Trabecular Bone Score (TBS) and Bone Metabolism in Patients Affected with Type 1 Neurofibromatosis (NF1).

In patients with neurofibromatosis type 1 (NF1), decreased bone mineral density (BMD) and low levels of 25-hydroxy vitamin D3 (25OHD) have been reported. Recently, the trabecular bone score (TBS) measurement has been proposed as index of bone microarchitecture and fracture risk. In 74 NF1 patients (48 females, 26 males, age 41 ± 12), we measured TBS and investigated clinical stage, lifestyle, vitamin D, serum bone turnover markers, vertebral and femoral BMD. A homogenous cohort of 61 healthy subjects was used as control group. TBS was lower in NF1 patients (1.266 ± 0.113 vs. 1.346 ± 0.105) without differences between sexes. No correlations with 25OHD, low exercise, low calcium intake, reduced sun exposure, and number of skin neurofibromas were observed. As expected, hypovitaminosis D was common (98.6%), as well as BMD reduction in hip and spine sites: In NF1 patients, bone texture evaluated by TBS was low in both sexes without any correlation with clinical or metabolic parameters, suggesting a direct role of the fibromin mutation.

The role of carboxy-terminal cross-linking telopeptide of type I collagen, dual x-ray absorptiometry bone strain and Romberg test in a new osteoporotic fracture risk evaluation: A proposal from an observational study.

The consolidated way of diagnosing and treating osteoporosis in order to prevent fragility fractures has recently been questioned by some papers, which complained of overdiagnosis and consequent overtreatment of this pathology with underestimating other causes of the fragility fractures, like falls. A new clinical approach is proposed for identifying the subgroup of patients prone to fragility fractures. This retrospective observational study was conducted from January to June 2015 at the Nuclear Medicine-Bone Metabolic Unit of the of the Fondazione IRCCS Ca' Granda, Milan, Italy. An Italian population of 125 consecutive postmenopausal women was investigated for bone quantity and bone quality. Patients with neurological diseases regarding balance and vestibular dysfunction, sarcopenia, past or current history of diseases and use of drugs known to affect bone metabolism were excluded. Dual X-ray absorptiometry was used to assess bone quantity (bone mineral density) and bone quality (trabecular bone score and bone strain). Biochemical markers of bone turnover (type I collagen carboxy-terminal telopeptide, alkaline phosphatase, vitamin D) have been measured. Morphometric fractures have been searched by spine radiography. Balance was evaluated by the Romberg test. The data were evaluated with the neural network analysis using the Auto Contractive Map algorithm. The resulting semantic map shows the Minimal Spanning Tree and the Maximally Regular Graph of the interrelations between bone status parameters, balance conditions and fractures of the studied population. A low fracture risk seems to be related to a low carboxy-terminal cross-linking telopeptide of type I collagen level, whereas a positive Romberg test, together with compromised bone trabecular microarchitecture DXA parameters, appears to be strictly connected with fragility fractures. A simple assessment of the risk of fragility fracture is proposed in order to identify those frail patients at risk for osteoporotic fractures, who may have the best benefit from a pharmacological and physiotherapeutic approach.

Estimated glomerular filtration rate by serum cystatin C correlates with cardiometabolic parameters in patients with primary hyperparathyroidism.

OBJECTIVE: Patients with primary hyperparathyroidism (PHPT) are at risk of chronic kidney disease (CKD). Cystatin C (Cys-C) is considered a more reliable tool to assess glomerular filtration rate (GFR) than creatinine. The study aimed to assess circulating Cys-C and its relationships with biochemical PHPT and cardiometabolic parameters. DESIGN AND METHODS: The present cross-sectional study was performed in academic endocrine units on PHPT patients (n=190) and non-hypertensive, non-diabetic, age- and sex-matched healthy controls (n=135) with no established CKD. The main outcomes were creatinine by alkaline picrate method, Cys-C by immunonephelometry and calculation of estimated GFR based on creatinine and Cys-C (eGFRcr-cys) using the CKD-EPI equation. RESULTS: In PHPT patients, circulating Cys-C ranged 0.45-3.13  mg/l and correlated with creatinine, age and BMI. Mean Cys-C level was higher in PHPT patients than in controls (0.93±0.02 vs 0.78±0.14  mg/l; P=0.03). Cys-C levels in PHPT patients were predicted by age, BMI, ionized calcium, hypertension and HDL-cholesterol, the most significant determinant being ionized calcium. Cys-C positively correlated with cardiovascular disease (CVD) occurrence. Overall, 18.4% of PHPT patients with eGFRcr >60  ml/min per 1.73  m(2) (n=169) had Cys-C levels higher than the 95th percentile in controls (1.03  mg/l), consistent with a preclinical CKD, which was associated with hypertension and insulin resistance. Considering eGFRcr-cys, CKD (stages G3a, G3b, 4) was diagnosed in 13.7% of PHPT patients. Estimated GFRcr-cys, but not eGFR based on creatinine, was predicted by insulin resistance and hypertension and positively correlated with CVD. CONCLUSIONS: Elevated Cys-C levels were associated with ionized calcium, cardiometabolic risk factors and CVD, and identified preclinical CKD in PHPT patients.

Hypercalciuria May Persist After Successful Parathyroid Surgery and It Is Associated With Parathyroid Hyperplasia.

CONTEXT: Hypercalciuria is frequently found in primary hyperparathyroidism (1HPT) and, although it generally normalizes after successful parathyroidectomy, may persist in some patients. The factors associated with persistent calcium renal leak (cRL) have not been clarified. OBJECTIVE: The purpose of this study was to determine the prevalence of cRL in our 1HPT population and investigate cRL-related factors. DESIGN: This was a retrospective longitudinal study. SETTING: The study was conducted in an outpatient setting. PATIENTS/INTERVENTION: The participants were 95 patients with 1HPT successfully operated on who had a normal estimated glomerular filtration rate. MAIN OUTCOME MEASURES: The biochemical parameters of calcium metabolism and bone mineral density (BMD) measured by dual-X-ray absorptiometry before and 24 months after surgery were assessed. All histological findings were recorded. RESULTS: The prevalence of hypercalciuria before and after surgery was 74% and 32%, respectively. Before, surgery patients with cRL showed lower calcium and higher phosphate levels than those without cRL (10.9 ± 0.6 vs 11.4 ± 0.8 mg/dL [2.7 ± 0.2 vs 2.8 ± 0.2 mmol/L], P = .01 and 2.6 ± 0.5 vs 2.4 ± 0.4 mg/dL [0.84 ± 0.2 vs 0.77 ± 0.1 mmol/L], P = .04, respectively), whereas 24-h calciuria levels and the prevalence of 1HPT complications (osteoporosis, renal stones, and hypertension) were comparable. After surgery, serum calcium, phosphate, and PTH levels were comparable between patients with and without cRL. The prevalence of the histological finding of parathyroid hyperplasia was higher in patients with cRL (50%) than in patients without cRL (22%) (P = .01). The presence of cRL was independently associated with presurgery hypercalciuria (odds ratio, 4.71; 95% confidence interval, 1.18-18.8; P = .03) and parathyroid hyperplasia (odds ratio, 3.52; 95% confidence interval, 1.31-9.43; P = .01). Only patients without cRL had improved BMD at the spine (P = .04), total femur (P = .01), and femoral neck (P = .01). CONCLUSIONS: cRL is present in 30% of patients with 1HPT after successful surgery, and it is associated with parathyroid hyperplasia before surgery and the lack of improvement in BMD after surgery.

Hypothalamic-Pituitary Axis in Non-Functioning Pituitary Adenomas: Focus on the Prevalence of Isolated Central Hypoadrenalism.

INTRODUCTION: Non-functioning pituitary adenomas (NFPA) account for about 40% of pituitary tumors. Pituitary deficiencies are present at diagnosis in 60-80% of NFPA, and, classically, growth hormone (GH) secretion is lost first, while adrenocorticotropic hormone is expected to disappear last. The aim of this study was to evaluate the incidence of multiple or isolated pituitary deficiencies in a large series of NFPA. MATERIALS AND METHODS: We retrospectively analyzed data on 218 NFPA cases (59% females, 59% with macroadenomas, average age: 50.2 ± 17 years) followed up at our center from 1990 to 2013. At diagnosis all patients had a complete evaluation of pituitary function in basal conditions and provocative tests for the hypothalamic-pituitary-adrenal axis, while tests for GH deficiency (GHD) were carried out in 38%. RESULTS: 52.3% of patients (65.6% of macroadenomas, 33.3% of microadenomas) presented at least 1 pituitary deficiency: isolated deficiency in 29.8%, multiple deficiencies in 30% and panhypopituitarism in 9%. Isolated deficiencies were hypogonadism in 11.5% of patients (8% in micro-, 14% in macroadenomas), hypoadrenalism in 10.1% (14% in micro-, 7% in macroadenomas) and GHD in 8.3% (8.9% in micro-, 7.8% in macroadenomas). About 30% of microadenomas had at least 1 pituitary deficiency at diagnosis, independently of tumor localization within the sellar region. CONCLUSIONS: The presence of isolated hypoadrenalism suggests that the order of appearance of hypopituitarism does not always follow the one expected. Given the relatively high prevalence of isolated hypoadrenalism even in microadenomas, we suggest a full assessment of basal and dynamic pituitary function in all NFPA regardless of tumor size.

Role of IGF1-(CA)19 promoter microsatellite in the clinical presentation of acromegaly.

BACKGROUND: A highly polymorphic Cytosine-Adenosine (CA) repeat sequence microsatellite has been identified in the promoter region of IGF1 gene. Several studies investigated the relationship between IGF1-(CA)n polymorphism and IGF1 levels, with conflicting results. Aim of this study was to investigate the influence of this polymorphism on clinical and biochemical characteristics of acromegalic patients. METHODS: Eighty-eight acromegalic patients and 104 normal subjects were included in the study. Blood DNA was extracted and analysed by microsatellite technique using capillary electrophoresis. Patients and controls were subdivided in 19/19 [homozygous for the (CA)19 allele], 19/X [heterozygous for the (CA)19 allele] and X/X (any other genotype). RESULTS: The genotype frequency was significantly different between patients and controls, the proportion of 19/19 being lower (28·4% vs. 50·0%) and 19/X and X/X higher in acromegalic patients than in controls (P = 0·004). There were no significant differences in age, gender, basal and nadir GH, IGF1-SDS, tumour size, metabolic parameters, outcome and treatment among the three groups. The different frequency of genotypes in acromegalic patients vs. controls, as well as the lack of relationship between IGF1-(CA)n polymorphism and clinical and biochemical data in acromegalic patients, was confirmed using an additional alternative genotyping considering (CA)19 and (CA)20 homozygotes and heterozygotes vs. alleles with more than 19 of 20 repeats or less. CONCLUSIONS: Our results do not support the hypothesis that IGF-(CA)n alleles may have a significant role in determining clinical, biochemical and outcome of patients with acromegaly. The possible role of IGF1 polymorphism on susceptibility to acromegaly remains to be investigated.

Diagnostic features and outcome of surgical therapy of acromegalic patients: experience of the last three decades.

OBJECTIVE: Transsphenoidal (TNS) surgery remains the primary therapeutic option for GH-secreting pituitary adenomas. The aims of this study were to verify the impact of TNS surgery on treatment of acromegaly before and after identification by a dedicated neurosurgical team and to enumerate diagnostic features of the disease described over three decades. DESIGN: 41 patients (group A) who underwent TNS surgery by a dedicated neurosurgical team (2000-2008) and 126 patients (group B) operated on by surgeons not specialized in pituitary surgery (1979-1999) were retrospectively analyzed. RESULTS: No significant differences were observed between the two groups in terms of delay of diagnosis, mean basal GH levels and GH nadir values, prevalence of hypopituitarism and hypertension. IGF-I SDS were significantly higher, while prevalence of IGT/diabetes was significantly lower in group B than in group A. Overall remission rate after surgery was 58.5% for group A (75% in microadenomas and 48% in macroadenomas, P=NS) and 37% for group B (P<0.05 vs group A; for microadenomas, 34% vs 75% of group A, P<0.05, for macroadenomas, 36% vs 48% of group A, P=NS). The mean delay of diagnosis was 4.9 and 5.9 years in group A and B, respectively. CONCLUSIONS: Our data confirm that a dedicated neurosurgical team is needed in order to improve remission rates in acromegalic patients. No changes in biochemical, clinical and neuroradiological presentation of disease were observed over the last three decades. As the high prevalence of macroadenomas negatively influences surgical cure, earlier diagnosis should be considered as mandatory to achieve a better outcome.

Quantitative analysis of methylation defects and correlation with clinical characteristics in patients with pseudohypoparathyroidism type I and GNAS epigenetic alterations.

CONTEXT: Pseudohypoparathyroidism type I (PHP-I) includes two main subtypes, PHP-Ia and -Ib. About 70% of PHP-Ia patients, who show Albright hereditary osteodystrophy (AHO) associated with resistance toward multiple hormones (PTH/TSH/GHRH/gonadotropins), carry heterozygous mutations in the α-subunit of the stimulatory G protein (Gsα) exons 1-13, encoded by the guanine nucleotide binding-protein α-stimulating activity polypeptide 1 (GNAS), whereas the majority of PHP-Ib patients, who classically display hormone resistance limited to PTH and TSH with no AHO sign, have methylation defects in the imprinted GNAS cluster. Recently methylation defects have been detected also in patients with PHP and different degrees of AHO, indicating a molecular overlap between the two forms. OBJECTIVES: The objectives of the study were to collect patients with the following characteristics: clinical PHP-I (with or without AHO), no mutation in Gsα coding sequence, but the presence of GNAS methylation alterations and to investigate the existence of correlations between the degree of the epigenetic defect and the severity of the disease. PATIENTS AND METHODS: We quantified GNAS methylation alterations by both PCR-pyrosequencing and methylation specific-multiplex ligation-dependent probe amplification assay in genomic DNA from 63 patients with PHP-I and correlated these findings with clinical parameters (age at diagnosis; calcium, phosphorus, PTH, TSH levels; presence or absence of each AHO sign). RESULTS: By both approaches, the degree of the imprinting defect did not correlate with the onset of the disease, the severity of endocrine resistances, or with the presence/absence of specific AHO signs. CONCLUSIONS: Similar molecular alterations may lead to a broad spectrum of diseases, from isolated PTH resistance to complete PHP-Ia, and the degree of methylation alterations does not reflect or anticipate the severity and the type of different PHP/AHO manifestations.

Reevaluation of Acromegalic Patients in Long-Term Remission according to Newly Proposed Consensus Criteria for Control of Disease.

Acromegaly guidelines updated in 2010 revisited criteria of disease control: if applied, it is likely that a percentage of patients previously considered as cured might present postglucose GH nadir levels not adequately suppressed, with potential implications on management. This study explored GH secretion, as well as hormonal, clinical, neuroradiological, metabolic, and comorbid profile in a cohort of 40 acromegalic patients considered cured on the basis of the previous guidelines after a mean follow-up period of 17.2 years from remission, in order to assess the impact of the current criteria. At the last follow-up visit, in the presence of normal IGF-I concentrations, postglucose GH nadir was over 0.4 µg/L in 11 patients (Group A) and below 0.4 µg/L in 29 patients (Group B); moreover, Group A showed higher basal GH levels than Group B, whereas a significant decline of both GH and postglucose GH nadir levels during the follow-up was observed in Group B only. No differences in other evaluated parameters were found. These results seem to suggest that acromegalic patients considered cured on the basis of previous guidelines do not need a more intensive monitoring than patients who met the current criteria of disease control, supporting instead that the cut-off of 0.4 mcg/L might be too low for the currently used GH assay.

Bone quality, as measured by trabecular bone score, in patients with primary hyperparathyroidism.

OBJECTIVE: In primary hyperparathyroidism (PHPT), vertebral fractures (VFx) occur regardless of bone mineral density (BMD) and may depend on decreased bone quality. Trabecular bone score (TBS) is a texture measurement acquired during a spinal dual-energy X-ray absorptiometry (DXA). Recently, TBS has been proposed as an index of bone micro-architecture. DESIGN: We studied 92 PHPT patients (74 females, age 62.1±9.7 years) and 98 control subjects. In all patients at baseline, in 20 surgically treated patients and in 10 conservatively treated patients after 24 months, TBS, spinal (lumbar spine (LS)) and femoral (total hip (TH) and femoral neck (FN)) BMD were assessed by DXA and VFx by spinal radiograph. RESULTS: PHPT patients had lower TBS (-2.39±1.8) and higher VFx prevalence (43.5%) than controls (-0.98±1.07 and 8.2% respectively, both P<0.0001). TBS was associated with VFx (odds ratio 1.4, 95% CI 1.1-1.9, P=0.02), regardless of LS-BMD, age, BMI and gender, and showed a better compromise between sensitivity (75%) and specificity (61.5%) for detecting VFx than LS-BMD, TH-BMD and FN-BMD (31 and 75%, 72 and 44.2%, and 64 and 65% respectively). In surgically treated patients, TBS, LS-BMD, TH-BMD and FN-BMD increased (+47±44.8,+29.2±34.1,+49.4±48.7 and +30.2±39.3% respectively, all P<0.0001). Among patients treated conservatively, TBS decreased significantly in those (n=3) with incident VFx (-1.3±0.3) compared with those without (-0.01±0.9, P=0.048), while BMD changes were not statistically different (LS 0.3±1.2 vs -0.8±0.9 respectively, P=0.19; TH 0.4±0.8 vs -0.8±1.4 respectively, P=0.13 and FN 0.4±0.9 vs -0.8±1.4 respectively, P=0.14). CONCLUSIONS: In PHPT, bone quality, as measured by TBS, is reduced and associated with VFx and improves after surgery.

Growth hormone-releasing hormone-producing pancreatic neuroendocrine tumor in a multiple endocrine neoplasia type 1 family with an uncommon phenotype.

The objective of this study was to describe a multiple endocrine neoplasia type 1 (MEN1) family characterized by primary hyperparathyroidism, in association with acromegaly because of ectopic growth hormone-releasing hormone (GHRH) secretion by a pancreatic neuroendocrine tumor in a young man and with a bronchial carcinoid in his mother. We investigate the clinical, radiological imaging, histopathologic findings, and therapy. An 18-year-old man successfully underwent subtotal parathyroidectomy for primary hyperparathyroidism. A subsequent genetic analysis showed a MEN1 gene mutation. Three years later, acromegaly because of ectopic GHRH secretion was diagnosed (pituitary MRI negative and elevated GHRH levels). A search for an ectopic tumor was unsuccessful and somatostatin analog therapy was started. Successively, scintigraphy with somatostatin analogs (68-Ga-DOTATOC-PET) showed three focal areas in the pancreatic tail. Distal pancreatectomy showed multiple pancreatic neuroendocrine tumors and hormonal status was normalized. Afterwards, the evaluation of the patient's mother, carrying the same mutation, indicated a primary hyperparathyroidism and a 4 cm lung mass. The patient underwent subtotal pneumonectomy and the histological analysis was consistent with the diagnosis of a typical bronchial carcinoid. In conclusion, an atypical phenotype may be recorded in MEN1 families, thus emphasizing the importance of the new imaging and surgical techniques in the diagnosis and treatment of such a rare disease.

Pharmacology of the calcium sensing receptor.

Calcium sensing receptor (CASR) is a G-protein couple receptor which plays a key role in calcium homeostasis in vertebrates. Its extracellular domain is sensitive to divalent cations, aminoacids and polyamines. In parathyroid glands, CASR activation causes parathyroid hormone (PTH) reduction and subsequently a decrease in blood calcium concentration. In PTH-dependent disorders, e.g. primary and secondary hyperparathyroidism (HPT), the need for therapeutic options other than surgery led to the synthesis of various allosteric CASR agonists (calcimimetics), such as cinacalcet. Cinacalcet is the only calcimimetic approved for HPT secondary to chronic kidney disease (CDK), parathyroid carcinoma, and, in some countries, primary HPT. Clinical trials showed that cinacalcet reduced PTH and calcemia both in CDK and primary HPT, lowering the risk of bone fractures, surgery, and cardiovascular complications in the former patients. Long-term safety and pharmacoeconomics have to be fully tested yet. Few both in vitro and in vivo studies showed an association between Arg990Gly-CASR polymorphism and cinacalcet sensitivity, though in patients with severe CASR inactivating mutations the drug substantially retained its positive clinical effects. Recently, a new class of allosteric antagonists of CASR, i.e. calcilytics, has been synthesized. Calcilytics are structurally similar to calcimimetics, but exert their effects acting on a different allosteric site. Infusion of calcilytics was followed by transient rise in PTH and calcium. One of these compounds, ronacaleret, was able to increase femur BMD in post menopausal women, but with induction of mild hyperparathyroidism. In the future, calcilytics may contribute to the osteoporosis treatment choice.

MEN1-related hyperparathyroidism: response to cinacalcet and its relationship with the calcium-sensing receptor gene variant Arg990Gly.

OBJECTIVE: Primary hyperparathyroidism (PHPT) is a challenging problem in type 1 multiple endocrine neoplasia (MEN1) due to the high postsurgery recurrence rate. The aim was to evaluate the efficacy of cinacalcet in MEN1 patients in comparison with patients with sporadic PHPT (sPHPT) and the effect of Arg990Gly calcium-sensing receptor (CASR) polymorphism on the response to treatment. DESIGN: This is a randomized, crossover, double-blind study carried out in the University Hospitals. METHODS: Fifteen MEN1 patients with PHPT were randomized to two groups, one administered with 30 mg daily cinacalcet, titrated until calcium normalization, and one with placebo. After 3 months, patients were reassessed and after washout switched to the other treatment. For comparison, 20 sPHPT patients with similar calcium levels were administered with cinacalcet for 3 months. Ionized and total calcium, phosphate, and parathyroid hormone (PTH) were evaluated. CASR Arg990Gly was genotyped on blood DNA by direct sequencing. RESULTS: Cinacalcet normalized calcium, increased phosphate, and reduced PTH levels in all patients. Cinacalcet dosage required to normalize calcium in MEN1 and sPHPT was not significantly different (45±21 vs 54±25 mg/day). Few mild adverse events, not requiring drug withdrawal, were observed in both the groups. No association between Arg990Gly CASR polymorphism and response to cinacalcet was found. CONCLUSIONS: This short-term prospective study demonstrated that the efficacy profile of cinacalcet in patients with MEN1-related PHPT and in those with sPHPT was similar and was not influenced by the 990 CASR variant. Although long-term safety and efficacy data are required, cinacalcet might be considered a treatment option in MEN1 patients who have contraindications to surgery or persistent PHPT after surgery.

Influence of the d3GH receptor polymorphism on the metabolic and biochemical phenotype of GH-deficient adults at baseline and during short- and long-term recombinant human GH replacement therapy.

OBJECTIVE: A common polymorphic variant of GH receptor (exon 3 deletion, d3GHR) has been linked with increased response to recombinant human GH (rhGH) in some patients with or without GH deficiency (GHD). The aim of the study was to investigate the impact of the GHR genotype on the phenotype of GHD adults and on the metabolic effect of rhGH therapy. DESIGN: Prospective study of GHD patients evaluated before and during short- (1 year, n=100) and long-term (5 years, n=50) rhGH therapy. METHODS: Effects of rhGH on IGF1 levels, body composition (body fat percentage, BF%), body mass index, lipid profile, and glucose homeostasis (fasting insulin and glucose, insulin sensitivity indexes) were evaluated according to the presence or the absence of the d3GHR variant. RESULTS: The different genotype did not influence basal phenotype of GHD. Short-term rhGH determined normalization of IGF1 levels, decrease in BF%, and worsening of insulin sensitivity, independently from the presence of the d3GHR allele. A significant increase in high-density lipoprotein cholesterol occurred in the d3GHR group. Normalization of IGF1 levels and decrease in BF% were maintained after 5 years. Insulin sensitivity restored to basal values, though in d3GHR patients fasting glucose remained significantly higher than at baseline. After both 1 and 5 years, percentage of subjects with impaired glucose tolerance, similar in the two groups at baseline, decreased in fl/fl while doubled in d3GHR patients. In this last group, a long-term significant reduction in total and low-density lipoprotein cholesterol was also observed. CONCLUSION: The functional difference of d3GHR may influence some metabolic effects of rhGH on GHD adults.

d3-Growth hormone receptor polymorphism in acromegaly: effects on metabolic phenotype.

OBJECTIVE: A common polymorphic variant of the growth hormone receptor (GHR) is because of genomic deletion of exon 3 and has been linked with increased responsiveness to exogenous GH. The impact of this polymorphism in acromegaly, a disease characterized by endogenous excess of GH and partial loss of IGF-I feedback on tumoural GH secretion, is not clear. The aim of this study was to investigate possible influences of d3GHR on the GH/IGF-I relationship and metabolic parameters in acromegaly. DESIGN AND METHODS: Retrospective study on 76 acromegalic patients. Genotype analysis was carried out on leucocyte DNA by multiplex PCR assay. Clinical, hormonal and biochemical parameters at diagnosis were collected from patients' medical records. RESULTS: Forty-two patients (55.3%) were homozygotes for the allele encoding the full-length GHR (fl/flGHR), 27 patients were heterozygotes (fl/d3) and seven homozygotes (d3/d3) for the genomic deletion of exon 3. Heterozygotes and homozygotes for the d3 allele were considered together (d3GHR) and compared with fl/flGHR patients. d3GHR and fl/flGHR patients showed no difference in GH and IGF-I levels or in the relationship between these two parameters. Patients bearing d3GHR had a lower body mass index (BMI) than patients bearing fl/flGHR (25.8 +/- 2.1 vs. 28.1 +/- 4.8 kg/m(2), P < 0.05). Diabetes mellitus and hypertension were equally distributed, but more d3GHR patients had a normal glucose tolerance (66.7%vs. 56.3%, P < 0.05). The presence of d3GHR allele, and not BMI or age, was a significant negative predictor of insulin levels 120 min after oral glucose load (beta = -80.8, P < 0.05). CONCLUSIONS: This study supports the hypothesis that the d3GHR is functionally different from the fl/fl variant mostly for the effects on body weight regulation and on glucose metabolism.

Fetal cell microchimerism in papillary thyroid cancer: studies in peripheral blood and tissues.

Fetal cell microchimerism (FCM) is defined as the persistence, for decades after pregnancy, of fetal cells in maternal organs and circulation without any apparent rejection. We recently reported evidence, in papillary thyroid cancer (PTC) tissues, supporting a possible role of FCM in tumor damage and repair. To extend those data at the peripheral level, 106 women with a previous male pregnancy, comprising 57 with PTC and 49 healthy controls were enrolled. The presence of circulating male DNA was assessed by the amplification of the Y chromosome-specific gene SRY, with a sensitivity of 1 male cell per 1 million female cells. Moreover, to compare the microchimeric status in blood and in tumors, the neoplastic tissues of 19 women were studied. At the blood level, a significantly lower frequency of FCM was found in parous women with PTC with respect to controls (49.1% vs. 77.6%; p = 0.002). By PCR, male DNA was identified in the tumor tissues of 6 patients, and FISH analyses confirmed the presence of microchimeric cells (range 2.1-6.9 cells/section). In some patients, FCM was negative in the blood, whereas microchimeric cells were identified in the tumor. In conclusion, the prevalence of FCM in peripheral blood was found to be significantly lower in patients than in healthy controls. The presence of microchimeric cells in the tumors, but not at the peripheral level, supports the hypothesis that fetal cells could reside in maternal niches and could be recruited to diseased areas, where they could differentiate to regenerate damaged tissues.

Two novel nonsense mutations in GALNT3 gene are responsible for familial tumoral calcinosis.

Ectopic periarticular calcifications associated with elevated levels of serum phosphate represent the principal clinical features of hyperphosphatemic familial tumoral calcinosis (HFTC), a rare autosomal recessive metabolic disorder. The disease can be caused by recessive mutations in at least two different genes: GalNAc transferase 3 (GALNT3), encoding a glycosyltransferase that initiates mucin-type O-glycosylation, and fibroblast growth factor 23 (FGF23), which encodes a regulator of phosphate circulating levels. In the current study, we performed mutation analyses of the GALNT3 gene in a subject with HFTC and in his relatives. Sequence analyses revealed that the proband was a compound heterozygote for two novel nonsense mutations in exon 4 (Y322X) and in exon 7 (Q481X). Cosegregation of the mutations with the disease within the family was confirmed by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis. This is the first report describing the simultaneous presence of two different stop codons in the coding sequence of the GALNT3 gene.

Biallelic expression of the Gsalpha gene in human bone and adipose tissue.

Mutations of the Gsalpha gene inherited from the mother lead to pseudohypoparathyroidism (PHP) type Ia (PHP Ia), in which Albright's hereditary osteodistrophy is associated to resistance to the action of different hormones, whereas the same mutations inherited from the father lead to isolated Albright's hereditary osteodistrophy [pseudo-PHP (PPHP)]. Accordingly, it has been suggested that Gsalpha is under tissue-specific imprinting control, and recent studies provided evidence for a predominant maternal origin of Gsalpha transcripts in different endocrine organs involved in the PHP Ia phenotype. To establish whether Gsalpha is imprinted also in tissues that are site of alteration both in PHP Ia and PPHP, we selected 20 bone and 10 adipose tissue samples, which were heterozygous for a known polymorphism in exon 5. Expression from both parental alleles was evaluated by RT-PCR and enzymatic digestion of the resulting fragments. By this approach, the great majority of the samples analyzed showed an equal expression of the two alleles. Our results provide evidence for the absence of Gsalpha imprinting in human bone and fat and suggest that the clinical finding of osteodystrophy and obesity in PHP Ia and PPHP patients despite the presence of a normal Gsalpha allele is likely due to Gsalpha haploinsufficiency in these tissues.

Cyclic nucleotide phosphodiesterases and their role in endocrine cell signaling.

The discovery that degradation and inactivation of the second messengers cAMP and cGMP are mediated by a complex enzymatic machinery has changed our perspective on cyclic nucleotide-mediated processes. In the cell, these second messengers are inactivated by no fewer than 11 distinct families of phosphodiesterases (PDEs). Much is known about the structure and function of these enzymes, their complex subcellular distribution and regulation. Yet, their potential as targets for therapeutic intervention in a broad range of endocrine abnormalities still needs to be investigated. This review explores the involvement of PDEs in the regulation of intracellular signaling and focuses on the known and potential roles that are of interest to endocrinologists.