Physiological Thyroid Hormone Levels Research Articles

SUN-LB054 Cilengitide Reverts Thyroid Hormones Effects in Bexarotene Treatment of Breast Cancer Cells

Bexarotene (Bex) is an oral retinoid-X-receptor agonist that is effective for the treatment of early and advanced-stage in cutaneous T cell lymphoma. There are ongoing clinical trials to determine its role in both breast cancer treatment and prevention. Unfortunately, hypothyroidism is recognized as an important side effect of such therapy, potentially manageable by thyroid hormones (THs) co-administration. We previously found (1) that physiological levels of THs increased cell proliferation of different subtypes of human T cell lymphoma by activating both the nuclear (TR) and membrane (integrin αVβ3) THs receptors. We first demonstrated that MDA-MB-231 triple negative breast cancer cells expressed both TR and integrin αvβ3 dimer. Also, we found that THs induced MDA-MB-231 cell proliferation (p<0.01; n=4). Interestingly, in THs-depleted culture conditions we observed a higher effect of Bex in decreasing cell viability (p<0.05; n=3) and THs impaired the action of this drug (p<0.05; n=3). Then, we performed the same experiments with THs and Bex in presence of the inhibitor of the non-canonical THs membrane receptor integrin αVβ3, cilengitide. We found that pharmacological inhibition with cilengitide improved the TH effect on Bex activity (p<0.01; n=3) in MDA-MB-231 cells, indicating that integrin αVβ3 plays a key role in THs-induced MDA-MB-231 cell proliferation. Our data suggest that the requirement of THs replacement therapy may counteract Bex antitumor effect but concomitant administration with cilengitide could be an effective strategy to improve Bex treatment in breast cancer. Reference: (1) Cayrol et al, Blood 2015 Jan 29;125(5):841-851. Unless otherwise noted, all abstracts presented at ENDO are embargoed until the date and time of presentation. For oral presentations, the abstracts are embargoed until the session begins. s presented at a news conference are embargoed until the date and time of the news conference. The Endocrine Society reserves the right to lift the embargo on specific abstracts that are selected for promotion prior to or during ENDO.

Inhibition of the Integrin αVβ3 Improves the Immune and Anti-Lymphoma Effects of Bexarotene in Cutaneous T-Cell Lymphoma (CTCL)

Bexarotene (Bex) is an oral RXR agonist that is effective for the treatment of early and advanced-stage CTCL. However, Bex is associated with the unavoidable side effect of hypothyroidism in about 95% of pts, which is prophylactically managed with the administration of thyroid hormone (TH). Paradoxically, we found that physiological levels of TH increase the proliferation of CTCL by activating both the nuclear TRA receptor and the membrane integrin αVβ3 in these cells. Here, we determined the influence of TH replacement therapy on the anti-lymphoma activity of Bex, an unknown topic with clinical implications.Results: In standard culture conditions, Bex decreases the proliferation and viability of CTCL cell lines HuT78 and MJ. We conducted RNA-sequencing in HuT78 cells treated with Bex (vs. vehicle) to understand the mechanism/s underpinning these effects. We found that Bex regulates pathways related to “cell proliferation and differentiation” (e.g. REL, SCD, CCND1) and “immune system” (e.g. TBX21, IFNG, MX1) , which we independently validated. This suggests that Bex treatment could also impact anti-neoplastic immunity by increasing INF-gamma release from CTCL cells. We then conducted the same experiment culturing HuT78 and MJ in TH-depleted culture conditions and observed a higher effect of Bex in decreasing proliferation and viability; supporting the notion that Bex should not be administered with TH replacement. However, hypothyroidism is associated with marked immunosuppression that could favour CTCL progression and potentially affect the immune modulator effect of Bex that we described above.We thus determined the impact of TH replacement on the anti-lymphoma effect of Bex using a murine CTCL model. We implanted murine EL4 TCL cells in the hypodermis of immunocompetent C57BL/6 mice. Once tumors developed, mice were randomized into treatment with vehicle (Veh), Bex with no TH replacement (Bex) and Bex with TH replacement (BexT4+). TH replacement was done with oral administration of T4. We measured T4 in these mice plasma by RIA to assure that mice with T4 replacement reach physiological levels and those with no T4 develop hypothyroidism. Compared to Veh mice, the administration of Bex significantly decreased lymphoma growth in both conditions although slightly better in mice without T4 replacement (p<0.001 and p<0.01) for Bex and BexT4+, respectively). However, mice with Bex alone that developed hypothyroidism showed a significant decrease of CD8+CD44+ T-cells (p<0.05 vs. BexT4+) and increase of myeloid-derived suppressor cells in the tumor microenvironment and draining lymph nodes. Since infiltration of CD8+ cells decreases with CTCL progression, our data indicates that Bex treatment should be administered with T4 replacement to avoid a negative immune microenvironment.Considering the cellular effects of TH are exerted through TRA and integrin αVβ3 that can be independently modulated pharmacologically, we investigated whether integrin αVβ3 inhibition would be sufficient to blunt the TH-induced decrease on the antineoplastic efficacy of Bex. The lack of expression of integrin αVβ3 in normal T-cells offers a rationale for a selective effect on CTCL cells while sparing immune cells. In HuT78 and MJ cell lines, either αVβ3 silencing by siRNAs or pharmacologic inhibition with the clinical drug cilengitide avoided the decrease in the anti-CTCL effect of Bex upon TH supplementation. Moreover, gene expression analysis (RNA-seq and PCR) demonstrated that αVβ3 silencing induced higher (vs. Bex) up-regulation of “immune ” genes like TBX21 and INFG, or down-regulation of “proliferation” genes like REL and CCND1 .We thus tested if this mechanism can be therapeutically capitalized to improve Bex treatment in our CTCL murine model. We implanted 24 C57BL/6 mice with murine EL4 TCL cells and randomized them to vehicle (Veh), Bex with TH replacement in combination with cilengitide 40mg/kg (BexT4+Cil) or without cilengitide (BexT4+). At end of treatment, BexT4+Cil mice showed significantly smaller tumors compared with Veh and BexT4+ mice (p<0.001 and p<0.05, respectively). Importantly, the anti-tumoral immune response of CD8+CD44+ T-cells in the CTCL microenvironment and draining lymph nodes remained similar in BexT4+Cil vs. BexT4+ mice. Our data indicates that inhibition of the integrin αVβ3 is an effective strategy to improve Bex-based treatments in CTCL. DisclosuresCerchietti:Celgene: Research Funding; Lymphoma Research Foundation: Research Funding; Leukemia and Lymphoma Society: Research Funding; Weill Cornell Medicine - New York Presbyterian Hospital: Employment.

Abstract 3132: Inhibition of the integrin αVβ3 improves the effect of bexarotene in the treatment of cutaneous T-cell lymphoma (CTCL)

Abstract CTCL are exposed to a complex paracrine and endocrine environment that influence their progression from skin to visceral disease. One of the most common treatments for CTCL is RXR agonists or “rexinoids” such as bexarotene (BEX). The most prevalent side effect of rexinoids (~95% of patients) is the development of hypothyroidism hence these patients are prophylactically treated with thyroid hormone (TH). Paradoxically, we recently found (PMID: 25488971) that TH can activate transcriptional programs required for proliferation of CTCL. We also found that the effects of TH on CTCL are mediated through the activation of two different receptors, the classical nuclear TR and a membrane receptor integrin αVβ3. Our aim was to study how the anti-lymphoma activity of BEX is modified by the activation of TR and αVβ3 during the TH replacement therapy, and unknown topic with clinical implications. We first evaluate the cell viability of CTCL cells HuT78 and MJ treated with BEX in presence and absence of physiological levels of TH. As expected, BEX decreased the viability of CTCL cells, however in presence of TH both effects decreased by 25-60% and 20-50% on HuT78 and MJ cells. However, hypothyroidism is associated with a higher dissemination of TCL cells in syngeneic mice. Thus, to evaluate the impact of TH replacement therapy in BEX-treated mice, we implanted TCL EL-4 cells subcutaneously in C57BL/6 mice. When tumors reached ~75 mm3, mice were randomized in three groups: vehicle, bexarotene alone (BEX) and bexarotene with T4 replacement (to reach euthyroidism) (BEX-T4). Tumor growth significantly decreased in both bexarotene arms (vs. vehicle). However, in contrast to mice receiving BEX-T4, mice receiving BEX alone showed a decreased infiltration of tumor-suppressive immune cells. Specifically, these tumors had decreased NK and CD3+CD8+ cells due to BEX-induced hypothyroidism. This indicates that although TH could decrease the effect of BEX in CTCL cells, the hypothyroidism would impact in the local antitumor response. Thus determine if the inhibition of the non-canonical TH membrane receptor integrin αVβ3 would decrease the pro-survival effect of TH on CTCL treated with BEX. We found that either, αVβ3 silencing or the pharmacological inhibitor Cilengitide, not only avoided the pro-survival effect of TH but increased the anti-lymphoma activity of BEX-T4 by 15-35% in CTCL cells. To evaluate the molecular program underpinning these effects, we conducted RNA-sequencing in HuT78 cells treated with BEX-T4 in presence of siRNA for TR, αVβ3 or controls. Among the top pathways we identified genes involved in “apoptosis”, “angiogenesis” and “IL6-JAK-STAT3 signaling” as differentially expressed in si-αVβ3 vs. the others. These genes were independently validated by using Cilengitide. Our data suggest that inhibition of the non-canonical TH receptor, the integrin αVβ3, could be an effective strategy to improve BEX treatment in CTCL Citation Format: Florencia Cayrol, Victoria Revueta, Alejandra Paulazo, Mercedes Debernardi, Maria Celeste Diaz Flaque, Helena Sterle, Leandro Cerchietti, Graciela Cremaschi. Inhibition of the integrin αVβ3 improves the effect of bexarotene in the treatment of cutaneous T-cell lymphoma (CTCL) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3132. doi:10.1158/1538-7445.AM2017-3132

Endometrial Polyps in Women Affected by Levothyroxine-Treated Hypothyroidism—Histological Features, Immunohistochemical Findings, and Possible Explanation of Etiopathogenic Mechanism: A Pilot Study

The aim of the study was to investigate the possible overexpression of estrogen (ERs) and progesterone (PRs) receptors both in EPs glandular and stromal cells in postmenopausal women with levothyroxine-treated hypothyroidism in comparison to EPs detected in women with physiological thyroid hormone levels. During the study period (January-February 2013) 22 patients were eligible (12 treated, 10 controls). The two groups were homogenous for general, EPs sonographic and hysteroscopic features. None of the cases of atypia was found. Immunohistochemistry showed that the two groups were similar for ERs and PRs intensity rates in EPs glandular cells despite a trend of ERs percentage expression more than 60% in 2/3 of treated patients versus 1/3 of controls. In stromal EPs components, ERs intensity was high positive in 10 (83,3%) treated cases while it was high positive in 1 control (10%). Percentage of ERs stromal expression showed a different trend between the two groups despite a borderline statistical significance. Our hypothesis is based on a possible double action of hypothyroidism and thyroxine intake: the subclinical TSH increased levels and its possible circadian oscillation could stimulate the endometrial TSHRs (increasing type 2 DIO activity); the circulating levels of exogenous thyroxine could be locally metabolized in active form by type 2 DIO stimulating ERs.

Physiological Thyroid Hormone Levels Regulate Numerous Skeletal Muscle Transcripts

ABSTRACT Context Skeletal muscle is an important target tissue for thyroid hormone (TH). It is currently unknown which genes are regulated by physiological TH levels. Objective We examined the effects of l-thyroxine on human skeletal muscle transcriptome. Design Microarray analysis of transcript levels was performed using skeletal muscle biopsies from patients under euthyroid and hypothyroid conditions. Setting The study was conducted in a university hospital laboratory. Patients We studied skeletal muscle obtained from 10 thyroidectomized patients with differentiated thyroid carcinoma on and after 4 wk off l-thyroxine replacement. Mean Outcome Measures Gene expression changes were measured using microarrays. Results were analyzed using dedicated statistical methods. Results We detected 607 differentially expressed genes on l-thyroxine treatment, of which approximately 60% were positively and approximately 40% were negatively regulated. Representative genes were validated by quantitative PCR. Genes involved in energy and fuel metabolism were overrepresented among the up-regulated genes, of which a large number were newly associated with thyroid state. l-thyroxine therapy induced a large down-regulation of the primary transcripts of the noncoding microRNA pair miR-206/miR-133b. Conclusion We demonstrated that physiological levels of TH regulate a myriad of genes in human skeletal muscle. The identification of novel putatively TH-responsive genes may provide the molecular basis of clinical effects in subjects with different TH status. The observation that TH regulates microRNAs reveals a new layer of complexity by which TH influences cellular processes.

Physiological Thyroid Hormone Levels Regulate Numerous Skeletal Muscle Transcripts

Skeletal muscle is an important target tissue for thyroid hormone (TH). It is currently unknown which genes are regulated by physiological TH levels. We examined the effects of l-thyroxine on human skeletal muscle transcriptome. Microarray analysis of transcript levels was performed using skeletal muscle biopsies from patients under euthyroid and hypothyroid conditions. The study was conducted in a university hospital laboratory. We studied skeletal muscle obtained from 10 thyroidectomized patients with differentiated thyroid carcinoma on and after 4 wk off L-thyroxine replacement. Gene expression changes were measured using microarrays. Results were analyzed using dedicated statistical methods. We detected 607 differentially expressed genes on L-thyroxine treatment, of which approximately 60% were positively and approximately 40% were negatively regulated. Representative genes were validated by quantitative PCR. Genes involved in energy and fuel metabolism were overrepresented among the up-regulated genes, of which a large number were newly associated with thyroid state. L-thyroxine therapy induced a large down-regulation of the primary transcripts of the noncoding microRNA pair miR-206/miR-133b. We demonstrated that physiological levels of TH regulate a myriad of genes in human skeletal muscle. The identification of novel putatively TH-responsive genes may provide the molecular basis of clinical effects in subjects with different TH status. The observation that TH regulates microRNAs reveals a new layer of complexity by which TH influences cellular processes.

Thyroid hormone is required for the phenotype transitions induced by the pharmacological inhibition of calcineurin in adult soleus muscle of rats

The present experiment was designed to examine the effects of hypothyroidism and calcineurin inhibition induced by cyclosporin A (CsA) administration on both contractile and metabolic soleus muscle phenotypes, with a novel approach to the signaling pathway controlling mitochondrial biogenesis. Twenty-eight rats were randomly assigned to four groups, normothyroid, hypothyroid, and orally treated with either CsA (25 mg/kg, N-CsA and H-CsA) or vehicle (N-Vh and H-Vh), for 3 wk. Muscle phenotype was estimated by the MHC profile and activities of oxidative and glycolytic enzymes. We measured mRNA levels of the peroxisome proliferator-activated receptor-gamma coactivator-1 alpha (PGC-1 alpha), the major regulator of mitochondrial content. We also studied the expression of the catalytic A-subunit of calcineurin (CnA) both at protein and transcript levels and mRNA levels of modulatory calcineurin inhibitor proteins (MCIP)-1 and -2, which are differentially regulated by calcineurin activity and thyroid hormone, respectively. CsA-administration induced a slow-to-fast MHC transition limited to the type IIA isoform, which is associated with increased oxidative capacities. Hypothyroidism strongly decreased both the expression of fast MHC isoforms and oxidative capacities. Effects of CsA administration on muscle phenotype were blocked in conditions of thyroid hormone deficiency. Changes in the oxidative profile were strongly related to PGC-1 alpha changes and associated with phosphorylation of p38 MAPK. Calcineurin and MCIPs mRNA levels were decreased by both hypothyroidism and CsA without additive effects. Taken together, these results suggest that adult muscle phenotype is primarily under the control of thyroid state. Physiological levels of thyroid hormone are required for the effects of calcineurin inhibition on slow oxidative muscle phenotype.

Thyroid control of sarcolemmal Na+/Ca2+ exchanger and SR Ca2+-ATPase in developing rat heart.

Thyroid hormone (TH) levels increase in the postnatal life and are essential for maturation of myocardial Ca2+ handling. During this time, the sarcolemmal (SL) Na+/Ca2+ exchanger (NCX) function decreases and the sarcoendoplasmic reticulum (SR) Ca2+-ATPase (SERCA2) function increases. We examined the effects of postnatal hypo- or hyperthyroidism on NCX and SERCA2 in rat hearts. Animals were rendered hypothyroid by 0.05% 6-n-propyl-2-thiouracil in drinking water given to nursing mothers from days 2 to 21 postpartum. Hyperthyroidism was induced by daily injections of 10 microg/100 g body weight of 3,3',5-triiodo-L-thyronine during this period. Ventricular steady-state mRNA and protein levels of NCX and SERCA2 were analyzed by Northern and Western blotting. These were compared with SL Na+ gradient-induced and SR oxalate-supported Ca2+ transports in isolated membranes. In hypothyroidism, NCX mRNA and protein were elevated by 66 and 80%, respectively, and SERCA2 mRNA and protein were reduced to 55 and 70%, respectively (P < 0.05 vs. euthyroid). Corresponding differences were observed in the respective Ca2+ transports. Conversely, reduced NCX (by 50%) and elevated SERCA2 (by 150%) activities were found in hyperthyroidism (P < 0.05). The levels of NCX and SERCA2 mRNA and protein were, however, unchanged in hyperthyroidism, indicating that functional changes are not due to altered NCX and SERCA2 expression. In this case, a decline in noninhibitory phosphorylated phospholamban is a likely explanation for the elevated SR Ca2+ transport. In conclusion, physiological TH levels appear to be essential for normal reciprocal changes in the expression and function of myocardial NCX and SERCA2 during postnatal development.

Hypothyroidism decreases the ATP sensitivity of KATP channels from rat heart.

The effects of thyroid status on the properties of ATP-sensitive potassium channels were investigated. Single-channel recordings were made using excised inside-out membrane patches from enzymatically dissociated ventricular myocytes from hearts of control and thyroidectomized rats and each group was studied with and without administration of thyroid hormone. In patches excised from hypothyroid myocytes the IC50 for ATP inhibition of KATP channels was 110 micro m. This value was 3-fold higher than the IC50 in control myocytes (43 micro m). Treatment of hypothyroid rats to restore physiological levels of thyroid hormone (tri-iodothyronine, T3), resulted in a return to normal ATP-sensitivity (IC50 = 46 micro M). In patches from animals rendered hyperthyroid, the IC50 for ATP was 50 micro M and this value was not significantly different from the control. There was no difference in the cooperativity of ATP-binding (Hill coefficient, nH) among control (nH = 2.2), hypothyroid (nH = 2.1), T3-treated (nH = 2.0) and hyperthyroid groups (nH = 2.4). The unitary conductance was unchanged and there was no apparent change in intraburst kinetics between examples of single KATP channels from control and hypothyroid rats. Action potentials recorded in myocytes from hypothyroid rats were significantly shortened by 50 micro M levcromakalim, a KATP channel opener (P < 0.001) but unchanged in control myocytes.We conclude that hypothyroidism significantly decreased the ATP-sensitivity of KATP channels, whereas the induction of hyperthyroid conditions did not alter the ATP-sensitivity of these channels. Thus, hypothyroidism is likely to have important physiological consequences under circumstances in which KATP channels are activated, such as during ischemia.

Relationship between circulating thyroid hormones and cell-mediated immunity in immature male chickens

The objective of this experiment was to examine the relationship between circulating triiodothyronine (T3), thryoxine (T4) and cell-mediated immunity in immature male chickens. Three week old Single Comb White Leghorn male chicks were used as the experimental animals. In order to produce a wide range of circulating thyroid hormone concentrations, birds were divided into groups and received one of nine treatments including surgical thyroidectomy; 0.1% propylthiouracil (PTU) in the feed; 1 ppm T3 and 10 ppm T4 in the feed. The graft versus host (GvH) response and response to phytohemagglutinin (PHA) were measured at 6 weeks of age. Total and differential white blood cells were counted. Birds were bled, weighed, sacrificed, and the lymphoid organs were removed and weighed. Concentrations of T3 and T4 were measured in the birds from all treatment groups. There were positive correlations between thyroid hormones, mainly T3, and weights of thymus and spleen. T4 but not T3 was positively correlated with number of circulating lymphocytes. There were no significant correlations between circulating T3 and T4 and either PHA or GvH responses. It was concluded that physiological levels of thyroid hormones are needed to maintain normal weights of thymus and spleen and a normal level of circulating lymphocytes. Furthermore, we conclude

The influence of nanomolar calcium ions and physiological levels of thyroid hormone on oxidative phosphorylation in rat liver mitochondria. A possible signal amplification control mechanism.

Using different conditions mitochondria from hypothyroid rats can show both unchanged ADP/O ratios and lowered ADP/O ratios without evidence of uncoupling when compared with euthyroid controls. Raising the free Ca2+ concentration to around 25 nM progressively lowered the ADP/O ratio in hypothyroid but not in euthyroid mitochondria. Ruthenium Red did not alter this behaviour and further increasing the Ca2+ concentration to levels below those which stimulate State 3 respiration had no additional effect. Measurements of the free Ca2+ concentration in the mitochondrial suspending medium using a Quin 2 fluorescence assay showed that the mitochondria did not buffer the free Ca2+ at these low concentrations. At 25 nM-free Ca2+, addition of 10-13) M-T3 to hypothyroid mitochondria produced an immediate and significant increase in the ADP/O ratio without altering the free Ca2+ concentration. The hormone effect was maximal by 10(-11) M. The concentration of ATP synthetase can be estimated to lie at about 10 nM in these experiments. Hence it appears possible that a substantial amplification of the hormone signal may have taken place. Comparison with binding studies suggests that T3 may have been maximally stimulating when somewhat less than half its receptor sites had been filled. The possible mechanisms by which this receptor mediated alteration of the ADP/O ratio might be achieved are discussed.

Relationship between circulating thyroid hormones and humoral immunity in immature male chickens

The objective of this experiment was to examine the relationship between levels of circulating T3, T4, and humoral immunity in immature male chickens. Three week old Single Comb White Leghorn male chicks were used as the experimental animals. In order to produce a wide range of circulating thyroid hormone concentrations, birds were divided into groups and received one of nine treatments including surgical thyroidectomy; 0.1% propylthiouracil (PTU) in the feed; 1 ppm T3 and 10 ppm T4 in the feed. Antibody production against sheep red blood cells (SRBC) (thymus-dependent antigen) and Brucella abortus (BA) (thymus-independent antigen) was tested at 6 weeks of age. Concentrations of T3 and T4 were measured in birds from each treatment group at 7 and 11 weeks of age. At 11 weeks of age, birds were weighed, sacrificed and lymphoid organs removed and weighed. There were positive correlations between circulating thyroid hormones and weights of bursa of Fabricius and spleens. There were no significant correlations between circulating thyroid hormones and antibody production. It was concluded that physiological levels of thyroid hormones are needed to maintain normal weights of bursa and spleen. Furthermore, we conclude that lower than physiological levels might be sufficient for normal antibody production. Finally, stimulation of antibody production using thyroid hormones may require different doses than what were utilized in this study.

Role of calmodulin in thyroid hormone stimulation in vitro of human erythrocyte Ca2+-ATPase activity.

Because human erythrocyte membrane Ca2+-ATPase is a calmodulin-dependent enzyme, and because physiological levels of thyroid hormone stimulate this enzyme system in vitro, we have studied the role of calmodulin in this model of extranuclear thyroid hormone action. Ca2+-ATPase activity in the absence of thyroid hormone ("basal activity") was increased by inclusion in the preassay incubation mixture of purified calmodulin or hypothyroid erythrocyte hemolysate that contained calmodulin (39 micrograms calmodulin/ml packed cells, determined by radioimmunoassay); addition of L-thyroxine or 3,5,3'-triiodo-L-thyronine (10(-10)M) significantly enhanced (P less than 0.001) enzyme activity in the presence of calmodulin or hemolysate. The stimulatory effects of thyroid hormone, calmodulin, and hemolysate were additive. At 5-10 microM, trifluoperazine, an antagonist of calmodulin, inhibited thyroid hormone stimulation of Ca2+-ATPase activity. Higher concentrations of trifluoperazine (50-100 microM) inhibited basal and hormone-stimulated enzyme activity, with or without added calmodulin. Anti-calmodulin antibody (10-50 micrograms antibody/mg membrane protein) inhibited basal, calmodulin-stimulated and thyroid hormone-stimulated Ca2+-ATPase activity. Membrane preparations were shown by radioimmunoassay to contain residual endogenous calmodulin (0.27 +/- 0.02 micrograms/mg membrane protein). The latter accounts for the effect of trifluoperazine and calmodulin antibody on membrane Ca2+-ATPase activity in the absence of added purified calmodulin. These results support the conclusion that the in vitro action of physiological levels of iodothyronines on human erythrocyte Ca2+-ATPase activity requires the presence of calmodulin.

Regulation of thyrotropin production by mouse pituitary thyrotropic tumor cells in vitro by physiological levels of thyroid hormones.

Primary suspension cultures from serially transplanted mouse pituitary thyrotropic tumors were shown to be regulated by physiological levels of thyroid hormones. TSH release was linear for up to 48 h in control cultures and was inhibited progressively after 10, 24, and 48 h in cultures exposed to triiodothyronine (T3), 4.0 nM. TSH release was inhibited up to 55% of control and glucose consumption was stimulated up to 2.6-fold in a dose-dependent fashion by T3 between 0.2 and 4.0 nM. A biphasic dose-dependent relationship for T3 and thyroxine (T4), and TSH production was demonstrated in two series of cultures. TSH production was stimulated progressively by T3 up to 0.1 nM and T4 up to 5 nM. At higher T3 and T4 levels, TSH production was progressively inhibited. Half-maximal inhibition occurred at total medium concentrations of 0.2 nM T3 and 15 nM T4 or free hormone levels of 8 x 10-12 M and 14 X 10-11 M, respectively. High affinity, low capacity nuclear binding sites were demonstrated for T3 and T4. The ap...

The behaviour of thyroid hormones in an infant with untreated neonatal thyrotoxicosis.

The clinical course from birth and serial measurements of serum T3, T4 and TSH in an infant with untreated neonatal thyrotoxicosis are reported. The thyroid hormone levels fell exponentially with time at rates very much slower than those previously reported for the maternally-transmitted thyroid stimulating antibody generally thought to cause the disorder. Steady physiological levels of thyroid hormones were achieved after 110 days (serum T3 = 3.4 NMOL/L, T4 = 118 nmol/l). TSH first rose to a measurable level after about 90 days.