Influence of HLA-D/DR antigen disparity in CTL generation in vitro.

This report describes the influence of HLA-D/DR antigen disparity upon the level of cytotoxicity in allogeneic in vitro cultures. Allogeneic cultures, between unrelated HLA-D/-DR full house donors, tested in CML gave three different levels of cytotoxicity, termed weak, intermediate and strong cytotoxicity. HLA-D/-DR compatibility predicts weak cytotoxicity and two HLA-B antigen incompatibility predicts strong cytotoxicity. On the contrary, HLA-A antigens have no major influence upon the strength of cytotoxicity. Accepting that the MLC/CML reaction is an in vitro parallel to the in vivo transplantation of allogeneic tissue, the observations are in accordance with the results of HLA-D/-DR matching for graft survival in human renal transplantation.

Activation of human alloreactive cytotoxic precursor T lymphocytes.

The requirements for allogeneic T-cell activation have been studied in experiments with T and/or B cells as stimulator. Although target determinants (TDs, defined by CTL effectors in CML) are present on B and T cells used as target cells, this study indicates that TDs are functionally different when expressed on B and T cells used as stimulator cells, as only B cells can activate CTL precursors. Further, the study confirms that inducing TDs and strong lymphocyte-activating determinants (LADs, defined by proliferation in MLC) can be distinct structures found on two different stimulator B cells. The study suggests that binding of cytotoxic precursor T cells to TDs per se does not allow any detectable activation or start of proliferation and differentiation but requires another function of the stimulator cells in the non-T-cell compartment. The nature of this function is unknown, but it is the background for the first signal received by the TD-specific clones of CTL precursors, resulting in the expression of growth receptors for T-cell growth factor or interleukin 2 which is the second signal necessary for clonal expansion and differentiation.

Competitive inhibition of T-cell mediated lympholysis by platelets.

Platelets are known to possess serologically detectable HLA-A, B, C (class I) antigens, but not HLA-DR (class II) antigens. We have used platelets as non-labelled (cold) competitors for cell-mediated lympholysis directed against determinants on PHA (3 days) and PWM (7 days) 51Cr-labelled (hot) lymphoblasts, i.e. T and B lymphoblasts, respectively. It is found that platelets are able to incompletely inhibit cytolysis against T-lymphoblasts, but not B-lymphoblasts. The inhibition is immunologically specific in the sense that only platelets autologous to the original responder cell do not. The immunogenetic specificity of platelet blocking is unknown at present, since no allogeneic third party platelets have been investigated. It is further found that platelets do not inhibit cytolysis by cytotoxic lymphocytes previously qualified to identify HLA-non A, B, C, D/DR determinants on T-lymphoblasts. Like in serology, platelets have to mature (greater than or equal to 7 days) in order to obtain optimal results. Since platelets are easy to procure and maintain their reactivity for months by simple storage in saline at 4 degrees C, platelets may be used to screen for cell-mediated lympholysis against HLA-class I, II or "new" determinants.

Human B-blast specific target determinants in CML: a family study.

Human B blast specific target determinants, selectively identified on PWM stimulated purified B lymphoblasts by in vitro generated CTLs, have previously been studied in the population and showed association to and inclusion of HLA-DR geneproducts. This report indicates that B blast target determinants are products of genes which in a codominant mendelain way segregate with the HLA haplotypes in 4 selected families. Furthermore tests of families with HLA-B/D, DR and HLA-D, DR/GLO recombinations show that human B blast specific target determinants are coded from loci (locus) in the HLA-D region, between HLA-B and GLO.

Typing by cytotoxic lymphocytes. Genetics of HLA-non-A, B, C determinants - family studies.

Some CTLs recognize HLA-structures in a different way than antibodies, i.e. they may identify other epitopes than antibodies or may define genetically distinct structures. From population studies, in vitro educated CTLs were selected giving rise to significant lysis on PHA-lymphoblasts in the absence of the sharing of HLA-A, B, (C) determinants between stimulator and target lymphocytes. Based on pair-wise correlations these CTLs form 3 clusters, identifying structures associated to HLA-markers. Subsequent testing in 14 complete families of greater than or equal to 4 children allowed the observations: (1) CML-traits assigned on the basis of single CTLs often segregated in a non-mendelian way or independent of HLA. (2) Assignment made on the basis of CTL-clusters segregated with HLA. (3) in 2 informative HLA-B/D, DR recombinant families, CML-traits segregated with HLA-B. (4) No haplotype could be assigned more than one CML-trait.