iTBS application provoked elevating of BDNF and stabilizing of GFAP and Ki67 expressions

Therapeutic effect of nucleoside analogs on experimental autoimmune encephalomyelitis in dark agouti rats

Experimental autoimmune encephalomyelitis (EAE) is a commonly used animal model of the human neurological disorder multiple sclerosis. The purpose of the present study was to investigate the effect of combined treatment with two nucleoside analogs, ribavirin and tiazofurin, on development of EAE actively induced in highly susceptible dark agouti rats. The obtained results showed that ribavirin and tiazofurin applied either separately or in combination from the onset of the firstsymptoms of EAE after its induction (therapeutic treatment) significantly suppressed EAE’s clinical symptoms. However, the most pronounced effect was gained with combined treatment, probably as a result of synergistic/additive action

Cell death of spinal cord ED1(+) cells in a rat model of multiple sclerosis

Infiltration of macrophages into the central nervous system and activation of microglia are hallmarks of multiple sclerosis and its animal model-experimental autoimmune encephalomyelitis (EAE). Cell death in EAE has been demonstrated as an essential mechanism in the local regulation of the inflammatory reaction, but also as one of the major factors contributing to the destruction of the nervous tissue. The focus of this study was on detection of cell death among ED1(+) cells (macrophages/activated microglia) in the spinal cord of Dark Agouti rats at the peak of EAE. Cell death was assessed using the TUNEL assay and immunostaining for cleaved caspase 3, as markers for cell death in general and ``classical{''} apoptosis, respectively. Major infiltrates of immune cells were detected both in white matter and gray matter of spinal cords in rats at the disease peak. ED1, TUNEL, and caspase 3 positive cells were detected within, but also outside the infiltrates. There were more dying ED1+ cells in white matter than in gray matter, both in the general population and in infiltrated regions. The observed discrepancy in the proportion of dying ED1+ cells in spinal cord gray and white matter indicated that in EAE rat macrophages/microglia within gray matter are less prone to cell death induction. This is of special interest in the context of the increasingly appreciated contribution of spinal cord gray matter inflammation to multiple sclerosis pathogenesis. Our findings suggest that activated macrophages/microglia of gray matter are less susceptible to cell death induction. Alternatively, it can be assumed that intrinsic cell death-inductive mechanisms of nervous tissue differ in white and gray matter. Thus, further research on the gray matter macrophages/microglia cell death during EAE is warranted. They should be aimed at identification of the reasons for the observed differences and finding suitable ways to stimulate gray matter activated macrophages/microglia death.Deutsche Forschungsgemeinschaft {[}DFG TR 1238/2-1, DFG PA1751/4-1]; Ministry of Education, Science and Technological Development, Republic of Serbia {[}OI173035, OI173013]; Kerstan Foundation; Alcon Research Institute; Open Access Publishing Fund of University of Tubinge

Brain cortical injury induces changes in peripheral lymphocyte ectonucleotidase activities

Injury and other pathological conditions induce a massive release of ATP and ADP that initiate an immune response. Extracellular nucleotides are degraded by ectonucleotidases: enzymes from E-NTPDase and E-NPP families sequentially hydrolyze ATP and ADP to AMP, which is further hydrolyzed by ecto-5'-nucleotidase to adenosine that exerts suppressive effects on immune cells. We investigated the ectonucleotidase activities of peripheral lymphocytes at different post-injury times after an unilateral brain injury in the rat. Significant and dynamic changes in the lymphocytic ectonucleotidase activities were obtained. ATP- and ADP-hydrolysis changes, together with their calculated ratios, indicate the major contribution of E-NTPDase 1 and its comparable upregulation between sham operation and injury. AMP hydrolysis changes were more brain-injury specific, with a longer-lasting lymphocytic response induced by cortical stab injury (CSI). In summary, CSI and sham operation induce the upregulation of the whole enzyme chain for adenine nucleotide hydrolysis in lymphocytes, suggesting an important roles of ectonucleotidases in the course of recovery after brain injury.Projekat ministarstva br. III4101