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Biomedical subjects

D G Williams

Publications and source records attributed to D G Williams.

At least 19 recordsLinked to original sources
PubMed1992

Anti-type II collagen ELISA. Increased disease specificity following removal of anionic contaminants from salt-fractionated type II collagen.

The purification of type II collagen, for the detection of anti-type II collagen antibodies by ELISA procedures, involves removal of proteoglycans by guanidine-HCl, followed by pepsin solubilisation and salt fractionation. However, type II collagen purified in this way may contain contaminants, despite the apparent purity on SDS-polyacrylamide gels. In this paper we demonstrate how additional purification by DEAE chromatography reduces the degree of background binding in the type II collagen ELISA, leading to an increase in disease specificity. The contaminants included proteoglycan and bound serum IgG from both rheumatoid arthritis (RA) patients and healthy controls in ELISA. Furthermore, positive correlations were observed in the sera (n = 24) between degree of reactivity to the contaminants and to (1) purified proteoglycan (r = 0.50, P = 0.01) and (2) pepsin (r = 0.65, P = 0.001). Thus, inadequate purification of type II collagen produces false positive reactions in the collagen ELISA and gives rise to a high background. A lack of specificity has been frequently associated with this assay.

Adolescent↗
PubMed1992

T and B cell responses following immunization with tetanus toxoid in IgA nephropathy.

The B and T cell responses were investigated in IgA nephropathy before and after immunization with tetanus toxoid (TT). Both IgA and IgG anti-tetanus toxoid antibodies were elicited, but the IgA antibodies were significantly greater in patients (92.6 +/- 11.7 ELISA units) than in the controls (49.2 +/- 7.5 ELISA units). This was associated with a significantly greater proportion of IgA+ B cells in patients than controls before immunization. However, a significant increase in the proportion of IgA1 binding CD4 and CD8 cells was also found. The proportion of CD3 cells with gamma delta T cell receptors (CD3+TCR gamma delta +), was significantly greater before immunization in the IgA nephropathy patients (37.0% +/- 2.4), compared with controls (10.0% +/- 2.3; P less than 0.001). Immunization with TT further enhanced the CD3+TCR gamma delta + cells in patients to 45.8% +/- 7.2 compared with controls (16.3% +/- 4.5), with a corresponding decrease in CD3+TCR alpha beta + cells in the patients (P less than 0.001). CD3+TCR gamma delta + cells are upregulated by common microbial antigens and clinical exacerbations of IgA nephropathy are frequently associated with mucosal infections and a rise in serum IgA concentration. The increased TCR gamma delta expression may be responsible for the enhanced IgA antibody response in IgA nephropathy. The increase in IgA antibodies may than exert a controlling effect by binding to augmenting T cells and thereby inhibiting their function.

Adult↗
PubMed1992

Successful transfer of collagen-induced arthritis to severe combined immunodeficient (SCID) mice.

We describe the adoptive transfer of erosive arthritis to an immunodeficient host. Spleen cells from arthritic DBA/1 mice (H-2q), immunized 4-6 weeks previously with bovine type II collagen in adjuvant, were transferred intraperitoneally into SCID mice (H-2d). SCID recipient mice also received native or denatured type II collagen (100 micrograms intraperitoneally) at the time of cell transfer. Arthritis developed in five out of five mice approximately 2 weeks after injection of cells plus native collagen, whereas animals injected with cells plus denatured collagen did not show any clinical or histological evidence of arthritis. The minimum graft size required for successful transfer of arthritis was established at 10(7) DBA/1 spleen cells. Histological examination of the joints of arthritic SCID recipient mice revealed synovitis, fibrosis and erosion of cartilage and underlying bone. Mean circulating levels of anti-type II collagen IgG were found to be significantly higher in mice injected with native collagen than those injected with denatured collagen (40 micrograms/ml and less than 1 microgram/ml, respectively). The ability to transfer collagen-induced arthritis adoptively should facilitate the study of the cellular requirement and pathological mechanisms involved in the induction of this arthropathy.

Animals↗
PubMed1992

Clinical associations of IgG antibodies to the ribonucleoprotein p67 polypeptide in patients with systemic lupus erythematosus.

The ribonucleoprotein (RNP) p67 antigen was purified from rabbit thymus and used in an enzyme linked immunosorbent assay (ELISA) with low interassay variability to detect IgG antibodies to p67 in patients with autoimmune connective tissue diseases. These antibodies were found in eight (80%) patients with a clinical diagnosis of mixed connective tissue disease (MCTD) but also in 27 (40%) patients with systemic lupus erythematosus (SLE). Sixty six per cent of the 12 patients with SLE with high levels of antibodies to p67 (> 50 U) had three or more features of MCTD, including myositis, fibrosing alveolitis, Raynaud's phenomenon, and sclerodactyly. Antibodies to the p67 RNP were not associated with the presence or absence of renal disease in the patients with SLE. This study suggests that antibodies against the p67 RNP are markers for clinical features of MCTD even in the context of SLE.

Autoantibodies↗
PubMed1992

Antigen-binding diversity of human hybridoma autoantibodies derived from splenocytes of patients with SLE.

The antigen-binding specificity of human hybridoma-derived monoclonal autoantibodies (mAb) was analysed with mAbs derived from the spleens of two patients with active systemic lupus erythematosus (SLE). From one patient 72 mAbs (RSP clones) and from the other 173 mAbs (RT clones) were obtained. The binding specificity of these mAbs was analysed by solid- and fluid-phase ELISA against the autoantigens ssDNA, dsDNA, cardiolipin, SmRNP, histones, Sm-D and SS-B (La) synthetic peptides, and foreign antigens including bacterial polysaccharides. In addition, antinuclear antibody activity and anti-dsDNA binding were confirmed by fluorescence staining methods. Reflecting the patient's serological profile, none of the antibodies from the RSP clones reacted with ssDNA or dsDNA but 12 reacted with cardiolipin. In addition, three mAbs reacted with H4, five with U1 RNP, two with Sm-D peptides and 12 with SS-B peptides. In contrast, from the RT fusion, nine mAbs reacted with ssDNA, HI and SS-B peptides, seven with cardiolipin, four with dsDNA, two with Sm-D peptides and one each with H2A, H3 and H4. In many cases one mAb showed reactivity with more than one antigen: for example, mAb RT 72 binds to ssDNA, dsDNA, cardiolipin, H1, H4 and an Sm-D peptide; RT 6 binds to H1, SmRNP and ubiquitinated histone H2A. However, none of the antibodies showed 'across the board' polyreactivity; indeed, the selectivity of the reactions was notable and marked variation in antibody affinity was recorded. Eight of the mAbs bound to Salmonella typhimurium and two to the Klebsiella polysaccharide K-30.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1991

Analysis of a positive feedback mechanism in the anti-Sm autoantibody response of MRL/MPJ-lpr/lpr mice.

The mechanism of induction of anti-Sm antibodies by passive transfer of anti-Sm mAb in MRL/lpr mice was investigated. No idiotypic relationship was detected between the inducing monoclonal antibody KSm2 and either the induced circulating anti-Sm antibodies or the products of anti-Sm-producing hybridomas derived from spleen cell fusion of treated mice. Treatment of mice with ribonucleoprotein Sm antigen, alone or as an immune complex, induced anti-Sm and anti-ribonucleoprotein antibodies similarly to treatment with KSm2. This suggests that autoantigen contributes to the development of the anti-Sm response in MRL mice.

Animals↗
PubMed1991

Cross reaction of antibodies to a glycine/alanine repeat sequence of Epstein-Barr virus nuclear antigen-1 with collagen, cytokeratin, and actin.

P62 is a synthetic peptide which corresponds to the glycine/alanine repeat sequence of Epstein-Barr virus nuclear antigen-1. It is the main epitope recognised by anti-rheumatoid arthritis nuclear antigen antibodies. It was shown previously that anti-P62 antibodies were raised fourfold in patients with rheumatoid arthritis compared with controls. To examine the possibility that this increase was due to cross reactive autoantibodies binding to P62, anti-P62 antibodies from serum samples taken from 10 patients with rheumatoid arthritis and five healthy controls were purified by affinity chromatography. Immunoglobulin G anti-P62 antibodies purified from four of 10 serum samples from patients with rheumatoid arthritis also reacted with human epidermal keratin, denatured collagen type II and actin, but not with influenza antigens, as determined by enzyme linked immunosorbent assay (ELISA). Anti-P62 antibodies in serum samples from healthy controls and patients with rheumatoid arthritis reacted with epidermal keratin by immunoblotting. It is suggested that antibodies to the glycine/alanine repeat sequence of Epstein-Barr nuclear antigen-1 recognise homologous epitopes on keratin, actin, and collagen. It is also possible that molecular mimicry between a major epitope on the Epstein-Barr virus and several autoantigens might contribute to the breakdown of tolerance and autoimmunity in patients with rheumatoid arthritis.

Actins↗
PubMed1990

Recurrence of focal segmental glomerulosclerosis in transplanted kidneys: analysis of incidence and risk factors in 59 allografts.

Fifty-nine allografts were placed in 43 patients with renal failure from focal segmental glomerulosclerosis (FSGS): 27 allografts were put into 16 children aged less than 15 years, and 32 allografts into 27 adolescents and adults. Recurrence of FSGS was noted histologically in 13 allografts, 10 in 8 children and 3 in adults. None of the 9 children and 24 adults who never developed an allograft nephrotic syndrome showed FSGS in their allograft biopsies. The age of onset was a strong risk factor for recurrence: recurrent FSGS developed in 8 of 16 children (50%) but only in 11% of adolescents and adults (3 of 27 patients). Although the time from apparent onset to renal replacement treatment was shorter in those with recurrence than those without in the children, there was no difference in the time spent on dialysis prior to transplantation. Mesangial prominence was observed in the original biopsy in 12 of 13 patients with recurrence, and recurrence rate was similar in living and cadaver donor allografts; class I MHC matching was similar in those with and without recurrence. Three allografts treated with cyclosporin A as well as 9 with azathioprine showed recurrence. Of 9 second or subsequent allografts placed in those with recurrence in the first allograft, only 3 showed further recurrence. In 3 re-grafted after 13, 11 and 5 years, normal function was seen.

Adolescent↗
PubMed1990

Antinuclear antibody determination methods.

Screening rheumatology patients for anti-nuclear and anti-cytoplasmic antibodies is easily done in a qualitative manner using the IF, CIE and ID assays. The immunoblot is of use for anti-La and anti-RNP assays but gives anomalous results for Sm binding by anti-RNP sera and is not easily quantitated. These deficiencies of the immunoblot do not apply to the ELISA. Advances in cloning of autoantigens will enable standardisation of antigen preparations used for these ELISAs. The quantitation of autoantibody appears significant since disease flares occur together with elevations in specific autoantibody. IgM anti-Sm autoantibody was detected with a different disease distribution to IgG anti-Sm but the prognostic implications for this remain to be determined.

Antibodies, Antinuclear↗
PubMed1990

A repeated proline-rich sequence in Sm B/B' and N is a dominant epitope recognized by human and murine autoantibodies.

Reactivity of a murine monoclonal anti-Sm B antibody was localized to a cDNA clone T3/2 encompassing codons 177-221 of Sm N and to a series of 10-residue synthetic peptides containing a proline-rich motif. Human sera containing anti-B antibodies also bound to this proline-rich motif in the form of a heptamer, PPGMRPP, and could distinguish the N/B sequence from similar sequences present in the RNP A and C polypeptides in a number of cases. Monoclonal antibodies binding to either this peptide or to a determinant present on both B and D polypeptides inhibited the majority of anti-B binding in 50% of anti-B sera. The region of N/B containing PPGMRPP also bears a second epitope, sterically blocked by monoclonal antibody KSm 5. This hotspot binds 40-60% of anti-B activity in 30% of anti-B sera. These data suggest that the anti-Sm B epitope recognition repertoire is restricted in SLE patients.

Amino Acid Sequence↗
PubMed1990

Genes encoding the beta-chains of HLA-DR7 and HLA-DQw2 define major susceptibility determinants for idiopathic nephrotic syndrome.

1. We have investigated the frequencies of the major histocompatibility complex class II alleles by restriction fragment length polymorphism analysis, using DR, DQ and DP complementary DNA probes, in 40 Caucasoid steroid-sensitive nephrotic children. 2. A significant association with HLA-DR7 was demonstrated (P = 2 x 10(-5); aetiological fraction 0.6). The DQB1 gene of HLA-DQw2 was present in 83% of our patients (P = 2 x 10(-4); aetiological fraction 0.7). We present evidence that it contributes a second susceptibility allele. 3. A weak association between the uncommon HLA-DP-Cp63 allele and the disease was also observed. 4. Our data suggest that the immune events in steroid-sensitive nephrotic syndrome are defined by a particular immunogenetic background involving the beta-chain genes of HLA-DR7 and HLA-DQw2.

Adolescent↗
PubMed1990

Regulated expression of the small nuclear ribonucleoprotein particle protein SmN in embryonic stem cell differentiation.

The SmN protein is a component of small nuclear ribonucleoprotein particles and is closely related to the ubiquitous SmB and B' splicing proteins. It is expressed in a limited range of tissues and cell types, including several undifferentiated embryonal carcinoma cell lines and undifferentiated embryonic stem cells. The protein declines to undetectable levels when embryonal carcinoma or embryonic stem cells are induced to differentiate, producing primitive endoderm or parietal endoderm or yielding embryonal bodies. This decline is due to a corresponding decrease in the level of the SmN mRNA. The potential role of SmN in the regulation of alternative splicing in embryonic cell lines and early embryos is discussed.

Actins↗
PubMed1990

Review of the teaching of medical ethics in London medical schools.

The study examined the influence of the Pond Report on the teaching of medical ethics in the London medical schools. A questionnaire was given to both medical students and college officers. All medical colleges reported that ethics was included in the curriculum. However, from students' replies, it seems that attendance of optional courses is low and that not all current final year medical students have had any formal teaching in medical ethics. Stronger guidelines are necessary to ensure appropriate ethical training in London medical schools.

Curriculum↗
PubMed1990

Recombinant autoantigens.

Highly specific serum autoantibodies directed at cellular macromolecules are detectable in the serum of many patients with autoimmune connective tissue diseases. This phenomenon has been useful in diagnosis and in identifying disease subsets. The mechanism whereby the immune system selects these specific targets, and their relation to the events of disease pathogenesis, remains unclear. The use of monoclonal autoantibodies has increased our knowledge of the molecular associations and components of the often complex heteropolymeric antigenic particles. The advent of antigen cDNA cloning now heralds the next stage in resolution of this understanding with the complete primary structure of the component proteins.

Autoantigens↗