Autoreactive T cell responses in insulin-dependent (Type 1) diabetes mellitus. Report of the first international workshop for standardization of T cell assays.
Type 1 diabetes is thought to result from a T cell-mediated destruction of the pancreatic beta-cells. Multiple and sometimes conflicting studies have identified a variety of aberrations in the cellular immune response to autoantigens in persons with the disease. Potential explanations for these discrepancies include incomparable techniques or culture conditions, diversity in the populations of patients or controls tested, and differences in autoantigen preparations. A T cell workshop was organized by the Immunology of Diabetes Society with the aim of appreciating and identifying problems associated with autoreactive T cell assays in type 1 diabetes. As a first phase, a series of candidate autoantigens were analysed by reference laboratories for quality. Subsequently, these preparations, as well as control stimuli, were distributed in a blind fashion to 26 laboratories worldwide, including all experienced centres, for analysis of T cell proliferation assays in 10 recent onset type 1 diabetes and 10 non-diabetic controls. For this analysis, participants used their own assays and references. The islet autoantigen quality control analyses performed prior to the distribution indicate that the quality of recombinant autoantigen preparations requires improvement. For example, several T cell clones specific for glutamic acid decarboxylase (GAD65) were unable to cross-react with GAD65 expressed in baculovirus, yeast or bacteria. Moreover, autoantigens expressed in E. coli interfered with autoantigen-specific proliferation of both T cell clones and peripheral blood mononuclear cells. Nonetheless, responses could be measured to all autoantigen preparations evaluated in the workshop. During the blind phase of the study, all centres were able to reproducibly measure T cell responses to two identical samples of tetanus toxoid, but there was significant interlaboratory variation in sensitivity and extent of the proliferative response measured. Third, the results using candidate autoantigens indicated that although a few laboratories could distinguish type 1 diabetes patients from non-diabetic controls in proliferative responses to individual islet autoantigens, in general, no differences in T cell proliferation between the two groups could be identified. This first T cell workshop on T cell autoreactivity in type 1 diabetes confirms that this was a difficult area for interlaboratory investigations, but provided insight towards future efforts focused on standardizing autoreactive T cell measurements. Some previously reported conflicting results can in part be explained by the observed interlaboratory variability. The inability to discriminate normal controls from new onset type 1 diabetes patients suggests that measuring proliferative responses in PBMC represents an incomplete picture of the immune response, perhaps complicated by difficulties in identifying suitable antigens and assays for standardized use.
Roep
B O
BO
Department of Immunohaematology and Blood Bank, Leiden University Medical Center, Leiden, The Netherlands. broep@pobox.LeidenUniv.NL
Atkinson
M A
MA
van Endert
P M
PM
Gottlieb
P A
PA
Wilson
S B
SB
Sachs
J A
JA
D000293:Adolescent; D000328:Adult; D001323:Autoantibodies / Q000097:blood; D001324:Autoantigens / Q000276:immunology; D002648:Child; D002675:Child, Preschool; D003922:Diabetes Mellitus, Type 1 / Q000276:immunology*; D005260:Female; D005968:Glutamate Decarboxylase / Q000276:immunology; D006801:Humans; D007328:Insulin / Q000276:immunology; D007527:Isoenzymes / Q000276:immunology; D008213:Lymphocyte Activation; D008297:Male; D008565:Membrane Proteins / Q000276:immunology; D008875:Middle Aged; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases / Q000276:immunology; D054635:Receptor-Like Protein Tyrosine Phosphatases, Class 8; D013601:T-Lymphocytes / Q000276:immunology*
D001323:Autoantibodies; D001324:Autoantigens; C087054:ICA1 protein, human; D007328:Insulin; D007527:Isoenzymes; D008565:Membrane Proteins; C516862:PTPRN protein, human; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases; D054635:Receptor-Like Protein Tyrosine Phosphatases, Class 8; D005968:Glutamate Decarboxylase; C401141:glutamate decarboxylase 2