Soluble Receptor Isoform of IFN-Beta (sIFNAR2) in Multiple Sclerosis Patients and Their Association With the Clinical Response to IFN-Beta Treatment

Alternative splicing; Soluble receptors; IFNAR; Interferon beta; Multiple sclerosisEmpalmament alternatiu; Receptors solubles; IFNAR; Interferó beta; Esclerosi múltipleSplicing alternativo; Receptores solubles; IFNAR; Interferón beta; Esclerosis múltiplePurpose: Interferon beta receptor 2 subunit (IFNAR2) can be produced as a transmembrane protein, but also as a soluble form (sIFNAR2) generated by alternative splicing or proteolytic cleavage, which has both agonist and antagonist activities for IFN-β. However, its role regarding the clinical response to IFN-β for relapsing-remitting multiple sclerosis (RRMS) is unknown. We aim to evaluate the in vitro short-term effects and after 6 and 12 months of IFN-β therapy on sIFNAR2 production and their association with the clinical response in MS patients. Methods: Ninety-four RRMS patients were included and evaluated at baseline, 6 and 12 months from treatment onset. A subset of 41 patients were classified as responders and non-responders to IFN-β therapy. sIFNAR2 serum levels were measured by ELISA. mRNA expression for IFNAR1, IFNAR2 splice variants, MxA and proteases were assessed by RT-PCR. The short-term effect was evaluated in PBMC from RRMS patients after IFN-β stimulation in vitro. Results: Protein and mRNA levels of sIFNAR2 increased after IFN-β treatment. According to the clinical response, only non-responders increased sIFNAR2 significantly at both protein and mRNA levels. sIFNAR2 gene expression correlated with the transmembrane isoform expression and was 2.3-fold higher. While MxA gene expression increased significantly after treatment, IFNAR1 and IFNAR2 only slightly increased. After short-term IFN-β in vitro induction of PBMC, 6/7 patients increased the sIFNAR2 expression. Conclusions: IFN-β administration induces the production of sIFNAR2 in RRMS and higher levels might be associated to the reduction of therapeutic response. Thus, levels of sIFNAR2 could be monitored to optimize an effective response to IFN-β therapy.This research was funded by grants from the Instituto de Salud Carlos III and co-funded by European Regional Development Fund (ERDF), Technological Development Project in health DTS/1800045 to BO-M. BO-M holds a contract from Red Andaluza de Investigacion Clínica y Traslacional en Neurología (Neuro-reca) (RIC-0111-2019). PA-G is supported by Promoción de Empleo Joven e Implantación de la Garantía Juvenil 2018 (PEJ2018-002719-A). JR-B is supported by grants from Red Temática de Investigación Cooperativa, Red Española de Esclerosis Multiple REEM (RD16/0015/0010). LL holds a Nicolás Monardes research contract (RC-002-2019) from the Andalusian Ministry of Health and Family. IB-M holds a pFIS contract (FI19/00139) from the Spanish Science and Innovation Ministry

Determinació del patró de miRNA en líquid cefaloraquidi de pacients amb esclerosi múltiple (estudi miEM)

Multiple sclerosis is a chronic neurological disease of the central nervous system with unknown, autoimmune mechanism, and a progressive and disabling course. Lipid-specific oligoclonal IgM bands in cerebrospinal fluid have been associated with worse clinical forms of the disease. MicroRNAs regulate genetic expression and participate in physiological processes and in different diseases. For this reason, they have been postulated as potential molecular biomarkers both of diagnosis and prognosis in many diseases. In the research for this doctoral thesis, the role of microRNAs in cerebrospinal fluid in the early phases of relapsing-remiting multiple sclerosis has been studied and some microRNAs have been identified as candidates to be biomarkers of the diagnosis of the diesease and are associated to the presence of lipid-specific oligoclonal IgM bands, which have been related with more aggressive forms of multiple sclerosis. The expression of these microRNAs is associated to certain clinical and radiological variables of affected patientsL’esclerosi múltiple és una malaltia neurològica crònica del sistema nerviós central, d’etiologia no ben coneguda, mecanisme autoinmmunitari, curs progressiu i pronòstic heterogeni. Les bandes oligoclonals d’IgM lípidesespecífiques en líquid cefaloraquidi s’han associat a formes clíniques de la malaltia de pitjor pronòstic. Els microRNA regules l’expressió genètica i intervenen en processos fisiològics i en moltes malalties. Per aquest motiu s’estan postulant com a potencials biomarcadors moleculars tant de diagnòstic com de pronòstic en moltes malalties. En aquest treball de tesi doctoral s’ha estudiat el paper dels microRNAs en líquid cefaloraquidi en les fases inicials de les formes remitent-recurrent d’esclerosi múltiple, i s’han identificat alguns microRNAs com a candidats a ésser biomarcadors del diagnòstic de la malaltia i associats a la presència de bandes oligoclonals d’IgM lípidespecífiques que s’han relacionat amb formes més agressives d’esclerosi múltiple. L’expressió d’aquests microRNAs s’ha associat a algunes variables clíniques i radiològiques dels pacients afectat

Evaluating the effects of white matter multiple sclerosis lesions on the volume estimation of 6 brain tissue segmentation methods

BACKGROUND AND PURPOSE: The accuracy of automatic tissue segmentation methods can be affected by the presence of hypointense white matter lesions during the tissue segmentation process. Our aim was to evaluate the impact of MS white matter lesions on the brain tissue measurements of 6 well-known segmentation techniques. These include straightforward techniques such as Artificial Neural Network and fuzzy C-means as well as more advanced techniques such as the Fuzzy And Noise Tolerant Adaptive Segmentation Method, fMRI of the Brain Automated Segmentation Tool, SPM5, and SPM8. MATERIALS AND METHODS: Thirty T1-weighted images from patients with MS from 3 different scanners were segmented twice, first including white matter lesions and then masking the lesions before segmentation and relabeling as WM afterward. The differences in total tissue volume and tissue volume outside the lesion regions were computed between the images by using the 2 methodologies. RESULTS: Total gray matter volume was overestimated by all methods when lesion volume increased. The tissue volume outside the lesion regions was also affected by white matter lesions with differences up to 20 cm3 on images with a high lesion load (≈50 cm3). SPM8 and Fuzzy And Noise Tolerant Adaptive Segmentation Method were the methods less influenced by white matter lesions, whereas the effect of white matter lesions was more prominent on fuzzy C-means and the fMRI of the Brain Automated Segmentation Tool. CONCLUSIONS: Although lesions were removed after segmentation to avoid their impact on tissue segmentation, the methods still overestimated GM tissue in most cases. This finding is especially relevant because on images with high lesion load, this bias will most likely distort actual tissue atrophy measurements

Quantifying brain tissue volume in multiple sclerosis with automated lesion segmentation and filling

Lesion filling has been successfully applied to reduce the effect of hypo-intense T1-wMultiple Sclerosis (MS) lesions on automatic brain tissue segmentation. However, a study of fully automated pipelines incorporating lesion segmentation and lesion filling on tissue volume analysis has not yet been performed. Here,we analyzed the % of error introduced by automating the lesion segmentation and filling processes in the tissue segmentation of 70 clinically isolated syndrome patient images. First of all, images were processed using the LST and SLS toolkits with different pipeline combinations that differed in either automated ormanual lesion segmentation, and lesion filling or masking out lesions. Then, images processed following each of the pipelines were segmented into gray matter (GM) andwhite matter (WM) using SPM8, and compared with the same images where expert lesion annotationswere filled before segmentation. Our results showed that fully automated lesion segmentation and filling pipelines reduced significantly the % of error inGMandWMvolumeon images ofMS patients, and performed similarly to the images where expert lesion annotations were masked before segmentation. In all the pipelines, the amount of misclassified lesion voxels was the main cause in the observed error inGMandWMvolume. However, the % of error was significantly lower when automatically estimated lesionswere filled and not masked before segmentation. These results are relevant and suggest that LST and SLS toolboxes allow the performance of accurate brain tissue volume measurements without any kind of manual intervention, which can be convenient not only in terms of time and economic costs, but also to avoid the inherent intra/inter variability between manual annotationsS. Valverde holds a FI-DGR2013 grant from the Generalitat de Catalunya. E. Roura holds a BR-UdG2013 grant. This work has been partially supported by “La Fundació la Marató de TV3” and by Retos de Investigación TIN2014-55710-

Effects of repeated long-term psychosocial stress and acute cannabinoid exposure on mouse corticostriatal circuitries: implications for neuropsychiatric disorders

Vulnerability to psychiatric manifestations is achieved by the influence of genetic and environment including stress and cannabis consumption. Here, we used a psychosocial stress model based on resident-intruder confrontations to study the brain corticostriatal-function, since deregulation of corticostriatal circuitries has been reported in many psychiatric disorders. CB1 receptors are widely expressed in the central nervous system and particularly, in both cortex and striatum brain structures. AIMS AND METHODS: The investigation presented here is addressed to assess the impact of repeated stress following acute cannabinoid exposure on behavior and corticostriatal brain physiology by assessing mice behavior, the concentration of endocannabinoid and endocannabinoid-like molecules and changes in the transcriptome. RESULTS: Stressed animals urinated frequently; showed exacerbated scratching activity, lower striatal N-arachidonylethanolamine (AEA) levels and higher cortical expression of cholinergic receptor nicotinic alpha 6. The cannabinoid agonist WIN55212.2 diminished locomotor activity while the inverse agonist increased the distance travelled in the center of the open field. Upon CB1 activation, N-oleoylethanolamide and N-palmitoylethanolamide, two AEA congeners that do not interact directly with cannabinoid receptors, were enhanced in the striatum. The co-administration with both cannabinoids induced an up-regulation of striatal FK506 binding protein 5. The inverse agonist in controls reversed the effects of WIN55212.2 on motor activity. When Rimonabant was injected under stress, the cortical levels of 2-arachidonoylglycerol were maximum. The agonist and the antagonist influenced the cortical expression of cholinergic receptor nicotinic alpha 6 and serotonin transporter neurotransmitter type 4 in opposite directions, while their co-administration tended to produce a null effect under stress. CONCLUSIONS: The endocannabinoid system had a direct effect on serotoninergic neurotransmission and glucocorticoid signaling. Cholinergic receptor nicotinic alpha-6 was shown to be deregulated in response to stress and following synthetic cannabinoid drugs thus could confer vulnerability to cannabis addiction and psychosis. Targeting the receptors of endocannabinoids and endocannabinoid-like mediators might be a valuable option for treating stress-related neuropsychiatric symptom

A new risk variant for multiple sclerosis at 11q23.3 locus is associated with expansion of CXCR5+ circulating regulatory T cells

Genome-wide association studies and meta-analysis have contributed to the identification of more than 200 loci associated with multiple sclerosis (MS). However, a proportion of MS heritability remains unknown. We aimed to uncover new genetic variants associated with MS and determine their functional effects. For this, we resequenced the exons and regulatory sequences of 14 MS risk genes in a cohort of MS patients and healthy individuals (n = 1,070) and attempted to validate a selection of signals through genotyping in an independent cohort (n = 5,138). We identified three new MS-associated variants at C-X-C motif chemokine receptor 5 (CXCR5), Ts translation elongation factor, mitochondrial (TSFM) and cytochrome P450 family 24 subfamily A member 1 (CYP24A1). Rs10892307 resulted in a new signal at the CXCR5 region that explains one of the associations with MS within the locus. This polymorphism and three others in high linkage disequilibrium mapped within regulatory regions. Of them, rs11602393 showed allele-dependent enhancer activity in the forward orientation as determined by luciferase reporter assays. Immunophenotyping using peripheral blood mononuclear cells from MS patients associated the minor allele of rs10892307 with increased percentage of regulatory T cells expressing CXCR5. This work reports a new signal for the CXCR5 MS risk locus and points to rs11602393 as the causal variant. The expansion of CXCR5+ circulating regulatory T cells induced by this variant could cause its MS association.Funding: The genotyping service was carried out at CEGEN-PRB2-ISCIII and was supported by grant PT13/0001, ISCIII-SGEFI/FEDER. This work was also supported by the following grants: BFU2016-77961-P (AEI/FEDER); 2017-SGR-00702 (Direcció General de Recerca, Generalitat de Catalunya); Unidad de Excelencia María de Maeztu funded by the MINECO (MDM-2014-0370); Proyectos de Investigación en Salud (FIS PI12/02229, PI15/00587, PI16/01259); Red Española de Esclerosis Múltiple (RD16/0015/0004 to M.C., RD16/0015/0002; RD16/0015/0005 to K.V.; RD16/0015/0016) integrated in the Plan Estatal I+D+I and cofunded by Instituto de Salud Carlos III and the Fondo Europeo de Desarrollo Regional (FEDER; “Otra forma de hacer Europa”); SAF2016-80595-C2-1-P (Ministerio de Economía, Industria y Competitividad

Neuromyelitis optica spectrum disorders: comparison according to the phenotype and serostatus

OBJECTIVE: To (1) determine the value of the recently proposed criteria of neuromyelitis optica (NMO) spectrum disorder (NMOSD) that unify patients with NMO and those with limited forms (NMO/LF) with aquaporin-4 immunoglobulin G (AQP4-IgG) antibodies; and (2) investigate the clinical significance of the serologic status in patients with NMO. METHODS: This was a retrospective, multicenter study of 181 patients fulfilling the 2006 NMO criteria (n = 127) or NMO/LF criteria with AQP4-IgG (n = 54). AQP4-IgG and myelin oligodendrocyte glycoprotein immunoglobulin G (MOG-IgG) antibodies were tested using cell-based assays. RESULTS: Patients were mainly white (86%) and female (ratio 6.5:1) with median age at onset 39 years (range 10-77). Compared to patients with NMO and AQP4-IgG (n = 94), those with NMO/LF presented more often with longitudinally extensive transverse myelitis (LETM) (p < 0.001), and had lower relapse rates (p = 0.015), but similar disability outcomes. Nonwhite ethnicity and optic neuritis presentation doubled the risk for developing NMO compared with white race (p = 0.008) or LETM presentation (p = 0.008). Nonwhite race (hazard ratio [HR] 4.3, 95% confidence interval [CI] 1.4-13.6) and older age at onset were associated with worse outcome (for every 10-year increase, HR 1.7, 95% CI 1.3-2.2). Patients with NMO and MOG-IgG (n = 9) had lower female:male ratio (0.8:1) and better disability outcome than AQP4-IgG-seropositive or double-seronegative patients (p < 0.001). CONCLUSIONS: In patients with AQP4-IgG, the similar outcomes regardless of the clinical phenotype support the unified term NMOSD; nonwhite ethnicity and older age at onset are associated with worse outcome. Double-seronegative and AQP4-IgG-seropositive NMO have a similar clinical outcome. The better prognosis of patients with MOG-IgG and NMO suggests that phenotypic and serologic classification is useful.This study was supported in part by Red Española de Esclerosis Múltiple (REEM) Instituto de Salud Carlos III, Spain (RD07/0060/01, P.V.; RD12/0032/0002, A.S.; Marató de TV3 [20141830], F.G.) and Instituto de Salud Carlos III, Spain (CM14/00081; T.A.)