Publications by Year: 1993

Intestinal intraepithelial lymphocytes (IEL) are a phenotypically distinct T cell population of unknown function. The majority of human intestinal IEL express the TCR-alpha beta, the CD8 accessory molecule, and the CD45RO Ag, suggesting that they are MHC class I-restricted memory T cells. Recent analyses of the TCR alpha- and beta-chains expressed by these cells have shown marked skewing toward one or several V region genes in individual donors and revealed the presence of clonally expanded cells. In addition, functional data has suggested that the MHC class I-like CD1 molecules may be the target ligands for some human intestinal IEL clones. This report examines in detail the TCR-beta repertoire of human jejunal IEL to determine what fraction of these cells are clonally expanded and to determine whether a particular subset of V beta genes are utilized by the clonally expanded cells. The results demonstrate that the majority of IEL are derived from the expansion of a relatively few T cell clones and that these clones can utilize a large number of different V beta genes. Oligoclonal expansion is also demonstrated among lamina propria lymphocytes (LPL), with overlapping but distinct clones detected in the LPL vs the IEL populations. These results indicate that most intestinal IEL-alpha beta, and a subpopulation of LPL, are specific for a limited number of Ag and place constraints on the possible roles played by IEL in the defense against diverse environmental pathogens or in the generation of oral tolerance.

CD4-CD8- (double negative [DN]) alpha/beta T cells are a largely uncharacterized subpopulation of unknown function. To investigate whether these cells are selected to recognize particular antigens or antigen-presenting molecules, DN alpha/beta T cells were purified from the peripheral blood of five normal donors and their T cell receptor (TCR) alpha and beta chains were examined. Random cloning of TCR alpha chains by single-sided polymerase chain reaction (PCR) amplification identified an invariant rearrangement between V alpha 24 and J alpha Q, with no N region diversity, which was expressed preferentially by DN alpha/beta T cells from all donors. Random cloning also identified a precise V alpha 7.2-J alpha (IGRJa14) rearrangement, with two variable amino acids encoded in the V-J junction, which was enriched in the DN alpha/beta T cell preparations from some, but not all, donors. Analysis of TCR beta chains by quantitative PCR amplification demonstrated that the expression of four V beta gene families, V beta 2, 8, 11, and 13, was markedly increased in these DN alpha/beta T cell preparations. The expression of particular TCRs by DN alpha/beta T cells from multiple donors indicates that these cells, or at least a subpopulation of cells with this phenotype, recognize a limited spectrum of antigens and suggests that they may use nonpolymorphic antigen-presenting molecules.

Analysis of immunoglobulin gene rearrangements by Southern blotting is a sensitive and specific method for detecting B cell malignancies but requires a relatively large amount of intact DNA. It cannot be utilized in many cases where only a small amount of tissue is available or where the tissue has been fixed. This report demonstrates that polymerase chain reaction (PCR) amplification in conjunction with single-strand conformational polymorphism (SSCP) analysis can be utilized to detect clonal immunoglobulin heavy chain (IgH) gene rearrangements. IgH gene rearrangements from a series of frozen or formalin-fixed B cell malignancies were PCR-amplified using oligonucleotide primers, based upon consensus sequences in the IgH variable and joining regions. Analysis of the single-stranded PCR products on nondenaturing polyacrylamide gels revealed discrete SSCPs corresponding to the malignant B cells. These SSCPs were detectable when the malignant cells represented as few as 0.2% of the total mononuclear cells in peripheral blood. PCR amplification in conjunction with SSCP analysis thus provides a sensitive and specific method to detect clonal IgH rearrangements from minute amounts of fresh, frozen, or fixed tissue.