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The two chemokines, monocyte chemoattractant protein (MCP)-1 and gamma -interferon inducible protein (IP)-10, are thought to be involved in the pathogenesis of multiple sclerosis (MS). We measured MCP-1 and IP-10 levels in serum and CSF samples from 38 acute and 25 stable MS patients and from 40 controls. The latter consisted in patients with other inflammatory neurological diseases (OIND) or with non-inflammatory neurological diseases, and healthy controls. CSF MCP-1 levels exceeded those found in serum in all the patients studied as well as in healthy controls. CSF MCP-1 levels were significantly Lower in acute MS [468 +/- (S.E.M.) 18 pg/ml] than in stable MS (857 +/- 104 pg/ml). When detectable, serum and CSF IP-10 levels were significantly higher in acute MS (serum 331 +/- 66 pg/ml; CSF 118 +/- 16 pg/ml) than in stable MS (serum 69 +/- 7 pg/ml; CSF 25 +/- 2 pg/ml). Among OIND patients, those with HIV-1-associated dementia showed high serum and CSF levels of both MCP-1 and IP-10. Those with encephalitis showed high serum and CSF levels of IP-10 and CSF mononuclear pleiocytosis. We also evaluated the effects of 6-methylprednisolone or IFN-beta 1a therapy on circulating MCP-1 and IP-10 levels. Neither MCP-1 nor IP-10 post-therapy levels varied significantly from baseline values. Our findings suggest that (a) MCP-1 could be constitutively produced within the brain; (b) MCP-1 and IP-10 CSF levels in acute MS vary significantly from those in stable MS. and these variations are inverse: and (c) current MS therapies do not modify circulating levels of MCP-1 and IP-10. (C) 2001 Elsevier Science B.V. All rights reserved.

Systemic administration of antiinflammatory molecules to patients affected by immune-mediated inflammatory demyelinating diseases of the central nervous system (CNS) has limited therapeutic efficacy due to the presence of the blood-brain barrier (BBB). We found that three of five rhesus monkeys injected intrathecally with a replication-defective herpes simplex virus (HSV) type 1-derived vector engineered with the human interleukin 4 (IL-4) gene were protected from an hyperacute and lethal form of experimental autoimmune encephalomyelitis induced by whole myelin. The intrathecally injected vector consistently diffused within the CNS via the cerebrospinal fluid and infected ependymal cells, which in turn sustained in situ production of IL-4 without overt immunological or toxic side effects. In EAE-protected monkeys, IL-4-gene therapy significantly decreased the number of brain as well as spinal cord inflammatory perivenular infiltrates and the extent of demyelination, necrosis, and axonal loss. The protective effect was associated with in situ downregulation of inflammatory mediators such as tumor necrosis factor alpha (TNF-alpha) and monocyte chemoattractant protein 1 (MCP-1), upregulation of transforming growth factor beta (TGF-beta), and preservation of BBB integrity. Our results indicate that intrathecal delivery of HSV-1-derived vectors containing antiinflammatory cytokine genes may play a major role in the future therapeutic armamentarium of inflammatory CNS-confined demyelinating diseases and, in particular, in the most fulminant forms where conventional therapeutic approaches have, so far, failed to achieve a satisfactory control of the disease evolution.

The peripheral delivery of drugs in patients affected by central nervous system (CNS)-confined diseases is therapeutically ineffective due to the presence of the blood-brain barrier which forms an inaccessible wall to the majority of CNS targeting molecules. When molecules with an anti-inflammatory profile have been systemically administered to patients affected by a chronic inflammatory demyelinating disease of the CNS, such as multiple sclerosis (MS), results have been disappointing. A successful therapeutic approach in MS should therefore consider the delivery of anti-inflammatory molecules directly into the CNS in order to inhibit blood-borne CNS-confined mononuclear cells which act as ultimate effector cells directly destroying oligodendrocytes and/or releasing myelinotoxic substances. Biological and physical vectors engineered with heterologous genes coding for immunomodulatory cytokines with an anti-inflammatory profile might represent the appropriate tool to deliver therapeutic genes into the CNS of patients with MS. So far, cytokine gene therapy has never been attempted in MS, but encouraging results have been obtained in the animal model of MS, experimental autoimmune encephalomyelitis (EAE), using viral vectors or plasmids engineered with cytokine genes and then injected systemically, either in the blood stream or circulating encephalitogenic T cells, or into the CNS. Here, we critically discuss the various attempts made in EAE using gene therapy protocols based on the delivery of immunomodulatory cytokine genes. Special emphasis is put on the use of non-replicative herpes simplex type-1 (HSV)-derived vectors engineered with the gene of the immunomodulatory cytokine interleukin (IL)-4.

Multiple sclerosis (MS) is considered an immune-mediated disease of the central nervous system (CNS) sustained by a chronic inflammatory process leading to patchy demyelination and axonal loss. However, the inflammatory triggering event as well as the target of the pathogenic process in MS are still partially unknown. We report evidence that a 'local' inflammatory process occurring in the CNS (considered as a reaction of blood vessels in vascularized living tissue to a local injury leading to the accumulation of fluid and blood cells) along with a concomitant, but possibly unrelated, peripheral inflammatory event may trigger a CNS-specific autoimmune reaction cascade sustaining the MS pathogenesis. In the CNS, inflammatory mediators (mainly cytokines) act either as regulatory (i.e. activation of glial cells, shaping the autoimmune response) or effector molecules (i.e. myelinotoxicity, oligodendrotoxicity). In the periphery, inflammatory cytokines induce, in a bystander fashion, activation of monocytes and T cells. Among this latter cell population there are myelin-specific T cells belonging to the normal 'autoimmune' repertoire that home to the CNS where they may trigger the continuous recruitment of effector cells (macrophages) from the periphery. The concept that two concomitant, but possibly unrelated, inflammatory events, occurring in the CNS and in the periphery, represent the crucial elements sustaining MS, might reveal a more comprehensive view (dual signal hypothesis) of the entire etiopathogenic process underlying this disease.

Pro-inflammatory cytokines play a crucial role in the regulatory and effector phase of the immune-mediated mechanism sustaining multiple sclerosis pathogenesis (MS) thus supporting the use of anti-inflammatory cytokines as a therapeutic option. Systemic administration of cytokines shows, however, limited therapeutic efficacy and undesirable/unpredictable side-effects. We have developed a non-toxic system to deliver cytokines within the central nervous system (CNS) based on the intrathecal (i.c.) administration of non-replicative herpes simplex (HSV) type-1-derived viral vectors engineered with heterologous cytokine genes. Compared to controls, mice affected by experimental autoimmune encephalomyelitis (EAE) and i.c. injected with an HSV-1-derived vector containing the gene of the anti-inflammatory cytokine IL-4 showed a significant amelioration of clinical and pathological EAE signs. A decreased mRNA expression of the monocyte chemoattractant protein-1 (MCP-1) by mononuclear CNS-infiltrating cells was also observed. Peripheral T cells from IL-4-treated mice were not affected both in their antigen-specific proliferative response and in the cytokine secretion pattern. Our results indicate that CNS cytokine delivery with HSV-1-derived vectors is a feasible therapeutic strategy and might represent an alternative approach for the treatment of immune-mediated demyelinating diseases. Advantages of this approach over systemic cytokine administration are the high cytokine level reached within the CNS and the absence of side-effects on the peripheral immune system. The short-lasting cytokine production in the CNS after a single vector administration (4 weeks) is the limiting factor of this novel technology which, although promising, has to be improved.

A method for intracellular iron labeling of human mononuclear cells (lymphocytes and monocytes) for magnetic resonance imaging (MRI) using simple incubation of cells with approved MRI iron contrast agents is presented. Labeled cells can be detected by MRI in vitro, and this suggests the possibility that the technique could become a marker for in vivo lymphocyte and monocyte trafficking studies in acute inflammatory lesions such as those in Multiple Sclerosis.

The state of Bahia in the northeastern coast of Brazil is a region in which HTLV-I infection is endemic. This study investigated the characteristics of 28 HTLV-I-associated lymphomas/leukemias in this region. HTLV-I-infection diagnosis was based on serologic study, Southern blot analysis, and polymerase chain reaction (PCR) in neoplastic tissue. The main clinical differences between these lymphomas and adult T-cell leukemia (ATL) cases from other endemic areas were as follows. The mean age was 47 years; 20% of the cases occurred in young adults; and a predominance was found among male subjects (2:1), blacks, and of those of mixed race (96%). Histologically, 20 cases were T-cell pleomorphic leukemia/lymphoma, 5 were Mycosis fungoides-like cutaneous lymphoma, and 3 were CD30+ large-cell anaplastic lymphoma. Immunohistochemistry demonstrated 4 cases of CD8+ lymphoma. Proviral genomic sequences were demonstrated by PCR in 9 lymph node biopsy specimens and in 3 skin biopsy specimens. Southern blot was performed and was positive in 8 cases.

Identification of reliable biomarkers in multiple sclerosis patients would be highly desirable to help disease activity and progression monitoring, and evaluate response to treatments. The field has gradually shifted from the aim to find the perfect surrogate marker to the construction of composite markers with higher performances, taking advantage of the technologies allowing unbiased screening (i.e., microarrays, proteomics, etc.). Despite the fact that very interesting results have been produced, no approach has yet made it to the patient's bedside.

Background Neuromyelitis Optica (NMO), also known as Devic’s disease, is a rare inflammatory and demyelinating disorder of the central nervous system characterized by recurrent attacks of optic neuritis (ON) and longitudinally extensive trasverse myelitis (LETM). Previously considered as a severe variant of multiple sclerosis, NMO has been recently recognized as a distinct disorder associated with peculiar pathogenic auto-antibodies to aquaporin-4 (AQP4). Despite transient control of disease activity, standard treatments are often followed by relapses with accumulating disability and ultimately a poor prognosis. Thus, new therapeutic strategies for NMO are warranted. Hematopoietic Stem Cell Transplantation (HSCT) can induce stable remissions in patients with severe treatment-refractory Autoimmune Diseases. Here we report our investigational experience on allogeneic HSCT in two patients with severe and refractory forms of the disease. Methods Both treated patients had aggressive forms of NMO and serum positivity for AQP4 antibodies. Upn#1 is a 30 yrs old male affected by severe relapsing LETM, with paraparesis and thoracic spinal cord lesion on magnetic resonance imaging (MRI). Upn#2 is a 28 yrs female with recurrent attacks of ON and LETM. Both had received several lines of treatment without benefit, including high-dose corticosteroids, cyclophosphamide, rituximab, natalizumab, alemtuzumab, plasma exchange, high-dose thiotepa/cyclophosphamide and/or BEAM with Anti-Thymocyte Globulin (ATG) and cyclosporine followed by autologous HSCT rescue. Before HSCT the two patients showed an Kurtzke Expanded Disability Status Score (EDSS) of 6 (assistance required to walk) and 8.5 (restricted to bed, paraplegia) respectively, while both of them presented active contrast enhancing lesions on MRI and positivity for the pathogenic AQP4 auto-antibodies. Upn#1 underwent allogeneic HSCT from his HLA-identical sibling, while Upn#2 from a matched unrelated donor. Conditioning regimen consisted of full-dose treosulfan and fludarabine. Graft versus host disease (GvHD) prophylaxis combined ATG-Fresenius with cyclosporine and a short course of methotrexate (Upn#1) or mycophenolate and rapamycin (Upn#2); B cell depletion of both patients and graft was obtained by rituximab. Results Hematopoietic recovery occurred in both patients within day 30, and was accompanied by rapid achievement of full donor chimerism. Each patient experienced an episode of febrile neutropenia and one of them a CMV reactivation, both responsive to medical therapy; no serious adverse events were reported. None of the patients experienced neither acute nor chronic GvHD. MRI demonstrated the disappearance of the inflammatory lesions, without evidence of new ones. The immunological data obtained from the follow-up of these two patients suggest that allogeneic HSCT can alter the course of NMO through several concurring mechanisms: the eradication of autoreactive cell clones by high-dose chemotherapy and by the in vivo T and B cell depletion used for conditioning, the disappearance of the pathogenetic anti-AQP4 antibodies and achievement of a stable full-donor hematopoietic chimerism by donor T cell-mediated alloreactivity; the re-establishment of thymic central tolerance and renewal of the immune repertoire. Strikingly, these results correlated with a marked improvement of neurological functions in both treated patients. In UPN#1 EDSS dropped from 6 to 3.5 (fully ambulatory but with moderate disability in one functional neurological system), while in UPN#2 EDSS decreased from 8.5 to 7.5 (unable to take more than a few steps, restricted to wheelchair). At last follow-up (48 and 36 months after HSCT, respectively), both patients are alive, well and relapse-free. Conclusions These findings suggest that allogeneic HSCT may be beneficial in patients who have aggressive forms of NMO, by renewing the immune repertoire of T- and B-cells, thus reducing disease activity and arresting disability progression. The preliminary evidence of long-term disease control in these two refractory patients paves the way for further prospective phase I-II clinical trials to validate allogeneic HSCT as promising novel option for aggressive forms of NMO. Disclosures: Bonini: MolMed S.p.A: Consultancy.

Chimpanzees infected with human immunodeficiency virus type 1 (HIV-1) are used to model acquired immunodeficiency syndrome (AIDS). Since the central nervous system (CNS) is involved in AIDS, we performed an immunovirological study in 18 chimpanzees inoculated up to 87 months prior to the study (mean, 45 months) with HIV-1 and 8 uninfected controls. Serum and cerebrospinal fluid (CSF) IgG and albumin levels of infected chimpanzees never exceeded those of controls. The CSF/serum albumin ratio was elevated in 1 of 18 infected chimpanzees compared to controls; however, all animals had an elevated ratio indicating a more open blood-brain barrier relative to humans. The intrathecal IgG production index was elevated in only 1 of 18 infected chimpanzees compared to controls. Identical serum and CSF IgG bands were found by isoelectric focusing in 2 of 8 controls and in 1 of 18 infected chimpanzees. None of these bands reacted with recombinant HIV-1 p24gag or gp 120env. HIV-1 was isolated from the peripheral blood of 4 of 18 infected chimpanzees but never from the paired CSF samples. Anti-HIV-1 antibody was detected by a enzyme-linked immunosorbent assay in 18 of 18 paired serum and CSF samples and by Western blot in 18 of 18 serum and 13 of 18 CSF samples from infected chimpanzees without a difference in pattern. Polymerase chain reaction analysis on brain tissue of one animal was negative for HIV-1 sequences. Our results demonstrate that, unlike human infection, chimpanzees inoculated with HIV-1 show no evidence of isolatable virus in the CSF and no evidence of intrathecal anti-HIV-1 antibody synthesis up to several years after experimental infection. The lack of CNS involvement may contribute to the delay or suppression of clinical disease in infected chimpanzees.

Widespread demyelination, axonal loss and permanent neurological impairment are hallmarks of chronic, inflammatory demyelinating disorders of the CNS, such as multiple sclerosis (MS). The multifocal, recurrent nature of MS complicates cell and gene therapy approaches aimed at restoring axonal conduction by inducing re-myelination, or transplanting myelin-forming cells, at the sites of injury. Previous studies have demonstrated that systemic delivery of the anti-inflammatory cytokine IL4 to the CNS through the cerebro-spinal fluid circulation can prevent demyelination in experimentally-induced autoimmune encephalomyelitis (EAE), a murine model of MS (Hum Gene Ther 8:13, 2001). We have established a cytokine-delivery system to the CNS by transducing the ependimal layer lining the ventricular system with fully deleted, helper-dependent adenoviral vectors (HD-Ad). A single injection of a HD-Ad vector expressing IL-4 and GFP (AdIL-4G) into the cisterna magna of immunocompetent mice is sufficient to transduce ependimal and leptomeningeal cells surrounding ventricular and sub-arachnoid spaces. Vector DNA and GFP expression were detected in ependimal and leptomeningeal cells by real-time PCR after laser-capture microdissection and immunohystochemistry up to five months after injection. IL-4 levels up to 340 pg/ml were measured by ELISA in the cerebrospinal fluid five months after treatment. Serum IL-4 levels were always below detection limits. To assess the therapeutic potential of intracysternal IL-4 delivery, 30 mice affected by myelin oligodendrocyte glycoprotein (MOG35-55)-induced EAE were injected the day of clinical onset with escalating doses (105 to 107 i.p./mouse) of AdIL-4G or a control HD-Ad vector expressing only GFP. Mice treated with >106 i.p. of AdIL-4G displayed a progressive clinical recovery, and 80 days after immunization had an average EAE score of 1.0 (±1.0) vs. 1.62 (±0.4) in control mice (p

Objective: to evaluate the correlation between 25(OH)D levels in CIS patients and the development of Clinically Defined Multiple Sclerosis (CDMS). Background the risk of developing MS is related to both genetic and environmental factors. Vitamin D could play a role in CDMS conversion. Design/Methods: this is a single-center retrospective study. 107 CIS patients hospitalized from 2000 to 2009 at San Raffaele Hospital were recruited. We evaluated baseline serum 25(OH)D levels as well as clinical, brain MRI, Multimodal Evoked Potential and CSF data. Results: 21% patients developed CDMS at 12-month follow-up, 36% at 2-year follow-up and 44% at 5 year follow-up. A statistically significant inverse correlation between 25(OH)D levels and the risk to develop CDMS (OR 2.197) was observed. The difference between serum 25(OH)D levels in patients who developed CDMS and patients who did not, is statistically higher during summertime (p Conclusions: the presence of at least 3 negative clinical/paraclinical prognostic factors highly predicts short term conversion to CDMS (OR 5.39, p Disclosure: Dr. Martinelli has received personal compensation for activities with Biogen Dompe, Merck Serono, Bayer Schering, Teva, and Sanofi Aventis as a speaker. Dr. Dalla Costa has nothing to disclose. Dr. Colombo has nothing to disclose. Dr. Dalla Libera has nothing to disclose. Dr. Leocani has nothing to disclose. Dr. Furlan has nothing to disclose. Dr. Rubinacci has nothing to disclose. Dr. Filippi has received personal compensation for activities with ECTRIMS, MSIF, MS Ireland, US NMSS, Bayer-Schering, Biogen-Dompe AG, Genmab, Merck Serono, Pepgen Corporation, Teva, and Sanofi-Aventis. Dr. Filippi has received research support from Teva, Bayer-Schering and Genmab. Dr. Comi has received personal compensation for activities with Novartis, Teva Neuroscience, Sanofi-Aventis Pharmaceuticals, Inc., Merck Serono, Bayer Schering, and Biogen Dompe.

In order to reproduce some of the pathological features of multiple sclerosis (MS) we transplanted peripheral blood lymphocytes (PBLs) from seven patients with MS into the peritoneal activity of 28 severe combined immunodeficiency (MS-SCID) mice. Seven SCID mice were also transplanted with PBLs from two healthy subjects (hu-SCID). Animals were sacrificed between 2 and 8 weeks after transplantation (a.t.). Polymerase chain reaction(PCR) using primers able to amplify the HLA-DQα region showed presence of human cells in neural tissues of MS-SCID mice. Immunocytochemical analysis revealed the scattered appearance of human lymphocytes (mostly CD45RO + T cells) in the meningeal space and choroid plexuses of MS-SCID brains. However, human lymphocytes were similarly found in brains of hu-SCID mice. Both groups of mice never showed signs or symptoms of neurological impairment. Our results indicate that the simple transplantation of lymphocytes from MS patients into SCID mice is not likely to produce an MS-like pathology.

To study the extent of systemic immunodysregulation in multiple sclerosis (MS) we measured immunoglobulin (Ig)G, A, and M levels and studied their migrational properties after agarose isoelectric focusing in serum, cerebrospinal fluid (CSF) and tear samples from 18 MS patients, 17 patients with other neurological diseases (OND), and tears and serum samples from ten normal controls (NC). A slight elevation of total IgG, IgM and IgA levels was detected in tears from patients with MS and OND compared to NC. Of the five patients (two MS, three OND) that showed IgG oligoclonal bands (OCB) in tears, only one MS patient showed unique bands in tears not seen in the paired CSF and serum. We never found IgA, and IgM OCB in serum, CSF or tear samples. Our results suggest that polyclonal Igs are systemically elevated during chronic neurological inflammatory diseases. Oligoclonal Ig in MS, although occasionally detectable in tears, is mainly confined to the central nervous system and appears restricted to class G.

Stiff-man syndrome is a rare neurological disorder characterized by skeletal muscle rigidity and spasms in which detection of circulating anti-glutamic acid decarboxylase antibodies has suggested an autoimmune pathogenesis. To further define the role of autoimmunity in the pathogenesis, we studied anti-glutamic acid decarboxylase antibodies, as well as organ- and non-organ-specific autoantibodies in 13 patients with stiff-man syndrome and 127 patients with other neurological disorders. Thyrogastric antibodies were more frequent in patients with stiff-man syndrome (46%) than in those with other neurological disorders (12%) (p < 0.05). Non-organ-specific antibodies were found at a similar frequency in the patients with stiff-man syndrome (61%) and those with other neurological disorders (65%). Islet-cell autoantibodies and anti-glutamic acid decarboxylase antibodies were more common in stiff-man syndrome patients (38% and 31%) compared to the patients with other neurological disorders (6% and 3%, respectively; p < 0.001). With the exception of 1 patient in the other neurological disorders group, anti-glutamic acid decarboxylase antibodies were always associated with islet-cell autoantibodies. Four patients with stiff-man syndrome had an associated solid tumor: 3 of them had antibodies recognizing a 125/130-kd protein and not glutamic acid decarboxylase.(ABSTRACT TRUNCATED AT 250 WORDS)

Some of the most prominent scientists in the field of neuroimmunology (C. Raine, C. C. Bernard, and W. Hickey) have reported in the past that C57BL/6 mice are resistant to experimental autoimmune encephalomyelitis (EAE) induced by active immunization with myelin basic protein (MBP) (for example, Ref

INTRODUCTION: Multiple sclerosis (MS) is characterized by the presence in the central nervous system (CNS) of perivascular inflammatory infiltrates containing auto-reactive T and B cells and activated macrophages thus indicating that MS is a T cell-mediated CNS-confined chronic inflammatory demyelinating disease in which the ultimate effector cell is the activated macrophage. DEVELOPMENT: The inflammatory process, leading to patchy demyelination and axonal loss, is mainly sustained by pro-inflammatory cytokines that modulate at different levels the pathogenic process underlying MS. Cytokines can 1. Sustain the 'putative' CNS-confined inflammatory process leading to the development of myelin-specific T cells; 2. Activate circulating myelin-specific T cells and shape their repertoire (Th1 versus Th2 pattern); 3. Induce the CNS recruitment of non antigen specific T cells and myelinotoxic effector cells (monocyte/macrophages) from the periphery, and 4. Cause direct oligodendrotoxicity (TNF alpha) or induce the secretion of myelinotoxic substances. The present chapter will focus on the mechanisms sustaining the activity of pro-inflammatory cytokines in MS pathogenesis.

A avaliação nutritiva de dietas contendo bagaço de cana submetido a diferentes tratamentos alcalinos foi realizada através de um ensaio de digestibilidade "in vivo", pelo método da coleta total de fezes, utilizando-se como unidades experimentais, oito bubalinos machos, com idade média de 18 meses, distribuídas em dois quadrados latinos de 4 x 4. As dietas continham na matéria original 40% de rolão de milho, 30% de farelo de algodão e 30% de bagaço submetido aos seguintes tratamentos: “in natura”, tratado com 4% de NaOH, 4% de Ca(OH)2 e 2% NaOH + 2% Ca(OH)2, respectivamente para as dietas A, B, C e D. Os consumos de matéria seca (6,58 e 7,80 kg/animal/dia) e consumo de matéria seca por unidade de peso metabólico (101,60 e 118,6 g/kg0,75, respectivamente para a dieta contendo bagaço “in natura” e para a média das dietas bagaço alcalinizado, não apresentaram dif'erenças estatisticamente significativas. Com relação ao consumo de água, não se observou efeito estatisticamente significativo das dietas sobre este parâmetro, estando o mesmo intimamente relacionado ao consumo de matéria seca, apresentando um valor médio de 5,83 l/kg de matéria seca consumida. Os coeficientes de digestibilidade aparente da matéria seca, proteína bruta, extrato etéreo, fibra bruta e extrativos não nitrogenados, cujos valores médios foram respectivamente 56,72; 61,63; 71,34; 47,01 e 60,94%, assim como os nutrientes digestivos totais, com valor médio de 57,37% não apresentaram diferenças estatisticamente significativas para as dietas experimentais. No tocante aos constituintes da parede celular, observaram-se maiores coeficientes de digestibilidade da fibra em detergente neutro e hemicelulose (P< 0,05) para as dietas contendo bagaço tratato com NaOH e mistura de NaOH + Ca(OH)2, dietas B e D, as quais apresentaram valores de 55,52 e 56,71%: 73,08 e 74,62; em comparação a dieta contendo bagaço "in natura", dieta A, com valores de 51,12 e 68,35%, respectivamente para fibra em detergente neutro e hemicelulose. Observou-se também um maior coeficiente de digestibilidade da fibra em detergente ácido (FDA) (P< 0,05), da dieta B (bagaço tratado com NaOH) em relação a dieta A (bagaço “in natura”), cujos valores observados foram respectivamente de 45,72 e 40,61%. No tocante à digestibilidade da celulose, não se observaram diferenças significativas entre as dietas, que apresentaram um valor médio de coeficiente de digestibilidade da ordem de 49,79%. Os valores de energia digestivel (kcal/kg) determinados evidenciaram uma superioridade (P< 0,05) da dieta D (bagaço tratado com NaOH + Ca(OH)2) em relação a dieta C (bagaço tratado com Ca(OH)2). Concluiu-se que os tratamentos alcalinos com 4% NaOH e 2% NaOH + 2% Ca(OH)2 foram mais eficazes na melhoria do valor nutritivo do bagaço de cana, pelo aumento que estes promoveram na digestibilidade da FDN e hemicelulose e que o método convencional de análise (Weende) não foi suficientemente preciso para determinar estas melhorias. Concluiu-se também que o nivel de 30% de bagaço de cana na dieta foi excessivo, pois o consumo e a digestibilidade da matéria seca das dietas estiveram abaixo dos niveis desejáveis. Nutritive value of diets contained alkali treated sugarcane bagasse, was realized through a "in vivo" digestibility trial, with buffaloes, by the total feces collection method. The diets were formulated to contain 40% ground ear corn with husks, 30% cottonseed meal, and 30% treated sugarcane bagasse: "in natura", treated with NaOH, 4% Ca(OH) and 2% NaOH plus 2% Ca(OH)2, respectively to diets A, B, C and D. Dry matter intake (6.58 and 7.80 kg/anímal/day) and dry matter intake per matabolic body size (101.60 and 118.60 g/kg0,75), respectively for the diet with sugarcane bagasse "in natura" and for the average of diets with treated sugarcane bagasse, were not statisticaly different. Water intake was not statisticaly different any the diets and averaged 5,8 l/kg DM. The apparent digestibility of dry matter, crude protein, ether extract, crude fiber, nitrogen free extract and the calculated values for the total digestible nutrients (TDN) were not statisticaly different for the experimental diets (56.72; 61.63; 71.34; 47.01; 60.94 and 57.37%, respectively). Digestibility of NDF and hemicellulose were higher (P< .05) for diets with bagasse plus NaOH and mixture of NaOH plus Ca(OH)2 (56.52 and 56.71%; 73.08 and 74.62%, respectively). It was observed a higher digestibility of ADF (P< .05) for the diet B (bagasse plus NaOH), as compared to the diet A, bagasse “in natura" (45.72 and 40.61%, respectively). Cellulose digestibility was not different any the diets and averaged 49.79%. The digestible energy (DE) was higher (P< .05) for diet B, as compared to diet C. It was concluded that the alkali treatments with 4% NaOH and 2% NaOH + 2% Ca(OH)2, were more efficient in improving the nutritive value of sugarcane bagasse, reflexted mostly bu the increase in the NDF andhemicellulose digestibility fractions. The proximate system (Weende) was not sufficiently accurated to determine this improvement. It was also concluded that the level of 30% of sugarcane bagasse in the diets, was harmfull to the animals, since the intake and dry matter digestibility were below expected levels.

The exclusive detrimental role of proinflammatory cytokines in demyelinating diseases of the CNS, such as multiple sclerosis, is controversial. Here we show that the intrathecal delivery of an HSV-1-derived vector engineered with the mouse IFN-γ gene leads to persistent (up to 4 wk) CNS production of IFN-γ and inhibits the course of a chronic-progressive form of experimental autoimmune encephalomyelitis (EAE) induced in C57BL/6 mice by myelin oligodendrocyte glycoprotein (MOG)35–55. Mice treated with the IFN-γ-containing vector before EAE onset showed an earlier onset but a milder course of the disease compared with control mice treated with the empty vector. In addition, 83% of IFN-γ-treated mice completely recovered within 25 days post immunization, whereas control mice did not recover up to 60 days post immunization. Mice treated with the IFN-γ-containing vector within 1 wk after EAE onset partially recovered from the disease within 25 days after vector injection, whereas control mice worsened. Recovery from EAE in mice treated with IFN-γ was associated with a significant increase of CNS-infiltrating lymphocytes undergoing apoptosis. During the recovery phase, the mRNA level of TNFR1 was also significantly increased in CNS-infiltrating cells from IFN-γ-treated mice compared with controls. Our results further challenge the exclusive detrimental role of IFN-γ in the CNS during EAE/multiple sclerosis, and indicate that CNS-confined inflammation may induce protective immunological countermechanisms leading to a faster clearance of encephalitogenic T cells by apoptosis, thus restoring the immune privilege of the CNS.

Multiple sclerosis (MS) is characterized by the presence in the central nervous system (CNS) of perivascular inflammatory infiltrates containing, among others, autoreactive T cells and activated macrophages. These observations indicate that MS is a T cell-mediated CNS-confined chronic inflammatory demyelinating disease in which the ultimate effector cell is the activated macrophage. The inflammatory process, leading to patchy demyelination and axonal loss, is mainly sustained by pro-inflammatory cytokines that, along with chemokines, adhesion molecules and metalloproteases, modulate at different levels the pathogenic process underlying MS. Due to their central role in MS pathogenesis, “inflammatory” molecules might represent suitable peripheral markers of disease (disease-trail) and/or disease activity (state-trait). However, reliable disease-trait or state-trait immunological markers for MS have not yet been identified. The intrinsic characteristics of these molecules (i. e. autocrine/paracrine activity, short half-life, redundancy) may in part explain their inconsistency as disease markers. Additionally, the unreliability of methodologies and the lack of careful patient stratification can also, at least in part, account for the unsatisfactory results so far obtained.

Multiple sclerosis (MS) is an immune-mediated inflammatory disease of the central nervous system (CNS) that might benefit from anti-inflammatory therapies. However, systemic delivery of anti-inflammatory drugs in MS patients has so far been disappointing, mostly due to the limited capacity of these molecules to enter the CNS. We injected into the cisterna magna (i.c.) of Biozzi AB/H mice affected by a relapsing–remitting form of experimental autoimmune encephalomyelitis (EAE), the animal model of MS, a non-replicative herpes simplex virus (HSV) type-1-derived vector containing the interleukin (IL)-4 gene (d120:LacZ:IL-4). CNS delivery of the d120:LacZ:IL-4 vector, after EAE onset, induced the in situ production of IL-4 by CNS-resident cells facing the cerebrospinal fluid (CSF) spaces and reduced by 47% (P

Background: Alterations of synaptic transmission induced by inflammatory activity have been linked to the pathogenic mechanisms of multiple sclerosis (MS). Regulated upon activation, normal T-cell expressed, and secreted (RANTES) is a pro-inflammatory chemokine involved in MS pathophysiology, potentially able to regulate glutamate release and plasticity in MS brains, with relevant consequences on the clinical manifestations of the disease. Objective: To assess the role of RANTES in the regulation of cortical excitability. Methods: We explored the association of RANTES levels in the cerebrospinal fluid (CSF) of newly diagnosed MS patients with magnetic resonance imaging (MRI) and laboratory measures of inflammatory activity, as well its role in the control of cortical excitability and plasticity explored by means of transcranial magnetic stimulation (TMS), and in hippocampal mouse slices in vitro. Results: CSF levels of RANTES were remarkably high only in active MS patients and were correlated with the concentrations of interleukin-1β. RANTES levels were associated with TMS measures of cortical synaptic excitability, but not with long-term potentiation (LTP)-like plasticity. Similar findings were obtained in mouse hippocampal slices in vitro, where we observed that RANTES enhanced basal excitatory synaptic transmission with no effect on LTP. Conclusion: RANTES correlates with inflammation and synaptic excitability in MS brains.

Context It has been reported that the opioid peptide β-endorphin (BE) has immunosuppressive effects. Interferon beta (IFN-β) is a well-established therapy for multiple sclerosis (MS), but immunological mechanisms underlying its beneficial effects in MS are partially undefined. Objectives To determine BE levels in peripheral blood mononuclear cells (PBMCs) of patients with relapsing-remitting MS during different phases of disease activity and the possible modulating effects of IFN-β treatment on PBMC BE synthesis in patients with MS. Design We measured BE levels in blood samples collected from 6 patients with MS who had not experienced clinical changes during the previous 3 months (patients with stable MS) and from 7 patients with MS during a clinical relapse. We also surveyed BE levels in PBMC samples from 8 patients with MS before treatment and for 6 months after the beginning of IFN-β administration. The control group was 13 healthy subjects. Results Low PBMC BE levels were detected in patients with stable MS and in those entering IFN-β treatment compared with control subjects. Increased BE concentrations were observed in MS patients experiencing a clinical relapse compared with patients with stable MS. During IFN-β treatment, the levels of BE in PBMC samples from patients with MS increased significantly (after 1 month, P = .02; after 3 months, P = .007; and after 6 months, P = .16). Conclusions A reduction of BE levels was present in patients with clinically inactive MS. Treatment with IFN-β seems to induce an increase of this opioid in PBMCs of MS patients. The increase of BE concentration during a clinical relapse may represent a possible control mechanism aimed at counterbalancing the inflammatory phase of the disease.

The cysteine protease caspase-1 plays a crucial part in the inflammatory process due to its ability to proteolitically activate proinflammatory cytokine precursors, such as interleukin (IL)-1β and IL-18. Multiple sclerosis is a chronic inflammatory demyelinating disease of the CNS in which the pathogenic process is mainly orchestrated by proinflammatory cytokines. The role of caspase-1 in multiple sclerosis was evaluated by measuring its mRNA levels in peripheral blood mononuclear cells (PBMCs) from seven patients with relapsing-remitting multiple sclerosis every 15 days over a 1 year period. The recorded levels were compared with clinical and MRI evidence of disease activity. Brain MRI was performed monthly in each patient. Caspase-1 mRNA levels were significantly increased in PBMCs from patients with multiple sclerosis compared with healthy controls (p

Alzheimer’s disease (AD) is a neurodegenerative disorder. The pathohistological features in AD are intracellular accumulation of neurofibrillary tangles and extracellular senile plaques. Plaque deposition leads to recruitment and activation of microglial cells, which induces neuroinflammation and drives neurodegeneration. Recent evidence show that soluble pre-fibrillar Aβ species, rather than insoluble fibrils, are highly neurotoxic and correlate with disease severity.Hence, preventing formation of soluble Aβ and its interaction with neurons is a major goal in AD. Despite massive efforts, how extracellular factors regulate assembly of Aβ peptide and neurotoxic activity of Aβ species is still largely undefined. Recent studies indicate that Extracellular Vesicles (EVs), including exosomes and PM-derived microvesicles (MVs), may influence Aβ neurotoxicity. Our findings reveal that production of microglial MVs (m-MVs) is strikingly high in patients with mild cognitive impairment and AD as compared to healthy controls and positively correlates with markers of neurodegeneration and hippocampal atrophy. Furthermore we found that MVs isolated from the CSF of AD patients are toxic to cultured hippocampal neurons. Through in vitro studies we demonstrate that the m-MVs promote generation of neurotoxic soluble species from almost inert Aβ aggregates, which is mediated by lipid components of MVs. Our findings suggest that m-MVs favor formation of neurotoxic Aβ species throughout the brain, possibly representing the mechanism behind transynaptic spread of Aβ in AD. On the other hand, studies conducted by Yuyama et al 2012, 2014 & An et al 2013 suggest that exosomes produced by neurons may exert opposite action by neutralizing neurotoxicity of soluble Aβ. To verify if the overall effect of exosomes and MVs on Aβ neurotoxicity may vary depending on parental cell type we are currently studying the influence of EVs (exosomes & MVs) derived from distinct brain cells on Aβ toxicity and assembly.

Background and purpose: Clinical outcomes of multiple sclerosis (MS) patients affected by coronavirus disease 2019 (COVID-19) have been thoroughly investigated, but a further analysis on main signs and symptoms and their risk factors still needs attention. The objective of this study was to group together and describe based on similarity the most common signs and symptoms of COVID-19 in MS patients and identify all factors associated with their manifestation. Method: Logistic and linear regression models were run to recognize factors associated with each pooled group of symptoms and their total number. Results: From March 2020 to November 2021, data were collected from 1354 MS patients with confirmed infection of COVID-19. Ageusia and anosmia was less frequent in older people (odds ratio [OR] 0.98; p=0.005) and more in smoker patients (OR 1.39; p=0.049). Smoke was also associated with an incremental number of symptoms (OR 1.24; p=0.031), substance abuse (drugs or alcohol), conjunctivitis and rash (OR 5.20; p=0.042) and the presence of at least one comorbidity with shortness of breath, tachycardia or chest pain (OR 1.24; p=0.008). Some disease-modifying therapies were associated with greater frequencies of certain COVID-19 symptoms (association between anti-CD20 therapies and increment in the number of concomitant symptoms: OR 1.29; p=0.05). Differences in frequencies between the three waves were found for flu-like symptoms (G1, p=0.024), joint or muscle pain (G2, p=0.013) and ageusia and anosmia (G5, p < 0.001). All cases should be referred to variants up to Delta. Conclusion: Several factors along with the choice of specific therapeutic approaches might have a different impact on the occurrence of some COVID-19 symptoms.

Diferentes tipos de terapias vêm sendo introduzidas como tratamento alternativo ao combate a diversos tipos de distúrbios humanos e entre eles, a utilização de chás com plantas medicinais vem se destacando pelo custo/benefício, facilidade de aquisição e administração. O objetivo deste estudo foi determinar os constituintes inorgânicos, avaliar a concentração dos elementos As, Ba, Br, Ca, Co, Cr, Cs, Fe, Hf, K, Mg, Mn, Na, Rb, Sb, Sc, Se, Ta, Th, Ti, U, V, Zn e Zr por análise por ativação neutrônica; o Cd, Cu, Ni e Pb pela espectrometria de emissão com plasma indutivamente acoplado e o Hg por espectrometria de absorção atômica com geração de vapor frio nas plantas medicinais e seus extratos cuja comercialização é regulamentada pela Agência Nacional de Vigilância Sanitária (ANVISA). A relevância dessas análises é justificada pela necessidade de contribuir para a recomendação destas plantas como fontes desses minerais na dieta. As técnicas utilizadas apresentaram sensibilidade adequada na obtenção da concentração de todos os elementos analisados, foram encontrados elementos considerados tóxicos em concentrações não prejudiciais ao organismo humano. Este estudo também possibilitou relacionar a concentração elementar nas espécies analisadas por meio das análises de coeficiente de correlação e da análise de agrupamento. Pode-se com isso, verificar a confirmação da similaridade no comportamento de grupos de elementos em relação à variação de suas concentrações observadas nas plantas e em seus extratos. Os elementos que exercem papel importante no metabolismo foram determinados em concentrações que podem auxiliar tanto no tratamento como na carência destes elementos no organismo, quando introduzido na dieta alimentar. Different types of therapies have been introduced as an alternative treatment to combat different types of human disorders. Among them, the use of herbal teas has been highlighted by the cost/benefit, easiness of acquisition and administration. The aim of this study was to determine the inorganic constituents, and evaluate the element concentrations of As, Ba, Br, Ca, Co, Cr, Cs, Fe, Hf, K, Mg. Mn, Na, Rb, Sb, Sc, Se, Ta, Th, Ti, U, V, Zn and Zr by neutron activation analysis; and Cd, Cu, Ni and Pb, by atomic emission spectrometry, with inductively coupled plasma source and Hg, by atomic absorption spectrometry, with cold vapor generation in medicinal plants and their extracts, whose marketing was recently regulated by National Health Surveillance Agency (ANVISA). The relevance of these analyses is justified by the need of contributing to the recommendation of such plants as sources of minerals in the diet and, also, to verify if their concentrations cam pose some harm to the organism. The techniques showed adequate sensitivity in determining the concentration for most of the elements. Toxic elements were found in concentration not harmful to the human body. The results, also, allowed possible to correlate the elemental concentration in the analyzed species, by the determination the correlation coefficients and applications of cluster analysis. From these results it was confirmers in the groups of elements, regarding the variation of the concentrations observed in some plants and their extracts. The elements that play important roles in the human metabolism were determined in concentrations that can help both, to avoid the lack of these elements in the organisms, from the diet, and in treatment of disease.