Advertisement

TNFRSF13C/BAFFR P21R and H159Y polymorphisms in multiple sclerosis

Published:September 30, 2019DOI:https://doi.org/10.1016/j.msard.2019.101422

      Highlights

      • BAFFR-signaling enhancing SNP H159Y is strongly associated with multiple sclerosis.
      • The age onset and severity of multiple sclerosis are not correlated with BAFFR SNPs.
      • Approaches interfering with BAFF system are less straight forward than anticipated.

      Abstract

      Recent studies implicate B cells in multiple sclerosis (MS) pathogenesis, and consequently, several molecules participating in B cell survival and proliferation, including B-cell activating factor (BAFF), have recently been analyzed in MS patients. BAFF mediates its function through binding to three receptors; among them, its interaction with the BAFF receptor (BAFFR) is crucial in mediating its survival function. Interestingly, two common polymorphisms of the TNFRSF13C gene, encoding BAFFR, P21R (rs77874543) and H159Y (rs61756766), have been reported to affect BAFFR assembly and signaling. In order to evaluate the possible contribution of BAFFR in MS pathogenesis and/or phenotype, we analyzed both TNFRSF13C/BAFFR polymorphisms in 486 MS patients in relation to their disease severity, their disability status and the age of disease onset and duration. As control group, we used allele frequencies extracted from the Exome Aggregation Consortium (ExAC) Browser. Interestingly, we found a higher prevalence of the H159Y polymorphism in MS patients, suggesting that enhanced BAFFR-signaling might contribute to the disease pathogenesis.

      Keywords

      1. Introduction

      Multiple sclerosis (MS) is a chronic, debilitating neurological disease that begins in early adulthood, with a strong autoimmune background, but no clear etiology. Inflammation is considered a hallmark of acute MS lesions, characterized by infiltration of T and B cells, activated macrophages and microglia (
      • Claes N.
      • Fraussen J.
      • Stinissen P.
      • Hupperts R.
      • Somers V.
      B cells are multifunctional players in multiple sclerosis pathogenesis: insights from therapeutic interventions.
      ;
      • Dendrou C.A.
      • Fugger L.
      Immunomodulation in multiple sclerosis: promises and pitfalls.
      ;
      • Hadjigeorgiou G.M.
      • Kountra P.M.
      • Koutsis G.
      • Tsimourtou V.
      • Siokas V.
      • Dardioti M.
      • Rikos D.
      • Marogianni C.
      • Aloizou A.M.
      • Karadima G.
      • Ralli S.
      • Grigoriadis N.
      • Bogdanos D.
      • Panas M.
      • Dardiotis E.
      Replication study of GWAS risk loci in Greek multiple sclerosis patients.
      ;
      • Hemmer B.
      • Archelos J.J.
      • Hartung H.P.
      New concepts in the immunopathogenesis of multiple sclerosis.
      ), resulting in tissue damage with axonal de- and re-myelination (
      • Stadelmann C.
      Multiple sclerosis as a neurodegenerative disease: pathology, mechanisms and therapeutic implications.
      ). Recent studies implicate B cells in disease pathogenesis, a notion that is also supported by the evidence of the therapeutic role of B cell depletion in the management of the disease (
      • Disanto G.
      • Morahan J.M.
      • Barnett M.H.
      • Giovannoni G.
      • Ramagopalan S.V.
      The evidence for a role of B cells in multiple sclerosis.
      ;
      • Franciotta D.
      • Salvetti M.
      • Lolli F.
      • Serafini B.
      • Aloisi F.
      B cells and multiple sclerosis.
      ).
      Consequently, several molecules participating in B cell proliferation and homeostasis have recently been analyzed in MS patients (
      • Srivastava P.
      • Mujtaba M.A.
      • Singhal M.
      Gene and cytokines expression of multiple sclerosis and its therapeutic regimen: a systemic review.
      ;
      • Steri M.
      • Orru V.
      • Idda M.L.
      • Pitzalis M.
      • Pala M.
      • Zara I.
      • Sidore C.
      • Faa V.
      • Floris M.
      • Deiana M.
      • Asunis I.
      • Porcu E.
      • Mulas A.
      • Piras M.G.
      • Lobina M.
      • Lai S.
      • Marongiu M.
      • Serra V.
      • Marongiu M.
      • Sole G.
      • Busonero F.
      • Maschio A.
      • Cusano R.
      • Cuccuru G.
      • Deidda F.
      • Poddie F.
      • Farina G.
      • Dei M.
      • Virdis F.
      • Olla S.
      • Satta M.A.
      • Pani M.
      • Delitala A.
      • Cocco E.
      • Frau J.
      • Coghe G.
      • Lorefice L.
      • Fenu G.
      • Ferrigno P.
      • Ban M.
      • Barizzone N.
      • Leone M.
      • Guerini F.R.
      • Piga M.
      • Firinu D.
      • Kockum I.
      • Lima Bomfim I.
      • Olsson T.
      • Alfredsson L.
      • Suarez A.
      • Carreira P.E.
      • Castillo-Palma M.J.
      • Marcus J.H.
      • Congia M.
      • Angius A.
      • Melis M.
      • Gonzalez A.
      • Alarcón Riquelme M.E.
      • da Silva B.M.
      • Marchini M.
      • Danieli M.G.
      • Del Giacco S.
      • Mathieu A.
      • Pani A.
      • Montgomery S.B.
      • Rosati G.
      • Hillert J.
      • Sawcer S.
      • D'Alfonso S.
      • Todd J.A.
      • Novembre J.
      • Abecasis G.R.
      • Whalen M.B.
      • Marrosu M.G.
      • Meloni A.
      • Sanna S.
      • Gorospe M.
      • Schlessinger D.
      • Fiorillo E.
      • Zoledziewska M.
      • Cucca F.
      Overexpression of the cytokine BAFF and autoimmunity risk.
      ). Amongst them is B-cell activating factor (BAFF), a TNF-family member which supports the survival of B cells (
      • Mackay F.
      • Schneider P.
      Cracking the BAFF code.
      ). BAFF is predominantly expressed by cells of myeloid origin, such as macrophages, monocytes and dendritic cells (
      • Woodland R.T.
      • Schmidt M.R.
      • Thompson C.B.
      BLyS and B cell homeostasis.
      ); however, non-hematopoietic cells, as astrocytes, can overexpress BAFF, either experimentally in mice following viral brain infection, or in pathological conditions, as primary central nervous system (CNS) lymphomas (
      • Krumbholz M.
      • Theil D.
      • Derfuss T.
      • Rosenwald A.
      • Schrader F.
      • Monoranu C.M.
      • Kalled S.L.
      • Hess D.M.
      • Serafini B.
      • Aloisi F.
      • Wekerle H.
      • Hohlfeld R.
      • Meinl E.
      BAFF is produced by astrocytes and up-regulated in multiple sclerosis lesions and primary central nervous system lymphoma.
      ;
      • Lokensgard J.R.
      • Mutnal M.B.
      • Prasad S.
      • Sheng W.
      • Hu S.
      Glial cell activation, recruitment, and survival of B-lineage cells following MCMV brain infection.
      ;
      • Mackay F.
      • Schneider P.
      Cracking the BAFF code.
      ). Interestingly, BAFF has been detected in MS lesions (
      • Krumbholz M.
      • Theil D.
      • Derfuss T.
      • Rosenwald A.
      • Schrader F.
      • Monoranu C.M.
      • Kalled S.L.
      • Hess D.M.
      • Serafini B.
      • Aloisi F.
      • Wekerle H.
      • Hohlfeld R.
      • Meinl E.
      BAFF is produced by astrocytes and up-regulated in multiple sclerosis lesions and primary central nervous system lymphoma.
      ;
      • Mackay F.
      • Schneider P.
      Cracking the BAFF code.
      ) and changes in BAFF concentrations in the cerebrospinal fluid have been associated with MS severity in humans (
      • Ragheb S.
      • Li Y.
      • Simon K.
      • VanHaerents S.
      • Galimberti D.
      • De Riz M.
      • Fenoglio C.
      • Scarpini E.
      • Lisak R.
      Multiple sclerosis: BAFF and CXCL13 in cerebrospinal fluid.
      ;
      • Thangarajh M.
      • Gomes A.
      • Masterman T.
      • Hillert J.
      • Hjelmstrom P.
      Expression of B-cell-activating factor of the TNF family (BAFF) and its receptors in multiple sclerosis.
      ;
      • Wang H.
      • Wang K.
      • Zhong X.
      • Qiu W.
      • Dai Y.
      • Wu A.
      • Hu X.
      Cerebrospinal fluid BAFF and APRIL levels in neuromyelitis optica and multiple sclerosis patients during relapse.
      ). Recent genetic studies revealed that defined genetic changes stabilizing BAFF mRNA and increasing its expression closely correlate with the incidence of MS (
      • Steri M.
      • Orru V.
      • Idda M.L.
      • Pitzalis M.
      • Pala M.
      • Zara I.
      • Sidore C.
      • Faa V.
      • Floris M.
      • Deiana M.
      • Asunis I.
      • Porcu E.
      • Mulas A.
      • Piras M.G.
      • Lobina M.
      • Lai S.
      • Marongiu M.
      • Serra V.
      • Marongiu M.
      • Sole G.
      • Busonero F.
      • Maschio A.
      • Cusano R.
      • Cuccuru G.
      • Deidda F.
      • Poddie F.
      • Farina G.
      • Dei M.
      • Virdis F.
      • Olla S.
      • Satta M.A.
      • Pani M.
      • Delitala A.
      • Cocco E.
      • Frau J.
      • Coghe G.
      • Lorefice L.
      • Fenu G.
      • Ferrigno P.
      • Ban M.
      • Barizzone N.
      • Leone M.
      • Guerini F.R.
      • Piga M.
      • Firinu D.
      • Kockum I.
      • Lima Bomfim I.
      • Olsson T.
      • Alfredsson L.
      • Suarez A.
      • Carreira P.E.
      • Castillo-Palma M.J.
      • Marcus J.H.
      • Congia M.
      • Angius A.
      • Melis M.
      • Gonzalez A.
      • Alarcón Riquelme M.E.
      • da Silva B.M.
      • Marchini M.
      • Danieli M.G.
      • Del Giacco S.
      • Mathieu A.
      • Pani A.
      • Montgomery S.B.
      • Rosati G.
      • Hillert J.
      • Sawcer S.
      • D'Alfonso S.
      • Todd J.A.
      • Novembre J.
      • Abecasis G.R.
      • Whalen M.B.
      • Marrosu M.G.
      • Meloni A.
      • Sanna S.
      • Gorospe M.
      • Schlessinger D.
      • Fiorillo E.
      • Zoledziewska M.
      • Cucca F.
      Overexpression of the cytokine BAFF and autoimmunity risk.
      ).
      BAFF binds to three receptors, namely BAFFR (BAFF receptor), BCMA (B cell maturation antigen), and TACI (transmembrane activator and calcium modulating ligand interactor) (
      • Kim H.M.
      • Yu K.S.
      • Lee M.E.
      • Shin D.R.
      • Kim Y.S.
      • Paik S.G.
      • Yoo O.J.
      • Lee H.
      • Lee J.O.
      Crystal structure of the BAFF-BAFF-R complex and its implications for receptor activation.
      ;
      • Mackay F.
      • Schneider P.
      Cracking the BAFF code.
      ). While TACI is expressed by mature and terminally differentiated B cells, such as switched memory B cells, marginal zone B cells and plasma cells, and BCMA only by plasma cells, BAFFR expression starts already at the stage of immature/transitional B cells, acting as essential pro-survival receptor of mature follicular and marginal zone B cells (
      • Mackay F.
      • Schneider P.
      Cracking the BAFF code.
      ;
      • Smulski C.R.
      • Eibel H.
      BAFF and BAFF-Receptor in B cell selection and survival.
      ). Interestingly, two functional common polymorphisms of the TNFRSF13C gene, encoding BAFFR, namely P21R (rs77874543) and H159Y (rs61756766), disturb either ligand-independent assembly of BAFFR into oligomers (P21R,
      • Pieper K.
      • Rizzi M.
      • Speletas M.
      • Smulski C.R.
      • Sic H.
      • Kraus H.
      • Salzer U.
      • Fiala G.J.
      • Schamel W.W.
      • Lougaris V.
      • Plebani A.
      • Hammarstrom L.
      • Recher M.
      • Germenis A.E.
      • Grimbacher B.
      • Warnatz K.
      • Rolink A.G.
      • Schneider P.
      • Notarangelo L.D.
      • Eibel H.
      A common single nucleotide polymorphism impairs B-cell activating factor receptor's multimerization, contributing to common variable immunodeficiency.
      ), or increase ligand-independent BAFFR signaling (H159Y,
      • Hildebrand J.M.
      • Luo Z.
      • Manske M.K.
      • Price-Troska T.
      • Ziesmer S.C.
      • Lin W.
      • Hostager B.S.
      • Slager S.L.
      • Witzig T.E.
      • Ansell S.M.
      • Cerhan J.R.
      • Bishop G.A.
      • Novak A.J.
      A BAFF-R mutation associated with non-Hodgkin lymphoma alters TRAF recruitment and reveals new insights into BAFF-R signaling.
      ). TNFRSF13C-P21R was found in a high frequency in patients with common variable immunodeficiency (CVID), but was low to undetectable in patients with chronic lymphocytic leukemia (
      • Jasek M.
      • Bojarska-Junak A.
      • Wagner M.
      • Sobczynski M.
      • Wolowiec D.
      • Rolinski J.
      • Karabon L.
      • Kuśnierczyk P.
      Association of variants in BAFF (rs9514828 and rs1041569) and BAFF-R (rs61756766) genes with the risk of chronic lymphocytic leukemia.
      ;
      • Losi C.G.
      • Silini A.
      • Fiorini C.
      • Soresina A.
      • Meini A.
      • Ferrari S.
      • Notarangelo L.D.
      • Lougaris V.
      • Plebani A.
      Mutational analysis of human BAFF receptor TNFRSF13C (BAFF-R) in patients with common variable immunodeficiency.
      ;
      • Pieper K.
      • Rizzi M.
      • Speletas M.
      • Smulski C.R.
      • Sic H.
      • Kraus H.
      • Salzer U.
      • Fiala G.J.
      • Schamel W.W.
      • Lougaris V.
      • Plebani A.
      • Hammarstrom L.
      • Recher M.
      • Germenis A.E.
      • Grimbacher B.
      • Warnatz K.
      • Rolink A.G.
      • Schneider P.
      • Notarangelo L.D.
      • Eibel H.
      A common single nucleotide polymorphism impairs B-cell activating factor receptor's multimerization, contributing to common variable immunodeficiency.
      ). On the other hand, the TNFFRSF13C-H159Y polymorphism results in a sustained overactivated BAFF-induced signaling and it was found in a high frequency in patients with Non-Hodgkin lymphomas and a Sjogren's syndrome-related lymphoproliferation (
      • Hildebrand J.M.
      • Luo Z.
      • Manske M.K.
      • Price-Troska T.
      • Ziesmer S.C.
      • Lin W.
      • Hostager B.S.
      • Slager S.L.
      • Witzig T.E.
      • Ansell S.M.
      • Cerhan J.R.
      • Bishop G.A.
      • Novak A.J.
      A BAFF-R mutation associated with non-Hodgkin lymphoma alters TRAF recruitment and reveals new insights into BAFF-R signaling.
      ;
      • Papageorgiou A.
      • Mavragani C.P.
      • Nezos A.
      • Zintzaras E.
      • Quartuccio L.
      • De Vita S.
      • Koutsilieris M.
      • Tzioufas A.G.
      • Moutsopoulos H.M.
      • Voulgarelis M.
      A BAFF receptor His159Tyr mutation in Sjogren's syndrome-related lymphoproliferation.
      ).
      Since increased BAFF levels were found to associate with susceptibility for MS, we started to study the possible contribution of BAFFR signaling by analyzing the correlation of both TNFRSF13C/BAFFR polymorphisms with MS pathogenesis and/or phenotype.

      2. Materials & methods

      2.1 Study population

      The study consisted of 486 consecutive patients with MS (male/female: 156/330, mean age: 44 years, range: 72 years), treated and followed-up in the University Hospital of Larissa, Greece. All patients fulfilled the 2010 revised McDonald diagnostic criteria for MS (
      • Polman C.H.
      • Reingold S.C.
      • Banwell B.
      • Clanet M.
      • Cohen J.A.
      • Filippi M.
      • Fujihara K.
      • Havrdova E.
      • Hutchinson M.
      • Kappos L.
      • Lublin F.D.
      • Montalban X.
      • O'Connor P.
      • Sandberg-Wollheim M.
      • Thompson A.J.
      • Waubant E.
      • Weinshenker B.
      • Wolinsky J.S.
      Diagnostic criteria for multiple sclerosis: 2010 revisions to the McDonald criteria.
      ) and the recent diagnostic MRI criteria (
      • Filippi M.
      • Rocca M.A.
      • Ciccarelli O.
      • De Stefano N.
      • Evangelou N.
      • Kappos L.
      • Rovira A.
      • Sastre-Garriga J.
      • Tintorè M.
      • Frederiksen J.L.
      • Gasperini C.
      • Palace J.
      • Reich D.S.
      • Banwell B.
      • Montalban X.
      • Barkhof F.
      MRI criteria for the diagnosis of multiple sclerosis: MAGNIMS consensus guidelines.
      ). Details relating to age of disease onset and disease duration were collected for all patients; disability status was assessed using the Expanded Disability Status Scale (EDSS) (
      • Kurtzke J.F.
      Rating neurologic impairment in multiple sclerosis: an expanded disability status scale (EDSS).
      ), while disease severity was evaluated by means of the Multiple Sclerosis Severity Score (MSSS) (
      • Roxburgh R.H.
      • Seaman S.R.
      • Masterman T.
      • Hensiek A.E.
      • Sawcer S.J.
      • Vukusic S.
      • Achiti I.
      • Confavreux C.
      • Coustans M.
      • le Page E.
      • Edan G.
      • McDonnell G.V.
      • Hawkins S.
      • Trojano M.
      • Liguori M.
      • Cocco E.
      • Marrosu M.G.
      • Tesser F.
      • Leone M.A.
      • Weber A.
      • Zipp F.
      • Miterski B.
      • Epplen J.T.
      • Oturai A.
      • Sοrensen P.S.
      • Celius E.G.
      • Lara N.T.
      • Montalban X.
      • Villoslada P.
      • Silva A.M.
      • Marta M.
      • Leite I.
      • Dubois B.
      • Rubio J.
      • Butzkueven H.
      • Kilpatrick T.
      • Mycko M.P.
      • Selmaj K.W.
      • Rio M.E.
      • Sa M.
      • Salemi G.
      • Savettieri G.
      • Hillert J.
      • Compston D.A.
      Multiple sclerosis severity score: using disability and disease duration to rate disease severity.
      ). MSSS scores were sorted into 3 categories, namely benign: 0–1.999, moderate: 2–6.999, and severe: 7–10 (
      • Dardiotis E.
      • Panayiotou E.
      • Provatas A.
      • Christodoulou K.
      • Hadjisavvas A.
      • Antoniades A.
      • Lourbopoulos A.
      • Pantzaris M.
      • Grigoriadis N.
      • Hadjigeorgiou G.M.
      • Kyriakides T.
      Gene variants of adhesion molecules act as modifiers of disease severity in MS.
      ). As a control, we used the publicly available data from the Exome Aggregation Consortium (ExAC; http://exac.broadinstitute.org) (
      • Lek M.
      • Karczewski K.J.
      • Minikel E.V.
      • Samocha K.E.
      • Banks E.
      • Fennell T.
      • O'Donnell-Luria A.H.
      • Ware J.S.
      • Hill A.J.
      • Cummings B.B.
      • Tukiainen T.
      • Birnbaum D.P.
      • Kosmicki J.A.
      • Duncan L.E.
      • Estrada K.
      • Zhao F.
      • Zou J.
      • Pierce-Hoffman E.
      • Berghout J.
      • Cooper D.N.
      • Deflaux N.
      • DePristo M.
      • Do R.
      • Flannick J.
      • Fromer M.
      • Gauthier L.
      • Goldstein J.
      • Gupta N.
      • Howrigan D.
      • Kiezun A.
      • Kurki M.I.
      • Moonshine A.L.
      • Natarajan P.
      • Orozco L.
      • Peloso G.M.
      • Poplin R.
      • Rivas M.A.
      • Ruano-Rubio V.
      • Rose S.A.
      • Ruderfer D.M.
      • Shakir K.
      • Stenson P.D.
      • Stevens C.
      • Thomas B.P.
      • Tiao G.
      • Tusie-Luna M.T.
      • Weisburd B.
      • Won H.-.H.
      • Yu D.
      • Altshuler D.M.
      • Ardissino D.
      • Boehnke M.
      • Danesh J.
      • Donnelly S.
      • Elosua R.
      • Florez J.C.
      • Gabriel S.B.
      • Getz G.
      • Glatt S.J.
      • Hultman C.M.
      • Kathiresan S.
      • Laakso M.
      • McCarroll S.
      • McCarthy M.I.
      • McGovern D.
      • McPherson R.
      • Neale B.M.
      • Palotie A.
      • Purcell S.M.
      • Saleheen D.
      • Scharf J.M.
      • Sklar P.
      • Sullivan P.F.
      • Tuomilehto J.
      • Tsuang M.T.
      • Watkins H.C.
      • Wilson J.G.
      • Daly M.J.
      • MacArthur D.G.
      Exome Aggregation, C.
      Analysis of protein-coding genetic variation in 60,706 humans.
      ), since they include much larger numbers of individuals analyzed yielding higher significance. These data do not differ from our cohort of 316 healthy individuals previously analyzed for the prevalence of TNFRSF13C/BAFFR polymorphisms in a Greek population (
      • Kompoti M.
      • Michopoulos A.
      • Michalia M.
      • Clouva-Molyvdas P.M.
      • Germenis A.E.
      • Speletas M.
      Genetic polymorphisms of innate and adaptive immunity as predictors of outcome in critically ill patients.
      ).
      The ExAC database lists 32 TNFRSF13C variants composed from 30 missense mutations and 2 copy number variants. As control group for the Greek MS cohort, we used then (Non-Finnish) European population, in which 455 of 65,566 total alleles carry the H159Y missense mutation encoded by the TNFSRF13C variant 22:42,321,451 G/A (dbSNP rs1756766). This corresponds to an allele frequency of 1/147 (0.006787). The allele frequencies of the other populations listed in the ExAC database are: Finnish 1/400, Latino 1/133, African 1/462, East Asian 0/8608, South Asian 1/179 alleles.
      Written informed consent was obtained from all the participants. The study was approved by the local institutional review board (University Hospital of Larissa, Greece) and was carried out in accordance with the principles of the Helsinki Declaration.

      2.2 Molecular studies

      Genomic DNA was extracted from peripheral blood using the QIAamp DNA Blood Mini Kit (Qiagen, Crawley, UK), according to manufacturer's instructions. The detection of P21R and H159Y polymorphisms was performed by PCR amplification of exon 1 and 3 of TNFRSF13C, respectively, followed by restriction fragment length polymorphism (RFLP) analysis, as previously described in detail (
      • Kompoti M.
      • Michopoulos A.
      • Michalia M.
      • Clouva-Molyvdas P.M.
      • Germenis A.E.
      • Speletas M.
      Genetic polymorphisms of innate and adaptive immunity as predictors of outcome in critically ill patients.
      ).

      2.3 Statistical analysis

      Allele frequencies were analyzed using Fischer's exact test. Pearson's chi-square test was used for comparison of categorical variables. For non-canonical variables, Mann-Whitney U non-parametric tests were performed. EDSS scores were analyzed as ordinal values. The dominant model of expression (
      • Pieper K.
      • Rizzi M.
      • Speletas M.
      • Smulski C.R.
      • Sic H.
      • Kraus H.
      • Salzer U.
      • Fiala G.J.
      • Schamel W.W.
      • Lougaris V.
      • Plebani A.
      • Hammarstrom L.
      • Recher M.
      • Germenis A.E.
      • Grimbacher B.
      • Warnatz K.
      • Rolink A.G.
      • Schneider P.
      • Notarangelo L.D.
      • Eibel H.
      A common single nucleotide polymorphism impairs B-cell activating factor receptor's multimerization, contributing to common variable immunodeficiency.
      ) was chosen to examine any association between disease manifestation and the presence of the polymorphisms. Multinomial logistic regression was performed to conduct the comparison between stages of disease severity; univariate and multivariate analysis was performed as appropriate. The statistical significance threshold was set at two-sided p < 0.05. The statistical package SPSS v22 was employed for the analysis.

      3. Results

      Fifty-nine MS patients (12.1%) carried the P21R polymorphism, with one being a homozygote and 22 of the these P21R carrying patients (4.5% out of the total, or 37.3% out of P21R-positive patients) also had in heterozygosity the H159Y mutation. The analysis revealed that both polymorphisms were in Hardy-Weinberg Equilibrium (P21R p = 0.262, and H159Y p = 0.260). While, the frequency of the P21R polymorphism did not differ significantly between MS patients and controls, the H159Y polymorphism was found more frequently in the MS patient group (2.2% vs. 0.68%), correlating with a significantly (p < 0.0001) higher relative risk (3.3, 95%CI: 2.168–4.913) and odds ratio (3.4, 95%CI: 2.170–5.170) for MS (Table 1).
      Table 1Allele frequencies of TNFRSF13C/BAFFR-P21R and TNFRSF13C/BAFFR-H159Y polymorphisms in the patients of the study.
      FrequenciesMS patients n 486Controls* n 933pOR (95%CI)RR (95%CI)
      n (%)n (%)
      wt/wt

      P21R/wt

      P21R/P21R
      912 (93.8%)

      58 (6.0%)

      2 (0.2%)
      1740 (93.2%)

      120 (6.4%)

      6 (0.32%)


      0.187


      0.801 (0.583–1.092)


      0.864 (0.690–1.061)
      wt/wt

      H159Y/wt

      H159Y/H159Y
      464 (97.7%)

      22 (2.3%)

      0 (0%)
      64,676 (99.3%)

      445 (0.7%)

      0 (0%)


      <0.001


      3.389 (2.170–5.170)


      3.270 (2.168–4.913)
      Abbreviations: CI = Confidence Interval, MS = Multiple Sclerosis, OR = Odds Ratio, RR = Relative Risk, wt: wild-type. *data for P21R: http://exac.broadinstitute.org/variant/22-42322716-G-C; data for H159Y: http://exac.broadinstitute.org/variant/22-42321451-G-A.
      To study whether the presence of P21R and H159Y polymorphisms could influence MS disease severity, we examined the impact of both polymorphisms on “age of disease onset” and “MSSS” (the later combines EDSS and disease duration). Finally, we conducted a subgroup analysis to examine any difference of the clinical characteristics in the P21R-positive population regarding the presence of the H159Y alteration.
      The mean age of MS patients at initial symptoms appearance was 32.3 years (range 56 years) and the mean duration of the disease was 11.7 years. The mean severity of MS as expressed by the MSSS was 4.5, while the median disability score (EDSS) was 3. No substantial differences regarding the clinical characteristics were noted between the genders (Supplementary Table 1).
      Frequencies of the P21R-carriers and P21R-wt patients with benign, moderate and severe MS disease (according to the MSSS score) are depicted in Table 2. Disease severity did not differ between patients with the P21R polymorphism and wt patients (p = 0.26). Frequencies of the H159Y-carriers and H159Y-wt patients with benign, moderate and severe MS disease (according to the MSSS score) are also depicted in Table 2. Disease severity did not differ between patients with the H159Y polymorphism and H159Y-wt patients (p = 0.41). Differences between severe and benign disease status were further examined, among the mutational groups, by means of multinominal logistic regression. Since P21R & H159Y are in linkage disequilibrium, they were entered separately in the multivariate model, so that multicollinearity would not arise. However, statistically significant differences were not observed between patients with severe and benign disease status, for neither P21R nor H159Y polymorphism, either in the univariate, or the multivariate model adjusted for gender and age of onset (p > 0.05, in all cases). Furthermore, no significant differences were found concerning “the age of disease onset” neither for the P21R nor for the H159Y polymorphism (Table 2).
      Table 2Association of BAFFR-P21R and BAFFR-H159Y polymorphisms with clinical characteristics of the patients of the study.
      P21R(+)P21R(-)pH159Y(+)H159Y(-)p
      Sex

      Male n(%)

      Female n(%)
      n 59

      18 (30.5%)

      41 (69.5%)
      n 427

      138 (32.3%)

      289 (67.7%)
      0.78*n 22

      6 (27.3%)

      16 (72.7%)
      n 464

      150 (32.3%)

      314 (67.7%)
      0.62*
      Age of Onset

      Mean

      Median

      Range
      n 59

      32.8

      31.5

      45
      n 427

      32.3

      31

      56
      0.78⁎⁎n 22

      29.8

      28

      34
      n 464

      32.5

      31

      56
      0.24⁎⁎
      MSSS

      Mean

      Median

      Range
      n 59

      4.44

      3.89

      9.47
      n 427

      4.55

      3.94

      9.73
      0.89⁎⁎n 22

      4

      3.89

      7.56
      n 464

      4.6

      3.94

      9.73
      0.32⁎⁎
      Benign MS n(%)

      Moderate MS n(%)

      Severe MS n(%)
      7 (11.9%)

      44 (74.6%)

      8 (13.6%)
      72 (16.9%)

      272 (63.7%)

      83 (19.4%)
      0.26*3 (13.6%)

      17 (77.3%)

      2 (9.1%)
      76 (16.4%)

      299 (64.4%)

      89 (19.2%)
      0.41*
      Abbreviations. SD: Standard Deviation, MS: Multiple Sclerosis, MSSS: Multiple Sclerosis Severity Score.
      The statistical analysis was performed according to Pearson's chi-square test (*) and Mann-Whitney-U test (**).
      Finally, further analysis in the P21R-positive subgroup revealed that patients who also harbored the H159Y polymorphism had an earlier age of disease onset compared to H159Y-negative patients (median value 28.0 vs 32.5 years respectively); however, the difference was found marginally non-significant (p = 0.096). No statistically significant difference was noted for the rest of the clinical characteristics (p > 0.05, in all cases).

      4. Discussion

      Analyzing a large cohort of MS patients, we did not find any significant association of TNFRSF13C/BAFFR-P21R polymorphism with MS. In contrast, the TNFRSF13C/BAFFR-H159Y polymorphism was clearly overrepresented in the MS cohort. H159Y was first found in a high prevalence in patients with non-Hodgkin lymphoma and the polymorphism has been suggested to increase the risk of developing the disease (
      • Hildebrand J.M.
      • Luo Z.
      • Manske M.K.
      • Price-Troska T.
      • Ziesmer S.C.
      • Lin W.
      • Hostager B.S.
      • Slager S.L.
      • Witzig T.E.
      • Ansell S.M.
      • Cerhan J.R.
      • Bishop G.A.
      • Novak A.J.
      A BAFF-R mutation associated with non-Hodgkin lymphoma alters TRAF recruitment and reveals new insights into BAFF-R signaling.
      ). Since the H159Y mutation was reported to increase the affinity of BAFFR for TRAF6 association resulting in a hyperactive receptor (
      • Hildebrand J.M.
      • Luo Z.
      • Manske M.K.
      • Price-Troska T.
      • Ziesmer S.C.
      • Lin W.
      • Hostager B.S.
      • Slager S.L.
      • Witzig T.E.
      • Ansell S.M.
      • Cerhan J.R.
      • Bishop G.A.
      • Novak A.J.
      A BAFF-R mutation associated with non-Hodgkin lymphoma alters TRAF recruitment and reveals new insights into BAFF-R signaling.
      ), the correlation between an increased risk for MS and BAFFR hyperreactivity fits well to the observation made by Steri et al. that a polymorphism in the BAFF gene, which leads to higher BAFF levels, correlates positively with the risk of developing MS (
      • Steri M.
      • Orru V.
      • Idda M.L.
      • Pitzalis M.
      • Pala M.
      • Zara I.
      • Sidore C.
      • Faa V.
      • Floris M.
      • Deiana M.
      • Asunis I.
      • Porcu E.
      • Mulas A.
      • Piras M.G.
      • Lobina M.
      • Lai S.
      • Marongiu M.
      • Serra V.
      • Marongiu M.
      • Sole G.
      • Busonero F.
      • Maschio A.
      • Cusano R.
      • Cuccuru G.
      • Deidda F.
      • Poddie F.
      • Farina G.
      • Dei M.
      • Virdis F.
      • Olla S.
      • Satta M.A.
      • Pani M.
      • Delitala A.
      • Cocco E.
      • Frau J.
      • Coghe G.
      • Lorefice L.
      • Fenu G.
      • Ferrigno P.
      • Ban M.
      • Barizzone N.
      • Leone M.
      • Guerini F.R.
      • Piga M.
      • Firinu D.
      • Kockum I.
      • Lima Bomfim I.
      • Olsson T.
      • Alfredsson L.
      • Suarez A.
      • Carreira P.E.
      • Castillo-Palma M.J.
      • Marcus J.H.
      • Congia M.
      • Angius A.
      • Melis M.
      • Gonzalez A.
      • Alarcón Riquelme M.E.
      • da Silva B.M.
      • Marchini M.
      • Danieli M.G.
      • Del Giacco S.
      • Mathieu A.
      • Pani A.
      • Montgomery S.B.
      • Rosati G.
      • Hillert J.
      • Sawcer S.
      • D'Alfonso S.
      • Todd J.A.
      • Novembre J.
      • Abecasis G.R.
      • Whalen M.B.
      • Marrosu M.G.
      • Meloni A.
      • Sanna S.
      • Gorospe M.
      • Schlessinger D.
      • Fiorillo E.
      • Zoledziewska M.
      • Cucca F.
      Overexpression of the cytokine BAFF and autoimmunity risk.
      ).
      However, the relation between BAFF, BAFFR signaling and the susceptibility to MS may be more complex than just a direct correlation between higher BAFF levels, a stronger BAFFR signaling and the increased risk of developing the disease. In this context, it has been reported that blocking the biological activity of BAFF through the administration of BCMA-Ig, suppressed the onset and severity of experimental autoimmune encephalomyelitis (EAE) (
      • Huntington N.D.
      • Tomioka R.
      • Clavarino C.
      • Chow A.M.
      • Linares D.
      • Mana P.
      • Rossjohn J.
      • Cachero T.G.
      • Qian F.
      • Kalled S.L.
      • Bernard C.C.
      • Reid H.H.
      A BAFF antagonist suppresses experimental autoimmune encephalomyelitis by targeting cell-mediated and humoral immune responses.
      ). Likewise, neutralization of BAFF and APRIL in a marmoset (Callithrix jacchus) EAE model by treating the animals with belimumab (an anti-BAFF mAb) and anti-APRIL antibodies, delayed the development of this relevant preclinical MS model with a higher efficiency by belimumab than by anti-APRIL antibodies (
      • Jagessar S.A.
      • Heijmans N.
      • Oh L.
      • Bauer J.
      • Blezer E.L.
      • Laman J.D.
      • Migone T.S.
      • Devalaraja M.N.
      • ‘t Hart B.A.
      Antibodies against human BLyS and APRIL attenuate EAE development in marmoset monkeys.
      ).
      In contrast to these experimental models, the treatment of MS patients with soluble TACI-Ig (atacicept), which neutralizes both BAFF and APRIL, had to be terminated prematurely, since the disease had aggravated in the group of atacicept-treated patients in ATAMS (
      • Kappos L.
      • Hartung H.P.
      • Freedman M.S.
      • Boyko A.
      • Radu E.W.
      • Mikol D.D.
      • Lamarine M.
      • Hyvert Y.
      • Freudensprung U.
      • Plitz T.
      • van Beek J.
      Atacicept in multiple sclerosis (ATAMS): a randomised, placebo-controlled, double-blind, phase 2 trial.
      and ATON (
      • Sergott R.C.
      • Bennett J.L.
      • Rieckmann P.
      • Montalban X.
      • Mikol D.
      • Freudensprung U.
      • Plitz T.
      • van Beek J.
      ATON: results from a Phase II randomized trial of the B-cell-targeting agent atacicept in patients with optic neuritis.
      ) trials. However, B cell depletion induced by treating MS patients with rituximab (an anti-CD20 mAb), suggested that the elimination of B cells in MS is safe and can ameliorate the disease (
      • Rommer P.S.
      • Dorner T.
      • Freivogel K.
      • Haas J.
      • Kieseier B.C.
      • Kumpfel T.
      • Paul F.
      • Proft F.
      • Schulze-Koops H.
      • Schmidt E.
      • Wiendl H.
      • Ziemann U.
      • Zettl U.K.
      Safety and clinical outcomes of rituximab treatment in patients with multiple sclerosis and neuromyelitis optica: experience from a national online registry (GRAID).
      ). Since the neutralization of BAFF and/or APRIL activity through the blocking of their receptors affects all circulating B cell subsets but spares a significant fraction of switched memory B cells, neutralization of BAFF and APRIL may deplete B cell subsets which are not involved into the pathology of MS leaving pathological switched memory B cells untouched (
      • Baker D.
      • Marta M.
      • Pryce G.
      • Giovannoni G.
      • Schmierer K.
      Memory B cells are major targets for effective immunotherapy in relapsing multiple sclerosis.
      ). The presence of such pathological B cells may interpret the failure of some BAFF/APRIL-blocking agents in the management of MS.
      Apart from BAFFR, Nogo is another BAFF-binding receptor that is expressed on neurons, inhibiting their outgrowth in response to BAFF binding (
      • Vincent F.B.
      • Saulep-Easton D.
      • Figgett W.A.
      • Fairfax K.A.
      • Mackay F.
      The BAFF/APRIL system: emerging functions beyond B cell biology and autoimmunity.
      ;
      • Zhang L.
      • Zheng S.
      • Wu H.
      • Wu Y.
      • Liu S.
      • Fan M.
      • Zhang J.
      Identification of BLyS (B lymphocyte stimulator), a non-myelin-associated protein, as a functional ligand for Nogo-66 receptor.
      ) and may contribute to the correlation between increased BAFF levels and MS. Nogo-BAFF interaction may be relevant for CNS injuries and diseases, as they may be enhanced by increased local production of BAFF from astrocytes (
      • Krumbholz M.
      • Theil D.
      • Derfuss T.
      • Rosenwald A.
      • Schrader F.
      • Monoranu C.M.
      • Kalled S.L.
      • Hess D.M.
      • Serafini B.
      • Aloisi F.
      • Wekerle H.
      • Hohlfeld R.
      • Meinl E.
      BAFF is produced by astrocytes and up-regulated in multiple sclerosis lesions and primary central nervous system lymphoma.
      ;
      • Vincent F.B.
      • Saulep-Easton D.
      • Figgett W.A.
      • Fairfax K.A.
      • Mackay F.
      The BAFF/APRIL system: emerging functions beyond B cell biology and autoimmunity.
      ). Obviously, the precise role of Nogo-BAFF interaction in MS should be addressed in further studies.
      Interestingly, there are no functional studies analyzing the contribution of BAFF/BAFFR polymorphisms in MS pathogenesis and/or phenotype, until now. As mentioned above, previous studies have demonstrated that the analyzed BAFFR polymorphisms P21R (rs77874543) and H159Y (rs61756766) have been associated with CVID, CLL and non-Hodgkin lymphoma pathogenesis (
      • Hildebrand J.M.
      • Luo Z.
      • Manske M.K.
      • Price-Troska T.
      • Ziesmer S.C.
      • Lin W.
      • Hostager B.S.
      • Slager S.L.
      • Witzig T.E.
      • Ansell S.M.
      • Cerhan J.R.
      • Bishop G.A.
      • Novak A.J.
      A BAFF-R mutation associated with non-Hodgkin lymphoma alters TRAF recruitment and reveals new insights into BAFF-R signaling.
      ;
      • Jasek M.
      • Bojarska-Junak A.
      • Wagner M.
      • Sobczynski M.
      • Wolowiec D.
      • Rolinski J.
      • Karabon L.
      • Kuśnierczyk P.
      Association of variants in BAFF (rs9514828 and rs1041569) and BAFF-R (rs61756766) genes with the risk of chronic lymphocytic leukemia.
      ;
      • Losi C.G.
      • Silini A.
      • Fiorini C.
      • Soresina A.
      • Meini A.
      • Ferrari S.
      • Notarangelo L.D.
      • Lougaris V.
      • Plebani A.
      Mutational analysis of human BAFF receptor TNFRSF13C (BAFF-R) in patients with common variable immunodeficiency.
      ;
      • Pieper K.
      • Rizzi M.
      • Speletas M.
      • Smulski C.R.
      • Sic H.
      • Kraus H.
      • Salzer U.
      • Fiala G.J.
      • Schamel W.W.
      • Lougaris V.
      • Plebani A.
      • Hammarstrom L.
      • Recher M.
      • Germenis A.E.
      • Grimbacher B.
      • Warnatz K.
      • Rolink A.G.
      • Schneider P.
      • Notarangelo L.D.
      • Eibel H.
      A common single nucleotide polymorphism impairs B-cell activating factor receptor's multimerization, contributing to common variable immunodeficiency.
      ). Moreover, other variants of BAFF gene have been associated with other diseases with autoimmune pathophysiology. More precisely, the rs9514828, which is located in BAFF promoter region, has been associated with high levels of serum BAFF increasing the risk of NHL, CLL and hepatitis C-related mixed cryoglobulinemia (
      • Ayad M.W.
      • Elbanna A.A.
      • Elneily D.A.
      • Sakr A.S.
      Association of BAFF -871C/T promoter polymorphism with hepatitis C-related mixed cryoglobulinemia in a cohort of egyptian patients.
      ;
      • Ferrer G.
      • Hodgson K.
      • Montserrat E.
      • Moreno C.
      B cell activator factor and a proliferation-inducing ligand at the cross-road of chronic lymphocytic leukemia and autoimmunity.
      ;
      • Jasek M.
      • Bojarska-Junak A.
      • Wagner M.
      • Sobczynski M.
      • Wolowiec D.
      • Rolinski J.
      • Karabon L.
      • Kuśnierczyk P.
      Association of variants in BAFF (rs9514828 and rs1041569) and BAFF-R (rs61756766) genes with the risk of chronic lymphocytic leukemia.
      ;
      • Lahiri A.
      • Pochard P.
      • Le Pottier L.
      • Tobon G.J.
      • Bendaoud B.
      • Youinou P.
      • Pers J.O.
      The complexity of the BAFF TNF-family members: implications for autoimmunity.
      ), the rs1041569 has been associated with CLL risk (
      • Jasek M.
      • Bojarska-Junak A.
      • Wagner M.
      • Sobczynski M.
      • Wolowiec D.
      • Rolinski J.
      • Karabon L.
      • Kuśnierczyk P.
      Association of variants in BAFF (rs9514828 and rs1041569) and BAFF-R (rs61756766) genes with the risk of chronic lymphocytic leukemia.
      ), while the AA genotype of the rs12583006 has been correlated with lupus and lupus related plaque formation (
      • Theodorou E.
      • Nezos A.
      • Antypa E.
      • Ioakeimidis D.
      • Koutsilieris M.
      • Tektonidou M.
      • Moutsopoulos H.M.
      • Mavragani C.P.
      B-cell activating factor and related genetic variants in lupus related atherosclerosis.
      ). In addition, the rs2893321 seems to increase the risk for the development of Grave's disease (
      • Lin J.D.
      • Yang S.F.
      • Wang Y.H.
      • Fang W.F.
      • Lin Y.C.
      • Lin Y.F.
      • Tang K.T.
      • Wu M.Y.
      • Cheng C.W.
      Analysis of associations of human BAFF gene polymorphisms with autoimmune thyroid diseases.
      ) and the rs375946 has been associated with response to rituximab in patients with antineutrophil cytoplasmic antibody–associated vasculitis (
      • Alberici F.
      • Smith R.M.
      • Fonseca M.
      • Willcocks L.C.
      • Jones R.B.
      • Holle J.U.
      • Wieczorek S.
      • Neumann T.
      • Martorana D.
      • Gregorini G.
      • Sinico R.A.
      • Bruchfeld A.
      • Gunnarsson I.
      • Ohlsson S.
      • Baslund B.
      • Tesar V.
      • Hruskova Z.
      • Cid M.C.
      • Vaglio A.
      • Lyons P.A.
      • Smith K.G.C.
      • Jayne D.R.W.
      Association of a TNFSF13B (BAFF) regulatory region single nucleotide polymorphism with response to rituximab in antineutrophil cytoplasmic antibody-associated vasculitis.
      ).
      A limitation of our study is the absence of data concerning the use of disease-modifying treatments, which have been reported to alter BAFF levels in blood of MS patients; particularly the use of INF-β is known to lead to a strong upregulation of BAFF in serum (
      • Kannel K.
      • Alnek K.
      • Vahter L.
      • Gross-Paju K.
      • Uibo R.
      • Kisand K.V.
      Changes in blood B cell-activating factor (BAFF) levels in multiple sclerosis: a sign of treatment outcome.
      ). Obviously, further studies may clarify also this point.
      Moreover, as presented in Table 1, we did not detect the H159Y/H159Y genotype in our cohort (both in MS patients and healthy controls). The latter could be attributed to the sample size that was genotyped, and it is a common phenomenon in candidate gene association studies examining low frequency variants (
      • Rikos D.
      • Siokas V.
      • Aloizou A.M.
      • Tsouris Z.
      • Aslanidou P.
      • Koutsis G.
      • Anagnostouli M.
      • Bogdanos D.P.
      • Grigoriadis N.
      • Hadjigeorgiou G.M.
      • Dardiotis E.
      TREM2 R47H (rs75932628) variant is unlikely to contribute to multiple sclerosis susceptibility and severity in a large Greek MS cohort.
      ). Moreover, bearing in mind that there is no consensus method for measuring disease severity in MS (
      • Roxburgh R.H.
      • Seaman S.R.
      • Masterman T.
      • Hensiek A.E.
      • Sawcer S.J.
      • Vukusic S.
      • Achiti I.
      • Confavreux C.
      • Coustans M.
      • le Page E.
      • Edan G.
      • McDonnell G.V.
      • Hawkins S.
      • Trojano M.
      • Liguori M.
      • Cocco E.
      • Marrosu M.G.
      • Tesser F.
      • Leone M.A.
      • Weber A.
      • Zipp F.
      • Miterski B.
      • Epplen J.T.
      • Oturai A.
      • Sοrensen P.S.
      • Celius E.G.
      • Lara N.T.
      • Montalban X.
      • Villoslada P.
      • Silva A.M.
      • Marta M.
      • Leite I.
      • Dubois B.
      • Rubio J.
      • Butzkueven H.
      • Kilpatrick T.
      • Mycko M.P.
      • Selmaj K.W.
      • Rio M.E.
      • Sa M.
      • Salemi G.
      • Savettieri G.
      • Hillert J.
      • Compston D.A.
      Multiple sclerosis severity score: using disability and disease duration to rate disease severity.
      ), we used the MSSS algorithm as a powerful method for comparing disease progression and severity. As presented, no significant associations were found between MSSS and BAFFR polymorphisms.
      In conclusion, we report a significant association of the TNFRSF13C/BAFFR-H159Y polymorphism with MS in a cohort of Greek patients, suggesting that an imbalanced BAFF/BAFFR system increases the risk of developing MS. Since BAFF can also induce IL-10 by regulatory B cells (
      • Yang M.
      • Sun L.
      • Wang S.
      • Ko K.H.
      • Xu H.
      • Zheng B.J.
      • Cao X.
      • Lu L.
      Novel function of B cell-activating factor in the induction of IL-10-producing regulatory B cells.
      ) that were found to inhibit EAE development and progression (
      • Matsushita T.
      • Yanaba K.
      • Bouaziz J.D.
      • Fujimoto M.
      • Tedder T.F.
      Regulatory B cells inhibit EAE initiation in mice while other B cells promote disease progression.
      ), treatment of MS patients with the BAFF-neutralizing antibodies, as belimumab, may not only lead to the enrichment of potentially pathogenic switched memory B cells, but also to the depletion of disease-modifying regulatory B cells. Thus, treatment approaches interfering with the BAFF/BAFFR system are clearly less straight forward than originally anticipated and should integrate the complex role of BAFF and its receptors in B cell development and homeostasis.

      Declaration of Competing Interest

      All the authors declare no conflicts of interest associated with this manuscript.

      Acknowledgments

      Study conception and design was performed within the context of the postgraduate program “Clinical Applications of Molecular Medicine”, Faculty of Medicine, School of Health Sciences, University of Thessaly, Greece. The authors are grateful to Katerina Dadouli for her assistance in the statistical analysis and to Adam Molyvdas and Evangelia Karamouti for their technical assistance. Finally, the authors would like to thank the Exome Aggregation Consortium and the groups that provided exome variant data for comparison. A full list of contributing groups can be found at http://exac.broadinstitute.org/about.

      Appendix. Supplementary materials

      References

        • Alberici F.
        • Smith R.M.
        • Fonseca M.
        • Willcocks L.C.
        • Jones R.B.
        • Holle J.U.
        • Wieczorek S.
        • Neumann T.
        • Martorana D.
        • Gregorini G.
        • Sinico R.A.
        • Bruchfeld A.
        • Gunnarsson I.
        • Ohlsson S.
        • Baslund B.
        • Tesar V.
        • Hruskova Z.
        • Cid M.C.
        • Vaglio A.
        • Lyons P.A.
        • Smith K.G.C.
        • Jayne D.R.W.
        Association of a TNFSF13B (BAFF) regulatory region single nucleotide polymorphism with response to rituximab in antineutrophil cytoplasmic antibody-associated vasculitis.
        J. Allergy. Clin. Immunol. 2017; 139 (e10): 1684-1687https://doi.org/10.1016/j.jaci.2016.08.051
        • Ayad M.W.
        • Elbanna A.A.
        • Elneily D.A.
        • Sakr A.S.
        Association of BAFF -871C/T promoter polymorphism with hepatitis C-related mixed cryoglobulinemia in a cohort of egyptian patients.
        Mol. Diagn. Ther. 2015; 19: 99-106https://doi.org/10.1007/s40291-015-0134-7
        • Baker D.
        • Marta M.
        • Pryce G.
        • Giovannoni G.
        • Schmierer K.
        Memory B cells are major targets for effective immunotherapy in relapsing multiple sclerosis.
        EBioMedicine. 2017; 16: 41-50https://doi.org/10.1016/j.ebiom.2017.01.042
        • Claes N.
        • Fraussen J.
        • Stinissen P.
        • Hupperts R.
        • Somers V.
        B cells are multifunctional players in multiple sclerosis pathogenesis: insights from therapeutic interventions.
        Front. Immunol. 2015; 6: 642https://doi.org/10.3389/fimmu.2015.00642
        • Dardiotis E.
        • Panayiotou E.
        • Provatas A.
        • Christodoulou K.
        • Hadjisavvas A.
        • Antoniades A.
        • Lourbopoulos A.
        • Pantzaris M.
        • Grigoriadis N.
        • Hadjigeorgiou G.M.
        • Kyriakides T.
        Gene variants of adhesion molecules act as modifiers of disease severity in MS.
        Neurol. Neuroimmunol. Neuroinflamm. 2017; 4: e350https://doi.org/10.1212/NXI.0000000000000350
        • Dendrou C.A.
        • Fugger L.
        Immunomodulation in multiple sclerosis: promises and pitfalls.
        Curr. Opin. Immunol. 2017; 49: 37-43https://doi.org/10.1016/j.coi.2017.08.013
        • Disanto G.
        • Morahan J.M.
        • Barnett M.H.
        • Giovannoni G.
        • Ramagopalan S.V.
        The evidence for a role of B cells in multiple sclerosis.
        Neurology. 2012; 78: 823-832https://doi.org/10.1212/WNL.0b013e318249f6f0
        • Ferrer G.
        • Hodgson K.
        • Montserrat E.
        • Moreno C.
        B cell activator factor and a proliferation-inducing ligand at the cross-road of chronic lymphocytic leukemia and autoimmunity.
        Leuk. Lymphoma. 2009; 50: 1075-1082https://doi.org/10.1080/10428190903013334
        • Filippi M.
        • Rocca M.A.
        • Ciccarelli O.
        • De Stefano N.
        • Evangelou N.
        • Kappos L.
        • Rovira A.
        • Sastre-Garriga J.
        • Tintorè M.
        • Frederiksen J.L.
        • Gasperini C.
        • Palace J.
        • Reich D.S.
        • Banwell B.
        • Montalban X.
        • Barkhof F.
        MRI criteria for the diagnosis of multiple sclerosis: MAGNIMS consensus guidelines.
        Lancet Neurol. 2016; 15: 292-303https://doi.org/10.1016/S1474-4422(15)00393-2
        • Franciotta D.
        • Salvetti M.
        • Lolli F.
        • Serafini B.
        • Aloisi F.
        B cells and multiple sclerosis.
        Lancet Neurol. 2008; 7: 852-858https://doi.org/10.1016/S1474-4422(08)70192-3
        • Hadjigeorgiou G.M.
        • Kountra P.M.
        • Koutsis G.
        • Tsimourtou V.
        • Siokas V.
        • Dardioti M.
        • Rikos D.
        • Marogianni C.
        • Aloizou A.M.
        • Karadima G.
        • Ralli S.
        • Grigoriadis N.
        • Bogdanos D.
        • Panas M.
        • Dardiotis E.
        Replication study of GWAS risk loci in Greek multiple sclerosis patients.
        Neurol. Sci. 2019; 40: 253-260https://doi.org/10.1007/s10072-018-3617-6
        • Hemmer B.
        • Archelos J.J.
        • Hartung H.P.
        New concepts in the immunopathogenesis of multiple sclerosis.
        Nat. Rev. Neurosci. 2002; 3: 291-301
        • Hildebrand J.M.
        • Luo Z.
        • Manske M.K.
        • Price-Troska T.
        • Ziesmer S.C.
        • Lin W.
        • Hostager B.S.
        • Slager S.L.
        • Witzig T.E.
        • Ansell S.M.
        • Cerhan J.R.
        • Bishop G.A.
        • Novak A.J.
        A BAFF-R mutation associated with non-Hodgkin lymphoma alters TRAF recruitment and reveals new insights into BAFF-R signaling.
        J. Exp. Med. 2010; 207: 2569-2579https://doi.org/10.1084/jem.20100857
        • Huntington N.D.
        • Tomioka R.
        • Clavarino C.
        • Chow A.M.
        • Linares D.
        • Mana P.
        • Rossjohn J.
        • Cachero T.G.
        • Qian F.
        • Kalled S.L.
        • Bernard C.C.
        • Reid H.H.
        A BAFF antagonist suppresses experimental autoimmune encephalomyelitis by targeting cell-mediated and humoral immune responses.
        Intern. Immunol. 2006; 18: 1473-1485https://doi.org/10.1093/intimm/dxl080
        • Jagessar S.A.
        • Heijmans N.
        • Oh L.
        • Bauer J.
        • Blezer E.L.
        • Laman J.D.
        • Migone T.S.
        • Devalaraja M.N.
        • ‘t Hart B.A.
        Antibodies against human BLyS and APRIL attenuate EAE development in marmoset monkeys.
        J. Neuroimmune Pharmacol. 2012; 7: 557-570https://doi.org/10.1007/s11481-012-9384-x
        • Jasek M.
        • Bojarska-Junak A.
        • Wagner M.
        • Sobczynski M.
        • Wolowiec D.
        • Rolinski J.
        • Karabon L.
        • Kuśnierczyk P.
        Association of variants in BAFF (rs9514828 and rs1041569) and BAFF-R (rs61756766) genes with the risk of chronic lymphocytic leukemia.
        Tumour Biol. 2016; 37: 13617-13626https://doi.org/10.1007/s13277-016-5182-z
        • Kannel K.
        • Alnek K.
        • Vahter L.
        • Gross-Paju K.
        • Uibo R.
        • Kisand K.V.
        Changes in blood B cell-activating factor (BAFF) levels in multiple sclerosis: a sign of treatment outcome.
        PLoS ONE. 2015; 10e0143393https://doi.org/10.1371/journal.pone.0143393
        • Kappos L.
        • Hartung H.P.
        • Freedman M.S.
        • Boyko A.
        • Radu E.W.
        • Mikol D.D.
        • Lamarine M.
        • Hyvert Y.
        • Freudensprung U.
        • Plitz T.
        • van Beek J.
        Atacicept in multiple sclerosis (ATAMS): a randomised, placebo-controlled, double-blind, phase 2 trial.
        Lancet Neurol. 2014; 13: 353-363https://doi.org/10.1016/S1474-4422(14)70028-6
        • Kim H.M.
        • Yu K.S.
        • Lee M.E.
        • Shin D.R.
        • Kim Y.S.
        • Paik S.G.
        • Yoo O.J.
        • Lee H.
        • Lee J.O.
        Crystal structure of the BAFF-BAFF-R complex and its implications for receptor activation.
        Nat. Struct. Biol. 2003; 10: 342-348https://doi.org/10.1038/nsb925
        • Kompoti M.
        • Michopoulos A.
        • Michalia M.
        • Clouva-Molyvdas P.M.
        • Germenis A.E.
        • Speletas M.
        Genetic polymorphisms of innate and adaptive immunity as predictors of outcome in critically ill patients.
        Immunobiology. 2015; 220: 414-421https://doi.org/10.1016/j.imbio.2014.10.006
        • Krumbholz M.
        • Theil D.
        • Derfuss T.
        • Rosenwald A.
        • Schrader F.
        • Monoranu C.M.
        • Kalled S.L.
        • Hess D.M.
        • Serafini B.
        • Aloisi F.
        • Wekerle H.
        • Hohlfeld R.
        • Meinl E.
        BAFF is produced by astrocytes and up-regulated in multiple sclerosis lesions and primary central nervous system lymphoma.
        J. Exp. Med. 2005; 201: 195-200https://doi.org/10.1084/jem.20041674
        • Kurtzke J.F.
        Rating neurologic impairment in multiple sclerosis: an expanded disability status scale (EDSS).
        Neurology. 1983; 33: 1444-1452https://doi.org/10.1212/WNL.33.11.1444
        • Lahiri A.
        • Pochard P.
        • Le Pottier L.
        • Tobon G.J.
        • Bendaoud B.
        • Youinou P.
        • Pers J.O.
        The complexity of the BAFF TNF-family members: implications for autoimmunity.
        J. Autoimmun. 2012; 39: 189-198https://doi.org/10.1016/j.jaut.2012.05.009
        • Lin J.D.
        • Yang S.F.
        • Wang Y.H.
        • Fang W.F.
        • Lin Y.C.
        • Lin Y.F.
        • Tang K.T.
        • Wu M.Y.
        • Cheng C.W.
        Analysis of associations of human BAFF gene polymorphisms with autoimmune thyroid diseases.
        PLoS. One. 2016; 11e0154436https://doi.org/10.1371/journal.pone.0154436
        • Lek M.
        • Karczewski K.J.
        • Minikel E.V.
        • Samocha K.E.
        • Banks E.
        • Fennell T.
        • O'Donnell-Luria A.H.
        • Ware J.S.
        • Hill A.J.
        • Cummings B.B.
        • Tukiainen T.
        • Birnbaum D.P.
        • Kosmicki J.A.
        • Duncan L.E.
        • Estrada K.
        • Zhao F.
        • Zou J.
        • Pierce-Hoffman E.
        • Berghout J.
        • Cooper D.N.
        • Deflaux N.
        • DePristo M.
        • Do R.
        • Flannick J.
        • Fromer M.
        • Gauthier L.
        • Goldstein J.
        • Gupta N.
        • Howrigan D.
        • Kiezun A.
        • Kurki M.I.
        • Moonshine A.L.
        • Natarajan P.
        • Orozco L.
        • Peloso G.M.
        • Poplin R.
        • Rivas M.A.
        • Ruano-Rubio V.
        • Rose S.A.
        • Ruderfer D.M.
        • Shakir K.
        • Stenson P.D.
        • Stevens C.
        • Thomas B.P.
        • Tiao G.
        • Tusie-Luna M.T.
        • Weisburd B.
        • Won H.-.H.
        • Yu D.
        • Altshuler D.M.
        • Ardissino D.
        • Boehnke M.
        • Danesh J.
        • Donnelly S.
        • Elosua R.
        • Florez J.C.
        • Gabriel S.B.
        • Getz G.
        • Glatt S.J.
        • Hultman C.M.
        • Kathiresan S.
        • Laakso M.
        • McCarroll S.
        • McCarthy M.I.
        • McGovern D.
        • McPherson R.
        • Neale B.M.
        • Palotie A.
        • Purcell S.M.
        • Saleheen D.
        • Scharf J.M.
        • Sklar P.
        • Sullivan P.F.
        • Tuomilehto J.
        • Tsuang M.T.
        • Watkins H.C.
        • Wilson J.G.
        • Daly M.J.
        • MacArthur D.G.
        • Exome Aggregation, C.
        Analysis of protein-coding genetic variation in 60,706 humans.
        Nature. 2016; 536: 285https://doi.org/10.1038/nature19057
        • Lokensgard J.R.
        • Mutnal M.B.
        • Prasad S.
        • Sheng W.
        • Hu S.
        Glial cell activation, recruitment, and survival of B-lineage cells following MCMV brain infection.
        J. Neuroinflamm. 2016; 13: 114https://doi.org/10.1186/s12974-016-0582-y
        • Losi C.G.
        • Silini A.
        • Fiorini C.
        • Soresina A.
        • Meini A.
        • Ferrari S.
        • Notarangelo L.D.
        • Lougaris V.
        • Plebani A.
        Mutational analysis of human BAFF receptor TNFRSF13C (BAFF-R) in patients with common variable immunodeficiency.
        J. Clin. Immunol. 2005; 25: 496-502https://doi.org/10.1007/s10875-005-5637-2
        • Mackay F.
        • Schneider P.
        Cracking the BAFF code.
        Nat. Rev. Immunol. 2009; 9: 491-502https://doi.org/10.1038/nri2572
        • Matsushita T.
        • Yanaba K.
        • Bouaziz J.D.
        • Fujimoto M.
        • Tedder T.F.
        Regulatory B cells inhibit EAE initiation in mice while other B cells promote disease progression.
        J. Clin. Invest. 2008; 18: 3420-3430https://doi.org/10.1172/JCI36030
        • Papageorgiou A.
        • Mavragani C.P.
        • Nezos A.
        • Zintzaras E.
        • Quartuccio L.
        • De Vita S.
        • Koutsilieris M.
        • Tzioufas A.G.
        • Moutsopoulos H.M.
        • Voulgarelis M.
        A BAFF receptor His159Tyr mutation in Sjogren's syndrome-related lymphoproliferation.
        Arthritis Rheumatol. 2015; 67: 2732-2741https://doi.org/10.1002/art.39231
        • Pieper K.
        • Rizzi M.
        • Speletas M.
        • Smulski C.R.
        • Sic H.
        • Kraus H.
        • Salzer U.
        • Fiala G.J.
        • Schamel W.W.
        • Lougaris V.
        • Plebani A.
        • Hammarstrom L.
        • Recher M.
        • Germenis A.E.
        • Grimbacher B.
        • Warnatz K.
        • Rolink A.G.
        • Schneider P.
        • Notarangelo L.D.
        • Eibel H.
        A common single nucleotide polymorphism impairs B-cell activating factor receptor's multimerization, contributing to common variable immunodeficiency.
        J. Allergy Clin. Immunol. 2014; 133: 1222-1225https://doi.org/10.1016/j.jaci.2013.11.021
        • Polman C.H.
        • Reingold S.C.
        • Banwell B.
        • Clanet M.
        • Cohen J.A.
        • Filippi M.
        • Fujihara K.
        • Havrdova E.
        • Hutchinson M.
        • Kappos L.
        • Lublin F.D.
        • Montalban X.
        • O'Connor P.
        • Sandberg-Wollheim M.
        • Thompson A.J.
        • Waubant E.
        • Weinshenker B.
        • Wolinsky J.S.
        Diagnostic criteria for multiple sclerosis: 2010 revisions to the McDonald criteria.
        Ann. Neurol. 2011; 69: 292-302https://doi.org/10.1002/ana.22366
        • Ragheb S.
        • Li Y.
        • Simon K.
        • VanHaerents S.
        • Galimberti D.
        • De Riz M.
        • Fenoglio C.
        • Scarpini E.
        • Lisak R.
        Multiple sclerosis: BAFF and CXCL13 in cerebrospinal fluid.
        Mult. Scler. 2011; 17: 819-829https://doi.org/10.1177/1352458511398887
        • Rikos D.
        • Siokas V.
        • Aloizou A.M.
        • Tsouris Z.
        • Aslanidou P.
        • Koutsis G.
        • Anagnostouli M.
        • Bogdanos D.P.
        • Grigoriadis N.
        • Hadjigeorgiou G.M.
        • Dardiotis E.
        TREM2 R47H (rs75932628) variant is unlikely to contribute to multiple sclerosis susceptibility and severity in a large Greek MS cohort.
        Mult. Scler. Relat. Disord. 2019; 35: 116-118https://doi.org/10.1016/j.msard.2019.07.007
        • Rommer P.S.
        • Dorner T.
        • Freivogel K.
        • Haas J.
        • Kieseier B.C.
        • Kumpfel T.
        • Paul F.
        • Proft F.
        • Schulze-Koops H.
        • Schmidt E.
        • Wiendl H.
        • Ziemann U.
        • Zettl U.K.
        Safety and clinical outcomes of rituximab treatment in patients with multiple sclerosis and neuromyelitis optica: experience from a national online registry (GRAID).
        J. Neuroimmune Pharmacol. 2016; 11: 1-8https://doi.org/10.1007/s11481-015-9646-5
        • Roxburgh R.H.
        • Seaman S.R.
        • Masterman T.
        • Hensiek A.E.
        • Sawcer S.J.
        • Vukusic S.
        • Achiti I.
        • Confavreux C.
        • Coustans M.
        • le Page E.
        • Edan G.
        • McDonnell G.V.
        • Hawkins S.
        • Trojano M.
        • Liguori M.
        • Cocco E.
        • Marrosu M.G.
        • Tesser F.
        • Leone M.A.
        • Weber A.
        • Zipp F.
        • Miterski B.
        • Epplen J.T.
        • Oturai A.
        • Sοrensen P.S.
        • Celius E.G.
        • Lara N.T.
        • Montalban X.
        • Villoslada P.
        • Silva A.M.
        • Marta M.
        • Leite I.
        • Dubois B.
        • Rubio J.
        • Butzkueven H.
        • Kilpatrick T.
        • Mycko M.P.
        • Selmaj K.W.
        • Rio M.E.
        • Sa M.
        • Salemi G.
        • Savettieri G.
        • Hillert J.
        • Compston D.A.
        Multiple sclerosis severity score: using disability and disease duration to rate disease severity.
        Neurology. 2005; 64: 1144-1151https://doi.org/10.1212/01.WNL.0000156155.19270.F8
        • Sergott R.C.
        • Bennett J.L.
        • Rieckmann P.
        • Montalban X.
        • Mikol D.
        • Freudensprung U.
        • Plitz T.
        • van Beek J.
        ATON: results from a Phase II randomized trial of the B-cell-targeting agent atacicept in patients with optic neuritis.
        J. Neurol. Sci. 2015; 351: 174-178https://doi.org/10.1016/j.jns.2015.02.019
        • Smulski C.R.
        • Eibel H.
        BAFF and BAFF-Receptor in B cell selection and survival.
        Front. Immunol. 2018; 9: 2285https://doi.org/10.3389/fimmu.2018.02285
        • Srivastava P.
        • Mujtaba M.A.
        • Singhal M.
        Gene and cytokines expression of multiple sclerosis and its therapeutic regimen: a systemic review.
        Int. J. Drug Dev. Res. 2012; 4: 55-66
        • Stadelmann C.
        Multiple sclerosis as a neurodegenerative disease: pathology, mechanisms and therapeutic implications.
        Curr. Opin. Neurol. 2011; 24: 224-229https://doi.org/10.1097/WCO.0b013e328346056f
        • Steri M.
        • Orru V.
        • Idda M.L.
        • Pitzalis M.
        • Pala M.
        • Zara I.
        • Sidore C.
        • Faa V.
        • Floris M.
        • Deiana M.
        • Asunis I.
        • Porcu E.
        • Mulas A.
        • Piras M.G.
        • Lobina M.
        • Lai S.
        • Marongiu M.
        • Serra V.
        • Marongiu M.
        • Sole G.
        • Busonero F.
        • Maschio A.
        • Cusano R.
        • Cuccuru G.
        • Deidda F.
        • Poddie F.
        • Farina G.
        • Dei M.
        • Virdis F.
        • Olla S.
        • Satta M.A.
        • Pani M.
        • Delitala A.
        • Cocco E.
        • Frau J.
        • Coghe G.
        • Lorefice L.
        • Fenu G.
        • Ferrigno P.
        • Ban M.
        • Barizzone N.
        • Leone M.
        • Guerini F.R.
        • Piga M.
        • Firinu D.
        • Kockum I.
        • Lima Bomfim I.
        • Olsson T.
        • Alfredsson L.
        • Suarez A.
        • Carreira P.E.
        • Castillo-Palma M.J.
        • Marcus J.H.
        • Congia M.
        • Angius A.
        • Melis M.
        • Gonzalez A.
        • Alarcón Riquelme M.E.
        • da Silva B.M.
        • Marchini M.
        • Danieli M.G.
        • Del Giacco S.
        • Mathieu A.
        • Pani A.
        • Montgomery S.B.
        • Rosati G.
        • Hillert J.
        • Sawcer S.
        • D'Alfonso S.
        • Todd J.A.
        • Novembre J.
        • Abecasis G.R.
        • Whalen M.B.
        • Marrosu M.G.
        • Meloni A.
        • Sanna S.
        • Gorospe M.
        • Schlessinger D.
        • Fiorillo E.
        • Zoledziewska M.
        • Cucca F.
        Overexpression of the cytokine BAFF and autoimmunity risk.
        N. Engl. J. Med. 2017; 376: 1615-1626https://doi.org/10.1056/NEJMoa1610528
        • Thangarajh M.
        • Gomes A.
        • Masterman T.
        • Hillert J.
        • Hjelmstrom P.
        Expression of B-cell-activating factor of the TNF family (BAFF) and its receptors in multiple sclerosis.
        J. Neuroimmunol. 2004; 152: 183-190https://doi.org/10.1016/j.jneuroim.2004.03.017
        • Theodorou E.
        • Nezos A.
        • Antypa E.
        • Ioakeimidis D.
        • Koutsilieris M.
        • Tektonidou M.
        • Moutsopoulos H.M.
        • Mavragani C.P.
        B-cell activating factor and related genetic variants in lupus related atherosclerosis.
        J. Autoimmun. 2018; 92: 87-92https://doi.org/10.1016/j.jaut.2018.05.002
        • Vincent F.B.
        • Saulep-Easton D.
        • Figgett W.A.
        • Fairfax K.A.
        • Mackay F.
        The BAFF/APRIL system: emerging functions beyond B cell biology and autoimmunity.
        Cytokine Growth Factor Rev. 2013; 24: 203-215https://doi.org/10.1016/j.cytogfr.2013.04.003
        • Wang H.
        • Wang K.
        • Zhong X.
        • Qiu W.
        • Dai Y.
        • Wu A.
        • Hu X.
        Cerebrospinal fluid BAFF and APRIL levels in neuromyelitis optica and multiple sclerosis patients during relapse.
        J. Clin. Immunol. 2012; 32: 1007-1011https://doi.org/10.1007/s10875-012-9709-9
        • Woodland R.T.
        • Schmidt M.R.
        • Thompson C.B.
        BLyS and B cell homeostasis.
        Semin. Immunol. 2006; 18: 318-326https://doi.org/10.1016/j.smim.2006.06.001
        • Yang M.
        • Sun L.
        • Wang S.
        • Ko K.H.
        • Xu H.
        • Zheng B.J.
        • Cao X.
        • Lu L.
        Novel function of B cell-activating factor in the induction of IL-10-producing regulatory B cells.
        J. Immunol. 2010; 184: 3321-3325https://doi.org/10.4049/jimmunol.0902551
        • Zhang L.
        • Zheng S.
        • Wu H.
        • Wu Y.
        • Liu S.
        • Fan M.
        • Zhang J.
        Identification of BLyS (B lymphocyte stimulator), a non-myelin-associated protein, as a functional ligand for Nogo-66 receptor.
        J. Neurosci. 2009; 29: 6348-6352https://doi.org/10.1523/JNEUROSCI.5040-08.2009