Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.11.1.8 (thyroid peroxidase)
3,116 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The immunoregulatory influence of transforming growth factor beta (TGF beta) was studied in patients with Graves' disease and in normal controls. Special attention was given to determine how TGF beta affects the interaction between thyroid epithelial cells and T lymphocytes. Human recombinant TGF beta 1 (rTGF beta 1) was immunosuppressive in patients with Graves' disease and in controls. In both groups it inhibited the proliferation of peripheral blood mononuclear cells and of peripheral and thyroid derived T cell lines and clones in response to non-specific stimuli. It also decreased the number of serine esterases expressing cytotoxic T cells and suppressed the recognition of thyroid epithelial cells by thyroid autoantigen specific T cell clones. Inhibition of autoantigen recognition was not only observed when rTGF beta 1 was added to the thyroid epithelial cell/lymphocyte co-culture, but was also found when thyroid epithelial cells were preincubated with rTGF beta 1, which was then removed before the initiation of co-culture. This was probably as a result of a decrease in the antigenicity of the target cells, as rTGF beta 1 also suppressed thyroid peroxidase as well as HLA class II autoantigen expression, in cultured thyroid epithelial cells. These results demonstrate that TGF beta may exert a variety of down-regulatory influences in Graves' disease. It may be of importance for the suppression of autoaggression in persons predisposed to autoimmunity; may be quantitatively overrun by immunostimulatory influences in the acute phase of the disease; and may be important for the induction of remission in patients with Graves' disease.
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PMID:The immunoregulatory influence of transforming growth factor beta in thyroid autoimmunity: TGF beta inhibits autoreactivity in Graves' disease. 177 15

To develop a model for endogenous thyroid autoantigen presentation, we transfected EBV-transformed B lymphoblastoid cell lines (EBV-LCL), established from patients with autoimmune thyroid disease and normal controls, with cDNA for the human thyroid autoantigen thyroid peroxidase (hTPO). hTPO-antigen presentation to patient peripheral blood T cells was demonstrated after stimulation in vitro for 7 d with irradiated hTPO-transfected or untransfected autologous EBV-LCL. Anti-hTPO-reactive T cells were subsequently cloned in the presence of irradiated, autologous hTPO-transfected EBV-LCL and IL-2.10 T cell-cloned lines exhibited specific hTPO-induced proliferation (stimulation indices of 2.1-7.9) towards autologous hTPO-transfected EBV-LCL, and were subjected to human T cell receptor (hTCR) V gene analysis, using the PCR for the detection of V alpha and V beta hTcR gene families. The results indicated a preferential use of hTCR V alpha 1 and/or V alpha 3 in 9 of the 10 lines. In contrast, hTCR V beta gene family use was more variable. These data demonstrate a model for the endogenous presentation of human thyroid peroxidase in the absence of other thyroid specific antigens. The high frequency of antigen-specific T cells obtained from PBMC using this technique will facilitate further studies at both the functional and hTCR V gene level.
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PMID:Endogenous antigen presentation by autoantigen-transfected Epstein-Barr virus-lymphoblastoid cells. I. Generation of human thyroid peroxidase-reactive T cells and their T cell receptor repertoire. 768 74

Autoimmune thyroid diseases (AITD) cluster in families, although the nature of this phenomenon is still poorly understood. One possible approach to the identification of genetic factors contributing to the pathogenesis of AITD is the study of association between polymorphic markers and AITD themselves. In the present study we have analyzed the allelic distribution of sRA-1, a TPO tetranucleotide repeat, among patients with AITD, in comparison with patients with nonautoimmune thyroid diseases and the general population. The polymorphic marker was analyzed by PCR followed by electrophoresis on polyacrylamide denaturant gel. Our data show that no association exists between AITD and any of sRA-1 alleles, despite the important role that TPO plays as a thyroid autoantigen.
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PMID:Thyroperoxidase microsatellite polymorphism in thyroid diseases. 880 96

Seven human T cell lines from a patient with Graves' disease were raised against endogenously generated human thyroid peroxidase (hTPO) with stimulation indices ranging from 2.1 to 7.6. Clonal expansion within these T cell lines was demonstrated by sequencing multiple bacterial colonies containing RT-PCR-generated fragments derived from the expressed hTcRs. Some lines had more than one human T cell receptor (hTcR) alpha and beta chain mRNAs as judged by RT-PCR. Stopcodons present in several hTcR sequences indicated that only one V alpha and one V beta gene were translated. Both the V alpha/beta gene families and the J alpha/beta gene segments differed amongst the lines and no characteristic recognition sequences were discernable in the CDR3 regions. Using Kyte-Doolittle analysis we found hydrophobic peaks in most N alpha-regions (but not N beta regions) suggesting that hydrophobic interactions may be important in the recognition of hTPO. However, increasing affinity values, as measured by SI, were strongly correlated with decreasing hydrophobicity in the N alpha region (1st order regression, r = -0.93138, p < 0.01). Thus, lower affinity, self-reactive, T cells may be more hydrophobic ('sticky') in their N alpha regions while higher affinity cells may be characterized by TcRs with lower hydrophobicity. These findings demonstrate a substantial role for hydrophobic interactions in hTPO-reactive T cell receptors and further support a role for the TcR alpha chain in the recognition of thyroid autoantigen.
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PMID:Endogenous antigen presentation by autoantigen-transfected Epstein-Barr virus-lymphoblastoid cells: T cell receptor N-region hydrophobicity relates to thyroid antigen recognition. 885 12

We have previously established that thyroperoxidase (TPO), a major thyroid antigen involved in autoimmune thyroid diseases, interacts with an idiotype present on human and mouse antibodies directed to thryoglobulin (TG), another thyroid autoantigen. In order to characterize the TPO-reactive idiotype, we selected a TG monoclonal antibody (mAb J7 B49.15) which bound to TPO and cross-reacted with human bispecific TG and TPO autoantibodies (TGPO aAb) for both TG and TPO binding. The TPO-reactive structure of the mAb J7 B49.15 was present on the F(ab')2, located next to the TG binding site and dependent on the association of the heavy and light chains. We found that mAb J7 B49.15 shared a TPO-reactive idiotypic structure with TG mAb from the same cluster of TG reactivity. All the mAb from this cluster were directed to an immunodominant region of TG, recognized by TG aAb. Natural anti-idiotypes from pooled normal human IgG used as a therapeutic intravenous preparation inhibited the TPO binding to mAb J7 B49.15. By homologous immunization in BALB/c mice, mAb J7 B49.15 induced an antiserum with both TG and TPO reactivities. Whereas TG reactivity decreased as early as day 14 post-immunization, TPO reactivity remained at a plateau value from day 21 to day 42. The TG and TPO reactive antisera were shown to inhibit the binding of mAb J7 B49.15 to TG and TPO, respectively. We concluded that mAb J7 B49.15 reacted with TPO through an interspecies idiotype which appeared conformational and related to the epitopic specificity of the mAb. Interestingly, this idiotype would carry the internal image of a TG structure able to induce, in homologous system, a TG antibody response followed by a TPO response without the need of any immunizing antigen.
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PMID:Idiotypic study of a bispecific thyroglobulin and thyroperoxidase monoclonal antibody. 893 81

Expression of a major thyroid autoantigen, thyroid peroxidase (TPO) was studied using the baculovirus-insect cell expression system. Human TPO cDNA modified so as to code for the extracellular fragment of the protein was placed under the control of the strong polyhedrin promoter in baculovirus transfer vector pBlueBacIII and cotransfected with linearized AcMNPV viral DNA. Expression in two insect cell lines Spodoptera frugiperda (Sf9) and Tricoplusia ni (High Five) was investigated and levels of recombinant TPO (rTPO) monitored by RIA and SDS-PAGE followed by Western blotting. Both insect cell lines expressed rTPO, but higher levels (30 mg/l culture medium) were obtained with High Five cells. Culture medium rTPO was purified and its glycosylation and immunoreactivity analysed. Lectin-affinity blotting and treatment with glycosidases indicated that both high mannose and complex-type sugar residues were associated with the recombinant protein. Studies with an ELISA based on biotinlabelled rTPO and an immunoprecipitation assay based on 125I-labelled rTPO indicated that the rTPO and native TPO showed similar reactivity to TPO autoantibodies (r = 0.96, P < 0.001, n = 50 and r = 0.99, P < 0.001, n = 80 respectively). In addition, rTPO expressed in High Five cells showed enzyme activity comparable with that of native TPO when the heme biosynthesis precursor delta-aminolevulinic acid was included in the culture medium. Overall, our studies indicate that the High Five insect cell line provides a useful system for the expression of relatively high levels of rTPO which should be suitable for structural analysis of TPO and TPO-TPO autoantibody complexes.
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PMID:High-level expression of recombinant immunoreactive thyroid peroxidase in the High Five insect cell line. 893 92

Structural studies on thyroid peroxidase (TPO), a major thyroid autoantigen, require milligram amounts of pure protein. We found that the human TPO ectodomain (amino acid residues 1-848) generated in insect cells did not remain in solution at high concentrations after affinity purification. In contrast, the TPO ectodomain secreted by mammalian (Chinese hamster ovary) cells, although generated to a lesser extent (1 vs. 8 mg/liter), remained in solution at high concentration (10 mg/ml) after purification to homogeneity. This purified material was well recognized by TPO autoantibodies, but lacked enzymatic activity. We attempted to restore activity by culturing the Chinese hamster ovary cells in the presence of added heme. TPO enzymatic activity was clearly detected in conditioned medium from cells cultured in hematin and hemin, but not in protoporphyrin IX (all at 1 mg/liter). Heme prosthetic group incorporation into affinity-purified TPO was highest for hematin and hemin, but unchanged for protoporphyrin IX (OD 410/280 nm ratios of 0.25, 0.23, and 0.14, respectively). Enzymatic activity was now evident with hemin (mean +/- SE, 27.2 +/- 2.6; n = 3; guaiacol units/mg protein), hematin (24.1 +/- 1.6), and, to a lesser extent, protoporphyrin IX (3.6 +/- 0.2). Culturing cells in 20 mg/liter hematin, the maximum concentration tolerated, increased enzymatic activity even further (45.6 +/- 0.6 guaiacol units/mg protein). All purified TPO preparations were homogeneous on PAGE and of similar size (105 kDa). Enzymatic deglycosylation showed a complex carbohydrate contribution of 13 kDa (unlike the 2.3 kDa in insect cell TPO). In conclusion, this is the first report on the purification to homogeneity of recombinant human TPO of mammalian cell origin. Unlike TPO generated in insect cells, mammalian TPO remains soluble at high concentration, possibly because of its greater carbohydrate content. This enzymatically active, recombinant human TPO may be useful for future structural studies.
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PMID:The greater glycan content of recombinant human thyroid peroxidase of mammalian than of insect cell origin facilitates purification to homogeneity of enzymatically protein remaining soluble at high concentration. 949 31

Stress has, for many years, been linked to the onset of autoimmune disease and, in particular, autoimmune thyroid disease (AITD). Whilst the exact mechanism of this association is unknown, it is clear that episodes of stress can induce profound changes in the immune system. More specifically, recent studies from several laboratories have shown an association between the expression of stress proteins and, particularly, the Hsp70 family with AITD. Our own studies describe a thyroid-specific Hsp70 which shares antigenicity with the key thyroid autoantigen, thyroid peroxidase. Further studies on the molecular basis for this observation are, however, hampered by the lack of a suitably validated thyroid cell model. In this paper we compare the response of primary cultures of human thyrocytes to hyperthermia with the response seen in the immortalized human thyroid cell line HTori3. Both cell types responded in a broadly similar manner, synthesizing proteins from two of the major stress protein families, Hsp70 and Hsp90. In the primary human thyrocyte cultures the 70 kDa proteins showed a 7.5-fold increase and the 90 kDa proteins a 2.7-fold increase with hyperthermia whilst in the HTori3 cells the increases in response to hyperthermia were 10- and 6.5-fold, respectively. We also show a dose-dependent stress response in HTori3 cells cultured in the presence of arsenite ions. We conclude that the response of this highly differentiated and stable thyroid cell line to stress is similar to that seen in primary cultures of human thyroid cells and that these immortalized cells will afford a convenient and effective model for the further study of the role of stress in the pathology of AITD.
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PMID:Stress protein expression in primary and immortalized cultures of human thyroid cells: a model system for the study of stress proteins in the pathogenesis of autoimmune thyroid disease. 967 43

The nature of the autoantibody repertoire to the dominant autoantigen in human autoimmune thyroid disease is controversial. There is evidence that autoantibodies to thyroid peroxidase (TPO) interact with overlapping conformational epitopes in an immunodominant region and binding to denatured (DN) protein is decreased. Contrary data demonstrate TPO autoantibody reactivity with DN-TPO or polypeptide fragments. However, none of the TPO-specific, human monoclonal autoantibodies isolated to date preferentially recognize denatured autoantigen. We therefore searched an immunoglobulin gene phage display library for human autoantibodies that bind TPO denatured by reduction and alkylation (DN-TPO). Thyroid-infiltrating B cells from a typical TPO autoantibody-positive patient were the source of mRNA for library construction. Surprisingly, the library enriched after panning on DN-TPO, as well as a panel of individual clones, preferentially bound native (N)-TPO. Of 13 clones selected using DN-TPO or N-TPO, 12 clones recognized the TPO immunodominant region. Moreover, regardless of selection with N-TPO or DN-TPO, their heavy and light chains were encoded by similar VDJ and Vkappa combinations. One clone (DN4), isolated using DN-TPO, did not interact with the TPO immunodominant region and its H chain derives from a different VH gene. Although DN4 binds specifically to TPO, its affinity is low, unlike the high affinities of other human TPO autoantibodies. In conclusion, human monoclonal autoantibodies that preferentially recognize denatured TPO could not be isolated from an immunoglobulin gene library despite selection with denatured protein. Our findings demonstrate the bias of the human B cell repertoire towards recognition of an immunodominant region on the conformationally intact form of a major thyroid autoantigen.
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PMID:Rarity of autoantibodies to a major autoantigen, thyroid peroxidase, that interact with denatured antigen or with epitopes outside the immunodominant region. 1040 11

We have demonstrated that Na+/I- symporter (NIS), a novel thyroid autoantigen, has local amino acid sequence homologies with the other thyroid autoantigens: Thyroglobulin (Tg), thyroid peroxidase (TPO) and thyrotropin receptor (TSH-R). These homologies concern the 4th, 5th, 6th extracellular loop and the beginning of the intracellular tail. We have expanded our studies and found that there are significant local homologies with other 11 proteins, most of them of bacterial or viral origin (e.g., Streptococcus or Herpes). These homologies concern the 2nd and 4th extracellular loop, and both the beginning and the end of the intracellular tail. These 11 homologies were retrieved by a computer-assisted search and extracted out of a database containing almost 300,000 amino acid sequences. These homologies were of magnitude greater than those concerning the three thyroid autoantigens [identities=51.1+/-7.3% vs 25.3+/-7.8% (mean+/-SD), p<0.001; similarities=70.6+/-10.7% vs 43.3+/-8.5%; p<0.001]. In addition, extensive, not local, homology was found with a number of unknown proteins from invertebrates (Drosophila melanogaster and Caenorhabditis elegans) and bacteria such as Bacillus subtilis and Xanthobacter. Previously, we had found that NIS has no extensive homology with Tg or TPO or TSH-R. This is the first demonstration of both extensive and local homologies between one thyroid autoantigen (NIS) and microbiological proteins. Taken together with data of the literature on the homologies between other thyroid antigens (Tg, TPO, TSH-R) and bacteria, the homologies we have now found reinforce the view that both bacterial and viral infections may trigger autoimmune thyroid diseases.
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PMID:Homologies of the thyroid sodium-iodide symporter with bacterial and viral proteins. 1047 51


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