William H. Marshall

Familial, EsD-linked, retinoblastoma with reduced penetrance and variable expressivity

SummaryWe report an example of four generation familial retinoblastoma in which there are three distinct categories of RB gene expression: frank retinoblastoma, unilateral or bilateral; retinoma; and no visible evidence of retinal pathology other than normal degeneration with age. Two large sibships derived from matings informative for RB and EsD provide strong confirmatory evidence for tight linkage between these loci (P=0.0002), and thus assignment of RB to chromosome 13q14. There is a striking difference (P(2%) in RB penetrance between the two principal generations, which suggests that an additional epistatic, host-resistance gene may also be segregating within the family.

Counts and characteristics of macrophage precursors in human peripheral blood

Abstract A method is described for counting the number of macrophages which develop from a sample of human venous blood. Because the macrophages are nondividing cells under these conditions of culture, it is possible to use the count to describe the size of a population of macrophage precursors in the blood. Counts on 43 healthy individuals show that on average there are some 200,000/ml of blood, which is 5–6% of the total white cell count. The precursor cells are mononuclear and the majority of them will adhere to plastic. These characteristics suggest that monocytes are the precursor cells.

Analysis of monoclonal antibodies specific for unique and shared determinants on HLA-DR4 molecules.

The specificities for seven mAbs to HLA-DR4 were determined initially using homozygous BCLs and L-cell transfectants expressing wild-type DR molecules. Three antibodies (NFLD.D1, NFLD.M1, and NFLD.D7) bound all DR4 molecules, but only one was specific for DR4. Four antibodies (NFLD.D2, NFLD.D3, NFLD.D8, and NFLD.D10) reacted with some but not all DR4 subtypes and had extra reactions, particularly with DR gene products associated with susceptibility to RA. To localize the antibody-binding epitopes on DR4 molecules, the antibodies were then analyzed on transfectants expressing hybrid genes, which were generated by exon shuffling of DRB1*0403 and DRB1*0701. Two of the pan-DR4 antibodies bound epitopes that require the beta 2 domain while the third mapped primarily to the HVR-I region. One antibody NFLD.D10 to subtypes of DR4 mapped to residues 40-97 on DR beta 1*0403 chains. Comparison of reaction patterns with amino acid sequences suggest that the antibodies against subtypes of DR4 are specific primarily for a region containing sequences postulated to determine susceptibility to RA.

Amino acids in the peptide-binding groove influence an antibody-defined, disease-associated HLA-DR epitope.

A shared amino‐acid sequence on the a helix of certain DRβ1 chains is predicted to generate a ‘shared epitope’ that is implicated in susceptibility to the development of rheumatoid arthritis (RA). Different relative risks (RR) for disease susceptibility and severity conferred by these DRβ31 chains suggest that their ‘shared epitopes’ are not equivalent. A set of monoclonal antibodies (MoAb) that map to the critical region, and for which optimal binding depends on DR context and cell lineage, was used to test this idea. Mapping experiments using mutated DRβ1* molecules showed that the antibody‐binding epitopes are overlapping; residue 70Q is pivotal for each, but neighbouring residues on the a helix and on the floor of the groove are also involved. Importantly, these epitopes are profoundly modified by peptide loading of DRβ31*0401 molecules. These data suggest that ‘shared epitopes’ on DR molecules that are associated with RA are influenced by their context; such structural modifications may be the basis for the varying susceptibilities conferred by these DR molecules for the development of RA.

HLA-DP Epitope Typing Using Monoclonal Antibodies

We have made a panel of murine anti-DP monoclonal antibodies for serological typing of HLA-DP polymorphisms; they can be used in microcytotoxicity (for 7 epitopes) and binding assays (for 8 epitopes). The antibodies detect polymorphic differences in both alpha and beta chains. As immunogens we sometimes used B-lymphoblastoid lines or purified DP molecules but mostly used mouse fibroblast transfectants expressing DP molecules. The DP beta genes were made from a cloned DPB1*0201 gene by replacing its major area of polymorphism with matching stretches of DNA amplified from other alleles; cloned DPA1*01 and DPA1*02 genes were used for transfection along with the beta chain genes. The monoclonal antibodies showed reaction patterns that correlated with the presence of particular amino-acid sequence motifs; thus none of the antibodies is allele-specific. They bind instead to epitopes which are found on a number of different HLA-DP types. We have constructed frequency tables so that the epitope (motif) data can be interpreted as the most likely genotype in each case. The basic assumption to justify this work is that HLA-DP matching or mismatching will likely influence transplant outcome, particularly in bone marrow transplantation. The present challenge is to define permissive and nonpermissive combinations of HLA-DP; it may be that matching for epitopes, rather than for full alleles, will help to resolve this issue.

Immunoglobulin concentration and Gm allotypes in a family with thirty‐three cases of myotonic dystrophy

Serum IgG, IgA, and IgM concentrations were measured in 120 members of a family with 33 cases of Dystrophia myotonica (Dm) and 27 members who were “possibly affected”. The Dm individuals had significantly lower serum concentrations of IgG and IgA (P<0.01), while the “possibly affected” did not differ from the matched pair controls. IgG subclass concentrations were measured and Gm and Am types determined. The lower concentration of IgA in the affected individuals was not associated with a particular Am type. The concentration of IgG3 was barely lower in the affected than in the controls (P=0.05), but there were no differences for IgGl. When IgG3 concentration was compared according to Gm haplotype, only the two affected individuals who were Gm gg had a statistically significant lower concentration than the 12 controls (P<0.02). Thus, there is no evidence that a particular subclass of IgG is being hypercatabolized in our Dm patients. A rare Gm haplotype, Gm(‐, n, b) had entered the family with two brothers; it is not known whether this codes for an IgGl‐IgG3 hybrid molecule or a normal IgGl molecule with an unknown Gm allele or a yl deleted Gm haplotype.

HLA-DR residues accessible under the peptide-binding groove contribute to polymorphic antibody epitopes

Many residues involved in polymorphic antibody-binding epitopes on class II molecules are located on the alpha-helix of DR beta chains. Although they have received less attention, residues in the peptide-binding groove and second domain of the DR beta chain may also be critical for polymorphic anti-DR antibody epitopes. In this study, we used transfectants expressing site-directed mutations at positions in the HLA-DR beta 1 and beta 2 domains and flow cytometry to define the epitopes of several polymorphic anti-DR antibodies. Both DR(beta 1*0403) residues 14 and 25 were shown to be involved in the epitopes of mAbs DA6. 164, HU-20, Q5/6, and 50D6, and DR(beta 1*0701) residue 14 was shown to be critical for the epitopes of two DR7-specific mAbs, SFR 16-DR7M and TAL13.1. Unlike most other residues shown to be important in antibody-binding epitopes, residue 14 is located in the floor of the peptide-binding groove and residue 25 is in an outer loop, each with their side chains pointing down, such that antibodies may directly contact these residues from below the binding groove. Two residues in the beta 2 domain, beta 180 and beta 181, were also shown to be involved in the epitopes of three polymorphic anti-DR mAbs, NFLD.D1, NFLD.M1, and LY9. Although these two residues are close to the transmembrane domain in the linear sequence, their solvent accessibility in the DR1 structures is quite impressive. Our data provide new evidence that residues accessible under the peptide-binding groove contribute to polymorphic antibody-binding epitopes.

Bf polymorphism. A very fast variant from Nigeria

SummaryIn a collection of Nigerian serum samples typed for alleles of factor B of the alternative complement pathway, a very high frequency of BfF was found (0.69). In addition, a new variant was found in two samples. This variant (F1.29) moved faster than BfF1. It was hemolytically active.

Immunoglobulins in Familial Hodgkin’s Disease and Immunodeficiency in Newfoundland

A familial aggregate of 7 cases of Hodgkin’s disease, 3 of common variable immunodeficiency, 8 of other lymphoreticular malignancies, and 4 of embryonic tumours is being studied in a multidiscipline f

Polymorphism of the Seventh Complement Component, C7, in Chinese

An allele of the seventh complement component, C7*2, which is very rare in Europeans, has been found at balanced polymorphic frequency (0.15) in Chinese. This marker may be useful for anthropological studies.