Citations

Abstract Some data suggest that human calcitonin (CT) secretion is lower in women than in men, decreases with age and the menopause, and is absent in thyroidectomized persons. To further explore CT secretory physiology, we have studied basal and calcium-stimulated plasma immunoreactive CT (iCT) and silica-extractable monomeric CT concentrations in 148 healthy volunteers and 33 patients with a history of thyroid damage (total or subtotal thyroidectomy, radioiodine treatment for thyrotoxicosis). Both whole-plasma iCT and extractable CT levels were lower basally and after calcium infusion in women than in men, basal levels being reduced about 50% and calcium-stimulated values about 75% from those of male subjects. There were no significant changes in basal iCT or extractable CT concentrations with age, and CT secretory capacity (CT response to calcium infusion) likewise did not change with age. Infusion of monomeric CT to constant concentration in 27 persons permitted estimates of CT metabolic clearance rates (MCRs) and secretion rates (SRs). Calculated MCRs of about 9 ml/min · kg−1 (persons aged 21–30 yr) and 6 ml/min · kg−1 (persons aged 54–70 yr) were in good agreement with published data, and did not differ between the sexes. SRs were dependent upon the assay method used to estimate basal plasma CT concentrations, being highest when whole-plasma iCT values were used. Based on estimates of plasma monomeric CT from the silica extraction procedure, the SR of CT was 59 ± 6 (SE) ng/d · kg−1 in men, and 22 ± 3 ng/d · kg−1 in women. Thyroid damage reduced, but did not abolish, apparent CT immunoreactivity, even in silica extracts of plasma. However, all subsets of thyroiddamaged patients had absent-to-markedly-impaired CT secretion in response to calcium infusion. We conclude that CT secretion is substantially lower both basally and after stimulation in women than in men, and that this difference in CT immunoreactivity probably reflects differences in circulating CT bioactivity. The sex difference in plasma CT concentrations probably results from lower rates of CT secretion in women, not increased MCR. There is no age-related decrease of plasma CT concentrations (or CT secretory reserve), calling into question the concept that a progressive deficiency of CT is partly resonsible for age-related (“senile”) osteoporosis. Surgical or radiation damage to the thyroid gland commonly abolishes C-cell response to calcium; such CT-deficient patients form a population suitable for determining whether or not reduced CT secretion can impair skeletal homeostasis.

Abstract Mammary glands are target tissues for 1,25-dihydroxyvitamin D3 (1,25(OH)2D3). We have examined a mouse mammary tumor cell line (GR) for receptors of 1,25(OH)2D3 and have examined alterations in the growth and morphology of these cells in response to 1,25(OH)2D3. GR cells contain a high affinity (Kd ∼ 10−11), lowcapacity receptor with a high specificity for 1,25(OH)2D3. The 1,25(OH)2D3 receptor in GR cells has a sedimentation coefficient of 3.5 and elutes from DEAE cellulose columns with ∼ 0.15 M KCI. These properties of the receptor are similar to those reported for other 1,25(OH)2D3 receptors. 1,25(OH)2D3 is internalized by GR cells in situ and specifically bound 1,25(OH)2D3 is found predominantly, if not entirely, in the nucleus as determined by cell fractionation and autoradiographic techniques. The incubation of GR cells in culture for 7 days with 1,25(OH)2D, markedly alters cell growth. Cell growth is retarded in a dose-dependent manner; physiologic concentrations (10−10M) of l,25(OH)2D3 retard cell growth by approximately 50%. In addition, GR cells incubated with 10−9 to 10−8M 1,25(OH)2D3 undergo marked morphological changes. The incubation of GR cells with other vitamin D metabolites such as 25-hydroxyvitamin D3 (25(OH)D3) at a concentration of 10−9M does not significantly alter cell growth or morphology. The presence of high affinity receptors for 1,25(OH)2D3, the specific internalization of 1,25(OH)2D3 predominantly into the nuclei, and the significant effects of physiological concentrations of 1,25(OH)2D3 on cell growth suggest a direct, specific, nuclear effect of 1,25(OH)2D3 on GR cells. The mouse mammary tumor model might be useful in examining the effect of 1,25(OH)2D3 on tumor formation.

Abstract We studied the mechanism of impaired calcium absorption with aging in 51 healthy women whose ages ranged from 26 to 88 years. Serum concentrations of 1,25-dihydroxyvitamin D [1,25-(OH)2D, mean of four measurements per subject] increased with age by 22% (P < 0.05) but, by split-point analysis, plateaued or decreased slightly after age 65. In a subset of 20 subjects, [3H]1,25-(OH)2D3 kinetic analysis showed that this increase with age resulted from both increased production and decreased metabolic clearance of 1,25-(OH)2D. Despite the increase in serum 1,25-(OH)2D concentration, true calcium absorption did not change with age. The expected inverse correlation between true fractional calcium absorption and dietary calcium intake, however, was easily demonstrated (r = 0.66, P < 0.001). Serum intact parathyroid hormone (PTH) increased with age by 35% (P < 0.02) and serum bone gla protein (BGP, osteocalcin) increased by 47% (P < 0.001); the increases in serum PTH and serum BGP were directly correlated (r = 0.32, P < 0.05). The data are consistent with the following hypothetical model: (1) intestinal resistance to 1,25-(OH)2D action accounts for the increase in serum 1,25-(OH)2D concentrations with aging with no change in true calcium absorption; (2) this results in a compensatory increase in PTH secretion and in 1,25-(OH)2D production that prevents true calcium absorption from decreasing; (3) the previously described defect in 25-hydroxyvitamin D (25-OHD) 1α-hydroxylase activity in aging animals and humans acccounts for the leveling off in serum 1,25-(OH)2D concentration after age 65 years; and (4) the secondary hyperparathyroidism leads to increased bone turnover and thus contributes to age-related bone loss.

Abstract A new bone cell line was established by transfecting normal adult human osteoblast-like (hOB) cells, derived from a 68-year-old woman, with the plasmid pSV3 neo. The plasmid included coding sequences and promotors for the large and small T antigens of the SV40 virus as well as resistance to the antibiotics neomycin and G418. A single antibiotic-resistant colony was located and cloned. Large tumor antigen production in the clonal cell line was confirmed by indirect immunofluorescence study. Treatment with 1,25-dihydroxy-vitamin D3 increased steady-state concentrations of protein and mRNA for osteocalcin and for alkaline phosphatase. Northern blot analyses also demonstrated the presence of mRNAs for α(I)-procollagen, osteopontin 1a, transforming growth factor β, and interleukin-1β. The plasma membrane calcium pump and osteonectin were identified by immunocytochemical analysis. These cells produced a matrix that mineralized when β-glycerophosphate was added to their cultures. As assessed by functional receptor assays, both estrogen and androgen receptors were present and functional, although at low concentrations. Treatment with parathyroid hormone did not stimulate adenylate cyclase activity. Thus, these cells are a well-differentiated, steroid-responsive clonal cell line that closely approximates the phenotype of the mature osteoblast. They should serve as an excellent model for the study of osteoblast biology.

Abstract Silicon in trace amounts enhances bone formation, and the silicon-containing compound zeolite A (ZA) increases eggshell thickness in hens. In the studies reported here, treatment of nearly homogeneous strains of normal human osteoblast-like cells for 48 h with ZA at 0.1–100 μg/ml induced a dose-dependent increase (r = 0.35, P < 0.001) in DNA synthesis (n = 31) to 162 ± 16% (mean ± SEM) of control and in the proportion of cells in mitosis (n = 4) from 9.1 ± 1.8 to 27.0 ± 4.5% (r = 0.69, P < 0.005). ZA treatment also increased alkaline phosphatase activity (P < 0.05) and osteocalcin release (P < 0.05) but did not significantly affect collagen production per individual cell. The mitogenic action of ZA was dependent on cell seeding density over the range of 1250–40,000 cells per cm2, which is consistent with induction of an autocrine factors). TGF-β is a potent mitogen for osteoblasts. ZA treatment increased the steady-state mRNA levels of transforming growth factor β1 (TGF-β1) and induced the release of the latent form of TGF-β protein into the conditioned medium within 6 h. We conclude that ZA induces the proliferation and differentiation of cells of the osteoblast lineage.

Abstract Vertebral fractures are the classic hallmark of osteoporosis, yet little is known of their epidemiology. The incidence of clinically diagnosed vertebral fractures was therefore directly assessed in the predominantly white (European descent) population of Rochester, Minnesota. Altogether, 341 Rochester residents were radiologically diagnosed for the first time with one or more vertebral fractures in the 5 year study period, 1985–1989. The overall age- and sex-adjusted incidence rate was 117 per 100,000 person-years (95% CI, 105 to 130). The age-adjusted rate in women (145 per 100,000 person-years) was almost twice that in men (73 per 100,000 person-years). Of all fractures, 47 (14%) followed severe trauma, 282 (83%) followed moderate or no trauma, and 12 (3%) were pathologic. Incidence rates for fractures following moderate trauma were higher in women than in men and rose steeply with age in both genders. In contrast, fractures following severe trauma were more frequent in men, and their incidence increased less with age. These Rochester rates are greater than those previously reported from studies in Britain and Sweden but lower than the incidence rates extrapolated from a prevalence study in this population.

Through the development of TGFβ-inducible early gene-1 (TIEG1) knockout (KO) mice, we have demonstrated that TIEG1 plays an important role in osteoblast-mediated bone mineralization, and in bone resistance to mechanical strain. To further investigate the influence of TIEG1 in skeletal maintenance, osteocytes were analyzed by transmission electron microscopy using TIEG1 KO and wild-type mouse femurs at one, three and eight months of age. The results revealed an age-dependent change in osteocyte surface and density, suggesting a role for TIEG1 in osteocyte development. Moreover, there was a decrease in the amount of hypomineralized bone matrix surrounding the osteocytes in TIEG1 KO mice relative to wild-type controls. While little is known about the function or importance of this hypomineralized bone matrix immediately adjacent to osteocytes, this study reveals significant differences in this bone microenvironment and suggests that osteocyte function may be compromised in the absence of TIEG1 expression.

Acute ischemic insults to the kidney are recognized complications of human sickle cell disease (SCD). The present study analyzed in a transgenic SCD murine model the early renal response to acute ischemia. Renal hemodynamics were profoundly impaired following ischemia in sickle mice compared with wild-type mice: glomerular filtration rate, along with renal plasma flow and blood flow rates, were markedly reduced, while renal vascular resistances were increased more than threefold in sickle mice following ischemia. In addition to these changes in renal hemodynamics, there were profound disturbances in renal signaling processes: phosphorylation of members of the MAPK and Akt signaling proteins occurred in the kidney in wild-type mice after ischemia, whereas such phosphorylation did not occur in the kidney in sickle mice after ischemia. ATP content in the postischemic kidney in sickle mice was less than half that observed in wild-type mice. Examination of the expression of candidate genes uncovered changes that may predispose to increased sensitivity of the kidney in sickle mice to ischemia: increased expression of inducible nitric oxide synthase and decreased expression of endothelial nitric oxide synthase, and increased expression of TNF-α. Inducibility of anti-inflammatory, cytoprotective genes, such as heme oxygenase-1 and IL-10, was not impaired in sickle mice after ischemia. We conclude that the kidney in SCD is remarkably vulnerable to acute ischemic insults. We speculate that such sensitivity of the kidney to ischemia in SCD may underlie the occurrence of acute kidney injury in patients with SCD and may set the stage for the emergence of chronic kidney disease in SCD.

AbstractPurpose: Over the past two decades, there has been significant interest in targeting HER-2/neu in immune-based approaches for the treatment of HER-2/neu+ cancers. For example, peptide vaccination using a CD8 T cell–activating HER-2/neu epitope (amino acids 369-377) is an approach that is being considered in advanced phase clinical trials. Studies have suggested that the persistence of HER-2/neu–specific CD8 T cells could be improved by incorporating human leukocyte antigen (HLA) class II epitopes in the vaccine. Our goal in this study was to identify broad coverage HLA-DR epitopes of HER-2/neu, an antigen that is highly expressed in a variety of carcinomas.Experimental Design: A combination of algorithms and HLA-DR–binding assays was used to identify HLA-DR epitopes of HER-2/neu antigen. Evidence of preexistent immunity in cancer patients against the identified epitopes was determined using IFN-γ enzyme-linked immunosorbent spot (ELIspot) assay.Results: Eighty-four HLA-DR epitopes of HER-2/neu were predicted, 15 of which had high binding affinity for ≥11 common HLA-DR molecules. A degenerate pool of four HLA-DR–restricted 15-amino acid epitopes (p59, p88, p422, and p885) was identified, against which >58% of breast and ovarian cancer patients had preexistent T-cell immunity. All four epitopes are naturally processed by antigen-presenting cells. Hardy-Weinberg analysis showed that the pool is useful in ∼84% of population. Lastly, in this degenerate pool, we identified a novel in vivo immunodominant HLA-DR epitope, HER-2/neu88-102 (p88).Conclusion: The broad coverage and natural immunity to this epitope pool suggests potential usefulness in HER-2/neu–targeting, immune-based therapies such as vaccines. Clin Cancer Res; 16(3); 825–34

OBJECTIVE Little is known regarding recent changes in glitazone use. RESEARCH DESIGN AND METHODS Interrupted time series analyses of nationally representative office-visit data using IMS Health's National Disease and Therapeutic Index. RESULTS From 2003 through 2005, glitazone use increased steadily. From February 2005 to January 2007, rosiglitazone use decreased by 16% (95% CI −20 to −11) annually; pioglitazone use increased at an annual rate of 14% (9–18). During a period of Food and Drug Administration (FDA) advisories, rosiglitazone use declined sharply from 0.42 million monthly treatment visits (February 2007) to 0.13 million monthly visits (May 2008). Pioglitazone use remained stable, accounting for ∼5.8 million physician visits (77% of all glitazone use) where a treatment was used during the final 12 months of observation. CONCLUSIONS The combined effect of scientific publications, advisories, and media exposure was associated with a substantial decrease in rosiglitazone use. Despite a class-level FDA advisory, pioglitazone use was not similarly affected.