Biomedical subjects
Publications and source records attributed to A So.
Characterization of T cells at sites of autoimmune damage has been difficult. Now, however, polymerase chain reaction (PCR)-based methods are being used to analyse T-cell receptor (TCR) gene expression in such lesions. Here, Christopher Marguerie, Claudio Lunardi and Alex So summarize and interpret the most recent findings and describe the current understanding of TCR usage in autoimmune diseases.
RATIONALE AND OBJECTIVES: The authors investigated whether iron oxide particles can be used as a magnetic resonance imaging (MRI) contrast agent to image abscesses in a two-stage experimental design. METHODS: Human buffy coat was incubated with iron oxide particles of different sizes and coatings. Smears of the incubation mixture were made on a glass slide and stained for iron. The percentage of iron oxide uptake was determined by counting 100 neutrophils and monocytes and scoring the number of cells that contain iron. Subcutaneous abscesses were created in the flanks of 18 Sprague-Dawley rats by injecting them with 0.1 mL of turpentine. Iron oxide was given intravenously, and the animals were imaged by MRI (1.5 T) 12 to 24 hours later. Different iron oxide coatings and doses were compared. RESULTS: The four different types of coating (constant fragment [Fc] of IgG, bovine serum albumin [BSA], lipid [Ferrosome], and dextran) had an uptake of 72% +/- 5.3%, 61% +/- 6.2%, 30.5% +/- 6.8%, and 5% +/- 2.5%, respectively. Comparison of two particle sizes (mean, 90 versus 35 nm) showed the large particles to have higher uptake (61% +/- 6.2%) compared with the small particles (6% +/- 1.8%) (P less than .001). Post-contrast imaging of the rats showed a hypointense ring around the abscess only in the animals injected with the lipid-coated agent. The effect was discernible within 12 hours after contrast injection and at a dose of 25 mumols iron/kg. Histologic sections showed phagocytic cells with iron granules in the periphery of the abscess. No hypointense ring on MRI or iron granules on histologic sections was seen around the abscess of the control animals or those injected with BSA-iron oxide or Fc-iron oxide. CONCLUSIONS: Lipid-coated iron oxide particles can be used to image abscesses by virtue of their phagocytosis into surrounding inflammatory cells. Positive uptake of these particles by human phagocytes in vitro suggests that similar results may be applicable in humans.
Stress proteins are frequently the target of humoral and cell-mediated immune responses to infection. These proteins belong to highly conserved gene families and there is substantial sequence homology between antigens produced by pathogenic organisms and the corresponding proteins from mammalian cells. Human T cells from sites of infectious and autoimmune lesions proliferate in response to stress proteins, and mapping of antigenic determinants on a mycobacterial stress protein shows that both species specific and highly conserved, 'self-like', regions of the molecule can take part in immune recognition. It is proposed that the lymphocyte population induced in response to stress proteins of pathogens during infection includes cells capable of autoimmune recognition of the corresponding self protein. Local accumulation of self stress proteins--in response to viral infection, for example--may subsequently provide a stimulus for proliferation of such autoreactive lymphocytes, thereby triggering a cycle of events which may contribute to the pathological damage associated with autoimmune disease.
An HLA-DX alpha gene polymorphism was analysed by Southern blotting in 49 British patients with Graves' disease and 61 control subjects. Two previously described allelic fragments of Taq I digested genomic DNA at 2.1 kb (U) and 1.9 kb (L) were found. The genotype frequencies for UU, UL and LL did not differ from the controls in Graves' disease, either for the whole groups or when subdivided into HLA-DR3-positive and -negative subjects. There was a significant association of the U allele with HLA-DR3 in both controls (P less than 0.05) and Graves' disease (P less than 0.025). The results indicate that DX alpha polymorphism is not primarily associated with Graves' disease. The findings differ from recent studies which showed that DX alpha polymorphisms may contribute to susceptibility in other DR3-associated autoimmune diseases.
The restriction fragment length polymorphism of the unrearranged T-cell antigen receptor (Tcr) alpha chain gene was investigated. Taq I digests, when probed with a Tcr alpha chain cDNA probe, revealed polymorphic bands of 7.0, 2.0, and 1.4 kb, due to variations around the C alpha gene, and the V gene cluster. Family studies confirmed the segregation of these polymorphic bands as allelic markers. These polymorphisms provide a new marker for the analysis of genetic variation of the Tcr alpha chain, and the influence of variation of the Tcr genes on the immune response.
Thirty-four lymphoblastoid cell lines that had been previously typed for HLA-DP antigens by primed lymphocyte typing (PLT) were tested by Southern blotting and by ELISA. Using two DP beta probes and a DP alpha probe with a series of enzymes, it is possible to identify restriction fragment length polymorphism (RFLP) patterns characteristic of DPw1, -2, -3, -4, and possibly -5. ELISA typing results, based on two polymorphic DP antibodies DP11.1 and ILR1, were compared with PLT-defined and RFLP-defined types. Thus, using a range of probes and enzymes it is possible to identify DP polymorphism. The value of monoclonal antibodies for such studies is demonstrated, and the molecular data can, in some cases, pinpoint the amino acids responsible for the specificity of the monoclonal antibodies.
Twelve patients with intractable rheumatoid arthritis were treated with antilymphocyte globulin (ALG), prednisolone and a cytotoxic agent, usually azathioprine, and were followed for 1 year. There was a significant (p less than 0.05) improvement in the mean score for early-morning stiffness, grip strength and the severity of nodules and vasculitis at 6 weeks and 3 months when compared to the initial visit. However, in most patients, this benefit was not sustained despite continued cytotoxic and steroid therapy. A rise in the haemoglobin and fall in ESR was maintained throughout the study period.
The restriction fragment length polymorphisms of the DR-beta, DQ-alpha and -beta genes in 11 DR4 homozygous cells of different Dw type were investigated. The results showed that, with the enzyme-probe combination studied, the DR beta restriction polymorphism patterns were constant between the different cell lines. With the DQ alpha and DQ beta probes, variations in restriction patterns were obtained. No absolute correlation was seen between RFLP and Dw types. These differences may be useful in the investigation of the association between RA and specific subsets of DR4.
Molecular analysis of the HLA-D region has uncovered a complex array of related genes encompassing a minimum of 6 alpha and 7 beta chain sequences. A high level of polymorphism is characteristic of the DQ alpha and beta genes, as well as DR beta. The DP genes, both alpha and beta, are also polymorphic, though to a lesser extent. The genes fit into the previously established loci: DP, DQ and DR, except for a newly-discovered sequence, DZ alpha, which is approximately equally related to all of the other alpha chain genes. Analysis of the polymorphism and evolution of the HLA-D region, by examination of the sequences, calls for several independent duplication events in the generation of this family of genes.
The suggestion that the intestinal mucosa may be abnormally permeable and a site of absorption of antigens in rheumatoid arthritis was tested by the use of a 51Cr-EDTA (edetic acid) absorption test. 24 patients with rheumatoid arthritis excreted significantly more 51Cr-EDTA than did 34 controls. Intestinal permeability was normal in untreated patients but almost invariably abnormal in patients treated with non-steroidal anti-inflammatory drugs. Studies in patients with osteoarthritis showed that the permeability abnormalities were due to an effect of NSAIDs on both the proximal and the distal intestine and that the effect was systemically mediated. Indium-111-labelled leucocyte scans showed ileocaecal inflammation in 6 of 9 patients on or recently on NSAIDs. Although increased intestinal permeability does not seem to be important in the pathogenesis of rheumatoid arthritis, the administration of NSAIDs may lead to loss of intestinal integrity, thus facilitating antigenic absorption and perhaps contributing to persistence of the disease.
Intravenous gammaglobulin was compared with the standard British intramuscular preparation in patients with hypogammaglobulinaemia and chronic bronchitis. Five patients were given six months' treatment with the weekly intramuscular preparation and six months' treatment with intravenous gammaglobulin given once every 18 days. During the trial they recorded symptoms of infection, absence from work, and sputum volume; lung function tests were performed during the intravenous treatment. The half life of the intravenous IgG and changes in serum IgG and C1q concentrations were also measured in seven other patients who received intravenous gammaglobulin every two weeks for 12 weeks. IgG concentrations, sputum volume, and infection scores were significantly better during intravenous treatment and there were no adverse effects from the intravenous gammaglobulin. These five patients were significantly more healthy when they received an intravenous gammaglobulin preparation, probably because the intravenous preparation increased serum IgG concentrations. Although longer studies are needed, intravenous gammaglobulin should be considered for patients with severe chest disease and those who cannot tolerate intramuscular injections.
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Recombinant interleukin-2 (rIL-2) therapy has been shown to be of value in the treatment of some cases of melanoma and renal cell carcinoma. However, its use can be limited by severe systemic toxicity. Targeting rIL-2 to the tumor should improve the antitumor immune response and decrease the systemic toxicity. With this aim, we have employed recombinant DNA techniques to construct a single-chain antibody interleukin-2 fusion protein (SCA-IL-2). The protein used in this model system consists of the variable domains of the antilysozyme antibody D1.3 fused to human IL-2 and is expressed in E. coli. It retains antigen-binding specificity and has the full biological activity of rIL-2. This approach can be taken to generate SCA-IL-2 proteins that bind to appropriate cellular antigens. In vivo administration of tumor-binding SCA-IL-2 should result in a localized high concentration of rIL-2 in the tumor tissues, maximizing the anti-tumor response while keeping systemic side effects to a minimum.