SummaryWe compared T cell receptor (TCK) V-segment frequencies in human leukocyte antigen (HLA) identical siblings to sibling pairs who differ at one or both HLA haplotypes using four Vflspecific and one Vot-specific monoclonal antibody. In every one of nine families HLA-identical sibs had the most similar patterns of V-segment frequencies in their peripheral blood, whereas totally mismatched sihs were, in general, the most dissimilar; HLA haploidentical sibs tended to be intermediate between the two groups. The degree of similarity among HLA-identical fibs was comparable to that observed among three pairs of identical twins suggesting that HLA is the major genetic component influencing TCK V-segment frequency. Consistent with this observation, it was found that the frequency of T ceils expressing particular Vfl segments was skewed towards either CD4 + or CD8 + cells indicating that T cells expressing some Vfl genes may be positively selected primarily by class I or class II major histocompatibility complex proteins. Finally, it was observed that individuals who express the HLA class I specificity, B38, tend to express high levels of Vcz2.3 + cells among their CD8 § T cells. These observations represent definitive proof that human V-segment frequencies are profoundly influenced by the HLA complex. I n mice, the H-2 complex plays a prominent role in determining TCR V-segment frequencies. Both class I and class II antigens influence the pattern of V-segment frequencies. This occurs either by negative sdection (1-5) where CD4 + 8 + thymocytes expressing TCK with high affinity for self MHC molecules plus endogenous peptide are clonaUy deleted or, by positive selection (6-13) where thymocytes with TCR expressing low affinity for the complex are permitted to mature into T cells capable of recognizing "foreign" peptides presented by self MHC molecules. A similar effect on human V-segment frequencies by the HLA complex has been difficult to demonstrate presumably because of the genetic heterogeneity and complexity of the HLA region in the outbred human population. In order to circumvent these difficulties we have analyzed V-segment frequencies within nine large families consisting of multiple siblings. In particular we have compared the V-segment frequency patterns of HLA-identical siblings to sibling pairs who differ at one or both HLA haplotypes. This experimental approach has allowed us to demonstrate the influence of HLA genes on V-segment frequencies in human peripheral blood T cells. Materials and MethodsCell Preparation. Peripheral blood was obtained from normal volunteers. Mononuclear cells were isolated by Ficoll/Hypaque gradient separation and T cells were isolated using magnetic beads to which anti-CD2 mAb was covalently coupled (Dynal Inc., Great Neck, NY). After overnight incubation, the spontaneously released cells were separated into CD4 + and CD8 + cells using magnetic beads covalently coupled with anti-CD4 and anti-CD8. The CD4 + and CDS+ cells obtained were >98% pure. Cold blood cells were purified by ...

To identify disease-specific T cell changes that occur in Crohn's disease (CD), the T cell receptor BV repertoires of lamina propria lymphocytes (LPL) isolated from both the inflamed and "disease-inactive" colons of seven CD patients were compared by the quantitative PCR and DNA sequence analysis. It was observed that the BV repertoires of LPL isolated from the disease-active and disease-inactive parts of the colon from the same individual were very different. Furthermore, nearly all of the differences occurred in CD4 ϩ LPL, with very few differences in the CD8 ϩ population of LPL. Although the pattern of BV segments that was increased in disease-active tissue relative to disease-inactive tissue was different for all seven CD patients, there were several BV segments that increased uniformly in the disease-active tissue of all seven individuals. CDR3 length analysis and DNA sequencing of these BV segments revealed that in six of the seven CD patients there was a striking degree of oligoclonality that was absent from disease-inactive tissue of the same individual. These observations suggest that at least some of the inflammation in CD is the result of responses by CD4 ϩ T cells to specific antigens.

NOD2/CARD15 contributes more to CD susceptibility in familial cases than in sporadic cases, and to an earlier AOO. There is no increased risk of CD for individuals carrying only a single copy of these NOD2/ CARD15 variants, whereas individuals carrying two copies have a 5-15-fold increased risk. The penetrance of the NOD2/CARD15 mutations was estimated at less than 1%.

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