Search PubMed⌕ Search

Biomedical subjects

I Roitt

Publications and source records attributed to I Roitt.

17 recordsLinked to original sources
PubMed1996

Immunochemical mapping of gonadotropins.

As a glycoprotein hormone, human chorionic gonadotropic (hCG) is not a single molecular entity but this term rather comprises an array of molecular variants such as hCG, hCG beta, hCGn, hCG beta n, hCG beta cf, -CTPhCG, hCG beta CTP, deglyhCG, asialohCG, hCGav and the closely related molecules hLH, hLH beta and hLH beta ef. The advent of monoclonal antibodies (MCA), the availability of ultrasensitive detection systems and the recent determination of the crystal structure of hCG, made it possible to design special purpose diagnostic and clinical research immunoassays for hCG-like molecules. For more than a decade we and others have tried to refine epitope maps for hCG and related molecules by means of a large panel of MCA, naturally occurring metabolic variants of hCG (hCGn, hCG beta, hCG alpha, hCG beta cf, hCG beta CTP), homologous hormones and subunits of various species (e.g. hLH, hLH beta, hFSH, hTSH, oLH, rLH beta), chemically modified molecules (deglyhCG, asialohCG, tryptic and chymotryptic hCG beta and hCG alpha fragments) and synthetic peptides (octapeptides and longer). It appeared that all epitopes on molecular hCG-variants recognized by our MCA are determined by the protein backbone. Except for the two major epitopes on hCG beta CTP and parts of two antigenic domains on hCG alpha, epitopes on hCG-derived molecules are determined by the tertiary and quarternary structure. Operationally useful descriptive epitope maps were designed including information on assay suitability of antigenic determinants. On this basis we established ultrasensitive time-resolved fluoroimmuno-assays for hCG, hCG and hCGn, hCG beta and hCG beta n and hCG beta cf, hCG alpha and additional assays recognizing different spectra of hCG-variants. Such assay have been applied by us and others to the detection of pregnancy, early pregnancy loss, choriocarcinoma, testicular cancer, other cancers and prenatal diagnosis. However, as the molecular structure of many epitopes utilized in immunoassays of different laboratories was not resolved, comparability of results was not satisfactory. Consequently, attempts were made to compare schematic epitope maps from different research institutions. The situation has been much improved by solving the three-dimensional (3D) structure of hCG. It has been shown that hCG is a member of the structural superfamily of cystine knot growth factors like NGF, PDGF-B and TGF-beta. Each of its subunits is stabilized in its topology by three disulfide bonds forming a cystine knot. Moreover, it turned out that the disulfide bridges in their majority have previously been wrongly assigned. Computer molecular modeling of crystallographic coordinates of hCG and subsequent selective combined--PCR-based and immunological--mutational analyses of hCG beta expressed via the transmembrane region of a MHC molecule made it possible to more precisely localize epitopes on hCG-derived molecules. Although the entire surface of hCG has to be regarded as potentially immunogenic there seems to be hot spots where epitopes are clustered in antigenic domains. These are located on the first and third loops protuding from the cystine knots of both subunits and are possibly centered around the knot itself. Ultimate answers on epitope localizations will be given by the crystal structure determination of hCG complexed with different Fabs.

Amino Acid Sequence↗
PubMed1996

Agalactosyl IgG [Gal(o)]--an analysis of its clinical utility in the long-term follow-up of patients with rheumatoid arthritis.

This long-term prospective study of patients with newly diagnosed RA assesses the relative value of certain clinical and laboratory measures at first consultation in order to determine factors that help to discriminate between patients likely to go into early remission and those with relapsing/remitting or persistent disease. Validation was sought in a similar group from a separate but comparable prospective study. Measures of clinical activity such as joint score, early morning stiffness (EMS) and acute phase (ESR) improved over 4 yr in both groups, whereas agalactosyl IgG [Gal(o)] levels increased. Using discriminant functional analysis in the first cohort to identify features at entry which would predict outcome at 4 yr, a combination of Gal(o), grip strength, age at onset and gender predicted the course of RA correctly in 95% of the patients. This combination was confirmed in the second cohort, although with reduced power (78% correct). Thus, we show that Gal(o) does not reflect activity of the disease like the ESR, but has greater potential as a prognostic index early in the course of disease.

Arthritis, Rheumatoid↗
PubMed1994

Mapping of T cell epitopes of the major fraction of rye grass using peripheral blood mononuclear cells from atopics and non-atopics. II. Isoallergen clone 5A of Lolium perenne group I (Lol p I).

Rye grass is the major cause of hay fever which currently affects 20% of the population. Lolium perenne group I (Lol p I) is a glycoprotein of 240 amino acid residues, representing the main allergen of rye grass. We have used peripheral blood mononuclear cells (PBMC) from controls and subjects allergic to rye grass and cultured them with L. perenne extract (LPE) and Lol p I and measured lymphocyte activation using thymidine incorporation. Patients were further studied against the 115 overlapping peptides of the iso-allergen clone 5A of Lol p I to see whether the 4 amino acid residue differences between clone 1A and clone 5A affect the T cell epitope and thus, lymphocyte activation. There are 24 peptide differences between isoallergen clone 1A and clone 5A occurring in pools 4, 13, 16 and 19 each one of which could be an immunodominant epitope. The PBMC from all allergic patients studied showed a strong proliferative response to LPE and Lol p I. Five immunogenic peptide pools, pool 6, 15, 16, 17 and 19 of the isoallergen clone 5A were also identified. Most of these pools are in the C-terminal region of Lol p I. Out of 20 pools tested in vitro 1 pool (pool-17) induced PBMC proliferation in five out of six patients who were not restricted to an HLA class II DR gene product. However, three out of the six subjects responded to various other peptide pools in addition to the immunodominant pool. In spite of the amino acid differences between the two clones, pool 17 still remains the immunodominant T cell epitope. Control subjects showed only weak responses to LPE and no detectable response to either Lol p I or peptide pools. From within the most active pool we have defined two peptides of the isoallergen clone 5A (identical in sequence with clone 1A) which stimulate lymphocytes from rye grass-sensitive patients in vitro. Previous studies with the two continuous sequences (193WGAVWRIDTPDK204 and 195AVWRIDTPDKLT206) tested in vivo by intradermal skin testing have shown typical delayed-type hypersensitivity reactions after 24-48 h in one patient. Comparison of amino acid sequences of Lol p I, Lol p II and Lol p III proteins revealed a significant level of structural similarity among them. Interestingly, 50% of the residues of the second peptide sequence are also present in Lol p II and Lol p III.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗
PubMed1993

Autoantigens in thyroid diseases.

The autoantigens involved in autoimmune thyroid disease have now been extensively characterised, and the autoantibodies they evoke provide important aids to diagnosis, leading to early treatment of thyroid autoimmunity. The next stage in the puzzle is to determine towards which epitopes on the autoantigens the immune response is directed. We have already come a long way in the identification of immunodominant epitopes and have been able to identify one T cell epitope which has pathogenic capabilities. Identification of other T cell and B cell epitopes will help us understand the cell-mediated and humoral responses in greater detail and in time lead to more specific therapeutic intervention. A greater understanding of the mechanisms underlying one particular autoimmune disease will give us insights into other diseases, due to the belief that there may well be common underlying defects that, due to a multitude of factors, manifest as different diseases. The susceptibility factors in autoimmune thyroidits and autoimmune disease in general are very complex. A greater understanding is required of HLA associations and how particular peptides are presented in vivo. Are susceptible MHC types the ones capable of presenting the pathogenic peptides? Our major T cell thyroiditogenic epitope contains a T4 residue which accounts for over half the molecular weight of the peptide. Its structure is large and consists of a double benzene ring structure with four iodine atoms. It will be interesting to see how such a peptide can be presented and which residues bind T cell receptor or MHC. In summary we can say that autoimmune disease is due to a cocktail of factors which all contrive to tip the delicate balance of the immune system into an autoimmune state. HLA association may play a role in conferring an enhanced ability to select from a restricted repertoire of pathogenic epitopes, those epitopes perhaps only becoming available for presentation after interaction with environmental agents, whatever they may be. Following this, the normal regulation of self presentation and tolerance mechanisms break down and autoimmunity supervenes.

Amino Acid Sequence↗
PubMed1993

T cell epitopes of the major fraction of rye grass Lolium perenne (Lol p I) defined using overlapping peptides in vitro and in vivo. I. Isoallergen clone1A.

One hundred and fifteen overlapping synthetic peptides spanning the entire sequence of the iso-allergen clone1A of Lol p I from rye grass Lolium perenne were synthesized by the multi-pin technique. The peptides were overlapping 12mers, offset by two residues and overlapping by 10 residues. Sets of six adjacent overlapping peptides (except pool-1, 15, 20) were pooled and were used in vitro and in vivo to map the T cell epitopes on Lol p I. Six atopics who were skin test and RAST positive to rye grass showed T cell responses to L. perenne extract (LPE) and its major fraction (Lol p I). Five out of six showed T cell responses in vitro to peptide pool-17, while five non-atopics did not respond to any of the peptide pools. By testing the individual peptides of pool-17, we have located the T cell epitope on Lol p I. Interestingly, when we tested pool-17 and its single peptides in vivo by intradermal skin testing we found in one patient a typical DTH after 24-48 h to pool-17 and its peptides (peptides 3 and 4) which exactly matched the in vitro responses. By defining the T cell epitopes in this way a greater understanding of the allergic response to pollen will be obtained, and a more effective and less dangerous vaccine may be possible for treating patients with hay fever.

Adult↗
PubMed1991

Agalactosyl IgG: an aid to differential diagnosis in early synovitis.

Sixty consecutive patients presenting with early-onset synovitis were studied by measuring rheumatoid factor (RF) titers and the percentage of oligosaccharide chains attached to the C gamma 2 domain of IgG that lack galactose (GAL[0]). After 2 years of followup, 39 patients (65%) had developed rheumatoid arthritis (RA), and 21 had developed a variety of other inflammatory joint diseases. A combination of RF positivity and GAL(0) levels above the age-corrected mean gave a positive predictive value for a diagnosis of RA in 94% of these patients. These observations may well have clinical utility.

Adult↗
PubMed1988

Age-related galactosylation of the N-linked oligosaccharides of human serum IgG.

In a study of 151 normal, healthy individuals of both sexes varying in age from 1-70 yr, it was found that the relative incidence of agalactosyl (with both outer arms terminating in N-acetylglucosamine) N-linked oligosaccharides on total serum IgG decreased from birth to a minimum (at 25 yr of age) and then increased with age. The relative incidence of digalactosyl structures varied inversely to this, and the relative incidence of monogalactosyl structures was constant. Galactosylation of the N-linked oligosaccharides of the human serum IgG of normal individuals is therefore an age-related molecular parameter. Several reports have suggested that rheumatoid arthritis is associated with a decreased galactosylation of serum IgG (3-5). The normal variation in galactosylation with age as described here allows a true assessment of disease-associated changes in this parameter, and raises the possibility that one of the lesions in rheumatoid arthritis is an accelerated aging of the immune system. In addition, heterogeneity within age groups may be due to intrinsic differences in genetic endowment, or may reflect the impact of extrinsic factors (8).

Adolescent↗
PubMed1988

A prognostic index for erosive changes in the hands, feet, and cervical spines in early rheumatoid arthritis.

Clinical and laboratory measurements taken at the onset of rheumatoid arthritis in 149 patients were compared with the severity of radiological changes seen at 3 years in the hands and feet, and cervical spine. The strongest association with the severity of peripheral radiological damage was rheumatoid factor (p less than 0.0001 for both the latex titre and RAHA titre). Subluxation of the cervical spine was associated only with the presence of HLA-Dw2 (p less than 0.02) and HLA-B7 cross-reacting group (p less than 0.02). Discriminant function analysis utilizing latex titre, RAHA titre, haemoglobin level, and platelet count predicted the development of erosive or nonerosive disease in 79%. This method was less successful in predicting the actual severity of erosive damage and was not improved by the addition of HLA data. Radiological outcome in the cervical spine was successfully predicted in 82% using HLA-Dw2, HLA-B27 and age of onset of disease. It is concluded that the best predictors of erosive disease were standard laboratory features measured at onset, but that more powerful discriminant factors are needed if these are to influence clinical practice. Further prospective studies will establish whether rheumatoid involvement of the cervical spine is an expression of the influence of HLA determinants in this disease.

Adult↗
PubMed1987

High efficiency antigen presentation by thyroglobulin-primed murine splenic B cells.

B cells primed in vivo with mouse or rat thyroglobulin present these antigens at very low concentrations to CH9, an Ly 1+2- T cell hybridoma specific for mouse and rat thyroglobulin. Presentation measured by interleukin 2 release from CH9 is sensitive to treatment with a monoclonal antibody eliminating splenic B cells but is unaffected by anti-Thy-1.2 or 33D1 (which destroy T cells and dendritic cells, respectively). Presentation is specific for the priming antigen and is blocked by preincubation of the B cells with sheep anti-mouse F(ab')2. We suggest that in this system, primed B cells present thyroglobulin and that this may represent a means by which an initial triggering event priming both B and T cells could allow maintenance of autoreactive responses in vivo in the presence of low concentrations of circulating antigen.

Animals↗
PubMed1987

A prospective study of early onset rheumatoid arthritis over fifteen years: prognostic features and outcome.

A total of 218 patients with early onset rheumatoid arthritis were entered into a prospective study over fifteen years. Of these, 151 remained in the study at three years (of whom 14 died), and a further 39 (23 deaths) defaulted after this time. The severity of disease was assessed by four different methods. Although persistently active joint disease was recorded in 26% of patients, only 13% had severe functional impairment, and only 14% developed severe erosions. An episode of RA lasting two to three years followed by complete remission with none or minimal sequelae ("non recurrent" RA) was present in 34%. Using discriminant analysis, which selected rheumatoid factor, haemoglobin level and platelet count from a number of clinical and laboratory variables measured at onset, it was possible to predict three mutually exclusive outcome categories correctly in 50-60% of patients depending on the method of assessment of outcome.

Adolescent↗
PubMed1986

HLA antigen associations with radiological changes in the hands, feet, and cervical spines in early rheumatoid arthritis.

Clinical, laboratory, and genetic features measured at the onset of rheumatoid arthritis in 100 patients were compared with the severity of radiological changes in the hands and feet and in the cervical spines at a mean of 7.7 years. HLA-Dw4 was associated with more severe (p = 0.009) and HLA-Dw2 with less severe (p = 0.02)radiological changes in the hands and feet but the single strongest correlation with the severity of peripheral erosions was rheumatoid factor (p = less than 0.0001). Although none of the standard clinical or laboratory parameters correlated with severity of cervical spine changes, the presence of HLA-Dw2 and/or HLA B7 cross-reactive group were associated with more severe radiological changes in the cervical spine (p less than 0.02). Discriminant analysis selected certain standard laboratory parameters which in combination provided the most powerful prognostic index of radiological outcome in the hands and feet which was correct in 82 per cent. The addition of HLA data did not improve this figure. Conversely, the combination of the presence of HLA-Dw2, B27, and older age of onset of disease was found to be the most powerful predictor of the development of cervical spine changes and successfully predicted this complication of RA in 73 per cent.

Adult↗
PubMed1985

Molecular analysis of induced idiotypes associated with autoanti-thyroglobulin.

Idiotype (Id) and autoanti-thyroglobulin were induced in different strains of mice by priming with anti-Id to monoclonal anti-thyroglobulins (D8 and G4) and challenged with a subimmunogenic dose of thyroglobulin (Tg). Both D8.Id and G4.Id were induced in CBA mice by priming with the appropriate anti-Id, but only priming with anti-D8.Id also induced an increase in anti-Tg. D8.Id was induced in other strains by the same schedule but it only appeared to be associated with anti-Tg in 129 and, to a lesser extent, BALB/c mice, both of which have the allotype Iga. The extent of the overlap between the D8 Id and the anti-Tg was estimated and shown to be greatest in the CBA strain from which the D8 clone was originally derived. Spectrotypic analysis of the induced Ids in CBA mice showed that some of the D8.Id, but none of the G4.Id, was identical to the original clonotype, implying that CBA mice normally have cells which can be induced to produce D8.Id-positive autoanti-Tg, which are normally weakly expressed or regulated. The observation that anti-D8.Id priming in some strains increased D8.Id-negative anti-Tg responses suggests that the D8.Id may also be associated with anti-Tg T-cells.

Animals↗
PubMed1983

The occurrence of defined idiotypes on autoantibodies to mouse thyroglobulin.

Anti-idiotypic antibodies were raised in rabbits to three monoclonal mouse autoanti-thyroglobulins. Of the idiotypes recognized, one was unique to that monoclonal, while the other two showed evidence of sharing idiotopes with other monoclonal anti-thyroglobulin (TG) idiotypes, the IgM idiotype in particular being found on nearly half of the hybridomas studied. Two of the idiotypes could be detected in the sera of mice with experimental allergic thyroiditis (EAT), but they constituted only a very small percentage of the total anti-TG present. In spite of its low levels in EAT mice the D8 idiotype could be induced by injection of anti-idiotype and TG and in this case it constituted up to 20% of the anti-TG activity in individual mice, showing a strain-related association between the idiotype and autoanti-TG.

Animals↗