F.76. NKT Cell Populations are Not Decreased in the Majority of Patients with XIAP Deficiency
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F.75. Molecular and Functional Studies in T Cells from Healthy Carriers of FOXP3 Mutations Rosa Bacchetta, 1 Sara Di Nunzio, 1 Lucia Beatrice Perroni, 2 Laura Passerini, 1 Sarah Allan, 3 Alicia Alstad, 3 Anne Junker, 3 Megan Levings, 3 Maria Grazia Roncarolo. 41 San Raffaele Hospital, Milan, Italy; 2 Galliera Hospital, Genoa, Italy; 3 University of British Columbia, Vancouver, BC, Canada; 4 VIta Salute San Raffaele University, Milan, Italy IPEX is a severe genetic autoimmune disease character- ized by immune dysregulation, polyendocrinopathy, entero- pathy and X-linked inheritance. The disease is due to mutations in FOXP3, a transcription factor crucial for differentiation and function of naturally occurring CD4+ CD25+ regulatory T (Treg) cells and for the maintenance of peripheral tolerance. FOXP3 is an X-linked gene that is subject to X-inactivation. Therefore, female carriers of FOXP3 mutations are expected to be mosaics of cells expressing the wild type (wt) or the mutated (mut) allele of FOXP3, although they are healthy with no clinical history of autoimmune manifestations. In order to investigate the wt/mut FOXP3 distribution in different peripheral T cell subsets of healthy carriers, we set up a method to evaluate, at cellular level, both the pattern of X-inactivation and the presence of FOXP3 mutation. We performed X-inactivation analysis on freshly isolated and/or in vitro expanded CD4+ CD25+/CD25- T cells, purified naive/ memory T cells and in vitro stimulated Ag-specific T cells derived from mothers of IPEX patients carrying FOXP3 mutations in different sites of the gene. Preliminary results demonstrate that a random distribution of wt/mutFOXP3 expressing cells does not always occur. For example, a predominant expression of wtFOXP3 was observed in the Treg cells from a mother carrier of a fork head binding domain mutation. This study will provide useful insights into the role of FOXP3 mutations in the development and function of Treg cells and Ag-specific T effector cells and will contribute to a better understanding of the genotype-phenotype correlation in IPEX. doi:10.1016/j.clim.2008.03.187 F.76. NKT Cell Populations are Not Decreased in the Majority of Patients with XIAP Deficiency Rebecca Marsh, Joyce Villanueva, Kejian Zhang, Kimberly Risma, Alexandra Filipovich, Jack Bleesing. Cincinnati Children's Hospital Medical Center, Cincinnati, OH NKT cells are a unique population of T cells that express an invariant TCR which recognizes glycosphingolipid antigens presented by the CD1d molecule. These cells are known to be absent in X-linked Lymphoproliferative Disease (XLP) due to SAP mutation, and were previously reported to be decreased in patients with XLP secondary to mutations in the BIRC4 gene which encodes the X-linked Inhibitor of Apoptosis (XIAP). This lack of normal NKT cell populations has been postulated to contribute to disease pathogenesis in patients with XLP. In order to further characterize NKT cell popula- tions in patients with XIAP deficiency, we studied whole blood NKT cell levels in four patients over several months by flow cytometric analysis. Identification was based on the co- expression of TCR-Vα24 and TCR-Vβ11, and results were compared to 50 pediatric and adult controls. We found that NKT cell populations were decreased in only 1 patient with XIAP deficiency. The other patients displayed normal or increased NKT cells compared to controls. We conclude that NKT cell populations are not decreased in the majority of patients with XIAP deficiency, and disease pathogenesis is likely not dependent on a lack of NKT cells. doi:10.1016/j.clim.2008.03.188 F.77. Can the Frequency of Memory B Cells be Used to Diagnose Primary Antibody Failure? Elizabeth Bateman, Mary Lucas, Kerri Packwood, Lisa Ayers, Daniel Harrison, John Burden, Ross Sadler, Berne Ferry, Helen Chapel. Oxford Radcliffe Hospital NHS Trust, Oxford, United Kingdom Since the reports of reduced numbers of CD27 memory B cells and increased immature CD21low B cells in patients with Common Variable Immune Deficiency (CVID), a number of classification systems using B cell phenotyping have been proposed. In a large European multi-centre trial using B cell phenotyping to define subgroups in CVID, the majority of patients were found to have reduced switched memory B cells (CD19+CD27+IgD-). Classification of individuals by measurement of B cell subpopulations may help identify the underlying immune defects and aid understanding of the aetiology of associated clinical manifestations. This study looked at a cohort of patients with recurrent infection, but no evidence of any primary antibody deficiency and compared them with patients with either CVID, IgG subclass deficiency, selective IgA deficiency or specific antibody deficiency. Memory B cell phenotypes were assessed by flow cytometry using the whole blood method. Switched (CD19+ CD27+IgD-) and unswitched (CD19+CD27+IgD+) memory B cells, as well as transitional and CD21low B cell populations were investigated. The aim of this study was to assess whether class switched memory B cell frequency could be used to distinguish those patients presenting with recurrent infections that had primary antibody failure, such as CVID, or partial antibody failure such as IgA deficiency or IgG subclass deficiency. This study will help in determining whether measuring memory B cell phenotype is a useful tool in the diagnosis of primary antibody failure. doi:10.1016/j.clim.2008.03.189 F.78. Study of the Selective Response to Different Glucocorticoids (GC) in Subject Affected by IPEX Marilena Granzotto, 1 Erica Valencic, 2 Elisa Piscianz, 1 Alberto Tommasini, 2 Giuliana Decorti. 21 Institute for Child Health Burlo Garofolo, Trieste, Italy; 2 University of Trieste, Trieste, Italy Immuodysregulation, polyendocrinopathy, enteropathy X- Linked syndrome (IPEX) is a genetic failure of immune S68 Abstracts
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Transcript of F.76. NKT Cell Populations are Not Decreased in the Majority of Patients with XIAP Deficiency
F.75. Molecular and Functional Studies in T Cellsfrom Healthy Carriers of FOXP3 MutationsRosa Bacchetta,1 Sara Di Nunzio,1 Lucia Beatrice Perroni,2
Laura Passerini,1 Sarah Allan,3 Alicia Alstad,3 Anne Junker,3
Megan Levings,3 Maria Grazia Roncarolo.4 1San RaffaeleHospital, Milan, Italy; 2Galliera Hospital, Genoa, Italy;3University of British Columbia, Vancouver, BC, Canada;4VIta Salute San Raffaele University, Milan, Italy
IPEX is a severe genetic autoimmune disease character-ized by immune dysregulation, polyendocrinopathy, entero-pathy and X-linked inheritance. The disease is due tomutations in FOXP3, a transcription factor crucial fordifferentiation and function of naturally occurring CD4+CD25+ regulatory T (Treg) cells and for the maintenance ofperipheral tolerance. FOXP3 is an X-linked gene that issubject to X-inactivation. Therefore, female carriers ofFOXP3 mutations are expected to be mosaics of cellsexpressing the wild type (wt) or the mutated (mut) alleleof FOXP3, although they are healthy with no clinical historyof autoimmune manifestations. In order to investigate thewt/mut FOXP3 distribution in different peripheral T cellsubsets of healthy carriers, we set up a method to evaluate,at cellular level, both the pattern of X-inactivation and thepresence of FOXP3 mutation. We performed X-inactivationanalysis on freshly isolated and/or in vitro expanded CD4+CD25+/CD25- T cells, purified naive/ memory T cells and invitro stimulated Ag-specific T cells derived from mothers ofIPEX patients carrying FOXP3 mutations in different sites ofthe gene. Preliminary results demonstrate that a randomdistribution of wt/mutFOXP3 expressing cells does notalways occur. For example, a predominant expression ofwtFOXP3 was observed in the Treg cells from amother carrierof a fork head binding domain mutation. This study willprovide useful insights into the role of FOXP3 mutations inthe development and function of Treg cells and Ag-specific Teffector cells and will contribute to a better understandingof the genotype-phenotype correlation in IPEX.
doi:10.1016/j.clim.2008.03.187
F.76. NKT Cell Populations are Not Decreased in theMajority of Patients with XIAP DeficiencyRebecca Marsh, Joyce Villanueva, Kejian Zhang, KimberlyRisma, Alexandra Filipovich, Jack Bleesing. CincinnatiChildren's Hospital Medical Center, Cincinnati, OH
NKTcells are a unique population of Tcells that express aninvariant TCR which recognizes glycosphingolipid antigenspresented by the CD1d molecule. These cells are known to beabsent in X-linked Lymphoproliferative Disease (XLP) due toSAP mutation, and were previously reported to be decreasedin patients with XLP secondary to mutations in the BIRC4gene which encodes the X-linked Inhibitor of Apoptosis(XIAP). This lack of normal NKT cell populations has beenpostulated to contribute to disease pathogenesis in patientswith XLP. In order to further characterize NKT cell popula-tions in patients with XIAP deficiency, we studied wholeblood NKT cell levels in four patients over several months by
flow cytometric analysis. Identification was based on the co-expression of TCR-Vα24 and TCR-Vβ11, and results werecompared to 50 pediatric and adult controls. We found thatNKT cell populations were decreased in only 1 patient withXIAP deficiency. The other patients displayed normal orincreased NKT cells compared to controls. We conclude thatNKT cell populations are not decreased in the majority ofpatients with XIAP deficiency, and disease pathogenesis islikely not dependent on a lack of NKT cells.
doi:10.1016/j.clim.2008.03.188
F.77. Can the Frequency of Memory B Cells be Usedto Diagnose Primary Antibody Failure?Elizabeth Bateman, Mary Lucas, Kerri Packwood, Lisa Ayers,Daniel Harrison, John Burden, Ross Sadler, Berne Ferry,Helen Chapel. Oxford Radcliffe Hospital NHS Trust,Oxford, United Kingdom
Since the reports of reduced numbers of CD27 memory Bcells and increased immature CD21low B cells in patientswith Common Variable Immune Deficiency (CVID), a numberof classification systems using B cell phenotyping have beenproposed. In a large European multi-centre trial using B cellphenotyping to define subgroups in CVID, the majority ofpatients were found to have reduced switched memory Bcells (CD19+CD27+IgD-). Classification of individuals bymeasurement of B cell subpopulations may help identifythe underlying immune defects and aid understanding of theaetiology of associated clinical manifestations. This studylooked at a cohort of patients with recurrent infection, butno evidence of any primary antibody deficiency andcompared them with patients with either CVID, IgG subclassdeficiency, selective IgA deficiency or specific antibodydeficiency. Memory B cell phenotypes were assessed by flowcytometry using the whole blood method. Switched (CD19+CD27+IgD-) and unswitched (CD19+CD27+IgD+) memory Bcells, as well as transitional and CD21low B cell populationswere investigated. The aim of this study was to assesswhether class switched memory B cell frequency could beused to distinguish those patients presenting with recurrentinfections that had primary antibody failure, such as CVID, orpartial antibody failure such as IgA deficiency or IgG subclassdeficiency. This study will help in determining whethermeasuring memory B cell phenotype is a useful tool in thediagnosis of primary antibody failure.
doi:10.1016/j.clim.2008.03.189
F.78. Study of the Selective Response to DifferentGlucocorticoids (GC) in Subject Affected by IPEXMarilena Granzotto,1 Erica Valencic,2 Elisa Piscianz,1
Alberto Tommasini,2 Giuliana Decorti.2 1Institute for ChildHealth Burlo Garofolo, Trieste, Italy; 2University of Trieste,Trieste, Italy
Immuodysregulation, polyendocrinopathy, enteropathy X-Linked syndrome (IPEX) is a genetic failure of immune
S68 Abstracts
