Search PubMed⌕ Search

Biomedical subjects

L J DeGroot

Publications and source records attributed to L J DeGroot.

At least 73 records · Page 4Linked to original sources
PubMed1995

Triiodothyroacetic acid has unique potential for therapy of resistance to thyroid hormone.

3,5,3,'-Triiodothyroacetic acid (Triac) has been used in therapy of resistance to thyroid hormone on an empirical basis and appears beneficial in some studies. We observed that the T3 analogs, Triac and 3,5,3'-triiodothyropropionic acid (Triprop), have a higher affinity for the thyroid hormone receptor-beta 1 (TR beta 1) than does T3 (2.7- and 1.8-fold, respectively), whereas the affinities of the three compounds for TR alpha 1 are the same. To evaluate whether T3 analogs would have a differential effect on TR beta 1 and TR beta 1 mutants and thus be a specific treatment for patients with resistance to thyroid hormone, we examined the induction of the transcriptional activation of wild-type (wt) TR alpha 1, TR beta 1, and mutant TR beta 1s by T3, Triac, and Triprop. The dose response of transcriptional activation by T3 analogs was measured by transient cotransfections with TRs and a rat malic enzyme-TRE fused to thymidine kinase (TK)-chloramphenicol acetyltransferase (CAT) in COS-1 cells. For TR alpha 1 wt, induction of CAT activity by T3 and Triac occurred at the same concentration. For TR beta 1 wt, Triac and Triprop showed a higher maximal activity than T3 (Tripro > Triac > T3) and reached 50% induction at a lower concentration than T3 (Tripro < Triac < T3). Induction of CAT activity in five mutant TR beta 1s (kindreds Mh, Mc, CL, Mf, and GH) was also analyzed. Even high levels of T3 analogs could not restore CAT activity to that of TR beta 1wt for any mutant. A dominant negative effect was produced by Mh, Mc, and Mf. Mutants CL and GH had a mild dominant negative effect depending on T3 analog concentrations and TREs. Cotransfection studies were performed using a rat malic enzyme-TK-CAT reporter plasmid to analyze the effects of hormones at near-physiological concentrations of T3 and Triac. Triac had a significantly higher transcriptional activation than T3 in Mc, CL, and GH, suggesting that Triac would have a beneficial effect to different degrees for different mutant TR beta 1s. Using mutants Mc and GH, further studies were carried out using rat GH and double palindromic and inverted palindromic TREs in COS-1 cells. On each TRE, 10 nmol/L Triac induced higher transcriptional activation in TR beta 1wt, mutant TR beta 1s, and TR beta 1wt plus mutant TR beta 1s (1:1 ratio) than the same dose of T3.(ABSTRACT TRUNCATED AT 400 WORDS)

Base Sequence↗
PubMed1995

Does thyroidectomy, radioactive iodine therapy, or antithyroid drug treatment alter reactivity of patients' T cells to epitopes of thyrotropin receptor in autoimmune thyroid diseases?

The effect of treatment on thyroid antibody production and T cell reactivity to thyroid antigens was studied in 15 patients with Graves' disease (GD) before and after thyroidectomy, 19 patients with GD before and after radioactive iodine (RAI) therapy, and 9 patients maintained euthyroid on antithyroid drugs (ATD). Twenty subjects matched for age and sex without known thyroid disease served as controls. In GD patients, the responses of peripheral blood mononuclear cells (PBMC) and TSH receptor (TSHR)-specific T cell lines to recombinant human TSHR extracellular domain, thyroglobulin, and TSHR peptides were examined on the day of surgery or RAI therapy (day 0) and also 6-8 weeks and 3-6 months thereafter. Reactivity to TSHR peptides before surgery was heterogeneous and spanned the entire extracellular domain. Six to 8 weeks after subtotal thyroidectomy, the number of patients' PBMC responding to any peptide and the average number of recognized peptides decreased. A further decrease in the T cell reactivity to TSHR peptides was observed 3-6 months after surgery. The responses of PBMC from Graves' patients before RAI therapy were less than those in the presurgical group. Six to 8 weeks after RAI therapy, the number of patients responding to any peptide and the average number of recognized peptides increased. Three to 6 months after RAI, T cell responses to TSHR peptides were less than those 6-8 weeks after RAI therapy, but still higher than the values on day 0. Responses of PBMC from patients with GD, maintained euthyroid on ATD, were lower than those before surgery or RAI therapy. The reactivity of T cell lines in different groups reflected a pattern similar to PBMC after treatment. TSHR antibody and microsomal antibody levels decreased after surgery, but increased after RAI therapy. The difference in the number of recognized peptides by patients' PBMC before RAI and surgery may reflect the effect of long term therapy with ATD in the patients before RAI vs. the shorter period in patients before surgery. The decreased T cell reactivity to thyroid antigens after thyroidectomy could be the result of removal of a major part of the thyroid gland or redistribution of suppressor-inducer T cells. The increased T cell response after RAI therapy is probably epitope specific, rather than a response to the whole TSHR molecule. Synchronous recognition of peptides 158-176 and 248-263 is important for the development of GD, and the loss of recognition of one of these epitopes may be an early sign of immune remission and a predictor of euthyroidism.

Adult↗
PubMed1995

The dominant negative effect of thyroid hormone receptor splicing variant alpha 2 does not require binding to a thyroid response element.

The functionally inactive thyroid hormone receptor splicing variant-alpha 2 (TRv alpha 2) can inhibit transcriptional activation by TR alpha 1 or beta 1, demonstrating a dominant negative effect (DNE). We examine here the three commonly proposed mechanisms, namely, competition for binding to thyroid hormone response elements (TREs), formation of inactive heterodimers, and squelching. A mutation introduced into the DNA-binding domain (DBD) of the TRv alpha 2 was designed to prevent its binding to TREs. In transient cotransfection studies, the DBD mutant has nearly the same DNE as does TRv alpha 2 on three different TRE-containing reporter genes. The DNE of TRv alpha 2 is also not reversed by cotransfection with excess retinoid X receptor-alpha. Extracts of COS cells cotransfected with TR alpha 1 and either TRv alpha 2 or DBD mutant at different ratios were analyzed by gel shift assays. Neither TRv alpha 2 or the mutant altered binding of TR alpha 1 to four radiolabeled TREs. TRv alpha 2 itself can inhibit constitutive transactivation by a thymidine kinase promoter-driven reporter construct. Our results suggest that TRv alpha 2 can function in a dominant negative manner without binding to a TRE, at least for certain TREs. It is concluded that the DNE of TRv alpha 2 may occur through another unrecognized mechanism, perhaps by binding to basal transcription factors.

Animals↗
PubMed1995

Importance of the most proximal GC box for activity of the promoter of human thyroid hormone receptor beta 1.

We wish to localize the sequences required for transcriptional expression of the thyroid hormone receptor beta 1 (TR beta 1). Constitutive activity of the promoter of human thyroid hormone receptor beta 1 was assessed by transient transfection of deletion constructs attached to luciferase as reporter, into P19, GH3, HepG2, H19-7, and COS1 cells. A 40-base pair fragment of the beta 1 promoter including the TATA box induced minimal luciferase activity, which was considered basal activity. The activities of various lengths of the beta 1 promoter were estimated relative to the minimal promoter in five cell lines. The region between -130 and -40 was crucial for constitutive activity in all cell lines. Further deletion analysis in HepG2 cells showed that two regions mainly augmented the transcriptional activity of the minimal 40 base pair fragment. One region located at -115 to -93, which is highly GC-rich, included the most proximal of five putative GC boxes present in the whole 1325-base pair promoter. A second region contributing to expression of TR beta 1 in HepG2 cells is at -70 to -40. Mutation of the most proximal GC box strongly suppressed transactivity of the whole promoter in P19 and HepG2 cells. In contrast, mutations in the other GC boxes did not suppress transactivation in P19 cells and slightly suppress activation in HepG2 cells. In Schneider cells, which do not express Sp1, transactivity of the region distal to -40 is positively regulated by cotransfection with a vector expressing Sp1.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Differential binding and activation of thyroid hormone response elements by TR alpha 1 and RXR alpha-trap heterodimers.

Thyroid hormone receptor (TR) forms homo- and heterodimers on various thyroid hormone response elements (TREs). We wished to clarify the relationship of homo- and heterodimer binding to TREs and their trans-activation. We investigated binding characteristics in gel mobility shift assays using synthetic direct repeat (DR) TREs having the consensus motifs separated by different oligonucleotide gaps, and we compared binding to trans-activation mediated via the direct repeat TRE. HTR alpha 1 purified from E. coli formed a monomer and homodimer on DR-TRE +0 to +5 but binding did not closely correlate with T3-dependent trans-activation. When RXR alpha expressed in COS 1 cell was added to purified TR alpha 1 in the gel shift assays, TR/RXR heterodimers were formed, and binding of heterodimers correlated highly with the level of trans-activation. These results strongly suggest that TR/TRAP heterodimers mediate the effect of thyroid hormone on DR-TREs. We also found T3-dependent disruption of homodimer formation on DR +0 to +2 and that T3 increased heterodimer formation on these TREs.

Base Sequence↗
PubMed1994

Proliferative responses of peripheral blood mononuclear cells from patients with Graves' disease to synthetic peptides epitopes of human thyrotropin receptor.

In Graves' disease thyrotropin receptor (TSH-R) autoantibodies cause hyperthyroidism. Production of TSH-R autoantibodies must be controlled by specific T cells. In this study we investigated T cell responses to 33 peptides corresponding to the sequence of the extracellular domain of human TSH-R. Peripheral blood mononuclear cells (PBMC) from 12 patients with Graves' disease and 9 healthy subjects were cultured with peptides for 3 days. The proliferative responses of PBMC were analyzed by measurement of [3H]thymidine incorporation. A stimulation index (SI; mean cpm in the presence of peptide/mean cpm in culture medium alone) of more than 3 was considered a positive response. When PBMC were stimulated with a pool containing all synthesized peptides, the mean SI of patients was significantly higher than that of controls (4.50 +/- 3.95 vs. 1.44 +/- 0.60; p < 0.05). When PBMC were cultured with individual peptides, PBMC from patients responded predominantly to two peptides, corresponding to sequence segments 152-157 (5 patients) and 207-222 (4 patients). No PBMC from controls responded to these two peptides. There was no clear correlation between the HLA-DR or HLA-DQ genotype and the stimulatory sequence segments. These results suggest that (a) TSH-R-specific T cells are present in peripheral blood of patients with Graves' disease, and (b) sequence segments 152-157 and 207-222 may be T cell epitopes of the human TSH-R in Graves' disease.

Amino Acid Sequence↗
PubMed1994

Diagnostic use of recombinant human thyrotropin in patients with thyroid carcinoma (phase I/II study).

Current diagnostic studies [radioiodine uptake and serum thyroglobulin (Tg) levels] for residual or metastatic thyroid tissue in patients with differentiated thyroid carcinoma require a hypothyroid status necessary for adequate endogenous TSH stimulation. However, almost all patients have symptoms of clinical hypothyroidism during this period. As shown in the present study, recombinant human TSH (rhTSH) allows stimulation of 131I uptake and Tg release from residual thyroid tissue in euthyroid patients. To assess safety, dosage, and preliminary efficacy, comparison was made of the stimulation of 131I uptake and Tg release after rhTSH administration and after T3 withdrawal in 19 patients after a recent thyroidectomy for differentiated thyroid carcinoma. Various doses (10-40 U) of rhTSH were injected im for 1-3 days in patients receiving suppressive doses of T3. Twenty-four hours after the last dose of rhTSH, 1-2 mCi 131I were administered, followed by a neck and whole body scan 48 h later. After discontinuing T3 for a median period of 19 days (range, 15-28), endogenous serum TSH levels were markedly elevated, and the patients were given a second dose of 131I and rescanned 48 h later. The injections of rhTSH were tolerated well. No major adverse effects were reported; nausea was reported in 3 (16%) and vomiting in 1 of the patients treated with high doses. The quality of life, as measured by two psychometric scales, was far better during rhTSH treatment than after T3 withdrawal. The peak levels of serum TSH (mean +/- SD) after a single dose of 10, 20, or 30 U were 127 +/- 19, 309 +/- 156, and 510 +/- 156 mU/L, respectively, and occurred 2-8 h after injection. Twenty-four hours after the injection, TSH levels decreased to 83 +/- 31, 173 +/- 73, and 463 +/- 148 mU/L in these treatment groups, respectively. The quality of the thyroid scans and the number of sites of abnormal 131I uptake were similar after rhTSH treatment and in the hypothyroid scans in 12 (63%) patients. Two additional sites of uptake in the chest and one in the thyroid bed, not visible on the hypothyroid scans, were identified in 3 (16%) patients after rhTSH. In 1 patient a focus of uptake was better visualized after rhTSH than after withdrawal. In 3 (16%) other patients, 1 lesion in the chest and 2 in the neck were seen only after T3 withdrawal.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗
PubMed1994

Strong association between HLA-DQA1*0501 and Graves' disease in a male Caucasian population.

Graves' disease (GD) is an autoimmune thyroid disease and is associated with human leukocyte antigen (HLA)-DR3 and -DQA1*0501 in caucasians. The incidence of GD is 5 times higher in females than in males, possibly due to their greater immune reactivity. Although many attempts have been made to correlate HLA phenotypes and clinical features of GD little attention has been paid to possible heterogeneous HLA distribution among patients of different sexes. To investigate this possibility, 133 (26 males) unrelated caucasian patients with GD and 104 (43 males) control subjects were typed for HLA-DRB1, -DQA1, and -DQB1, using sequence-specific oligonucleotide probes to analyze polymerase chain reaction-amplified DNA. There were no significant differences in HLA distribution between male and female controls. The frequencies of HLA-DQA1*0501 were increased in both [88.5%; relative risk (RR) = 9.13; P = 0.000015] and female patients (66.4%; RR = 2.66; P = 0.00046) compared to that in the entire control group (42.3%). The frequencies of DR11 (RR = 2.83; P = 0.019) and DQB1*0301 (RR = 2.50; P = 0.034) were increased only in male patients, whereas that of DR3 was higher in female (RR = 2.39; P = 0.0066) and male (RR = 2.54; not significant, P = 0.051) patients, suggesting the possible heterogeneous HLA distribution between the sexes. When the male and female patients were compared, a significant difference was found only for DQA1*0501. The prevalence of DQA1*0501 was significantly higher in males than in females (P = 0.019). As females have augmented immune responsiveness, we speculate that considerable numbers of females may develop GD without a strong HLA susceptibility allele, whereas only a few males develop GD without it.

DNA↗
PubMed1994

Serum thyroglobulin in the follow-up of patients with treated differentiated thyroid cancer.

To determine the significance of serum thyroglobulin (Tg) level in terms of presence or absence of thyroid cancer, we evaluated available serum Tg data on and off T4 therapy in 180 patients with differentiated thyroid cancer who have now been followed up to 18 yr. The presence of cancer was established by radioiodine scans, x-rays, and clinical examination. Thirty-two patients with detectable serum Tg autoantibodies were excluded from this analysis. Tg was measured by RIA with a sensitivity of 1 ng/mL. Patients who had all stages of cancer, but who had no evidence of active disease after treatment, were grouped according to operative and 131I ablative therapy. In patients with a partial thyroidectomy with or without ablation, the presence of Tg did not indicate the presence of cancer since levels were often above either a 5 ng/mL or a 10 ng/mL cutoff. The presence of residual normal thyroid tissue decreases the diagnostic value of serum Tg assay. In patients who underwent near total (NTT) or total thyroidectomy (TT) and 131I ablation, 3 of 55 (5.5%) patients had Tg greater than 5 ng/mL and 1 of 55 (1.8%) patients had Tg greater than 10 ng/mL during therapy, whereas off therapy 13 of 57 (22.8%) patients had Tg greater than 5 ng/mL and 6 of 57 (10.5%) patients had Tg levels greater than 10 ng/mL. In this group of patients, a Tg level less than 10 ng/mL during suppressive therapy indicated the absence of apparent tumor in 54 of 55 (98.2%) of patients. Whereas sensitivity of the assay was increased by withdrawal of hormone, "false positives" increased especially at lower (3-6 ng/mL) cut-off levels. No cut-off value properly categorized all patients. These data suggest, that even in patients who underwent 131I ablation and total thyroidectomy and were thought to be cured, small foci of thyroid tissue which are undetectable by standard 2 mCi 131I scans may exist and produce some Tg. However, these residual cells do not appear to cause an adverse prognosis in most patients. In patients with recurrent or continued disease, during T4 treatment, Tg levels ranged between 2-21,000 ng/mL and 5 of 11 patients had a Tg less than 5 ng/mL. Off treatment, Tg levels ranged between 6-10,700 ng/mL and 3 of 13 patients had a Tg less than 10 ng/mL. In 4 patients Tg levels were less than 10 ng/mL on treatment but greater than 10 ng/mL off therapy.(ABSTRACT TRUNCATED AT 400 WORDS)

Adolescent↗
PubMed1994

Two thyroid hormone response elements are present in the promoter of human thyroid hormone receptor beta 1.

Autoregulation of the human thyroid hormone receptor beta 1 (hTR beta 1) promoter was assessed by chloramphenicol acetyltransferase and luciferase reporter assays of transient transfections into COS1 and GH3 cells, DNase I footprinting, and gel shift assays. A 5'-deletional analysis of the promoter showed that the region between -906 and -839 and the sequence from -438 to -130 were positively regulated by T3 in COS1 cells cotransfected with an hTR beta 1 expression vector. We also transfected deletion constructs into GH3 cells and showed similar effects of T3 on the trans-activation of the reporters. DNase I footprinting showed a protected inverted palindromic thyroid response element (TRE) at position -890 to -866 in the distal fragment and a direct repeat at position -190 to -166 in the proximal fragment, which were protected by TRs. Mutation of each TRE significantly decreased the trans-activation of the promoter by T3. Gel mobility shift assays showed both proximal and distal TREs formed a retarded band with hTR alpha 1 or hTR beta 1 expressed in COS1 cells and reticulocyte lysates. The bands formed on the distal TRE and the proximal TRE appear to be preferentially formed by a TR homodimer and a heterodimer, respectively. Furthermore, the band formed on the distal TRE disappeared after adding T3 but that on the proximal TRE did not. These results indicate that hTR beta 1 expression is directly regulated by hTR alpha 1, beta 1, and their ligand through two TREs. The different structure of the TREs in this promoter suggests their physiological role in transcriptional regulation may be different.

Animals↗
PubMed1994

Evaluation of the ontogeny of thyroid hormone receptor isotypes in rat brain and liver using an immunohistochemical technique.

We performed an immunohistochemical study on rat brain and liver during fetal and neonatal life using rabbit antipeptide polyclonal antibodies able to recognize each thyroid hormone receptor (TR) isoform. The expression of TR alpha-1, alpha-2 and beta-1 proteins from 14 days of gestation to 21 days after birth was evaluated. Frozen tissues from 14 (F14), 17 (F17) and 21 (F21)-day-old fetuses and from 5 (N5), 16 (N16) and 21 (N21)-day old newborn rats were stained with anti-TR antibodies using an avidin-biotin-peroxidase system. The antipeptide antibodies utilized in the present study were characterized previously: alpha-144 antibody recognizes both TR alpha-1 and alpha-2; alpha-2-431 antibody is specific for TR variant alpha-2, and beta-62 antibody specifically reacts with the TR beta-1 isoform. The expression of TR alpha-1 was deduced by comparing the staining obtained with alpha-144 and alpha-2-431 antibodies. We demonstrated that each TR isoform is expressed in rat brain from 14 days of gestation and that the alpha isoform was predominant in the early stage. The three TR isoforms were expressed in both neural cell nuclei and in glial cell nuclei. As far as the liver is concerned, at F14 the expression of TR isoforms was weaker in hepatocytes when, on the contrary, TR alpha was clearly detected in hematopoietic cells. The expression of TRs in hepatocytes becomes evident later. The data that we obtained, although not quantitative, emphasize the presence of each TR isoform in brain and liver from 14 days of fetal rat life.

Animals↗
PubMed1994

Thyroid neoplasia.

Explore the source record for details and available documents.

Antineoplastic Agents↗