UNCORRECTED PROOF 1 Q1 Guidelines for standard preclinical experiments in the mouse model of 2 myasthenia gravis induced by acetylcholine receptor immunization 3 Q2 Erdem Tuzun a , Sonia Berrih-Aknin b,c,d,e , Talma Brenner f , Linda L. Kusner g , Rozen Le Panse b,c,d,e , 4 Huan Yang h , Socrates Tzartos i , Premkumar Christadoss j, ⁎ 5 a Department of Neuroscience, Institute for Experimental Medical Research (DETAE), Istanbul University, Istanbul, Turkey 6 b Q3 INSERM U974, Paris, France 7 c CNRS UMR 7215, Paris, France 8 d UPMC Univ Paris 6, Paris, France 9 e AIM, Institute of Myology, Paris, France 10 f Department of Neurology, The Agnes Ginges Center for Human Neurogenetics, Hadassah-Hebrew University Medical Center, Jerusalem, Israel 11 g Department of Pharmacology and Physiology, George Washington University, Washington, DC, USA 12 h Department of Neurology, Xiangya Hospital, Central South University, Changsha, Hunan, PR China 13 i Department of Pharmacy, School of Health Sciences, University of Patras, Patras 26504, Greece 14 j Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA abstract 15 article info 16 Article history: 17 Received 24 October 2014 18 Revised 13 January 2015 19 Accepted 4 February 2015 20 Available online xxxx 21 Keywords: 22 Myasthenia gravis 23 Experimental animal model 24 Standard operating procedures 25 Pre-clinical experiments 26 Myasthenia gravis (MG) is an autoimmune disorder characterized by generalized muscle weakness due to 27 neuromuscular junction (NMJ) dysfunction brought by acetylcholine receptor (AChR) antibodies in most cases. 28 Although steroids and other immunosuppressants are effectively used for treatment of MG, these medications 29 often cause severe side effects and a complete remission cannot be obtained in many cases. For pre-clinical 30 evaluation of more effective and less toxic treatment methods for MG, the experimental autoimmune myasthenia 31 gravis (EAMG) induced by Torpedo AChR immunization has become one of the standard animal models. Although 32 numerous compounds have been recently proposed for MG mostly by using the active immunization EAMG 33 model, only a few have been proven to be effective in MG patients. The variability in the experimental design, 34 immunization methods and outcome measurements of pre-clinical EAMG studies make it difficult to interpret 35 the published reports and assess the potential for application to MG patients. In an effort to standardize the active 36 immunization EAMG model, we propose standard procedures for animal care conditions, sampling and 37 randomization of mice, experimental design and outcome measures. Utilization of these standard procedures 38 might improve the power of pre-clinical EAMG experiments and increase the chances for identifying promising 39 novel treatment methods that can be effectively translated into clinical trials for MG. 40 © 2015 Published by Elsevier Inc. 41 42 43 44 45 Q4 Introduction 46 Myasthenia gravis (MG) is an autoimmune disorder characterized by 47 fluctuating generalized muscle weakness due to impaired neuromuscu- 48 lar junction (NMJ) transmission. In most MG cases, autoantibodies to the 49 muscle acetylcholine receptor (AChR) cause NMJ dysfunction by activat- 50 ing the complement system and thus giving rise to accumulation of 51 membrane attack complexes that destroy the postsynaptic muscle 52 membrane and reduce the number of functional AChR (Cavalcante 53 et al., 2013; Jayawant et al., 2013). Experimental autoimmune myasthe- 54 nia gravis (EAMG) induced by AChR immunization is the most common- 55 ly used animal model of MG. The active model is mostly performed by 56 injection of purified native AChR in complete Freund's adjuvant (CFA) 57 into mice or rats. Administration of AChR in adjuvant results in production 58 of antibodies to AChR, cytokine expression, accumulation of complement 59 deposits at the NMJ, lysis of the postsynaptic muscle membrane and sub- 60 sequent muscle weakness. Mice immunized with recombinant AChR sub- 61 units or peptides obtained from human, mouse or rat AChR also display 62 IgG and complement deposits and reduced AChR at their NMJs. EAMG 63 induced by active immunization closely mimics the clinical, immuno- 64 pathological and electrophysiological features of MG (Christadoss et al., 65 2000; Tüzün et al., 2011). 66 AChR immunization-based models engage the afferent stage charac- 67 terized by sensitization of AChR reactive T and B cells as well as the effec- 68 tor stage, which involves AChR antibodies and the complement system. 69 Mouse and rat models are available for the active immunization-based 70 EAMG. One of the advantages of the mouse model is that the disease 71 progression is much slower than the rat model allowing researchers to Experimental Neurology xxx (2015) xxx–xxx ⁎ Corresponding author at: Department of Microbiology and Immunology, UTMB, Galveston, TX, USA. Fax: +1 409 747 6869. E-mail address: pchrista@utmb.edu (P. Christadoss). YEXNR-11929; No. of pages: 7; 4C: http://dx.doi.org/10.1016/j.expneurol.2015.02.009 0014-4886/© 2015 Published by Elsevier Inc. Contents lists available at ScienceDirect Experimental Neurology journal homepage: www.elsevier.com/locate/yexnr Please cite this article as: Tuzun, E., et al., Guidelines for standard preclinical experiments in the mouse model of myasthenia gravis induced by acetylcholine receptor immuniza..., Exp. Neurol. (2015), http://dx.doi.org/10.1016/j.expneurol.2015.02.009