NT5E DNA methylation in papillary thyroid cancer: Novel opportunities for precision oncology.

The ectoenzyme CD73, encoded by the NT5E gene, has emerged as a potential prognostic and therapeutic marker for papillary thyroid carcinoma (PTC), which has increased in incidence in recent decades. Here, from The Cancer Genome Atlas Thyroid Cancer (TCGA-THCA) database, we extracted and combined clinical features, levels of NT5E mRNA, and DNA methylation of PTC samples and performed multivariate and random forest analyses to evaluate the prognostic relevance and the potential of discriminating between adjacent non-malignant and thyroid tumor samples. As a result, we revealed that lower levels of methylation at the cg23172664 site were independently associated with BRAF-like phenotype (p = 0.002), age over 55 years (p = 0.012), presence of capsule invasion (p = 0.007) and presence of positive lymph node metastasis (LNM) (p = 0.04). The methylation levels of cg27297263 and cg23172664 sites showed significant and inversely correlations with levels of NT5E mRNA expression (r = -0.528 and r = -0.660, respectively), and their combination was able to discriminate between adjacent non-malignant and tumor samples with a precision of 96%-97% and 84%-85%, respectively. These data suggest that combining cg23172664 and cg27297263 sites may bring new insights to reveal new subsets of patients with papillary thyroid carcinoma.

The gene expression of GPER1 is low in fresh samples of papillary thyroid carcinoma (PTC), and in silico analysis.

Papillary thyroid cancer (PTC), whose incidence has been increasing in the last years, occurs more frequently in women. Experimental studies suggested that estrogen could be an important risk factor for the higher female incidence. In fact, it has been demonstrated that 17ß-estradiol (E2) could increase proliferation and dedifferentiation in thyroid follicular cells. Genomic estrogen responses are typically mediated through classical estrogen receptors, the α and ß isoforms, which have been described in normal and abnormal human thyroid tissue. Nevertheless, effects mediated through G protein estrogen receptor 1 (GPR30/GPER/GPER1), described in some thyroid cancer cell lines, could be partially responsible for the regulation of growth in normal cells. In this study, GPER1 gene and protein expression are described in non-malignant and in papillary thyroid cancer (PTC), as well as its association with clinical features of patients with PTC. The GPER1 expression was lower in PTC as compared to paired non-malignant thyroid tissues in fresh samples of PTC and in silico analysis of GEO and TCGA databases. In PTC cases of TCGA database, low GPER1 mRNA expression was independently associated with metastatic lymph nodes, female gender, and BRAF mutation. Besides, GPER1 mRNA levels were positively correlated with mRNA levels of thyroid differentiation genes. These results support the hypothesis that GPER1 have a role in PTC tumorigenesis and might be a potential target for its therapy. Further studies are needed to determine the functionality of these receptors in normal and diseased thyroid.

Seasonal variation of vitamin D among healthy adult men in a subtropical region.

OBJECTIVE: To evaluate seasonal variation of 25(OH)vitamin D [25(OH)D3] levels, and factors associated with it, in healthy adult men, who exercised outdoors for 50 min., at least twice a week, from 10AM to 4PM, in a Brazilian semitropical region. METHODS: Blood samples were collected at the end of each season for 25(OH)D3, measured by liquid chromatography with tandem mass spectrometry. Ultraviolet irradiation was estimated by radiometer, calculating the daily photobiological response to vitamin D synthesis in human skin (D-VitD). The prevalence of 25(OH)D3 <20ng/mL changed with the seasons (p=0.000): 8.7% (n=6/69), 1.5% (n=1/66), 0 (n=0/64), and 21.7% (n=13/60), respectively, at the end of winter, spring, summer, and autumn. The prevalence, adjusted for multiple comparisons, was higher in winter than summer (p=0.026), and in autumn than spring (p=0.001) and summer (p=0.000). There were no associations of 25(OH) D3 levels with BMI (p=0.207), body fat (p=0.064), and phototype (p=0.485), in univariate analysis. It was associated with D-VitD in the 30 days before blood sampling (p=0.000), after adjustment to body fat. The prevalence of 25(OH)D3 <30ng/mL varied seasonally (p=0.000): 69.6% (n=48/69), 68.2% (n=45/66), 43.8% (n=28/64), and 88.4% (n=53/60), respectively, in winter, spring, summer, and autumn. CONCLUSIONS: In a Brazilian subtropical region, a seasonal variation in 25(OH)D3 was observed in healthy adult males, although they spent at least 50 min outdoors twice a week, wearing shorts and T-shirts. 25(OH)D3 <20ng/mL was 21.7% in autumn; D-vitD 30 days prior to blood sampling was the only factor independently associated with 25(OH)D3 levels.

Seasonal variation of vitamin D among healthy adult men in a subtropical region

SUMMARY OBJECTIVE: To evaluate seasonal variation of 25(OH)vitamin D [25(OH)D3] levels, and factors associated with it, in healthy adult men, who exercised outdoors for 50 min., at least twice a week, from 10AM to 4PM, in a Brazilian semitropical region. METHODS: Blood samples were collected at the end of each season for 25(OH)D3, measured by liquid chromatography with tandem mass spectrometry. Ultraviolet irradiation was estimated by radiometer, calculating the daily photobiological response to vitamin D synthesis in human skin (D-VitD). The prevalence of 25(OH)D3 <20ng/mL changed with the seasons (p=0.000): 8.7% (n=6/69), 1.5% (n=1/66), 0 (n=0/64), and 21.7% (n=13/60), respectively, at the end of winter, spring, summer, and autumn. The prevalence, adjusted for multiple comparisons, was higher in winter than summer (p=0.026), and in autumn than spring (p=0.001) and summer (p=0.000). There were no associations of 25(OH) D3 levels with BMI (p=0.207), body fat (p=0.064), and phototype (p=0.485), in univariate analysis. It was associated with D-VitD in the 30 days before blood sampling (p=0.000), after adjustment to body fat. The prevalence of 25(OH)D3 <30ng/mL varied seasonally (p=0.000): 69.6% (n=48/69), 68.2% (n=45/66), 43.8% (n=28/64), and 88.4% (n=53/60), respectively, in winter, spring, summer, and autumn. CONCLUSIONS: In a Brazilian subtropical region, a seasonal variation in 25(OH)D3 was observed in healthy adult males, although they spent at least 50 min outdoors twice a week, wearing shorts and T-shirts. 25(OH)D3 <20ng/mL was 21.7% in autumn; D-vitD 30 days prior to blood sampling was the only factor independently associated with 25(OH)D3 levels.

RESUMO OBJETIVOS: Avaliar a sazonalidade da 25(OH)vitamina D3 [25(OH)D3] e fatores associados em homens adultos saudáveis, que se exercitavam ao ar livre pelo menos 50 min duas vezes por semana, das 10 às 16h, em uma região subtropical. MÉTODOS: Sangue foi colhido no fim das estações para medir 25(OH)D3, por cromatografia líquida em tandem com espectroscopia de massas. A radiação ultravioleta foi estimada por radiômetro, calculando diariamente a resposta fotobiológica para sintetizar vitamina D na pele humana (D-VitD). RESULTADOS: A prevalência de 25(OH)D3 <20ng/mL foi sazonal (p=0.000): 8.7% (n=6/69), 1.5% (n=1/66), 0% (n= 0/64), e 21.7% (n=13/60), respectivamente, no final do inverno, primavera, verão e outono. A prevalência, ajustada para comparações múltiplas, foi maior no inverno do que no verão (p=0.026) e no outono do que na primavera (p=0.001) e verão (p=0.000). A 25(OH)D3 não se associou com o índice de massa corporal (p=0.207), gordura corporal (p=0.064) ou fototipo (p=0.485), na análise univariada. Associou-se à D-VitD nos 30 dias antes da coleta de sangue (p=0.000), ajustada para gordura corporal. Houve sazonalidade na prevalência de 25(OH)D3 <30ng/mL (p=0.000): 69.6% (n=48/69), 68.2% (n=45/66), 43.8% (n=28/64), e 88.4% (n=53/60), respectivamente, no inverno, primavera, verão e outono. CONCLUSÕES: Em uma região subtropical, houve sazonalidade na 25(OH)D3 em homens adultos, saudáveis, embora se exercitassem ao ar livre pelo menos 50 minutos duas vezes por semana, usando shorts e camiseta. 25(OH)D3 <20ng/mL foi 21.7% no outono e a D-vitD 30 dias antes da coleta do sangue foi o único fator associado de modo independente à 25(OH)D3.

GPER1 in the thyroid: A systematic review.

BACKGROUND: Thyroid cancer incidence has been increasing, acquiring a greater importance in health, especially of women, who are more frequently affected. As 17-ß-estradiol (E2) has been shown to have a proliferative effect on benign and malignant thyroid cells, G protein-coupled estrogen receptor (GPER1) could have a role on the pathogenesis of thyroid cancer. OBJECTIVE: To evaluate data on GPER1 in the thyroid. DATA SOURCES: PubMed, Scielo and Cochrane Library databases were searched, using the keywords GPER1 or GPR30 or GPER and thyroid, since the inception until Jun, 2019. Other sources were used, as cross-referencing. STUDY SELECTION: All studies which evaluated GPER1 GPER1 or GPR30 or GPER in the thyroid. DATA EXTRACTION: From 23 articles identified, eight studies were included: one in commercial samples of human thyroid, four in human thyroid cancer cell lines, and three in human samples of benign and/or malignant thyroid diseases. DATA SYNTHESIS: GPER1 gene and protein expression were described, respectively, in six and five studies, and the results varied according to the study. In three studies, increased proliferation of four thyroid cancer cell lines were induced by E2, with evidences suggesting that GPER1 at least partially mediated growth in these cells. GPER1 was identified in the cell membrane, in three studies, and in the cytoplasm in two studies. CONCLUSIONS: The paucity of studies about GPER1 in the thyroid, as well as methodological differences between them, precludes firm conclusions about GPER1 role in the thyroid, although there are some evidences of GPER1-induced proliferation of thyroid cancer cells.

Activity of ecto-5'-nucleotidase (NT5E/CD73) is increased in papillary thyroid carcinoma and its expression is associated with metastatic lymph nodes.

The incidence of papillary thyroid carcinoma (PTC) has been increasing, which raised the interest in its molecular pathways. Although the high expression of ecto-5'-nucleotidase (NT5E) gene expression and NT5E enzymatic activity in several types of cancer is associated with tumor progression, its role in PTC remains unknown. Here, we investigated the AMP hydrolysis in human normal thyroid cells and PTC cells, in primary culture, and the association of NT5E expression with clinical aspects of PTC patients. AMPase activity was higher in thyroid cells isolated from PTC, as compared to normal thyroid (P = 0.0063). Significant correlation was observed between AMPase activity and NT5E levels in primary thyroid cell cultures (r = 0.655, P = 0.029). NT5E expression was higher in PTC than in the adjacent non-malignant thyroid tissue (P = 0.0065) and were positively associated with metastatic lymph nodes (P = 0.0007), risk of recurrence (P = 0.0033), tumor size (P = 0.049), and nodular hyperplasia in the adjacent thyroid parenchyma, when compared to normal thyroid or lymphocytic thyroiditis (P = 0.0146). After adjusting for potential confounders, the malignant/non-malignant paired expression ratio of NT5E mRNA was independently associated with metastatic lymph nodes (P = 0.0005), and tumor size (P=0.0005). In addition, the analysis of PTC described in the TCGA database also showed an association between higher expression of NT5E and metastatic lymph nodes, and tumor microinvasion. These results support the hypothesis that NT5E have a role in PTC microenvironment and might be a potential target for PTC therapy.

Extracellular ATP is Differentially Metabolized on Papillary Thyroid Carcinoma Cells Surface in Comparison to Normal Cells.

The incidence of differentiated thyroid cancer has been increasing. Nevertheless, its molecular mechanisms are not well understood. In recent years, extracellular nucleotides and nucleosides have emerged as important modulators of tumor microenvironment. Extracellular ATP is mainly hydrolyzed by NTPDase1/CD39 and NTPDase2/CD39L1, generating AMP, which is hydrolyzed by ecto-5'-nucleotidase (CD73) to adenosine, a possible promoter of tumor growth and metastasis. There are no studies evaluating the expression and functionality of these ectonucleotidases on normal or tumor-derived thyroid cells. Thus, we investigated the ability of thyroid cancer cells to hydrolyze extracellular ATP generating adenosine, and the expression of ecto-enzymes, as compared to normal cells. We found that normal thyroid derived cells presented a higher ability to hydrolyze ATP and higher mRNA levels for ENTDP1-2, when compared to papillary thyroid carcinoma (PTC) derived cells, which had a higher ability to hydrolyze AMP and expressed CD73 mRNA and protein at higher levels. In addition, adenosine induced an increase in proliferation and migration in PTC derived cells, whose effect was blocked by APCP, a non-hydrolysable ADP analogue, which is an inhibitor of CD73. Taken together, these results showed that thyroid follicular cells have a functional purinergic signaling. The higher expression of CD73 in PTC derived cells might favor the accumulation of extracellular adenosine in the tumor microenvironment, which could promote tumor progression. Therefore, as already shown for other tumors, the purinergic signaling should be considered a potential target for thyroid cancer management and treatment.

Intestinal absorption of vitamin D: a systematic review.

Context: Vitamin D is frequently prescribed as a supplement, yet its absorption remains poorly understood. Objective: This systematic review was performed to evaluate data on mechanisms involved in the intestinal absorption of vitamin D. Data Sources: PubMed, Embase, and Cochrane Library databases were searched. Study Selection: The following studies were included: experimental laboratory studies of vitamin D absorption through the enterocyte brush-border membrane; absorption tests that used radiolabeled vitamin D; and clinical trials in adults that investigated a single dose of cholecalciferol or ergocalciferol and reported at least 2 measurements of serum cholecalciferol, ergocalciferol, or 25-hydroxyvitamin D. Data Extraction: From 2069 articles identified, 46 met the inclusion criteria. Results: Different methods were employed to evaluate vitamin D absorption. Recent research suggests that vitamin D absorption is not an exclusive simple diffusion process. Vitamin D was better absorbed when it was consumed with fat-containing meals, but absorption also occurred without fat or oily vehicles. Factors that modified cholesterol absorption also altered vitamin D absorption. Conclusion: Vitamin D is probably absorbed through passive diffusion and a mechanism involving membrane carriers, especially cholesterol transporters, although data remain scarce. Some data suggest that fat, when consumed concomitantly with vitamin D, improves vitamin D absorption.

Aberrant Activation of Notch Signaling Inhibits PROX1 Activity to Enhance the Malignant Behavior of Thyroid Cancer Cells.

Papillary thyroid cancer (PTC) is one of the most common endocrine malignancies associated with significant morbidity and mortality. Although multiple studies have contributed to a better understanding of the genetic alterations underlying this frequently arising disease, the downstream molecular effectors that impact PTC pathogenesis remain to be further defined. Here, we report that the regulator of cell fate specification, PROX1, becomes inactivated in PTC through mRNA downregulation and cytoplasmic mislocalization. Expression studies in clinical specimens revealed that aberrantly activated NOTCH signaling promoted PROX1 downregulation and that cytoplasmic mislocalization significantly altered PROX1 protein stability. Importantly, restoration of PROX1 activity in thyroid carcinoma cells revealed that PROX1 not only enhanced Wnt/ß-catenin signaling but also regulated several genes known to be associated with PTC, including thyroid cancer protein (TC)-1, SERPINA1, and FABP4. Furthermore, PROX1 reexpression suppressed the malignant phenotypes of thyroid carcinoma cells, such as proliferation, motility, adhesion, invasion, anchorage-independent growth, and polyploidy. Moreover, animal xenograft studies demonstrated that restoration of PROX1 severely impeded tumor formation and suppressed the invasiveness and the nuclear/cytoplasmic ratio of PTC cells. Taken together, our findings demonstrate that NOTCH-induced PROX1 inactivation significantly promotes the malignant behavior of thyroid carcinoma and suggest that PROX1 reactivation may represent a potential therapeutic strategy to attenuate disease progression.

An unexpected cause of severe and refractory PTH-independent hypercalcemia: case report and literature review.

Hypercalcemia is a common condition in the internal medicine practice. Sometimes its cause is not readily apparent, so extensive investigation is appropriate. Here we report an unexpected cause for hypercalcemia in an elderly woman. The case of an 82-years old woman with PTH-independent hypercalcemia, lymphocytosis, normal serum 1,25 (OH)vitamin D levels, and low serum PTHrp levels, is described. Medical history and complementary investigation were unremarkable, except for increased metabolic activity in the glutei regions, as measured by whole body 18F-FDG PET-CT. Reviewing her medical history, her sister recalled that she had been submitted to intramuscular methylmethacrylate injections, for cosmetic purposes, five years before presentation, which was confirmed by muscle biopsy. Low calcium intake, parenteral bisphosphonates, calcitonin, and glucocorticoids were used to control serum calcium levels. Methylmethacrylate injections, used cosmetically, are a new cause for hypercalcemia, even after many years. Hypercalcemia was probably due to calcitriol overproduction in foreign body granulomas. Persistent reactive lymphocytosis could be a clue for this inflammatory cause of hypercalcemia.

Impact of a cholesterol membrane transporter's inhibition on vitamin D absorption: A double-blind randomized placebo-controlled study.

Oral supplements are important to prevent and treat vitamin D deficiency. Despite the growing number of prescriptions, vitamin D's absorptive mechanisms are not clearly elucidated. By evaluating the effect of ezetimibe on vitamin D absorption, we aim to determine if the cholesterol transporter Niemann-Pick C1-Like 1 transporter contributes to it. This randomized, double-blind, placebo-controlled trial (ClinicalTrials.govNCT02234544) was developed in a South Brazilian University Hospital. Fifty-one medical students were randomized to ezetimibe 10mg/day or placebo for 5 days. On the fifth and 19th days, blood samples for 25-hydroxycholecalciferol (25OHD), parathyroid hormone (PTH), calcium, and albumin were collected. After the first blood sample collection, all participants received a single oral 50,000 IU cholecalciferol dose during a 15 g-fat meal. Serum 25OHD levels were measured by the immunoassay Diasorin Liaison®. Measurements were compared in a general linear model adjusted for multiple comparisons by the Bonferroni test. Before cholecalciferol administration, 25OHD was <30 ng/mL and <20 ng/mL, respectively, in all and in 82.3% of the participants. Fourteen days after a single 50,000 IU oral dose of cholecalciferol, mean (SD) changes in serum 25OHD were similar in both groups, after adjustment to BMI and 25OHD levels before cholecalciferol administration (p=0.26): 8.7 (3.7) ng/mL in the ezetimibe group, versus 10.0 (3.8) ng/mL in the placebo group. Mean serum 25OHD, PTH, calcium and albumin levels remained similar in both groups. We conclude that ezetimibe had no effect on the mean change in serum 25OHD after a single oral dose of cholecalciferol, in these healthy and young adults.

Progesterone Upregulates Gene Expression in Normal Human Thyroid Follicular Cells.

Thyroid cancer and thyroid nodules are more prevalent in women than men, so female sex hormones may have an etiological role in these conditions. There are no data about direct effects of progesterone on thyroid cells, so the aim of the present study was to evaluate progesterone effects in the sodium-iodide symporter NIS, thyroglobulin TG, thyroperoxidase TPO, and KI-67 genes expression, in normal thyroid follicular cells, derived from human tissue. NIS, TG, TPO, and KI-67 mRNA expression increased significantly after TSH 20 µUI/mL, respectively: 2.08 times, P < 0.0001; 2.39 times, P = 0.01; 1.58 times, P = 0.0003; and 1.87 times, P < 0.0001. In thyroid cells treated with 20 µUI/mL TSH plus 10 nM progesterone, RNA expression of NIS, TG, and KI-67 genes increased, respectively: 1.78 times, P < 0.0001; 1.75 times, P = 0.037; and 1.95 times, P < 0.0001, and TPO mRNA expression also increased, though not significantly (1.77 times, P = 0.069). These effects were abolished by mifepristone, an antagonist of progesterone receptor, suggesting that genes involved in thyroid cell function and proliferation are upregulated by progesterone. This work provides evidence that progesterone has a direct effect on thyroid cells, upregulating genes involved in thyroid function and growth.

An unexpected cause of severe and refractory PTH-independent hypercalcemia: case report and literature review

Hypercalcemia is a common condition in the internal medicine practice. Sometimes its cause is not readily apparent, so extensive investigation is appropriate. Here we report an unexpected cause for hypercalcemia in an elderly woman. The case of an 82-years old woman with PTH-independent hypercalcemia, lymphocytosis, normal serum 1,25 (OH)vitamin D levels, and low serum PTHrp levels, is described. Medical history and complementary investigation were unremarkable, except for increased metabolic activity in the glutei regions, as measured by whole body 18F-FDG PET-CT. Reviewing her medical history, her sister recalled that she had been submitted to intramuscular methylmethacrylate injections, for cosmetic purposes, five years before presentation, which was confirmed by muscle biopsy. Low calcium intake, parenteral bisphosphonates, calcitonin, and glucocorticoids were used to control serum calcium levels. Methylmethacrylate injections, used cosmetically, are a new cause for hypercalcemia, even after many years. Hypercalcemia was probably due to calcitriol overproduction in foreign body granulomas. Persistent reactive lymphocytosis could be a clue for this inflammatory cause of hypercalcemia. Arch Endocrinol Metab. 2015;59(3):277-80.

Decreased Expression of GPER1 Gene and Protein in Goiter.

Goiter is more common in women, suggesting that estrogen could be involved in its physiopathology. The presence of classical estrogen receptors (ERα and ERß) has been described in thyroid tissue, suggesting a direct effect of estrogen on the gland. A nonclassic estrogen receptor, the G-protein-coupled estrogen receptor (GPER1), has been described recently in several tissues. However, in goiter, the presence of this receptor has not been studied yet. We investigated GPER1 gene and protein expressions in normal thyroid and goiter using reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot, respectively. In normal thyroid (n = 16) and goiter (n = 19), GPER1 gene was expressed in all samples, while GPER1 protein was expressed in all samples of normal thyroid (n = 15) but in only 72% of goiter samples (n = 13). When comparing GPER1 gene and protein levels in both conditions, gene expression and protein levels were higher in normal thyroid than in goiter, suggesting a role of this receptor in this condition. Further studies are needed to elucidate the role of GPER1 in normal thyroid and goiter.

Does serum 25-hydroxyvitamin D decrease during acute-phase response? A systematic review.

Low levels of 25-hydroxyvitamin D, or 25(OH)D, are commonly associated with inflammatory diseases. These associations could be due to an increased prevalence of inflammatory diseases in hypovitaminosis D, although reverse causality cannot be excluded. We aimed to systematically review the longitudinal studies that reported serum 25(OH)D during an acute inflammatory response in humans. Using Ovid MEDLINE, EMBASE, and the Cochrane Library, an electronic search of the literature was conducted from database inception until January 2014 by combining the MeSH terms: vitamin D and acute-phase reactants. Other sources for obtaining articles were used as cross-referencing texts. Based on 670 titles and abstracts, 40 articles were selected for full-text review, and 8 of these studies met the final inclusion criteria. In 6 of the reviewed studies, 25(OH)D dropped after the inflammatory insult; this decrease was abrupt in the studies that measured 25(OH)D early after the insult. In 2 studies, there was no change of 25(OH)D during the course of the disease, but baseline levels were measured in both after days of symptoms onset. One study suggested that hemodilution decreased 25(OH)D, with no effect on inflammation. Serum C-reactive protein concentrations were used as inflammatory markers in almost all studies. The metabolic meaning and the functional importance of these changes are unknown. In light of the current evidence, the 25(OH)D measured during acute-phase response should be interpreted with care. Future research, including other markers of vitamin D adequacy, could help to clarify if hypovitaminosis D might be the cause or the consequence of inflammatory diseases.

Effect of fat on serum 25-hydroxyvitamin D levels after a single oral dose of vitamin D in young healthy adults: a double-blind randomized placebo-controlled study.

PURPOSE: This double-blind placebo-controlled trial evaluated serum 25-hydroxyvitamin D [25(OH)D] levels after the oral intake of a single dose of cholecalciferol during one of the three meals, containing different amounts of fat or placebo. METHODS AND RESULTS: Sixty-four healthy medical residents or students of a university hospital in Porto Alegre, latitude 30° S, Brazil, were divided into four groups. Three groups received a single 50,000 IU oral dose of cholecalciferol during a meal containing 0 g (Group 1), 15 g (Group 2) or 30 g (Group 3) of fat, and one group received placebo (Group 4), according to randomization. Serum 25(OH)D, parathyroid hormone, total calcium, albumin, magnesium, and creatinine levels, and urinary calcium, magnesium, and creatinine levels were measured at baseline and after 14 days. Baseline mean serum 25(OH)D levels were low in all groups. Vitamin D given during breakfast increased the mean change of serum 25(OH)D levels, when compared to placebo. Furthermore, the intake of fat with vitamin D increased the mean change of serum 25(OH)D levels. CONCLUSION: A single oral dose of vitamin D given with food increased mean serum 25(OH)D levels, after 2 weeks, and the mean increase was larger, when the meal had at least 15 g of fat. These findings can have important implications to oral vitamin D supplementation.

Validation of reference genes for normalization gene expression in reverse transcription quantitative PCR in human normal thyroid and goiter tissue.

Reverse transcription quantitative polymerase chain reaction (RT-qPCR) has been recognized as the most accurate method for quantifying mRNA transcripts, but normalization of samples is a prerequisite for correct data interpretation. So, this study aimed to evaluate the most stable reference gene for RT-qPCR in human normal thyroid and goiter tissues. Beta-actin (ACTB); glyceraldehyde-3-phosphate dehydrogenase (GAPDH); succinate dehydrogenase, subunit A, flavoprotein (Fp) (SDHA); hypoxanthine phosphoribosyltransferase I (HPRTI); tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein, zeta polypeptide (YWHAZ); and beta-2-microglobulin (B2M) were evaluated in 14 thyroid tissue samples (7 normal and 7 goiter tissues) by RT-qPCR. The mean Cq and the maximum fold change (MFC) and NormFinder software were used to assess the stability of the genes. As a result, ACTB gene was more stable than GAPDH, SDHA, HPRTI, YWHAZ, and B2M. In conclusion, ACTB could be used to normalize RT-qPCR data in normal thyroid and goiter tissues.

Hypovitaminosis D in patients with cystic fibrosis: a cross-section study in South Brazil.

INTRODUCTION: Cystic fibrosis (CF) patients have a susceptibility to vitamin D deficiency because of nutrient malabsorption. OBJECTIVES: To evaluate the prevalence of hypovitaminosis D in CF patients and the factors associated with serum 25-hydroxyvitamin D levels. METHODS: We evaluated the prevalence of vitamin D deficiency defined as 25-hydroxyvitamin D <30 ng/mL, as suggested recently by the Cystic Fibrosis Foundation, and factors associated with its serum levels. Patients with confirmed CF were included. Nutritional status and hospital admissions were evaluated. Serum C-reactive protein, calcium, phosphate, magnesium, albumin, 25-hydroxyvitamin D and parathyroid hormone levels were measured. Lung function was evaluated by spirometry, and clinical and chest radiographic scores were assessed. Statistical significance level was set at P < 0.05. RESULTS: Fifty-nine patients were included. Prevalence of hypovitaminosis D was 61%. Patients with pancreatic insufficiency had a trend to have higher vitamin D levels. Sixteen patients had severe lung disease with percentage of forced expiratory volume in 1 s predicted below 40%. After multivariate analysis, body mass index and hospitalization in the last month remained significantly associated with serum vitamin D levels. CONCLUSIONS: Vitamin D insufficiency is still a problem in CF patients, even in those receiving supplementation.

A short stay unit decreased mean length of stay in a hospital in south Brazil

Introduction: Public health services in Brazil are periodically overcrowded. Since reducing the length of stay (LOS) could increase the availability of hospital beds, this study evaluated the impact of a short stay unit (SSU) on LOS, early readmission rates, and intra-hospital mortality rates. Methods Data were evaluated retrospectively in the 12 months before and after the establishment of a multidisciplinary SSU in a tertiary care hospital in south Brazil. All admissions of adult patients through the Emergency Department for causes in nine groups of the World Health Organization International Code of Diseases-10 were included. Results Mean LOS decreased 1.42 days in the first year after the implementation of a SSU with no change in 7-day readmission rates or intra-hospital mortality rates. This decrease may be partially explained by the decrease in the mean LOS in other areas of the hospital, although the amount of hospital-bed days saved in the second year was 5,668 days in the Internal Medicine Division and 1,595 days in all other clinical or surgical areas. Mortality rates after discharge were not evaluated. Conclusions A SSU decreased mean LOS of selected patients admitted through the Emergency Department without increasing 7-day readmission rates or intra-hospital mortality.