No abstract

Hyperthyroidism is associated with increased bone resorption but the mechanisms by which thyroid hormone (T3) affects bone cell metabolism remain unclear. Recently it has been suggested that T3 stimulates osteoclastic resorption indirectly through the release of soluble mediators from osteoblasts. The aim of the present study was to investigate whether the T3-induced increase in bone resorption could be due to the regulation of cytokine production by human osteoblasts (hOb).The effects of T3 (1, 10, 100 nM) and IL-1 (100 U/ml) as the positive control were examined on cytokine protein release and mRNA levels in cultured hOb cell lines (MG63, SaOs-2), primary hOb and human bone marrow stromal (hBMS) cells. T3 increased IL-6 and IL-8 mRNA levels as well as IL-6 and IL-8 protein release into the culture media from MG63 and hBMS cells in a time-and dose-dependent manner. The maximal effect on protein release in hBMS cells occurred at 24 h with a dose of T3 10 nM (IL-6 5·5 1·1-fold above controls; IL-8 3·7 0·5-fold above controls, P<0·05). At the same time, mRNA levels in hBMS cells were increased 6·2 0·8-fold for IL-6 (P<0·05) and 5·7 0·8-fold for IL-8 (P<0·05). Similar results were obtained in MG63 cells but no response was seen in SaOs-2 or hOb cells despite measurable basal production. Nor was there detectable regulation of IL-1 , IL-3, IL-11, IL-4 or granulocyte macrophage-colony stimulating factor by T3 in any cell type.In conclusion, T3 increases IL-6 and IL-8 production by MG63 and hBMS cells, suggesting that IL-6 and IL-8 may be T3-regulated genes in osteoblasts.

Whether biochemical markers can predict improvement in reduced bone mineral density (BMD) associated with thyrotoxicosis is unclear. We investigated the relationship between serum osteocalcin (OC), bone-specific alkaline phosphatase (b-ALP), serum deoxypyridinoline (Sdpd) and pyridinoline (Spyr), 24-hour urinary deoxypyridinoline (Udpd), and BMD in 17 thyrotoxic patients during 1 yr of treatment.Coinciding with euthyroidism at 4 -8 weeks, there was a peak in b-ALP and OC and a prompt fall into the normal range in Udpd and Sdpd, but not Spyr, levels. Mean b-ALP continued to be raised at week 52 when it was inversely correlated with BMD. Mean BMD rose approximately 6%, P Ͻ 0.01, over 1 yr. Coupling indices were calculated as a measure of bone balance and, at diagnosis, was [minus4.26 in favor of bone resorption and rose with treatment in favor of bone formation: weeks 2: Ϫ0.23; 4: ϩ4.01; 8: ϩ4.37; 12: ϩ4.44; 24: ϩ2.32; and 52: ϩ1.56.Bone turnover is balanced within 2 weeks of starting treatment for thyrotoxicosis. Udpd accurately indicates thyrotoxic bone resorption. . Furthermore, osteoblastic activity mediates T 3 activation of osteoclasts (2), leading to bone resorption and the release of markers such as the collagen cross-links pyridinoline and deoxypyridinoline. ␤-adrenergic receptor-mediated acceleration of bone activation frequency further exacerbates net bone loss (6). Whether treatment merely halts the damage or actually reverses the changes has been uncertain.Retrospective and cross-sectional studies suggest that a significant proportion of patients with a history of thyrotoxicosis continue to be at risk of reduced bone mineral density (BMD) up to 18 yr later (7). Longitudinal studies have been few, and estimation of BMD has been hindered by imprecise and indirect techniques. Of the four studies reported, one has used calcium bone index (8), and the remaining three, dual photon ( 153 Gd) absorptiometry for either all (9, 10) or at least one (11) of the serial estimations of bone mineral content (BMC). Although it is established that thyrotoxicosis leads to reduced BMC or density, the issues that remain unclear are: 1) the capacity of antithyroid medication to restore bone mass; and 2) the prediction of which individuals are more likely to suffer a long-term deficit in BMD.Changes in BMD occur only slowly, making it difficult to judge the benefits, if any, of antithyroid medication. Early identification of those individuals at risk of long-term reduction in BMD caused by thyrotoxicosis may enable early therapeutic intervention. A number of biochemical markers are thought to reflect the rapid bone turnover in thyrotoxicosis, although their predictive value remains to be established.Such markers include serum ALP, OC, and serum and urinary collagen cross-links. Previous studies of these markers in thyrotoxicosis have been either cross-sectional (12-14) or limited by cumbersome techniques (15, 16) and absence of simultaneous BMD measurements (17,18). Total ALP and OC are raised in approximately 30%...

Thyroid hormones are important regulators of GH synthesis and secretion. In this study we have made a detailed examination of the time-course of the effects of hypothyroidism and tri-iodothyronine (T3) replacement in the intact rat on GH gene expression in the anterior pituitary gland. Changes in pituitary cytoplasmic GH messenger (m)RNA levels were compared with total pituitary GH content and serum GH concentration during the development of hypothyroidism and following short-term T3 replacement in vivo. Hypothyroidism was associated with a fall in pituitary GH mRNA levels. Treatment of hypothyroid animals with T3 rapidly stimulated GH mRNA levels to values above those seen in euthyroid controls. The reduction in GH mRNA levels seen during the development of hypothyroidism was accompanied by a fall in serum GH and pituitary GH content, both of which were partially restored by T3 replacement. Thus thyroid hormone replacement in hypothyroidism rapidly stimulates GH mRNA synthesis, which is followed by the gradual restoration of pituitary GH stores and serum GH concentration.

The role of oestrogen in the regulation of TSH gene expression is unclear. We have examined the effect of administration of oestrogen in the rat on serum TSH, pituitary TSH content and pituitary cytoplasmic concentrations of mRNA encoding the TSH beta and alpha subunits, thus deriving measures of hormone release and synthesis. In addition, we have examined the effect of oestrogen on the binding of tri-iodothyronine (T3) to nuclear receptors in the anterior pituitary. Administration of oestrogen did not affect serum concentrations of TSH in euthyroid or untreated hypothyroid rats, but did augment the effects of T3 (1 and 2 micrograms) on serum TSH in hypothyroid animals 6 h after injection of T3. No influence of oestrogen or of thyroid status on pituitary content of TSH was seen. A marked increase in the concentrations of TSH beta and alpha mRNA in pituitary cytoplasm was found in hypothyroidism, compared with those in the euthyroid state. No effect of oestrogen on TSH mRNA was seen in euthyroid animals but concentrations of TSH beta and alpha mRNA were lower in hypothyroid animals than in vehicle-treated controls. A stimulatory influence of T3 on TSH mRNA was seen 6 h after injection of T3; this stimulation was absent in oestrogen-treated rats. No effect of oestrogen on the action of T3 was evident 72 h after beginning treatment with T3. In addition to effects on serum TSH and TSH mRNA, an increase in the number of pituitary nuclear receptors for T3 was seen after oestrogen treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Overproduction of thyroid hormones promotes bone resorption in vivo and in vitro, and we have evaluated whether mediators of such effects could include the osteotropic cytokines. Previous studies have demonstrated raised serum interleukin (IL)-6 in thyrotoxic patients, but differentiating the contribution of the elevated thyroid hormones from that of the autoimmune inflammation present in Graves' disease (GD) has been difficult. We undertook a longitudinal study of 34 patients (19-45 yr old) with GD, toxic nodular goiter (TNG), or a history of thyroid carcinoma but no evidence of disease recurrence, receiving sufficient T4 to suppress TSH. Controls were 12 euthyroid females. The following measurements were made basally and for 6 months after carbimazole treatment: serum free T4, T3, bone-specific alkaline phosphatase (b-ALP), IL-6, IL-8, IL-1beta, tumor necrosis factor-alpha, IL-11, and urinary deoxypyridinoline (Udpd). Compared with controls (IL-6, 1.1 +/- 0.3 ng/L; IL-8, 3.2 +/- 0.8 ng/L), untreated patients with GD and TNG had elevated IL-6 (GD, 7.11 +/- 0.88 ng/L; TNG, 7.30 +/- 0.77 ng/L; P < 0.001) and IL-8 (GD, 10.3 +/- 1.23 ng/L; TNG, 9.81 +/- 1.27 ng/L; P < 0.001). These levels fell after treatment and were then indistinguishable from those in control subjects. Thyroid carcinoma patients on TSH suppressive therapy also had significantly raised levels of IL-6 (2.5 +/- 0.42 ng/L) and IL-8 (4.4 +/- 0.63 ng/L). When data from all the patients were pooled, the levels of IL-6 and IL-8 correlated with serum T3 and free T4 but not with Udpd or b-ALP. IL-1beta, IL-11, and tumor necrosis factor-alpha were not raised in any patient. The elevations in serum IL-6 and -8 that occur in hyperthyroidism seem to result from the chronic effects of thyroid hormone excess rather than the accompanying autoimmune inflammatory condition produced by Graves' thyroid or eye disease. The site of the presumed increased production of IL-6 and -8 is most likely from bone osteoblasts, despite the inability of bone markers (such as Udpd and b-ALP) to correlate with acute changes in thyroid hormone status produced by antithyroid therapy.

SUMMARY. Neural cell adhesion molecules (NCAMs) are found predominantly in neural, muscle and endocrine cells. Recent interest has focused on their potential role in tumorigenesis. We have analysed the expression and secretion of NCAM in a series of 48 human pituitary adenomas. Immunocytochemical analysis of 19 adenomas demonstrated NCAM expression in all tumours with, in each case, diffuse cytoplasmic staining being found with variable membrane accentuation. There were no apparent differences in the expression of immunoreactivity seen on sections between individual tumours. Cell culture media from 43 dispersed human pituitary tumours were analysed by immunoassay for the secretion of soluble NCAM and all the pituitary hormones. In contrast to the immunocytochemical studies, soluble NCAM was released from only 27% of human pituitary tumours, but this was not related to tumour type nor was the amount of soluble NCAM released correlated with the amount of pituitary hormone secreted by each adenoma. NCAM expression is common to all human pituitary adenoma types and the observed differences in release of soluble NCAM between individual tumours may reflect different molecular mechanisms, altering adhesive interactions between normal and adenomatous tissue.

Hormone release in culture in response to pituitary adenylate cyclase-activating polypeptide (PACAP) was examined in 28 human pituitary adenomas: 10 null cell adenomas, 4 gonadotropin-, 6 GH-, 6 ACTH-, and 2 PRL-producing adenomas. The effects of PACAP38 were compared with those of the classical hypothalamic releasing hormones and other activators of intracellular signaling pathways. PACAP38 significantly stimulated GH release from 1 somatotrope tumor (125 +/- 3% of control; P < 0.05) and ACTH release from 3 corticotrope tumors (134 +/- 6%, 136 +/- 7%, and 137 +/- 9% of control; P < 0.05). The effects of PACAP38 were less potent than either GHRH on GH release in the somatotrope tumor or CRH on ACTH release in the corticotrope tumors but similar to the responses seen with the cAMP analog 8-bromo-cAMP (8-Br-cAMP). No detectable effects of PACAP38 on hormone release from null cell, gonadotropin-, or PRL-producing adenomas were observed. Of the 5 somatotrope tumors that failed to respond to PACAP38, all also failed to respond to either 8-Br-cAMP, TRH, or GHRH. Of the corticotrope tumors that failed to respond, 2 of the 3 also failed to respond to CRH. In addition to eliciting hormone release appropriate to the tumor type, PACAP38 also stimulated glycoprotein hormone alpha-subunit (alpha SU) release from one somatotrope tumor (229 +/- 35% of control, P < 0.01) and one corticotrope tumor (149 +/- 4% of control; P < 0.01). This response was not mimicked by 8-Br-cAMP in the somatotrope tumor, but in the corticotrope tumor a significant alpha SU release was also seen after stimulation with the protein kinase C activator 12-O-tetradecanoyl-phorbol-13-acetate and 8-Br-cAMP. These results suggest that the novel hypothalamic peptide PACAP38 has a modest role in the regulation of GH, ACTH, and alpha SU secretion from some human tumourous pituitary corticotropes and somatotropes. Further studies are needed to elucidate the intracellular signaling pathways that mediate the effects of PACAP on hormone secretion by these tumor types.