Given the complexity of SLE, the study will focus on immunological traits that occur associated with the disease, but which are likely to be under a less complex genetic regulation, and therefore more amenable to genetic mapping. In particular, immune phenotypes related to T cell apoptosis, which are common denominators among autoimmune disorders, will be investigated. Abnormalities in apoptosis regulation have been implicated in the breakdown of immunologic tolerance to self-antigens in SLE, possibly through the production of autoantibodies directed to apoptotic cells which deposit in the kidney leading to lupus nephritis, a very severe manifestation of the disease. Patients and relatives will be screened for clinical and immunological phenotypes, and interesting phenotypes will be mapped. The possibility of characterizing identified genes in a mouse model, and conversely of searching for human genes previously identified in mice will be a main asset for lupus research.
Specific objectives and work plan:
The proposed study aims at the identification and characterization of genes involved in the susceptibility to SLE.
1. Phenotype identification and characterization in the NWB/NZB mice models.
These mice models will be screened for immunological phenotypes, namely lymphocyte activation, apoptosis resistance and abnormalities in apoptosis related molecules. This strategy will permit the identification and characterization of some disease associated traits that can then be investigated in human SLE patients. Conversely, the role in the physiopathology of the disease of genes identified human SLE will be evaluated in these mouse models.
2. Sample collection and organization of data and biological specimens.
The ADL has nearly 1800 associated patients that are being evaluated following a predefined comprehensive protocol (that includes a structured interview, a clinical examination and a classical battery of serological tests for SLE), providing an accurate overall clinical assessment. The same evaluation protocol will be used for the assessment of patients from the HPD in S. Miguel. In mainland Portugal, 82 familiar cases have been identified out of the associated patients. A number of families with members suffering from SLE and other autoimmune disorders such as diabetes and rheumatoid arthritis have also been identified, and will be included in the study. Relatives are assessed using a standard protocol to identify autoimmune disease, and relatives with SLE undergo the previously mentioned protocol.
The candidate will actively participate in all stages of sample and data collection, including the establishment of a cell/DNA/serum bank. She will be involved in the genealogical reconstruction of pedigrees using family history, church registers and civic records. She will also collaborate in the organization of the clinical and genetic data collected throughout the study in a database specific for genetic analysis.
3. Identification and characterization of phenotypes contributing to the disease in patients and relatives.
Patients and relatives will be screened for subphenotypes for SLE, with particular focus on clinical patterns, prevalence of different types of autoantibodies, and lymphocyte physiology. Particular attention will be devoted to alterations in T cell regulation, apoptosis and apoptosis signaling pathways. The role of molecules involved in apoptosis, namely the CTLA4 and CD28 regulators of T cell activation, will be analyzed in patients and relatives, searching for alterations associated with the disease.
4. Mapping of susceptibility loci for SLE or disease associated phenotypes, and progress to fine mapping, cloning and characterization of candidate genes for these traits.
A genome wide screening will be conducted for SLE and for identified and characterized sub-phenotypes. Automated genotyping using 400 polymorphic markers throughout the genome will be performed using the ABI Prism genotyping system. Common genetic analysis strategies will be employed: linkage and family-based association studies according to the population structure. Fine mapping will follow identification of susceptibility loci. Candidate genes in the disease critical region(s) will be analyzed for mutations specific for affected individuals. Emphasis will be given to genes that map in regions syntenic with the loci identified in mouse models of Lupus.
References:
1. Lindqvist AK and Alarcon-Riquelme ME. The genetics of systemic lupus erythematosus. Scand J Immunol. 50: 562-571, 1999.
2. Vyse TJ and Kotzin BL. Genetic susceptibility to systemic lupus erythematosus. Annu Rev Immunol. 16: 261-292, 1998.
3. Harley JB, Moser KL, Gaffney PM and Behrens TW. The genetics of human systemic lupus erythematosus. Curr Opin Immunol. 10: 690-696, 1998.
4.Balomenos D, MartinCaballero J, Garcia M, Prieto I, Flores J, Serrano M, Martinez C. A T cell cycle inhibitor p21 controls T cell proliferation and sex-linked Lupus development. Nature Med 6(2): 171-176, 2000.
5. Shai R, Quismorio FP Jr, Li L, Kwon OJ, Morrison J, Wallace DJ, Neuwelt CM, Brautbar C, Gauderman WJ and Jacob CO. Genome-wide screen for systemic lupus erythematosus susceptibility genes in multiplex families. Hum Mol Genet
8: 639-644, 1999.
6. Gaffney PM, Kearns GM, Shark KB, Ortmann WA, Selby SA, Malmgren ML, Rohlf KE, Ockenden TC, Messner RP, King RA, Rich SS and Behrens TW. A genome-wide search for susceptibility genes in human systemic lupus erythematosus sib-pair families. Proc Natl Acad Sci USA. 95: 14875-1479, 1998.
7. Jacobsen S, Petersen J, Ullman S, Junker P, Voss A, Rasmussen JM, Tarp U, Poulsen LH, van Overeem Hansen G, Skaarup B, Hansen TM, Podenphant J and Halberg P. A multicentre study of 513 Danish patients with systemic lupus erythematosus. II. Disease mortality and clinical factors of prognostic value. Clin Rheumatol. 17: 4784-84, 1998.
8. Morel L, Mohan C, Yu Y, Schiffenbauer J, Rudofsky UH, Tian N, Longmate JA and Wakeland EK. Multiplex inheritance of component phenotypes in a murine model of lupus. Mamm Genome. 10: 176-181, 1999.
9. Craft J, Peng S, Fujii T, Okada M, Fatenejad S. Autoreactive T cells in murine lupus: origins and roles in autoantibody production. Immunol Res. 19: 245-257, 1999.
10. Morel L, Tian X, Croker B, Wakeland E. Epistatic modifiers of autoimmunity in a murine model of Lupus nephritis. Immunity 11:131-139, 1999.
11. Lindqvist A, Steinsson K, Johanneson B, Kristjánsdóttir H, Àrnasson A, Gröndal G, Jonasson I, Nagnusson V, Sturfelt G, Truedsson L, Svenungsson E, Lundberg I, Terwilliger J, Gyllensten U, Alarcón-Riquelme M. A susceptibility locus for Human Systemic Lupus Erythematosus (hSLE1) on chromosome 2q. J Autoimmun 14(2):169-178, 2000.
.jpg)
