Biomedical subjects
Publications and source records attributed to D Middleton.
Five new restriction fragment length polymorphisms (RFLPs) using TaqI and a DR beta probe have been found in Caucasian individuals. The families of these individuals have been tissue typed by serological and RFLP methods. The new RFLPs are similar to previously established RFLPs except for the size of one fragment in each instance.
DNA from multiple sclerosis (MS) patients and healthy control individuals from the Northern Ireland population, was assessed by restriction fragment length polymorphism (RFLP) analysis to look for disease-associated polymorphisms. HLA-DR and -DQ allogenotyping was performed using a single enzyme (TaqI)/Multiple probe system. The TaqI/DR beta RFLPs correlate well with serologically defined specificities and in addition detect further subtypes of these associated with DQ or Dw specificities. The results confirm an association of MS with DR beta 15 and show a decreased frequency of DR beta 4 in the patients. An increased frequency of DR beta 17 in patients negative for DR beta 15 was also found. There is a decrease in the frequency of the TaqI/DQ beta 3 (TA10 + ve) RFLP and the TaqI/DQ alpha 2LL genotype in the MS patients, these not being associated with DR beta 15. In addition RFLP analysis of the DPA1 and DPB1 genes reveals an MspI/DP beta 3.0-kb fragment which occurs at a higher frequency in the patients than in the controls.
Previous reports of an association between HLA tissue type and Wegener's granulomatosis are contradictory. By using for the first time a highly sensitive restriction fragment-length polymorphism (RFLP) analysis in addition to standard microcytotoxicity assays, the largest series yet investigated (41 patients) was tissue typed. No association was found between any specific HLA antigen and Wegener's granulomatosis. Although the condition appears to be immunologically mediated, this study indicates that the HLA antigens do not have a major role.
The immunoglobulin (IgM) heavy chain constant region gene of the channel catfish, Ictalurus punctatus, has been cloned and characterized. The gene contains four constant region domain-encoding exons (CH1 to CH4) expressed in the secreted form of the immunoglobulin, and two exons encoding the transmembrane (TM) domain utilized in the lymphocyte membrane receptor form of the immunoglobulin. The sequence of a cDNA clone encoding the 3' region of the message for the membrane receptor form of the mu chain indicates that the TM1 exon is spliced directly to the CH3 exon, and not into a site within the CH4 exon, as occurs in the mammals, a shark and an amphibian. This unusual pattern of splicing, which produces a membrane heavy chain that is characteristically smaller than the secreted heavy chain, may be common to all teleost fish.
Mineralization of the supraspinatus tendon is proposed as a cause of forelimb lameness in dogs. A new radiographic position (cranioproximal-cranio-distal) is advanced to detect abnormalities of the craniomedial aspect of the proximal portion of the humerus. Four dogs had surgery to remove mineralized debris from the supraspinatus tendon insertion and have improved limb function. Two dogs were evaluated in depth, with no signs of relapse 2 to 4 years after surgery. Other dogs, without apparent forelimb lameness, have had mineralization detected by use of radiography, thus emphasizing the need for thorough forelimb evaluation before determining that the mineralization is indeed the cause of lameness.
Multiple sclerosis has been shown to be associated with the presence of certain major histocompatibility (MHC) tissue antigens which are coded on chromosome 6. There are racial differences in the antigens associated with MS. The strength of the associations vary in different communities in Western Europe. We have investigated the association between MS and MCH antigens in Northern Ireland in a group of 104 patients. There is a particularly strong association between MS and HLA-DR2, 65.4% compared with 25.5% in 184 controls. A weaker association has been demonstrated with HLA-A3 (44.2% vs 26.5% in controls). There have been conflicting reports concerning an association of HLA-DW2 and HLA-DR2 with a rapidly progressive form of MS. Our data do not support that hypothesis.
HLA-DR allogenotyping was performed on a panel of 1019 Caucasian individuals from the Northern Ireland population, of whom 150 were also HLA-DQ allogenotyped, using a single enzyme (TaqI)/multiple probe system. TaqI/DR beta restriction fragment length polymorphisms correlate well with the serologically defined specificities and in addition detect further subtypes of these associated with DQ or Dw specificities. The most frequent DR beta allogenotypes were DR beta 15, DR beta 4, and DR beta 17 (Dw24). The frequency of the subtypes of DR beta 4 associated with DQ beta 3 (TA10+) was higher than that reported elsewhere for Caucasian populations, whereas the frequency of the subtypes of DR4 beta associated with DQ beta 3(TA10-) was subsequently reduced. The subtypes of DR beta 4 associated with DQ beta 3 (TA10+) were strongly associated with B44. The TaqI/DX alpha gene polymorphisms, as revealed by the DQ alpha cDNA probe, were found to be associated with DR beta allogenotypes.
After primary successful PTA, 199 patients were randomized into one of three treatment groups, namely, placebo or a combination of 75 mg dipyridamole with either 330 mg (high dose) or 100 mg (low dose) acetylsalicylic acid (ASA) tid. The duration of treatment was six months. Of the 199 patients admitted to the study, 156 completed the six-month trial period. Not all patients had a second angiogram, and in these cases clinical findings were used in the evaluation. Evaluation of the combined angiographic and clinical results showed improvement or no deterioration in 37% of patients in the placebo group compared with 49% in the low-dose and 61% in the high-dose ASA groups respectively. The only statistically significant difference observed was between the placebo group and the group treated with dipyridamole and high-dose ASA (p = 0.01). This difference remained statistically significant at p = 0.039 if only the angiographic findings were considered for group comparison. It cannot, however, be concluded from this study that 75 mg dipyridamole in combination with 100 mg ASA tid is more effective in preventing reocclusion after PTA than in combination with 330 mg ASA tid.
Fifteen women with positive islet cell antibodies were identified in a group of 115 consecutive patients found to have impaired glucose tolerance in pregnancy. These subjects were postulated to be at increased risk of later developing type 1 diabetes mellitus. They were examined post--partum for HLA types known to be associated with this disease and for any increase in Interleukin 2 receptor expression or alteration of T cell subsets of possible relevance to its pathogenesis. Fifteen women negative for islet antibodies and with normal glucose tolerance during previous pregnancy and 15 women with a normal fasting plasma glucose who had never been pregnant were studied as controls. Using flow cytometric techniques a significant increase in both the number and proportion of activated (Interleukin 2 receptor, CD25) lymphocytes in the peripheral blood of women who had islet cell antibodies and previous impaired glucose tolerance in pregnancy was found (0.14 +/- SE 0.03 x 10(9)/l; 7.1 +/- 1.1%) when compared with normal parous controls (0.09 +/- 0.01 x 10(9)/l; 4.2 +/- 0.6%), p less than 0.01 x 10(9)/l; showed significant increases when compared with nulliparous controls (0.04 +/- 0.01 x 10(9)/l; 2.1 +/- 0.2%), p less than 0.01. No differences were detected between the three groups with respect to total T-lymphocytes (CD3), helper T-lymphocytes (CD4), suppressor cytotoxic T-lymphocytes (CD8), or the inducer of suppressor (Leu 3+/Leu 8+) subset of T-lymphocytes. Three women persistently islet cell antibody positive, two of whom were HLA DR4, showed impaired glucose tolerance at the time of lymphocyte subset analysis, while two further patients, one DR3 and the other DR4, had developed type 1 (insulin-dependent) diabetes. No correlation between increased Interleukin 2 receptor expression and glucose intolerance was demonstrated. We conclude that islet cell antibody positive women with impaired glucose tolerance during pregnancy are at increased risk of later developing type 1 diabetes but that heightened immune activation present in these women is in part a post-pregnancy phenomenon.
Live donor renal transplants are often preferred to cadaver grafts because of better graft survival. In a retrospective study of 41 live donor transplants performed in the Belfast Renal Unit from 1971 until November 1988, actual graft survival at 2 and 5 years was 84% and 69% respectively. Corresponding patient survival rates were 87% and 81%. These results are no better than those of cadaver grafts. A subdivision of patients considered to be relatively poor risk for transplantation showed less favourable results than those who were good risk. 15% of the donors suffered post-operative complications, which occurred more often in older donors. Live donor transplantation is not necessarily preferable to cadaver organ graft, and is not recommended for poor risk recipients or donors aged over 50 years.
During a five month period in the summer of 1987, seven cases of reactive arthritis were seen at Musgrave Park Hospital, Belfast. Of the seven patients, five were male and two female. Their ages ranged from 13 to 44 (mean: 26 +/- 4) years. Five patients gave a preceding short history of diarrhoea which was usually mild. All patients presented with acute inflammatory oligoarthritis which predominantly affected the lower limbs. The knees were involved in 100%, the ankles in 70% with enthesopathy in 30% of cases. The diagnosis was confirmed in all cases by positive serology for Salmonella enteritidis. In three patients, stool cultures grew a particular strain of Salmonella enteritidis (phage type 4). Six patients were HLA-B27 positive. It is not possible to estimate the prevalence of reactive arthritis in this outbreak because there were different sources of infection and the total number of gastroenteritis cases was unknown. However, gut infections with Salmonella enteritidis are becoming increasingly common in this community and throughout the Western world. Over the past two years, in our area, there has been an almost thirty-fold increase in the number of recorded cases of infections due to this organism. By far the commonest source of infection appears to be poultry and egg products.
Extensive analysis of restriction fragment length polymorphism using HLA class II and T-cell receptor gene probes has been carried out in an attempt to identify genetic markers more strongly associated with multiple sclerosis than the classically defined antigens DR2, Dw2, and DQw1. The use of DNA pooled from groups of patients and controls from northeast Scotland enabled the screening of 14 restriction endonucleases with five HLA-D region probes (DP alpha, DP beta, DQ alpha, DQ beta, DR beta) and two T-cell antigen receptor probes. Restriction fragment length polymorphisms which discriminated between multiple sclerosis and control pools were identified with four restriction enzymes: Msp1 (DQ alpha), BamH1, Bgl11, and Taq1 (DQ beta). No discriminatory polymorphism was seen with any of the other enzyme/probe combinations. Subsequent Southern blot analysis of individual DNA samples was carried out using these enzymes and probes in two independently conducted studies, in Northern Ireland and northeast Scotland. Following Msp1-digestion and hybridization to DQ alpha, a 3.25-kb fragment was observed in 31% of Scottish patients but in only 4% of controls from the same population. Furthermore, when only DR2-positive individuals were analyzed, there was a significant excess of this fragment in patients from both Scotland (28, or 2.9%) and Northern Ireland (20, or 3.4%). Although the DQ alpha gene characterized by this fragment remains to be determined, this fragment exhibits apparent allelism to DQw1. Therefore, these data raise the possibility that two different DQ alleles, one on each haplotype, may jointly contribute to disease susceptibility.
93 patients received 102 renal transplants between 1968 and 1977. 99 grafts were from cadavers and 3 were from live donors; 93 were first grafts, 7 were second, and 2 were third. At 10 years total actual patient survival was 66.6%. 50 (55.5%) of 90 first cadaver grafts, and 52.2% of all cadaver grafts, survived at 10 years. Cardiovascular disease was the commonest cause of death, being responsible for 18 of 31 deaths (58%).
DNA techniques were applied to typing a population of renal patients in order to assess the number of discrepancies between this technique and the serological technique. Five patients had been given an incorrect type by serology, and in 22 instances allogenotypes were found where previously there had been a serological blank. DNA typing was also able to determine allogenotypic subtypes that correlated with DR antigens difficult to split by serology and to determine allogenotypic subtypes correlating with DR antigens not known to have a split by serology. Whereas DNA typing provided a result first time of testing on all but 3 patients, the average number of serological testings for each patient was 1.9.
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Nucleotide sequences for two immunoglobulin heavy chain variable region (VH) genes and one pseudogene in the goldfish (Carassius auratus) and the family relationships and distribution of these genes in individual fish are presented. Comparison of the nucleotide and inferred amino acid sequences of goldfish and other vertebrate VH genes indicates that goldfish VH genes show the major VH gene regulatory and structural features (5'-putative promoter region, split hydrophobic leader, three framework and two complementarity-determining regions, and 3'-recombination signals for VH to diversity region joining) and that goldfish VH genes are not more closely related to one another than they are to VH genes of evolutionarily distant vertebrates such as the mammals. Goldfish VH genes appear to exist in distinct families, and individual goldfish can carry from none to apparently greater than 15 genes of a given family. These results suggest that whereas the basic structure of VH genes has been conserved in evolution, there may be substantial variation in the nature and population distribution of VH gene families in the vertebrates.
A single enzyme/multiple probe system of HLA-DR and DQ typing using restriction fragment-length polymorphism (RFLP) analysis is presented. TaqI-digested genomic DNAs are hybridized sequentially with short DR beta, DQ beta, and DQ alpha cDNA probes. The DR beta probe discriminates between the DR allelic specificities DR1 to DRw14, with the two exceptions of some DR3/DRw13 and some DR7/DRw9 combinations. We describe the positive identification of a DRw10-specific RFLP and demonstrate its segregation in families. The DQ beta probe defines an allelic system that identifies the alleles DQw1, DQw2, and DQw3. This permits the resolution of DR3/DRw13 and DR7/DRw9 alleles by defining the DR/DQ association caused by linkage disequilibrium. The DQ alpha probe defines another allelic series interrelated with, but independent from, the DQ beta series. Specific DQ beta/DQ alpha RFLP combinations correlate with known Dw splits of DR2, DRw6, and DR7. Combined use of the three probes permits the identification of HLA-DR, DQ, and certain Dw specificities and provides an effective and easily interpretable system for major histocompatibility complex class II allogenotyping.
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