Nuclear trafficking of the G-protein-coupled parathyroid hormone receptor

G-protein-coupled receptors are a family of receptors that signal primarily through heterotrimeric G proteins. However, new evidence has emerged to show that the signaling capabilities of the receptors are beyond those of traditional signaling cascades. One such example is that the parathyroid hormone (PTH) type 1 receptor is found not only at the plasma membrane but also in the nucleus of cells in cell lines and tissues. This review discusses the emerging concepts of nuclear PTH signaling and relates this information to the growing field of nuclear G-protein-coupled receptors. We review recently published studies on the mechanism of PTH nuclear localization, its role in the cell nucleus, and the contrasting roles ligands play in regulating the receptors' nuclear localization. The review also discusses the importance of nuclear G-protein-coupled receptors and future directions for research in this field.     

Expression of parathyroid hormone-related protein and the parathyroid hormone/parathyroid hormone-related protein receptor in rat thymic epithelial cells

Thymic epithelial cells are an important source of cytokines and other regulatory peptides which guide thymocyte proliferation and maturation. Parathyroid hormone-related protein (PTHrP), a cytokine-like peptide, has been reported to affect the proliferation of lymphocytes in vitro. The studies presented here were undertaken to test the hypotheses that PTHrP is produced locally within the thymus where it could influence thymocyte maturation and, more specifically, that thymic epithelial cells (TEC) could be the intrathymic source of PTHrP expression. To this end, immunohistochemical studies were performed to localise PTHrP and the PTH/PTHrP receptor within the adult rat thymus. Antibodies directed against 2 different PTHrP epitopes, PTHrP(1–34) and PTHrP(34–53), demonstrated prominent specific PTHrP immunoreactivity in both subcapsular and medullary TEC. In addition, faint but specific staining for PTHrP was seen in the cortex, interdigitating between cortical lymphocytes while sparing epithelial-free subcapsular areas, thus suggesting that cortical TEC could also be a source of PTHrP immunoreactivity. In contrast, PTH/PTHrP receptor immunoreactivity was only seen in medullary and occasional septal TEC; no evidence of cortical or lymphocytic PTH/PTHrP receptor immunoreactivity was detected. Immunohistochemical studies of cultured cytokeratin-positive rat TEC confirmed the results of these in situ studies as cultured TEC were immunoreactive both for PTHrP and the PTH/PTHrP receptor. Thus these results demonstrate that PTHrP is produced by the epithelial cells of the mature rat thymus. This suggests that PTHrP, a peptide with known cytokine, growth factor and neuroendocrine actions, could exert important intrathymic effects mediated by direct interactions with TEC, or indirect effects on PTH/PTHrP receptor-negative thymocytes.     

Postresection parathyroid hormone and parathyroid hormone decline accurately predict hypocalcemia after thyroidectomy

Hypocalcemia is the most frequent complication after total thyroidectomy. Parathyroid hormone (PTH) measurement has been proposed as an early predictor of this condition. Total thyroidectomy was performed in 39 patients. Hypocalcemia was present in 15 cases (38%). Patients undergoing hemithyroidectomy (n = 13) were considered control subjects not developing hypocalcemia. PTH was measured before surgery and 10 minutes after resection of the gland using a rapid (15 minutes) chemiluminescent immunometric assay. Patients developing hypocalcemia had lower calcium and postresection PTH levels and higher PTH decline than patients not developing hypocalcemia (P < .0001). PTH decline (cutoff value, 62.5%) had the better sensitivity (93.3%) for predicting hypocalcemia, allowing for a fairly safe early discharge. However, the best overall results corresponded to the combination of postresection PTH level (< or = 18 pg/mL [< or = 1.9 pmol/L]) and PTH decline (>62.5%), with a sensitivity of 90% and a specificity of 97.9%. Perioperative PTH measures can accurately predict hypocalcemia after thyroidectomy, granting the laboratory a key role in the immediate decision about calcium supplementation for patients at risk.     

Immune function of thyroid stimulating hormone and receptor

Thyroid stimulating hormone (TSH) is a central component of the hypothalamus-pituitary-thyroid axis. Although TSH is known for its important biological effects as a neuroendocrine used to regulate thyroid hormone activity and subsequent metabolic functions, TSH also has been shown to be produced and used by cells ofthe mammalian immune system. Moreover, recent findings have linked the use of TSH by cells of the immune system in humans and mice to a group of monocytic cells and lymphocytes--primarily dendritic cells, macrophages, and subset of na?ve peripheral T cells. Other studies have demonstrated the capacity of dendritic cells and monocytes to produce biologically active TSH, thereby pointing to a process of paracrine or autocrine TSH-mediated communication during the earliest stages of an immune response to antigen. In this article, these and other features of TSH immune-endocrine interactions are discussed in the context of an intrinsic TSH immunological pathway. Additionally, a hypothesis is proposed in which TSH produced by cells of the immune system during acute antigen exposure plays a dual role, consisting on the one hand of TSH communication during antigen-driven immune activation while concomitantly serving to regulate physiological homeostasis by modulating and adjusting thyroid hormone activity.     

Immunoelectron microscopical evidence of a calcium receptor function in parathyroid, placenta and kidney.

The parathyroid, kidney and placenta have been investigated for ultrastructural signs of immunostaining with a murine monoclonal IgG 1-antibody denoted E11. Dispersed cells and tissue sections revealed a finely granular, electron dense precipitate after fixation with periodate-lysine-paraformaldehyde and indirect immunoperoxidase staining with the native or biotinylated antibody. This precipitate was confined to the surface membrane of parathyroid chief cells of normal and adenomatous human glands as well as the bovine and rat parathyroid parenchyma, preferentially the brush border membrane of proximal tubular cells in the human, rat and mouse kidney, cytotrophoblast cells of the human placenta, and trophoblast cells lining fetal blood vessels in the rat placenta. Since the E11 antibody recognizes a large glycoprotein regulating intracellular calcium mobilization and cation fluxes across the cell membrane, the present findings suggest the existence of a similar calcium receptor function on cells involved in different aspects of the calcium homeostasis within a variety of species.     

Monoclonality and abnormal parathyroid hormone genes in parathyroid adenomas

Previous work based on the relative tissue content of glucose-6-phosphate dehydrogenase isoenzymes suggested that parathyroid adenomas, like primary hyperplasia, may be multicellular (not clonal) in origin. We have reexamined this issue by using two independent molecular genetic methods. We report tumor-cell-specific restriction-fragment-length alterations involving the parathyroid hormone gene from two human parathyroid adenomas. These abnormal restriction fragments indicate that in each case a clonal proliferation of cells was present and also suggest that DNA alterations involving the parathyroid hormone locus may be important in the tumorigenesis or clonal evolution of some parathyroid adenomas. In addition, we used a restriction-fragment-length polymorphism in an X-linked gene (hypoxanthine phosphoribosyltransferase) to examine the clonality of eight parathyroid adenomas in women. Of these eight adenomas, six had the DNA hybridization pattern of monoclonality, and two had an equivocal pattern. None of five hyperplastic parathyroid glands had a monoclonal pattern. We conclude that some (and perhaps many) single parathyroid adenomas are monoclonal neoplasms. Our observations suggest that there is a fundamental biologic difference between parathyroid adenomas and primary hyperplasia--a difference that could prove useful in distinguishing these entities clinically.     

巨噬细胞甘露糖受体的结构与功能

巨噬细胞甘露糖受体属于哺乳动物类C型凝集素超家族成员,主要表达于巨噬细胞和未成熟树突状细胞表 面,不仅在抵抗病原体感染的天然免疫防御中起重要作用,还参与抗原提呈和获得性免疫应答。     

Immunocytochemical localization of parathyroid hormone in bovine parathyroid glands and human parathyroid adenomas.

Light and electron microscopic localization of parathyroid hormone (PTH) in human and bovine parathyroid tissue has been achieved using an indirect peroxidase labeled antibody method. Granular deposition of the reaction product was found throughout the chief cell cytoplasm. There was no nuclear staining. At the ultrastructural level, parathyroid hormone localized by this method appeared to be largely confined to the secretory granules in the cytoplasm of cells. Mitochondria and nuclei were free of reaction product. Aggregated sacs of granular endoplasmic reticulum were minimally reactive, and Golgi apparatuses did not show reaction product.     

Interrelationship of chlorothiazide and parathyroid hormone: a micropuncture study.

Sodium and calcium are normally reabsorbed in parallel in the renal tubule. Both parathyroid hormone (PTH) and thiazide diuretics may influence this relationship. This study was designed to show whether the dissociation of Na from Ca transport produced by thiazides is dependent upon the presence of PTH. Hydropenic thyroparathyroidectomized (TPTX) dogs were given chlorothiazide alone and together with PTH. Chlorothiazide alone significantly increased fractional excretion of sodium (0.5 +/- 0.3-5.6 +/- 0.3%) and calcium (0.74 +/- 0.18-1.4 +/- 0.24%). However, the Ca/Na excretion ratio fell markedly from 1.57 to 0.24%. Micropuncture revealed this dissociation to occur at the distal tubule. Proximal reabsorption of water, sodium, and calcium were inhibited to an equal extent. However, distal fractional sodium reabsorption fell 10% whereas calcium reabsorption remained unchanged following chlorothiazide administration in TPTX animals. When phosphaturic doses of PTH were administered with chlorothiazide, no significant changes were observed in calcium or sodium reabsorption. It is concluded that PTH plays no role in the dissociation of sodium from calcium reabsorption resulting from acute chlorothiazide administration.     

Structure and flexibility of the multiple domain proteins that regulate complement activation

In this review we summarise more than 10 years of biophysical exploration into the structural biology of the regulators of complement activation (RCA). The five human proteins responsible for regulation of the early events of complement are homologous and are composed largely from building blocks called “complement control protein (CCP) modules”. Unlike most multiple domain proteins they do not contain any of the other widely occurring module types. This apparent simplicity of RCA structure, however, is belied by their sophistication of function. In fact, the structures of the individual CCP modules exhibit wide variations on a common theme while the extent and nature of intermodular connections is diverse. Some neighbouring modules within a protein stabilise each other and some co‐operate to form specific binding surfaces. The degree of true “modularity” of CCPs is open to debate. The study of RCA proteins clearly illustrates the value of combining complementary structural biology techniques. The results could have implications for folding, evolution, flexibility and structure–function relationships of other molecules in the large, diverse and little understood category of multiple domain proteins. Work in the Barlow laboratory was supported by the Medical Research Council and the Biotechnology and Biological Sciences Research Council of the UK and by the Wellcome Trust. MDK was supported by the Human Frontiers Science Program.     

Delineation of regions in the extracellular domain of follicle-stimulating hormone receptor involved in hormone binding and signal transduction

PROBLEM: To use antipeptide antibodies to potential surface-oriented regions of the extracellular domain (ECD) of the human follicle-stimulating hormone receptor (hFSHR) to delineate regions involved in FSH binding and FSH-induced signal transduction. METHOD OF STUDY: We developed and characterized antipeptide antibodies to different, potentially surface-oriented regions of the ECD of hFSHR. The ability of these antibodies to recognize the receptor was studied by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by Western blotting. The ability to modulate FSH binding and cAMP generation was studied by the radioreceptor assay and in vitro FSH bioassay respectively. RESULTS: Antipeptide antibodies to regions 15-31, 216-235, 285-300 and 327-341 hFSHR inhibited both FSH binding and cAMP production. Regions 15-31 and 216-235 were accessible to their cognate antipeptide antibodies both before and after FSH binding, while regions 285-300 and 327-341 hFSHR were accessible only prior to FSH binding. CONCLUSIONS: Based on the observations made with respect to accessibility to antipeptide antibodies, ability of antibodies to inhibit FSH binding and the subsequent cAMP generation and kinetics of antibody binding, regions 285-300 and 327-341 hFSHR appear to be the chief FSH-binding sites, while regions 15-31 and 216-235 hFSHR serve as ancillary FSH-binding sites.     

Co-secretion of parathyroid hormone and parathyroid-hormone-related protein via a regulated pathway in human parathyroid adenoma cells.

Parathyroid-hormone-related protein (PTHrP) is widely expressed not only in malignant tumors but also in both epithelial and nonepithelial cells of normal tissues. Secreted PTHrP is suspected to act as a paracrine or autocrine regulator. However, little is known about its secretory pathway. To cast light on this question, we studied the intracytoplasmic distribution of parathyroid hormone (PTH) and PTHrP immunohistochemically and immunoelectron microscopically in 10 surgically resected parathyroid adenomas. Double immunostaining was performed using anti-PTH antibody and a newly established anti-PTHrP antibody to reveal the relationship between their two distributions. Additional examination by cell immunoblot assay was performed to determine whether both PTH and PTHrP are secreted simultaneously. Both PTH and PTHrP were actually secreted from individual parathyroid cells simultaneously on cell immunoblot assay. Immunohistochemically, there were two different types of adenoma cells, i.e., one positive only for PTH and the other positive for both PTH and PTHrP. PTH was distributed linearly or fine granularly along the cytoplasmic membrane, whereas PTHrP was distributed diffusely or coarse granularly in the cytoplasm. The intracytoplasmic distributions of PTH and PTHrP often overlapped. Immunoelectron microscopical examination demonstrated that PTHrP co-localized with PTH in the same secretory granules. The results clearly demonstrated that PTHrP can be co-secreted with PTH via a regulated pathway using secretory granules.     

Immunohistochemical analysis of low-grade and high-grade prostate carcinoma: relative changes of parathyroid hormone-related protein and its parathyroid hormone 1 receptor, osteoprotegerin and receptor activator of nuclear factor-kB ligand

AIM: To investigate multiple bone cytokines produced by prostate carcinoma (PCa) as a novel strategy to differentiate potential aggressiveness in localised PCa using immunohistochemical analysis. METHODS: A total of 47 cases of PCa undergoing radical prostatectomy or transurethral prostatic resection at our institution (Fundación Jiménez Díaz (Grupo Capio), Madrid, Spain) between January 1991 and June 1998 were identified as low-grade (< or =4; n = 22) or high-grade (> or =7, excluding 7 (3+4) cases; n = 25) PCa according to Gleason grade. PCa specimens were immunostained for: parathyroid hormone (PTH)-related protein (PTHrP), the PTH1 receptor, osteoprotegerin and receptor activator of nuclear factor-kappa B ligand (RANKL), as well as Ki67 (a proliferation marker) and CD34 (an angiogenesis marker). RESULTS: PCa samples showed an increased immunostaining for both osteoprotegerin and RANKL, associated with tumour grade and PTHrP positivity, in the tumoral epithelium. Using a score value of 4-corresponding to moderate staining - as cut-off, the best sensitivity value was for PTHrP (with C-terminal antiserum C6; 100 %); wheras the best specificity value was for RANKL (95 %). CONCLUSIONS: All the evaluated factors are overexpressed mainly in the high-grade tumours. Our findings indicate that, in most patients with PCa (with Ki67 values between 1% and 9%), sequential determination of C-terminal PTHrP and RANKL immunoreactivities is a useful approach to discriminate low-grade and high-grade tumours.     

The effect of histamine2 and muscarine receptor antagonists on plasma levels of parathyroid hormone and calcitonin

Summary Long-term administration of cimetidine, a histamine₂ receptor antagonist, has been reported to normalize elevated parathyroid hormone (PTH) concentrations in patients with secondary [1] and primary hyperparathyroidism [2] and even to improve the clinical symptoms. We have compared the effect of cimetidine and pirenzepine on PTH and calcitonin (CT) plasma levels in a short-term trial on patients with secondary hyperparathyroidism. After cimetidine a significant effect on PTH was seen within 30 min lasting 30 min and after pirenzepine, within 60 min and lasting 60 min. The effect on CT was only significant after cimetidine.