Sudhaker D. Rao

Development of a novel immunoradiometric assay exclusively for biologically active whole parathyroid hormone 1-84: Implications for improvement of accurate assessment of parathyroid function

We developed a novel immunoradiometric assay (IRMA; whole parathyroid hormone [PTH] IRMA) for PTH, which specifically measures biologically active whole PTH(1–84). The assay is based on a solid phase coated with anti‐PTH(39–84) antibody, a tracer of125I‐labeled antibody with a unique specificity to the first N‐terminal amino acid of PTH(1–84), and calibrators of diluted synthetic PTH(1–84). In contrast to the Nichols intact PTH IRMA, this new assay does not detect PTH(7–84) fragments and only detects one immunoreactive peak in chromatographically fractionated patient samples. The assay was shown to have an analytical sensitivity of 1.0 pg/ml with a linear measurement range up to 2300 pg/ml. With this assay, we further identified that the previously described non‐(1–84)PTH fragments are aminoterminally truncated with similar hydrophobicity as PTH(7–84), and these PTH fragments are present not only in patients with secondary hyperparathyroidism (2°‐HPT) of uremia, but also in patients with primary hyperparathyroidism (1°‐HPT) and normal persons. The plasma normal range of the whole PTH(1–84) was 7–36 pg/ml (mean ± SD: 22.7 ± 7.2 pg/ml, n = 135), whereas over 93.9% (155/165) of patients with 1°‐HPT had whole PTH(1–84) values above the normal cut‐off. The percentage of biologically active whole PTH(1–84) (pB%) in the pool of total immunoreactive “intact” PTH is higher in the normal population (median: 67.3%; SD: 15.8%; n = 56) than in uremic patients (median:53.8%; SD: 15.5%; n = 318; p < 0.001), although the whole PTH(1–84) values from uremic patients displayed a more significant heterogeneous distribution when compared with that of 1°‐HPT patients and normals. Moreover, the pB% displayed a nearly Gaussian distribution pattern from 20% to over 90% in patients with either 1°‐HPT or uremia. The specificity of this newly developed whole PTH(1–84) IRMA is the assurance, for the first time, of being able to measure only the biologically active whole PTH(1–84) without cross‐reaction to the high concentrations of the aminoterminally truncated PTH fragments found in both normal subjects and patients. Because of the significant variations of pB% in patients, it is necessary to use the whole PTH assay to determine biologically active PTH levels clinically and, thus, to avoid overestimating the concentration of the true biologically active hormone. This new assay could provide a more meaningful standardization of future PTH measurements with improved accuracy in the clinical assessment of parathyroid function.

Effects of Age and Serum 25-OH-Vitamin D on Serum Parathyroid Hormone Levels

CONTEXT Several studies define optimal serum 25-hydroxyvitamin D (25-OHD) levels based on serum PTH level reaching an asymptote. However, results differ widely, ranging from 25-OHD levels of 12-44 ng/ml: many studies are constrained by small sample size. OBJECTIVE The objective of the study was to determine the relationship between serum PTH and 25-OHD levels and age in a very large reference laboratory database. DESIGN This was a detailed cross-sectional analysis of 312,962 paired serum PTH and 25-OHD levels measured from July 2010 to June 2011. RESULTS Median PTH levels and the proportion of patients (PTH > 65 pg/ml), from 63 successive 25-OHD frequency classes of 5000 patients, provide smooth, exceptionally well-fitted curves (R(2) = 0.994 and R(2) = 0.995, respectively) without discernible inflection points or asymptotes but with striking age dependencies. Serum 25-OHD was below the recent Institute of Medicine sufficiency guidance of 20 ng/ml in 27% (85,000) of the subjects. More importantly, 40 and 51% of subjects (serum 25-OHD <20 and 10 ng/ml, respectively) had biochemical hyperparathyroidism (PTH > 65 pg/ml). CONCLUSIONS This analysis, despite inevitable inherent limitations, introduces several clinical implications. First, median 25-OHD-dependent PTH levels revealed no threshold above which increasing 25-OHD fails to further suppress PTH. Second, the large number of subjects with 25-OHD deficiency and hyperparathyroidism reinforces the Third International Workshop on Asymptomatic Primary Hyper parathyroidisms recommendations to test for, and replete, vitamin D depletion before considering parathyroidectomy. Third, strong age dependency of the PTH-25-OHD relationship likely reflects the composite effects of age-related decline in calcium absorption and renal function. Finally, this unselected large population database study could guide clinical management of patients based on an age-dependent, PTH-25-OHD continuum.

Reduced vitamin D receptor expression in parathyroid adenomas: implications for pathogenesis

Parathyroid adenomas discovered fortuitously grow very slowly and their cell birth rate greatly declines, features explicable by an initial increase in secretory set‐point. In the nodules of severe uraemic parathyroid hyperplasia, there is an increased set‐point and decreased expression of both the calcium sensing receptor (CaSR) and the vitamin D receptor (VDR). Accordingly, we examined VDR and CaSR expression in parathyroid adenomas.

Methylation status of the CpG islands in vitamin d and calcium-sensing receptor gene promoters does not explain the reduced gene expressions in parathyroid adenomas

AIM The exact mechanism causing decreased expression of the vitamin D receptor (VDR) and calcium-sensing receptor (CASR) genes in parathyroid adenoma is not known, but methylation of promoter regions is often detected during epigenetic downregulation of gene expression. We investigated whether epigenetic silencing is involved in the decreased expression of VDR and CASR. EXPERIMENTAL DESIGN Real-time PCR and immunohistochemistry confirmed the downregulation of the VDR and CASR genes at transcriptional and translational levels. Bisulfite-converted DNA samples from parathyroid adenomas with control samples were analyzed for methylation in the promoter region of VDR and CASR genes. RESULTS There was no significant methylation in the promoter regions of VDR and CASR genes in parathyroid adenomatous tissues. CONCLUSIONS Methylation-mediated silencing of VDR and CASR promoter does not appear to be associated with reduced expression, indicating the involvement of other factors in specific suppression of VDR and CASR in parathyroid adenomas.

A Longitudinal Study of Skeletal Histomorphometry at 6 and 24 Months Across Four Bone Envelopes in Postmenopausal Women With Osteoporosis Receiving Teriparatide or Zoledronic Acid in the SHOTZ Trial.

Previously, we reported the effects of teriparatide (TPTD) and zoledronic acid (ZOL) on bone formation based on biochemical markers and bone histomorphometry of the cancellous envelope at month 6 in postmenopausal women with osteoporosis who participated in the 12‐month primary Skeletal Histomorphometry in Subjects on Teriparatide or Zoledronic Acid Therapy (SHOTZ) study. Patients were eligible to enter a 12‐month extension on their original treatment regimen: TPTD 20 μg/day (s.c. injection) or ZOL 5 mg/year (i.v. infusion). A second biopsy was performed at month 24. Here we report longitudinal changes between and within each treatment group in the cancellous, endocortical, intracortical, and periosteal bone envelopes in patients with evaluable biopsies at months 6 and 24 (paired data set: TPTD, n = 10; ZOL, n = 9). Between‐group differences are also reported in the larger set of patients with evaluable biopsies at month 6 (TPTD, n = 28; ZOL, n = 30). Data from the cancellous envelope at month 6 or month 24 provided a reference to compare differences across envelopes within each treatment group. The 24‐month results extend our earlier report that TPTD and ZOL possess different tissue‐level mechanisms of action. Moreover, these differences persisted for at least 2 years in all four bone envelopes. Few longitudinal differences were observed within or across bone envelopes in ZOL‐treated patients, suggesting that the low bone formation indices at month 6 persisted to month 24. Conversely, the magnitude of the effect of TPTD on bone formation varied across individual envelopes: median values for mineralizing surface (MS/BS) and bone formation rate (BFR/BS) at month 6 were approximately 3‐fold to 5‐fold higher in the endocortical and intracortical envelopes compared to the cancellous envelope. Although MS/BS and BFR/BS declined in these envelopes at month 24, median values continued to exceed, or were not significantly different from, those in the cancellous envelope. This study demonstrates for the first time that bone formation indices are higher with TPTD treatment than with ZOL in all four bone envelopes and the difference persists for at least 2 years. Moreover, the magnitude of the effect of TPTD in cortical bone remains robust at 24 months.

Sclerostin: Recent advances and clinical implications

Purpose of reviewDiscovery of the Wnt signaling pathway and understanding the central role of osteocyte in skeletal homeostasis have been the major advances in skeletal biology over the past decade. Sclerostin, secreted mainly (but not exclusively) by osteocytes, has emerged as a key player in skeletal homeostasis. This review highlights the most relevant recent advances. Recent findingsSclerostin by inhibiting Wnt signaling pathway decreases bone formation and osteoblast differentiation and promotes osteoblast apoptosis. Ability to measure serum sclerostin levels better clarified the role of sclerostin in various physiologic and pathologic states. Early clinical trials with antibodies to sclerostin have produced robust increases in bone mineral density, and fracture prevention trials are underway. SummarySince the discovery of Wnt signaling pathway and sclerostins association with high bone mass, there has been a remarkable progress. Clinical trials with fracture endpoints, already underway, should expand osteoanabolic therapeutic horizon in the very near future. Measurement of sclerostin levels in a number of conditions has advanced our knowledge about pathophysiology of skeletal and nonskeletal disorders in an altogether new light.

Differential Effects of Teriparatide and Denosumab on Intact PTH and Bone Formation Indices: AVA Osteoporosis Study.

We compared effects of teriparatide and denosumab on PTH, bone turnover markers, and bone histomorphometry in osteoporotic postmenopausal women. The findings were inconsistent with an early indirect anabolic effect of denosumab.

Comparative Effectiveness of Ultrasonography, 99mTc-Sestamibi, and 18F-Fluorocholine PET/CT in Detecting Parathyroid Adenomas in Patients With Primary Hyperparathyroidism

Purpose Accurate preoperative localization of parathyroid lesion(s) is crucial for successful surgical management of primary hyperparathyroidism (PHPT). This study was conducted to compare the effectiveness of ultrasonography (USG) of the neck, 99mTc-sestamibi (MIBI) imaging with or without SPECT/CT, and 18F-fluorocholine (FCH) PET/CT imaging in the preoperative localization of parathyroid lesions in patients with PHPT. Methods Fifty-four consecutive patients with PHPT were included in this prospective study who underwent preoperative localization of the parathyroid lesion(s) using 3 diagnostic modalities followed by surgery. The sensitivity, positive predictive value, and accuracy of the 3 imaging procedures to accurately detect abnormal parathyroid glands were determined using histopathology as criterion standard with postoperative biochemical response confirmation. Results 18F-fluorocholine PET/CT detected 52 of 54 patients and 52 of 56 lesions with histopathologically proven parathyroid adenomas on patient-based and lesion-based analysis, respectively. Preoperative USG, MIBI, and FCH PET/CT localized abnormal parathyroid gland(s) in 39 (72.2%), 43 (79.6%), and 54 (100%) patients, respectively. The sensitivity and positive predictive value were 69.3% and 87.1% for USG, 80.7% and 97.6% for MIBI, and 100% and 96.3% for FCH PET/CT. The accuracy was 62.9%, 79.6%, and 96.3% for USG, MIBI, and FCH PET/CT, respectively, in patient-wise analysis. In 6 patients with ectopic lesions, FCH PET/CT demonstrated higher sensitivity and accuracy than MIBI and USG (100% vs 66.6% and 16.7%, respectively). Conclusions Among the 3 imaging techniques tested simultaneously, FCH PET/CT was superior for accurate preoperative localization of parathyroid adenomas, especially for ectopic or small parathyroid lesions.

Parathyroid Growth: Normal and Abnormal

The functional performance of every endocrine gland requires delivery of the right number of hormone molecules into the circulation during each successive interval of time. Total hormone secretion comprises the aggregate contribution of each cell and so depends not only on the average secretion per cell, but also on the number of contributing cells. The regulation of cell number should receive as much attention as the regulation of individual cell behavior, but in practice it is almost entirely ignored. Cell number is never considered explicitly in the description of feedback relationships. There are several reasons for this neglect. The rules of development normally ensure that each organ grows to the right size, so that adult cell number varies only over a two- to three-fold range. By contrast, hormone secretion by many endocrine glands can vary acutely over as much as a 100-fold range, which must reflect changes in the performance of individual cells. Another reason is that the endocrine glands are among the smallest of organs.

Deliberate Total Parathyroidectomy: A Potentially Novel Therapy for Tumor-Induced Hypophosphatemic Osteomalacia

BACKGROUND Tumor-induced osteomalacia (TIO) is an acquired hypophosphatemic metabolic bone disorder that can be cured by removing or ablating the offending tumor. However, when the tumor cannot be localized, lifelong therapy with oral phosphate and calcitriol or cinacalcet with close monitoring is required. CASE REPORT A 56-year-old man was diagnosed with TIO in 1990. Initial therapy consisted of oral phosphate and calcitriol with symptomatic and biochemical improvement and healing of osteomalacia. Eight years later, hypercalcemic hyperparathyroidism developed, requiring subtotal parathyroidectomy with a transient increase in serum phosphate and normalization of serum calcium and PTH. Recurrent hypercalcemic hyperparathyroidism developed after 10 years of medical therapy. A deliberate total parathyroidectomy produced a prompt rise in serum phosphate into the normal range > 3.0 mg/dL and remained normal during the next 4 years of follow-up, despite continued very high serum fibroblast growth factor-23 levels throughout the 23-year follow-up. CONCLUSION We report an unusual case of a TIO patient with long-term follow-up who developed recurrent hypercalcemic hyperparathyroidism on long-term oral phosphate therapy. Deliberate total parathyroidectomy normalized serum phosphate despite persistently elevated fibroblast growth factor-23 levels. Total parathyroidectomy offers a potentially novel therapy in some patients with TIO in whom medical therapy is not feasible or the tumor is unresectable.