Nathali Kaushansky - Academia.edu (original) (raw)

Papers by Nathali Kaushansky

Research paper thumbnail of Cannabidiol effects on mRNA levels and signaling pathways in MOG-35–55 activated encephalitogenic T cells

Journal of Neuroimmunology, 2014

Research paper thumbnail of Cannabinoids Decrease the Th17 Inflammatory Autoimmune Phenotype

Journal of Neuroimmune Pharmacology, 2013

Cannabinoids, the Cannabis constituents, are known to possess anti-inflammatory properties but th... more Cannabinoids, the Cannabis constituents, are known to possess anti-inflammatory properties but the mechanisms involved are not understood. Here we show that the main psychoactive cannabinoid, Δ-9-tetrahydrocannabinol (THC), and the main nonpsychoactive cannabinoid, cannabidiol (CBD), markedly reduce the Th17 phenotype which is known to be increased in inflammatory autoimmune pathologies such as Multiple Sclerosis. We found that reactivation by MOG35-55 of MOG35-55-specific encephalitogenic T cells (cells that induce Experimental Autoimmune Encephalitis when injected to mice) in the presence of spleen derived antigen presenting cells led to a large increase in IL-17 production and secretion. In addition, we found that the cannabinoids CBD and THC dose-dependently (at 0.1-5 μM) suppressed the production and secretion of this cytokine. Moreover, the mRNA and protein of IL-6, a key factor in Th17 induction, were also decreased. Pretreatment with CBD also resulted in increased levels of the anti-inflammatory cytokine IL-10. Interestingly, CBD and THC did not affect the levels of TNFα and IFNγ. The downregulation of IL-17 secretion by these cannabinoids does not seem to involve the CB1, CB2, PPARγ, 5-HT 1A or TRPV1 receptors. In conclusion, the results show a unique cannabinoid modulation of the autoimmune cytokine milieu combining suppression of the pathogenic IL-17 and IL-6 cytokines along with boosting the expression of the anti-inflammatory cytokine IL-10.

Research paper thumbnail of Aggrecan Turnover in Human Articular Cartilage: Use of Aspartic Acid Racemization as a Marker of Molecular Age

Proteoglycans are major constituents of the extracel-Aggrecan is a key component of the cartilage... more Proteoglycans are major constituents of the extracel-Aggrecan is a key component of the cartilage matrix. lular matrix of articular cartilage and are largely re-During aging, many changes occur in its composition sponsible for the high resistance to compression of this and structure; in particular, there is an increase in the load-bearing tissue. This resistance to compression is proportion of lower molecular weight monomers and mainly due to the high osmotic pressure of the nega-of the ''free'' binding region. An important question tively charged glycosaminoglycan (GAG) 2 chains. The has been whether these changes represent alterations proteoglycans (aggrecan) are organized in the matrix in biosynthesis or whether they are due to the accumu-through a highly specific interaction to form large ag-lation with age of the partially degraded fragments gregates consisting of a central filament of hyaluronan, of the originally synthesized large monomer. In the to which aggrecan monomers are attached via the hya-present work we have used an independent tool, viz., luronan-binding domain (G1) of their core proteins, the extent of racemization of aspartic acid to study the with the interaction being stabilized by the presence molecular ''age'' of different buoyant density fractions of a link protein. Aggregate formation is very important of the aggrecan of human articular cartilage, as well from the physiological point of view, since it ensures as of isolated free binding region and link protein. By the retention of aggrecan within the collagen network. measuring the D/L Asp ratio of the different aggrecan

Research paper thumbnail of Post-CNS-inflammation expression of CXCL12 promotes the endogenous myelin/ neuronal repair capacity following spontaneous recovery from multiple sclerosis-like disease

Background: Demyelination and axonal degeneration, hallmarks of multiple sclerosis (MS), are asso... more Background: Demyelination and axonal degeneration, hallmarks of multiple sclerosis (MS), are associated with the central nervous system (CNS) inflammation facilitated by C-X-C motif chemokine 12 (CXCL12) chemokine. Both in MS and in experimental autoimmune encephalomyelitis (EAE), the deleterious CNS inflammation has been associated with upregulation of CXCL12 expression in the CNS. We investigated the expression dynamics of CXCL12 in the CNS with progression of clinical EAE and following spontaneous recovery, with a focus on CXCL12 expression in the hippocampal neurogenic dentate gyrus (DG) and in the corpus callosum (CC) of spontaneously recovered mice, and its potential role in promoting the endogenous myelin/neuronal repair capacity.

Research paper thumbnail of HU-446 and HU-465, derivatives of the non-psychoactive cannabinoid cannabidiol, decrease the activation of encephalitogenic T cells

Chemical biology & drug design, Jan 11, 2015

Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease ... more Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease peripheral inflammation and neuroinflammation in mouse experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). Here we have studied the anti-inflammatory effects of newly synthesized derivatives of natural (-)-CBD ((-)-8,9-dihydro-7-hydroxy-CBD; HU-446) and of synthetic (+)-CBD ((+)-8,9-dihydro-7-hydroxy-CBD; HU-465), on activated myelin oligodendrocyte glycoprotein (MOG)35-55 specific mouse encephalitogenic T cells (TMOG ) driving EAE/MS-like pathologies. Binding assays followed by molecular modelling revealed that HU-446 has negligible affinity towards the cannabinoid CB1 and CB2 receptors while HU-465 binds to both CB1 and CB2 receptors at the high nanomolar concentrations (Ki =76.7 ± 5.8 nM and 12.1 ± 2.3 nM, respectively). Both, HU-446 and HU-465, at 5 and 10 μM (but not at 0.1 and 1 μM) inhibited the MOG35-55-induced proliferation of autoreactive TMOG cell...

Research paper thumbnail of Cannabidiol inhibits pathogenic T cells, decreases spinal microglial activation and ameliorates multiple sclerosis-like disease in C57BL/6 mice

British Journal of Pharmacology, 2011

Cannabis extracts and several cannabinoids have been shown to exert broad anti-inflammatory activ... more Cannabis extracts and several cannabinoids have been shown to exert broad anti-inflammatory activities in experimental models of inflammatory CNS degenerative diseases. Clinical use of many cannabinoids is limited by their psychotropic effects. However, phytocannabinoids like cannabidiol (CBD), devoid of psychoactive activity, are, potentially, safe and effective alternatives for alleviating neuroinflammation and neurodegeneration.

Research paper thumbnail of A highly conserved sequence associated with the HIV gp41 loop region is an immunomodulator of antigen-specific T cells in mice

Research paper thumbnail of An Immunomodulating Motif of the HIV-1 Fusion Protein Is Chirality-independent: IMPLICATIONS FOR ITS MODE OF ACTION

Journal of Biological Chemistry, 2013

Recently, an immunosuppressive motif was identified in the HIV-1 envelope glycoprotein complex. R... more Recently, an immunosuppressive motif was identified in the HIV-1 envelope glycoprotein complex. Results: Both D-and L-stereoisomers of the motif inhibit T-cell receptor activation and preferentially bind T-cells over B-cells.

Research paper thumbnail of Highlight in Nature Immuno (1)

Research paper thumbnail of Cannabidiol, a non-psychoactive cannabinoid, leads to EGR2-dependent anergy in activated encephalitogenic T cells

Background: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown b... more Background: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown by us to ameliorate clinical symptoms and to decrease inflammation in myelin oligodendrocyte glycoprotein (MOG)35-55-induced mouse experimental autoimmune encephalomyelitis model of multiple sclerosis as well as to decrease MOG35-55-induced T cell proliferation and IL-17 secretion. However, the mechanisms of CBD anti-inflammatory activities are unclear. Methods: Here we analyzed the effects of CBD on splenocytes (source of accessory T cells and antigen presenting cells (APC)) co-cultured with MOG35-55-specific T cells (T MOG) and stimulated with MOG35-55. Using flow cytometry, we evaluated the expression of surface activation markers and inhibitory molecules on T cells and B cells. T MOG cells were purified using CD4 positive microbead selection and submitted for quantitative PCR and microarray of mRNA transcript analyzes. Cell signaling studies in purified T MOG were carried out using immunoblotting. Results: We found that CBD leads to upregulation of CD69 and lymphocyte-activation gene 3 (LAG3) regulatory molecules on CD4 + CD25 − accessory T cells. This subtype of CD4 + CD25 − CD69 + LAG3 + T cells has been recognized as induced regulatory phenotype promoting anergy in activated T cells. Indeed, we observed that CBD treatment results in upregulation of EGR2 (a key T cell anergy inducer) mRNA transcription in stimulated T MOG cells. This was accompanied by elevated levels of anergy promoting genes such as IL-10 (anti-inflammatory cytokine), STAT5 (regulatory factor), and LAG3 mRNAs, as well as of several enhancers of cell cycle arrest (such as Nfatc1, Casp4, Cdkn1a, and Icos). Moreover, CBD exposure leads to a decrease in STAT3 and to an increase in STAT5 phosphorylation in T MOG cells, positive and negative regulators of Th17 activity, respectively. In parallel, we observed decreased levels of major histocompatibility complex class II (MHCII), CD25, and CD69 on CD19 + B cells following CBD treatment, showing diminished antigen presenting capabilities of B cells and reduction in their pro-inflammatory functions.

Research paper thumbnail of HU-446 and HU-465, derivatives of the non-psychoactive cannabinoid cannabidiol, decrease the activation of encephalitogenic T cells

Chemical biology & drug design, Jan 11, 2015

Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease ... more Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease peripheral inflammation and neuroinflammation in mouse experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). Here we have studied the anti-inflammatory effects of newly synthesized derivatives of natural (-)-CBD ((-)-8,9-dihydro-7-hydroxy-CBD; HU-446) and of synthetic (+)-CBD ((+)-8,9-dihydro-7-hydroxy-CBD; HU-465), on activated myelin oligodendrocyte glycoprotein (MOG)35-55 specific mouse encephalitogenic T cells (TMOG ) driving EAE/MS-like pathologies. Binding assays followed by molecular modelling revealed that HU-446 has negligible affinity towards the cannabinoid CB1 and CB2 receptors while HU-465 binds to both CB1 and CB2 receptors at the high nanomolar concentrations (Ki =76.7 ± 5.8 nM and 12.1 ± 2.3 nM, respectively). Both, HU-446 and HU-465, at 5 and 10 μM (but not at 0.1 and 1 μM) inhibited the MOG35-55-induced proliferation of autoreactive TMOG cell...

Research paper thumbnail of The disease susceptibility in a new ‘humanized’ multiple sclerosis model in HLA-DR15 (DRB1*1501;DQB1*0602) transgenic mice is determined by HLA-DQB1*0602

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS) characterize... more Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS) characterized by perivascular inflammation accompanied by primary demyelination, axonal damage, and neuronal loss. Numerous studies show that both genetic and environmental factors play a role in the etiology of MS. Although the genetic and environmental risk factors have been extensively studied in the context of MS, the major histocompatibility complex (MHC) is the only genomic region consistently associated with MS and several MHC alleles predispose to the disease. In patients of North European, Caucasian origin, the MHC class II alleles that are most prevalent among MS patients come from the HLA-DR15 haplotype. The ‘DR15 haplotype’ encodes three functional HLA-class II heterodimers, DR15 (DRA1*0101/DRB1*1501 pair), DRB5 (DRA1*0101/DRB5*0101 pair), and the DQ6 (DQA1*0102/DQB1*0602 pair). Although several other HLA and non-HLA disease predisposing alleles have been identified, alleles of the HLA-D...

Research paper thumbnail of Role of a novel HLA-DQA1*01:02;DRB1*15:01 mixed-isotype heterodimer in the pathogenesis of 'humanized' multiple sclerosis-like disease

The Journal of biological chemistry, Jan 24, 2015

Gene-wide-association and candidate gene studies indicate that the greatest effect on multiple sc... more Gene-wide-association and candidate gene studies indicate that the greatest effect on multiple sclerosis (MS) risk is driven by the HLA-DRB1*15:01 allele within the HLA-DR15-haplotype (HLA-DRB1*15:01-DQA1*01:02-DQB1*0602-DRB5*01:01). Nevertheless, linkage-disequilibrium makes it difficult to define, without functional studies, whether the functionally relevant effect derives from DRB1*15:01 only, its neighboring DQA1*01:02-DQB1*06:02 or DRB5*01:01 genes of HLA-DR15-haplotype, or from their combinations or epistatic interactions. Here, we analyzed the impact of the different HLA-DR15-haplotype alleles on disease-susceptibility in a new humanized model of MS induced in HLA-transgenic (Tg) mice by human oligodendrocyte-specific protein (OSP)/claudin-11 (hOSP), one of the bone fide potential primary target antigens in MS. We show that the hOSP-associated MS-like disease is dominated by DRB1*15:01 allele not only as the DRA1*01:01;DRB1*15:01 isotypic-heterodimer, but also, unexpectedly, ...

Research paper thumbnail of Cannabidiol, a non-psychoactive cannabinoid, leads to EGR2-dependent anergy in activated encephalitogenic T cells

Journal of Neuroinflammation, 2015

Background: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown b... more Background: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown by us to ameliorate clinical symptoms and to decrease inflammation in myelin oligodendrocyte glycoprotein (MOG)35-55-induced mouse experimental autoimmune encephalomyelitis model of multiple sclerosis as well as to decrease MOG35-55-induced T cell proliferation and IL-17 secretion. However, the mechanisms of CBD anti-inflammatory activities are unclear. Methods: Here we analyzed the effects of CBD on splenocytes (source of accessory T cells and antigen presenting cells (APC)) co-cultured with MOG35-55-specific T cells (T MOG ) and stimulated with MOG35-55. Using flow cytometry, we evaluated the expression of surface activation markers and inhibitory molecules on T cells and B cells. T MOG cells were purified using CD4 positive microbead selection and submitted for quantitative PCR and microarray of mRNA transcript analyzes. Cell signaling studies in purified T MOG were carried out using immunoblotting. Results: We found that CBD leads to upregulation of CD69 and lymphocyte-activation gene 3 (LAG3) regulatory molecules on CD4 + CD25 − accessory T cells. This subtype of CD4 + CD25 − CD69 + LAG3 + T cells has been recognized as induced regulatory phenotype promoting anergy in activated T cells. Indeed, we observed that CBD treatment results in upregulation of EGR2 (a key T cell anergy inducer) mRNA transcription in stimulated T MOG cells. This was accompanied by elevated levels of anergy promoting genes such as IL-10 (anti-inflammatory cytokine), STAT5 (regulatory factor), and LAG3 mRNAs, as well as of several enhancers of cell cycle arrest (such as Nfatc1, Casp4, Cdkn1a, and Icos). Moreover, CBD exposure leads to a decrease in STAT3 and to an increase in STAT5 phosphorylation in T MOG cells, positive and negative regulators of Th17 activity, respectively. In parallel, we observed decreased levels of major histocompatibility complex class II (MHCII), CD25, and CD69 on CD19 + B cells following CBD treatment, showing diminished antigen presenting capabilities of B cells and reduction in their pro-inflammatory functions.

Research paper thumbnail of The “multi-epitope-targeting” approach to antigen-based immunotherapy of multiple sclerosis-like disease is more efficacious through a specifically engineered multi-epitope protein than through peptides

Multiple sclerosis (MS) is a chronic inflammatory disease of the CNS, associated with complex ant... more Multiple sclerosis (MS) is a chronic inflammatory disease of the CNS, associated with complex anti-myelin autoimmunity. Among all approaches proposed for MS therapy, an approach that neutralizes only the pathogenic T cells reacting against myelin, while leaving the innocent immune cells intact, is the ultimate goal in the immune-specific therapy for MS. The multiplicity of primary target antigens, along side the dynamic nature of autoimmunity in MS, whereby the specificity of anti-myelin pathogenic autoreactivities may shift or expand in the same patient with disease progression, impose major difficulties in devising immune-specific therapy to MS. To overcome this multiplicity and the potential complexity of pathogenic autoreactivities in MS, we have put forward the concept of concomitant multi-antigen/multi-epitope targeting as, a conceivably more effective approach to immunotherapy of MS. We constructed an EAE/MS-related synthetic human Target Autoantigen Gene (MS-shMultiTAG) desi...

Research paper thumbnail of Peptides derived from HIV gp41 for treating T-cell mediated pathologies

Research paper thumbnail of Epitope specificity of autoreactive T and B cells associated with experimental autoimmune encephalomyelitis and optic neuritis induced by oligodendrocyte-specific protein in SJL/J mice

Journal of immunology (Baltimore, Md. : 1950), Jan 15, 2006

The encephalitogenic potential of oligodendrocyte-specific protein (OSP) in mice, its specific lo... more The encephalitogenic potential of oligodendrocyte-specific protein (OSP) in mice, its specific localization in the intralamellar tight junctions in CNS myelin, and the detection of autoreactivity against OSP in multiple sclerosis (MS) strongly suggest the relevance of autoreactivity against OSP in the pathogenesis of MS. In this study, we have characterized the autoimmune T and B cells that are associated with clinicopathological manifestations of OSP-induced MS-like disease in mice by using recombinant soluble mouse OSP (smOSP) and synthetic overlapping peptides spanning smOSP. SJL/J mice immunized with smOSP developed chronic relapsing clinical experimental autoimmune encephalomyelitis accompanied with intense perivascular and parenchymal inflammatory infiltrates, widespread demyelination, axonal loss, and remarkable optic neuritis. The smOSP-primed lymph node cells reacted predominantly against OSP55-80 and to a lesser extent also to OSP22-46 and OSP179-207. Unexpectedly, in vitr...

Research paper thumbnail of DQB1*06:02-Associated Pathogenic Anti-Myelin Autoimmunity in Multiple Sclerosis-Like Disease: Potential Function of DQB1*06:02 as a Disease-Predisposing Allele

Frontiers in Oncology, 2014

Susceptibility to multiple sclerosis (MS) has been linked mainly to the HLA-DRB1 locus, with the ... more Susceptibility to multiple sclerosis (MS) has been linked mainly to the HLA-DRB1 locus, with the HLA-DR15 haplotype (DRB1*1501-DQA1*0102-DQB1*0602-DRB5*0101) dominating MS risk in Caucasians. Although genes in the HLA-II region, particularly DRB1*1501, DQA1*0102-DQB1*0602, are in tight linkage disequilibrium, genome-wide-association, and gene candidate studies identified the DRB1*15:01 allele as the primary risk factor in MS. Many genetic and immune-functional studies have indicated DRB1*15:01 as a primary risk factor in MS, while only some functional studies suggested a diseasemodifying role for the DRB5*01 or DQB1*06 alleles. In this respect, the susceptibility of DRB1*15:01-transgenic (Tg) mice to myelin basic protein-or myelin oligodendrocyte glycoprotein-induced MS-like disease is consistent with primary contribution of DRB1*15:01 to HLA-DR15+ MS. The studies summarized here show that susceptibility to MS-like disease, induced in HLA-"humanized" mice by myelin oligodendrocytic basic protein or by the proteolipid protein, one of the most prominent encephalitogenic target antigens implicated in human MS, is determined by DQB1*06:02, rather than by the DRB1*15:01 allele. These findings not only offer a rationale for a potential role for DQB1*06:02 in predisposing susceptibility to MS, but also suggest a more complex and differential functional role for HLA-DR15 alleles, depending on the primary target myelin antigen. However, the conflict between these findings in HLA-Tg mice and the extensive genome-wide-association studies, which could not detect any significant effect from the DQB1*06:02 allele on MS risk, is rather puzzling. Functional analysis of MS PBLs for DQB1*06:02-associated anti-myelin autoimmunity may indicate whether or not DQB1*06:02 is associated with MS pathogenesis.

Research paper thumbnail of ‘Multi-Epitope-Targeted’ Immune-Specific Therapy for a Multiple Sclerosis-Like Disease via Engineered Multi-Epitope Protein Is Superior to Peptides

PLoS ONE, 2011

Antigen-induced peripheral tolerance is potentially one of the most efficient and specific therap... more Antigen-induced peripheral tolerance is potentially one of the most efficient and specific therapeutic approaches for autoimmune diseases. Although highly effective in animal models, antigen-based strategies have not yet been translated into practicable human therapy, and several clinical trials using a single antigen or peptidic-epitope in multiple sclerosis (MS) yielded disappointing results.

Research paper thumbnail of DQB1*0602 rather than DRB1*1501 confers susceptibility to multiple sclerosis-like disease induced by proteolipid protein (PLP)

Journal of Neuroinflammation, 2012

Background: Multiple sclerosis (MS) is associated with pathogenic autoimmunity primarily focused ... more Background: Multiple sclerosis (MS) is associated with pathogenic autoimmunity primarily focused on major CNSmyelin target antigens including myelin basic protein (MBP), proteolipidprotein (PLP), myelin oligodendrocyte protein (MOG). MS is a complex trait whereby the HLA genes, particularly class-II genes of HLA-DR15 haplotype, dominate the genetic contribution to disease-risk. Due to strong linkage disequilibrium in HLA-II region, it has been hard to establish precisely whether the functionally relevant effect derives from the DRB1*1501, DQA1*0102-DQB1*0602, or DRB5*0101 loci of HLA-DR15 haplotype, their combinations, or their epistatic interactions. Nevertheless, most genetic studies have indicated DRB1*1501 as a primary risk factor in MS. Here, we used 'HLAhumanized' mice to discern the potential relative contribution of DRB1*1501 and DQB1*0602 alleles to susceptibility to "humanized" MS-like disease induced by PLP, one of the most prominent and encephalitogenic target-antigens implicated in human MS.

Research paper thumbnail of Cannabidiol effects on mRNA levels and signaling pathways in MOG-35–55 activated encephalitogenic T cells

Journal of Neuroimmunology, 2014

Research paper thumbnail of Cannabinoids Decrease the Th17 Inflammatory Autoimmune Phenotype

Journal of Neuroimmune Pharmacology, 2013

Cannabinoids, the Cannabis constituents, are known to possess anti-inflammatory properties but th... more Cannabinoids, the Cannabis constituents, are known to possess anti-inflammatory properties but the mechanisms involved are not understood. Here we show that the main psychoactive cannabinoid, Δ-9-tetrahydrocannabinol (THC), and the main nonpsychoactive cannabinoid, cannabidiol (CBD), markedly reduce the Th17 phenotype which is known to be increased in inflammatory autoimmune pathologies such as Multiple Sclerosis. We found that reactivation by MOG35-55 of MOG35-55-specific encephalitogenic T cells (cells that induce Experimental Autoimmune Encephalitis when injected to mice) in the presence of spleen derived antigen presenting cells led to a large increase in IL-17 production and secretion. In addition, we found that the cannabinoids CBD and THC dose-dependently (at 0.1-5 μM) suppressed the production and secretion of this cytokine. Moreover, the mRNA and protein of IL-6, a key factor in Th17 induction, were also decreased. Pretreatment with CBD also resulted in increased levels of the anti-inflammatory cytokine IL-10. Interestingly, CBD and THC did not affect the levels of TNFα and IFNγ. The downregulation of IL-17 secretion by these cannabinoids does not seem to involve the CB1, CB2, PPARγ, 5-HT 1A or TRPV1 receptors. In conclusion, the results show a unique cannabinoid modulation of the autoimmune cytokine milieu combining suppression of the pathogenic IL-17 and IL-6 cytokines along with boosting the expression of the anti-inflammatory cytokine IL-10.

Research paper thumbnail of Aggrecan Turnover in Human Articular Cartilage: Use of Aspartic Acid Racemization as a Marker of Molecular Age

Proteoglycans are major constituents of the extracel-Aggrecan is a key component of the cartilage... more Proteoglycans are major constituents of the extracel-Aggrecan is a key component of the cartilage matrix. lular matrix of articular cartilage and are largely re-During aging, many changes occur in its composition sponsible for the high resistance to compression of this and structure; in particular, there is an increase in the load-bearing tissue. This resistance to compression is proportion of lower molecular weight monomers and mainly due to the high osmotic pressure of the nega-of the ''free'' binding region. An important question tively charged glycosaminoglycan (GAG) 2 chains. The has been whether these changes represent alterations proteoglycans (aggrecan) are organized in the matrix in biosynthesis or whether they are due to the accumu-through a highly specific interaction to form large ag-lation with age of the partially degraded fragments gregates consisting of a central filament of hyaluronan, of the originally synthesized large monomer. In the to which aggrecan monomers are attached via the hya-present work we have used an independent tool, viz., luronan-binding domain (G1) of their core proteins, the extent of racemization of aspartic acid to study the with the interaction being stabilized by the presence molecular ''age'' of different buoyant density fractions of a link protein. Aggregate formation is very important of the aggrecan of human articular cartilage, as well from the physiological point of view, since it ensures as of isolated free binding region and link protein. By the retention of aggrecan within the collagen network. measuring the D/L Asp ratio of the different aggrecan

Research paper thumbnail of Post-CNS-inflammation expression of CXCL12 promotes the endogenous myelin/ neuronal repair capacity following spontaneous recovery from multiple sclerosis-like disease

Background: Demyelination and axonal degeneration, hallmarks of multiple sclerosis (MS), are asso... more Background: Demyelination and axonal degeneration, hallmarks of multiple sclerosis (MS), are associated with the central nervous system (CNS) inflammation facilitated by C-X-C motif chemokine 12 (CXCL12) chemokine. Both in MS and in experimental autoimmune encephalomyelitis (EAE), the deleterious CNS inflammation has been associated with upregulation of CXCL12 expression in the CNS. We investigated the expression dynamics of CXCL12 in the CNS with progression of clinical EAE and following spontaneous recovery, with a focus on CXCL12 expression in the hippocampal neurogenic dentate gyrus (DG) and in the corpus callosum (CC) of spontaneously recovered mice, and its potential role in promoting the endogenous myelin/neuronal repair capacity.

Research paper thumbnail of HU-446 and HU-465, derivatives of the non-psychoactive cannabinoid cannabidiol, decrease the activation of encephalitogenic T cells

Chemical biology & drug design, Jan 11, 2015

Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease ... more Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease peripheral inflammation and neuroinflammation in mouse experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). Here we have studied the anti-inflammatory effects of newly synthesized derivatives of natural (-)-CBD ((-)-8,9-dihydro-7-hydroxy-CBD; HU-446) and of synthetic (+)-CBD ((+)-8,9-dihydro-7-hydroxy-CBD; HU-465), on activated myelin oligodendrocyte glycoprotein (MOG)35-55 specific mouse encephalitogenic T cells (TMOG ) driving EAE/MS-like pathologies. Binding assays followed by molecular modelling revealed that HU-446 has negligible affinity towards the cannabinoid CB1 and CB2 receptors while HU-465 binds to both CB1 and CB2 receptors at the high nanomolar concentrations (Ki =76.7 ± 5.8 nM and 12.1 ± 2.3 nM, respectively). Both, HU-446 and HU-465, at 5 and 10 μM (but not at 0.1 and 1 μM) inhibited the MOG35-55-induced proliferation of autoreactive TMOG cell...

Research paper thumbnail of Cannabidiol inhibits pathogenic T cells, decreases spinal microglial activation and ameliorates multiple sclerosis-like disease in C57BL/6 mice

British Journal of Pharmacology, 2011

Cannabis extracts and several cannabinoids have been shown to exert broad anti-inflammatory activ... more Cannabis extracts and several cannabinoids have been shown to exert broad anti-inflammatory activities in experimental models of inflammatory CNS degenerative diseases. Clinical use of many cannabinoids is limited by their psychotropic effects. However, phytocannabinoids like cannabidiol (CBD), devoid of psychoactive activity, are, potentially, safe and effective alternatives for alleviating neuroinflammation and neurodegeneration.

Research paper thumbnail of A highly conserved sequence associated with the HIV gp41 loop region is an immunomodulator of antigen-specific T cells in mice

Research paper thumbnail of An Immunomodulating Motif of the HIV-1 Fusion Protein Is Chirality-independent: IMPLICATIONS FOR ITS MODE OF ACTION

Journal of Biological Chemistry, 2013

Recently, an immunosuppressive motif was identified in the HIV-1 envelope glycoprotein complex. R... more Recently, an immunosuppressive motif was identified in the HIV-1 envelope glycoprotein complex. Results: Both D-and L-stereoisomers of the motif inhibit T-cell receptor activation and preferentially bind T-cells over B-cells.

Research paper thumbnail of Highlight in Nature Immuno (1)

Research paper thumbnail of Cannabidiol, a non-psychoactive cannabinoid, leads to EGR2-dependent anergy in activated encephalitogenic T cells

Background: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown b... more Background: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown by us to ameliorate clinical symptoms and to decrease inflammation in myelin oligodendrocyte glycoprotein (MOG)35-55-induced mouse experimental autoimmune encephalomyelitis model of multiple sclerosis as well as to decrease MOG35-55-induced T cell proliferation and IL-17 secretion. However, the mechanisms of CBD anti-inflammatory activities are unclear. Methods: Here we analyzed the effects of CBD on splenocytes (source of accessory T cells and antigen presenting cells (APC)) co-cultured with MOG35-55-specific T cells (T MOG) and stimulated with MOG35-55. Using flow cytometry, we evaluated the expression of surface activation markers and inhibitory molecules on T cells and B cells. T MOG cells were purified using CD4 positive microbead selection and submitted for quantitative PCR and microarray of mRNA transcript analyzes. Cell signaling studies in purified T MOG were carried out using immunoblotting. Results: We found that CBD leads to upregulation of CD69 and lymphocyte-activation gene 3 (LAG3) regulatory molecules on CD4 + CD25 − accessory T cells. This subtype of CD4 + CD25 − CD69 + LAG3 + T cells has been recognized as induced regulatory phenotype promoting anergy in activated T cells. Indeed, we observed that CBD treatment results in upregulation of EGR2 (a key T cell anergy inducer) mRNA transcription in stimulated T MOG cells. This was accompanied by elevated levels of anergy promoting genes such as IL-10 (anti-inflammatory cytokine), STAT5 (regulatory factor), and LAG3 mRNAs, as well as of several enhancers of cell cycle arrest (such as Nfatc1, Casp4, Cdkn1a, and Icos). Moreover, CBD exposure leads to a decrease in STAT3 and to an increase in STAT5 phosphorylation in T MOG cells, positive and negative regulators of Th17 activity, respectively. In parallel, we observed decreased levels of major histocompatibility complex class II (MHCII), CD25, and CD69 on CD19 + B cells following CBD treatment, showing diminished antigen presenting capabilities of B cells and reduction in their pro-inflammatory functions.

Research paper thumbnail of HU-446 and HU-465, derivatives of the non-psychoactive cannabinoid cannabidiol, decrease the activation of encephalitogenic T cells

Chemical biology & drug design, Jan 11, 2015

Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease ... more Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease peripheral inflammation and neuroinflammation in mouse experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). Here we have studied the anti-inflammatory effects of newly synthesized derivatives of natural (-)-CBD ((-)-8,9-dihydro-7-hydroxy-CBD; HU-446) and of synthetic (+)-CBD ((+)-8,9-dihydro-7-hydroxy-CBD; HU-465), on activated myelin oligodendrocyte glycoprotein (MOG)35-55 specific mouse encephalitogenic T cells (TMOG ) driving EAE/MS-like pathologies. Binding assays followed by molecular modelling revealed that HU-446 has negligible affinity towards the cannabinoid CB1 and CB2 receptors while HU-465 binds to both CB1 and CB2 receptors at the high nanomolar concentrations (Ki =76.7 ± 5.8 nM and 12.1 ± 2.3 nM, respectively). Both, HU-446 and HU-465, at 5 and 10 μM (but not at 0.1 and 1 μM) inhibited the MOG35-55-induced proliferation of autoreactive TMOG cell...

Research paper thumbnail of The disease susceptibility in a new ‘humanized’ multiple sclerosis model in HLA-DR15 (DRB1*1501;DQB1*0602) transgenic mice is determined by HLA-DQB1*0602

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS) characterize... more Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS) characterized by perivascular inflammation accompanied by primary demyelination, axonal damage, and neuronal loss. Numerous studies show that both genetic and environmental factors play a role in the etiology of MS. Although the genetic and environmental risk factors have been extensively studied in the context of MS, the major histocompatibility complex (MHC) is the only genomic region consistently associated with MS and several MHC alleles predispose to the disease. In patients of North European, Caucasian origin, the MHC class II alleles that are most prevalent among MS patients come from the HLA-DR15 haplotype. The ‘DR15 haplotype’ encodes three functional HLA-class II heterodimers, DR15 (DRA1*0101/DRB1*1501 pair), DRB5 (DRA1*0101/DRB5*0101 pair), and the DQ6 (DQA1*0102/DQB1*0602 pair). Although several other HLA and non-HLA disease predisposing alleles have been identified, alleles of the HLA-D...

Research paper thumbnail of Role of a novel HLA-DQA1*01:02;DRB1*15:01 mixed-isotype heterodimer in the pathogenesis of 'humanized' multiple sclerosis-like disease

The Journal of biological chemistry, Jan 24, 2015

Gene-wide-association and candidate gene studies indicate that the greatest effect on multiple sc... more Gene-wide-association and candidate gene studies indicate that the greatest effect on multiple sclerosis (MS) risk is driven by the HLA-DRB1*15:01 allele within the HLA-DR15-haplotype (HLA-DRB1*15:01-DQA1*01:02-DQB1*0602-DRB5*01:01). Nevertheless, linkage-disequilibrium makes it difficult to define, without functional studies, whether the functionally relevant effect derives from DRB1*15:01 only, its neighboring DQA1*01:02-DQB1*06:02 or DRB5*01:01 genes of HLA-DR15-haplotype, or from their combinations or epistatic interactions. Here, we analyzed the impact of the different HLA-DR15-haplotype alleles on disease-susceptibility in a new humanized model of MS induced in HLA-transgenic (Tg) mice by human oligodendrocyte-specific protein (OSP)/claudin-11 (hOSP), one of the bone fide potential primary target antigens in MS. We show that the hOSP-associated MS-like disease is dominated by DRB1*15:01 allele not only as the DRA1*01:01;DRB1*15:01 isotypic-heterodimer, but also, unexpectedly, ...

Research paper thumbnail of Cannabidiol, a non-psychoactive cannabinoid, leads to EGR2-dependent anergy in activated encephalitogenic T cells

Journal of Neuroinflammation, 2015

Background: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown b... more Background: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown by us to ameliorate clinical symptoms and to decrease inflammation in myelin oligodendrocyte glycoprotein (MOG)35-55-induced mouse experimental autoimmune encephalomyelitis model of multiple sclerosis as well as to decrease MOG35-55-induced T cell proliferation and IL-17 secretion. However, the mechanisms of CBD anti-inflammatory activities are unclear. Methods: Here we analyzed the effects of CBD on splenocytes (source of accessory T cells and antigen presenting cells (APC)) co-cultured with MOG35-55-specific T cells (T MOG ) and stimulated with MOG35-55. Using flow cytometry, we evaluated the expression of surface activation markers and inhibitory molecules on T cells and B cells. T MOG cells were purified using CD4 positive microbead selection and submitted for quantitative PCR and microarray of mRNA transcript analyzes. Cell signaling studies in purified T MOG were carried out using immunoblotting. Results: We found that CBD leads to upregulation of CD69 and lymphocyte-activation gene 3 (LAG3) regulatory molecules on CD4 + CD25 − accessory T cells. This subtype of CD4 + CD25 − CD69 + LAG3 + T cells has been recognized as induced regulatory phenotype promoting anergy in activated T cells. Indeed, we observed that CBD treatment results in upregulation of EGR2 (a key T cell anergy inducer) mRNA transcription in stimulated T MOG cells. This was accompanied by elevated levels of anergy promoting genes such as IL-10 (anti-inflammatory cytokine), STAT5 (regulatory factor), and LAG3 mRNAs, as well as of several enhancers of cell cycle arrest (such as Nfatc1, Casp4, Cdkn1a, and Icos). Moreover, CBD exposure leads to a decrease in STAT3 and to an increase in STAT5 phosphorylation in T MOG cells, positive and negative regulators of Th17 activity, respectively. In parallel, we observed decreased levels of major histocompatibility complex class II (MHCII), CD25, and CD69 on CD19 + B cells following CBD treatment, showing diminished antigen presenting capabilities of B cells and reduction in their pro-inflammatory functions.

Research paper thumbnail of The “multi-epitope-targeting” approach to antigen-based immunotherapy of multiple sclerosis-like disease is more efficacious through a specifically engineered multi-epitope protein than through peptides

Multiple sclerosis (MS) is a chronic inflammatory disease of the CNS, associated with complex ant... more Multiple sclerosis (MS) is a chronic inflammatory disease of the CNS, associated with complex anti-myelin autoimmunity. Among all approaches proposed for MS therapy, an approach that neutralizes only the pathogenic T cells reacting against myelin, while leaving the innocent immune cells intact, is the ultimate goal in the immune-specific therapy for MS. The multiplicity of primary target antigens, along side the dynamic nature of autoimmunity in MS, whereby the specificity of anti-myelin pathogenic autoreactivities may shift or expand in the same patient with disease progression, impose major difficulties in devising immune-specific therapy to MS. To overcome this multiplicity and the potential complexity of pathogenic autoreactivities in MS, we have put forward the concept of concomitant multi-antigen/multi-epitope targeting as, a conceivably more effective approach to immunotherapy of MS. We constructed an EAE/MS-related synthetic human Target Autoantigen Gene (MS-shMultiTAG) desi...

Research paper thumbnail of Peptides derived from HIV gp41 for treating T-cell mediated pathologies

Research paper thumbnail of Epitope specificity of autoreactive T and B cells associated with experimental autoimmune encephalomyelitis and optic neuritis induced by oligodendrocyte-specific protein in SJL/J mice

Journal of immunology (Baltimore, Md. : 1950), Jan 15, 2006

The encephalitogenic potential of oligodendrocyte-specific protein (OSP) in mice, its specific lo... more The encephalitogenic potential of oligodendrocyte-specific protein (OSP) in mice, its specific localization in the intralamellar tight junctions in CNS myelin, and the detection of autoreactivity against OSP in multiple sclerosis (MS) strongly suggest the relevance of autoreactivity against OSP in the pathogenesis of MS. In this study, we have characterized the autoimmune T and B cells that are associated with clinicopathological manifestations of OSP-induced MS-like disease in mice by using recombinant soluble mouse OSP (smOSP) and synthetic overlapping peptides spanning smOSP. SJL/J mice immunized with smOSP developed chronic relapsing clinical experimental autoimmune encephalomyelitis accompanied with intense perivascular and parenchymal inflammatory infiltrates, widespread demyelination, axonal loss, and remarkable optic neuritis. The smOSP-primed lymph node cells reacted predominantly against OSP55-80 and to a lesser extent also to OSP22-46 and OSP179-207. Unexpectedly, in vitr...

Research paper thumbnail of DQB1*06:02-Associated Pathogenic Anti-Myelin Autoimmunity in Multiple Sclerosis-Like Disease: Potential Function of DQB1*06:02 as a Disease-Predisposing Allele

Frontiers in Oncology, 2014

Susceptibility to multiple sclerosis (MS) has been linked mainly to the HLA-DRB1 locus, with the ... more Susceptibility to multiple sclerosis (MS) has been linked mainly to the HLA-DRB1 locus, with the HLA-DR15 haplotype (DRB1*1501-DQA1*0102-DQB1*0602-DRB5*0101) dominating MS risk in Caucasians. Although genes in the HLA-II region, particularly DRB1*1501, DQA1*0102-DQB1*0602, are in tight linkage disequilibrium, genome-wide-association, and gene candidate studies identified the DRB1*15:01 allele as the primary risk factor in MS. Many genetic and immune-functional studies have indicated DRB1*15:01 as a primary risk factor in MS, while only some functional studies suggested a diseasemodifying role for the DRB5*01 or DQB1*06 alleles. In this respect, the susceptibility of DRB1*15:01-transgenic (Tg) mice to myelin basic protein-or myelin oligodendrocyte glycoprotein-induced MS-like disease is consistent with primary contribution of DRB1*15:01 to HLA-DR15+ MS. The studies summarized here show that susceptibility to MS-like disease, induced in HLA-"humanized" mice by myelin oligodendrocytic basic protein or by the proteolipid protein, one of the most prominent encephalitogenic target antigens implicated in human MS, is determined by DQB1*06:02, rather than by the DRB1*15:01 allele. These findings not only offer a rationale for a potential role for DQB1*06:02 in predisposing susceptibility to MS, but also suggest a more complex and differential functional role for HLA-DR15 alleles, depending on the primary target myelin antigen. However, the conflict between these findings in HLA-Tg mice and the extensive genome-wide-association studies, which could not detect any significant effect from the DQB1*06:02 allele on MS risk, is rather puzzling. Functional analysis of MS PBLs for DQB1*06:02-associated anti-myelin autoimmunity may indicate whether or not DQB1*06:02 is associated with MS pathogenesis.

Research paper thumbnail of ‘Multi-Epitope-Targeted’ Immune-Specific Therapy for a Multiple Sclerosis-Like Disease via Engineered Multi-Epitope Protein Is Superior to Peptides

PLoS ONE, 2011

Antigen-induced peripheral tolerance is potentially one of the most efficient and specific therap... more Antigen-induced peripheral tolerance is potentially one of the most efficient and specific therapeutic approaches for autoimmune diseases. Although highly effective in animal models, antigen-based strategies have not yet been translated into practicable human therapy, and several clinical trials using a single antigen or peptidic-epitope in multiple sclerosis (MS) yielded disappointing results.

Research paper thumbnail of DQB1*0602 rather than DRB1*1501 confers susceptibility to multiple sclerosis-like disease induced by proteolipid protein (PLP)

Journal of Neuroinflammation, 2012

Background: Multiple sclerosis (MS) is associated with pathogenic autoimmunity primarily focused ... more Background: Multiple sclerosis (MS) is associated with pathogenic autoimmunity primarily focused on major CNSmyelin target antigens including myelin basic protein (MBP), proteolipidprotein (PLP), myelin oligodendrocyte protein (MOG). MS is a complex trait whereby the HLA genes, particularly class-II genes of HLA-DR15 haplotype, dominate the genetic contribution to disease-risk. Due to strong linkage disequilibrium in HLA-II region, it has been hard to establish precisely whether the functionally relevant effect derives from the DRB1*1501, DQA1*0102-DQB1*0602, or DRB5*0101 loci of HLA-DR15 haplotype, their combinations, or their epistatic interactions. Nevertheless, most genetic studies have indicated DRB1*1501 as a primary risk factor in MS. Here, we used 'HLAhumanized' mice to discern the potential relative contribution of DRB1*1501 and DQB1*0602 alleles to susceptibility to "humanized" MS-like disease induced by PLP, one of the most prominent and encephalitogenic target-antigens implicated in human MS.