Interaction Effects of FTO and MC4R Polymorphisms on Total Body Weight Loss, Post-Surgery Weight, and Post-Body Mass Index after Bariatric Surgery.

Bariatric surgery (BS) is considered the most effective intervention for patients with severe obesity and is used to maintain long-term weight loss and glycemic control. The aim of this study was to analyze the effects of genotypes and haplotypes of the fat mass and obesity-associated (FTO) and melanocortin 4 receptor (MC4R) genes on total body weight loss (TBWL), post-surgery weight, and post-BMI after bariatric surgery. We retrospectively selected 101 patients from Bajio High Specialty Regional Hospital, León Guanajuato, México, who underwent Roux-en-Y gastric bypass (RYGB) to determine their body mass index (BMI), blood pressure, biochemical characteristics, and comorbidities. Post-surgery, patients were referred for registered anthropometry and blood pressure. Glucose, lipid and hepatic profiles, and insulin, leptin, and ghrelin levels were measured, and rs9939609, rs9930506, and rs1421085 FTO and rs17782313 MC4R polymorphisms were genotyped. Six (4-8) years after BS, post-surgery weight was greater in carriers of the rs9939609 and rs1421085 risk genotypes. TBWL was lower for the rs9930506 and rs1421085 risk genotypes. Insulin and HOMA-IR were greater in patients with the three FTO polymorphisms. There were significant interaction effects of the rs9930506 and rs1421085 FTO risk genotypes on weight and BMI in response to BS. No association was found with the MC4R polymorphism. The genotypes and haplotypes of the FTO gene influence post-surgery weight, TBWL, insulin levels, and HOMA-IR.

Analysis of Factors Associated with Outcomes of Bariatric Surgery: rs1800497 ANKK1, rs1799732 DRD2 Genetic Polymorphisms, Eating Behavior, Hedonic Hunger, and Depressive Symptoms.

INTRODUCTION: A therapeutic approach to severe obesity is bariatric surgery (BS), which is considered an effective intervention for ameliorating comorbidities such as T2DM, hypertension, dyslipidemia, and cardiovascular diseases. Some polymorphisms are considered markers for addictive disorders and hedonic hunger. We analyzed factors associated with the outcomes of BS, including rs1800497 ANKK1 and rs1799732 DRD2 polymorphisms, eating behavior, hedonic hunger, and depressive symptoms. METHODS: We retrospectively selected 101 patients who underwent BS and agreed to participate. The previous conditions to BS, such as body mass index (BMI), systolic blood pressure (SBP), diastolic blood pressure (DBP), and comorbidities, were registered; the scholarship value was evaluated as the total number of years of scholarly education. To evaluate the post-surgery conditions of the participants, we took blood samples, anthropometric measures, and 3 questionnaires to evaluate eating behavior (TFEQ-R18), hedonic hunger (PFS), and depressive symptoms (PHQ-9). The ANKK1 rs1800497 and rs1799732 DRD2 polymorphisms were genotyped. RESULTS: The median total weight loss (TWL) was 34.7 kg, with a BMI of 33.8 kg/m2, 6 (4-8) years after BS. The TWL was positively associated with the TFEQ-R18 score (p = 0.006) and negatively associated with triglycerides (p = 0.011). rs1800497 ANKK1 was associated with TFEQ-R18 (OR = 1.13 (1.02-1.25), p = 0.009). We also found a negative correlation of pre-surgery BMI with scholarship (r = - 0.27, p < 0.05). CONCLUSION: The patients showed an improvement in metabolic and anthropometric parameters post-surgery. Interestingly, the ANKK1 Taq1A polymorphism was associated with eating behavior and scholarship with pre-surgery BMI, which may be considered predictors of BS outcomes.

Profile of urinary exosomal microRNAs and their contribution to diabetic kidney disease through a predictive classification model.

AIM: Evaluate the expression of exomiRs-126, -146, and -155 in urinary exosomes of patients with T2DM and diabetic kidney disease to establish a predictive classification model with exomiRs and clinical variables in order to determine their contribution to DKD. METHODS: The study group included 92 subjects: 64 patients diagnosed with T2DM subclassified into two groups with albuminuria (T2DM with albuminuria, n = 30) and without albuminuria (TD2M, n = 34) as well as 28 healthy, non-diabetic participants. Exosomes were isolated from urine and identified by TEM and flow cytometry. Profile expression of exomiRs-126, -146 and -155 was evaluated by RT-qPCR. Data were analysed by permutational multivariate analysis of variance (PERMANOVA), similarity percentage (SIMPER), principal coordinate analysis (PCO), and canonical analysis of principal coordinates (CAP). RESULTS: T2DM patients with and without albuminuria showed higher levels of miR-155 and miR-146 compared with controls. In addition, T2DM patients with albuminuria presented a significant increase in miR-126 contrasted to controls and patients without albuminuria. PCO analysis explained 34.6% of the total variability of the data (PERMANOVA; p < .0001). Subsequently, SIMPER analysis showed that miR-146, miR-155, and miR-126 together, with some clinical parameters, contributed to 50% of the between-group significance. Finally, the CAP analysis developed showed a correct classification of 89.01% with the analysed parameters. CONCLUSION: A platform using a combination of clinical variables and exomiRs could be used to classify individuals with T2D as risk for developing DKD.

Associations of nicotidamide-N-methyltransferase, FTO, and IRX3 genetic variants with body mass index and resting energy expenditure in Mexican subjects.

The enzyme nicotidamide-N-methyltransferase (NNMT) regulates adipose tissue energy expenditure through increasing nicotinamide adenosine dinucleotide (NAD+) content. NNMT methylates nicotinamide to N1-methylnicotidamide (MNA-1) using S-adenosyl methionine. The rs694539 NNMT polymorphism is associated with non-alcoholic steatohepatitis, and rs1941404 is associated with hyperlipidemia. The rs1421085 FTO is related to poor eating behaviors, and rs3751723 IRX3 is associated with obesity. To investigate the association of rs694539 and rs1941404 NNMT, rs140285 FTO and rs3751723 IRX3 polymorphisms with MNA-1 concentrations, resting energy expenditure (REE) and BMI, we included clinically healthy Mexican subjects 30 to 50 years old, 100 subjects (35 men/65 women) with BMI > 30 kg/m2 and 100 subjects (32 men/68 women) with BMI < 25 kg/m2. Glucose, lipid profile, insulin, leptin, acylated ghrelin, and MNA-1 (LC-MS) were quantified. Resting energy expenditure (REE) was estimated using indirect calorimetry with a Fitmate instrument. Genotyping was performed using PCR-RFLP, and allelic discrimination was examined using TaqMan probes. MNA-1 concentrations and REE were significantly higher in obese subjects. Subjects with the rs694539AA NNMT genotype (recessive model) had lower weight, BMI, and REE. BMI showed an association with HDL-C, triglycerides, MNA-1, acetylated ghrelin, leptin, insulin concentrations, HOMA-IR, REE, and rs1421085. Subjects with the TC or CC genotypes of rs1421085 FTO showed 6 kg and 2 units of BMI more than did those with the TT wild type. The CG of the rs1421085 and rs3751723 haplotypes was associated with BMI. These findings showed that BMI was strongly associated with REE, rs1421085 FTO and the CG rs1421085 FTO and rs3751723 IRX3 haplotypes. We used the GMDR approach in obesity phenotype to show the interaction of four SNPs and metabolic variables.

Identification of metabolic markers in patients with type 2 Diabetes by Ultrafast gas chromatography coupled to electronic nose. A pilot study.

Metabolomics is a potential tool for the discovery of new biomarkers in the early diagnosis of diseases. An ultra-fast gas chromatography system equipped to an electronic nose detector (FGC eNose) was used to identify the metabolomic profile of Volatile Organic Compounds (VOCs) in type 2 diabetes (T2D) urine from Mexican population. A cross-sectional, comparative, and clinical study with translational approach was performed. We recruited twenty T2D patients and twenty-one healthy subjects. Urine samples were taken and analyzed by FGC eNose. Eighty-eight compounds were identified through Kovats's indexes. A natural variation of 30% between the metabolites, expressed by study groups, was observed in Principal Component 1 and 2 with a significant difference (p < 0.001). The model, performed through a Canonical Analysis of Principal coordinated (CAP), allowed a correct classification of 84.6% between healthy and T2D patients, with a 15.4% error. The metabolites 2-propenal, 2-propanol, butane- 2,3-dione and 2-methylpropanal, were increased in patients with T2D, and they were strongly correlated with discrimination between clinically healthy people and T2D patients. This study identified metabolites in urine through FGC eNose that can be used as biomarkers in the identification of T2D patients. However, more studies are needed for its implementation in clinical practice.

Possible role of n-hexane as an endocrine disruptor in occupationally exposed women at reproductive age.

Human exposure to n-hexane has been associated with subfertility and, experimentally, with a decrease in follicular development. In order to assess occupational exposure to n-hexane on ovarian function and gonadotropic hormones, we studied Mexican women labouring in a leather shoe factory (n = 34). Individual environmental levels for seven solvents, n-hexane included, were measured; also, urinary 2,5-hexanedione (2,5-HD) was determined. For ovarian function and hormonal status, FSH, LH, oestradiol and anti-Müllerian hormone (AMH) levels were determined. We performed all determinations also in a reference group, administrative workers with no exposure to solvents (n = 32). Results: N-hexane and urinary 2,5-HD levels were higher in exposed group (p < 0.001). More cases of oligomenorrhea as well as longer time for getting pregnant were observed in exposed women compared with controls; a positive association was found between menstrual cycle length and "time for getting pregnant" (p = 0.010); significant associations between FSH serum levels and 2,5-HD urinary levels (post-shift sample) were observed in non-smokers participants presenting oligomenorrhea from exposed group. Also, we found a trend for lower oestradiol levels in exposed participants with current smoking habit (p = 0.059). Conclusions: 2,5-HD urinary levels are associated with decreased gonadotropins levels; hence, n-hexane should be considered an endocrine disruptor in reproductive-age women.

Analysis of the percentages of monocyte subsets and ILC2s, their relationships with metabolic variables and response to hypocaloric restriction in obesity.

PURPOSE: Obesity results from excess energy intake over expenditure and is characterized by chronic low-grade inflammation involving circulating monocytes (Mo) and group 2 innate lymphoid cells (ILC2s) imbalance. We analyzed circulating Mo subsets and ILC2s percentages and ß2-adrenergic receptor (ß2AR) expression in lean and obese subjects, and the possible effect of hypocaloric restriction on these innate immune cells. METHODS: In 139 individuals aged 45 to 57 years, classified in 74 lean individuals (>18.9kg/m2 BMI <24.9kg/m2) and 65 with obesity (n = 65), we collected fasting blood samples to detect Mo subsets, ILC2s number, and ß2AR expression by flow cytometry. Lipids, insulin, leptin, and acylated-ghrelin concentrations were quantified. Resting energy expenditure (REE) was estimated by indirect calorimetry. These measurements were repeated in obese subjects after 7-weeks of hypocaloric restriction. RESULTS: Non-classical monocytes (NCM) and ß2AR expression on intermediate Mo (IM) were increased in obese individuals (p<0.001, in both cases), whereas the percent of ILC2s was decreased (p<0.0001). Stepwise regression analysis showed significantly negative associations of ILC2s with caloric intake, ß2AR expression on IM with REE, but a positive relationship between NCM and HOMA-IR. Caloric restriction allowed a significant diminution of NCM and the ß2AR expression on IM, as well as, an increase in the percent of classical Mo (CM), and ILC2s. ΔREE was related to ΔCD16+/CD16- ratio. CONCLUSIONS: These findings show that in obesity occur changes in NCM, ILC2s and ß2AR expression, which contribute to the low-grade inflammation linked to obesity and might revert with caloric restriction.

Sleep Extension Increases the Effect of Caloric Restriction Over Body Weight and Improves the Chronic Low-Grade Inflammation in Adolescents With Obesity.

PURPOSE: Diminution of sleep may be associated with obesity. However, evidence that extending sleep duration might favor weight loss is insufficient. The aim of this study was to compare the effect of dietary restriction with or without prescription of sleep extension on weight loss in adolescents with obesity. METHODS: A total of 52 adolescents with obesity (24 males and 28 females) received a diet with 500 calories restriction, randomly allocated to groups without (n = 27) and with sleep extension (n = 25) for 4 weeks. We collected data on anthropometry, caloric intake, and self-reported sleep diaries. Serum interleukin 6, tumor necrosis factor α, leptin, and insulin levels were quantified by enzyme-linked immunosorbent assay. Cortisol and 6-sulfatoxymelatonin excretions were measured in the first urine collection in the morning by liquid chromatography-mass spectrometry. Measurements were carried out at baseline and at the end of the intervention. RESULTS: After diet, weight decreased in both groups. Sleep extension, improved weight loss (p < .00001), and waist girth reduction (p = .00003), with diminution of insulin (p = .002) and interleukin 6 levels (p = .02). Caloric restriction was less effective in adolescent females. No differences in cortisol or 6-sulfatoxymelatonin excretion were found. CONCLUSIONS: A sleep extension favors weight loss in adolescents under caloric restriction and improves inflammation and metabolic conditions, thus supporting a possible additional benefit to diet in the treatment of obesity in adolescents.

Effects of sleeve gastrectomy and rs9930506 FTO variants on angiopoietin/Tie-2 system in fat expansion and M1 macrophages recruitment in morbidly obese subjects.

Angiogenesis in inflammation are hallmarks for adipose tissue expansion in obesity. The role of angiopoietin/Tie-2 system in adipose tissue expansion and immune cell recruitment is unclear. We studied the effect of sleeve gastrectomy and the influence of FTO rs9930506 polymorphism on Tie-2, angiopoietin-1 and angiopoietin-2 expression in morbid obesity. Fifteen morbidly obese subjects (4 men and 11 women) aged 24-55 years were followed-up 3 and 6 months after sleeve gastrectomy. Serum sTie-2, angiopoietin-1, angiopoietin-2, and hypoxia-inducible factor-1α concentrations were determined by ELISA. Tie-2 and its ligands in visceral and subcutaneous adipose tissue were localized by immunohistochemistry. Tie-2 expression was measured by flow cytometry in circulating monocytes and infiltrated macrophages. Comparisons before and after sleeve gastrectomy were carried out using ANOVA for repeated measures. rs9930506FTO genotyping was performed by PCR-RFLP. Circulating sTie-2 and angiopoietin-2 were higher before sleeve gastrectomy. Tie-2 and angiopoietin-2 mRNA levels were higher in subcutaneous adipose tissue than visceral and both decreased after surgery. Monocytes and infiltrated macrophages showed a pro-inflammatory phenotype, with increased Tie-2 expression that decreased 3 and 6 months after sleeve gastrectomy. Baseline sTie-2 correlated inversely with adiponectin levels. At baseline the rs9930506FTO AG ó GG genotypes carriers had more 34 kg than genotype carriers of rs9930506 AA. Weight and body mass index decreased at 6 months. We found that angiopoietin/Tie-2 system is mainly expressed in subcutaneous adipose tissue, contributing to expandability, fat accumulation, and monocytes attachment in obesity. Bariatric surgery favorably modifies the pro-angiogenic profile, allowed a reduced angiogenic expression in the circulation and adipose tissue.

Alterations in adhesion molecules, pro-inflammatory cytokines and cell-derived microparticles contribute to intima-media thickness and symptoms in postmenopausal women.

Menopause, the cessation of menses, occurs with estrogens decline, low-grade inflammation, and impaired endothelial function, contributing to atherosclerotic risk. Intima-media thickness (IMT) is an early subclinical biomarker of atherosclerosis. Inflammation may have a role on symptoms: hot flashes, anxiety, and depressive mood, which also are related to endothelial dysfunction, increased IMT and cardiovascular risk. In this study we compared several inflammatory markers in early vs. late postmenopausal women and studied the association of IMT and symptoms with these markers in the full sample. In a cross-sectional design including 60 women (53.1 ± 4.4 years old) at early and late postmenopause, we evaluated the expression of CD62L, ICAM-1, PSGL-1, CD11b, CD11c, and IL-8R on PBMC by flow cytometry. Serum soluble ICAM-1, sVCAM-1, sCD62E, sCD62P, CXCL8, IL-1ß, IL-6, and TNF-α levels were quantified by ELISA. Plasma levels of microparticles (MPs) were determined by FACS. Finally, carotid intima-media thickness (IMT) was measured by ultrasound. We observed that ICAM-1 expression by lymphocytes and serum sVCAM-1 levels were augmented at late postmenopause. Late postmenopause women with severe hot flashes had increased expression of CD62L and IL-8R on neutrophils. By multivariate analysis, the carotid IMT was strongly associated with membrane-bound TNF-α, CD11b expression, Annexin V(+) CD3(+) MPs, LPS-induced NO production, HDL-cholesterol and age. Depressive mood was associated negatively with PSGL-1 and positively with LPS-induced NO. Finally, Log(AMH) levels were associated with carotid IMT, IL-8R expression and time since menopause. IMT and depressive mood were the main clinical features related to vascular inflammation. Aging, hormonal changes and obesity were also related to endothelial dysfunction. These findings provide further evidence for a link between estrogen deficiency and low-grade inflammation in endothelial impairment in mature women.

Influence of heart rate variability and psychosocial factors on carotid stiffness, elasticity and impedance at menopause.

BACKGROUND AND AIMS: The risk for cardiovascular diseases (CVD) increases after menopause. Heart rate variability (HRV), a measure of autonomic control, is a strong predictor of CVD. We undertook this study to test the association of ultrasound indices of early carotid atherosclerosis with HRV, symptoms, hormonal conditions, metabolic state, indicators of stress, and psychosocial factors in women at peri- and postmenopause, registering ambulatory R-R interval monitoring. METHODS: In a cross-sectional design we studied 100 women at peri- and early postmenopause collecting anthropometry, symptoms, stress-related measurements, metabolic variables, cortisol, FSH and estradiol. We evaluated carotid ultrasonographic indices, and HRV was recorded for 4 h calculating time (SDNN, pNN50, rMSSD) and frequency domains (LF, HF, LF/HF) in women according to menopausal stage, estradiol levels, body mass index and waist circumference. RESULTS: Carotid indices were similar in peri- and postmenopausal women. For HRV measurements, SDNN was increased at postmenopause. Women with estradiol levels <109.2 pmol/L had increased intima-media thickness (IMT), resistive index, and systolic diameter. Using multivariate analysis, we found the associations of IMT positively with non-HDL-cholesterol, resistive index positively with LF-HRV, but negatively with effort/reward imbalance, carotid ß stiffness index inversely with estradiol, and arterial distensibility positively with HF-HRV and creatinine concentrations, but negatively with non-HDL-cholesterol. CONCLUSIONS: Carotid thickness was related mainly with lipid alterations. Indices of early carotid damage were related with various components of HRV as a manifestation of autonomic imbalance, indicating CVD risk. Other factors involved were time since last menses and psychological stress. Low creatinine was associated with diminished carotid distensibility. This suggests that estrogen, lifestyle, behavior and autonomic regulation participate in vascular damage.

Relationship of sleep alterations with perimenopausal and postmenopausal symptoms.

OBJECTIVE: Sleep disturbance is an important change in menopause because it affects quality of life and can lead to other conditions such as depression. This study measured sleep alterations and explored associated physical, emotional, hormonal, and lifestyle factors during perimenopause and postmenopause. METHODS: This cross-sectional study enrolled 160 women who were classified as perimenopausal (n = 85) or postmenopausal (n = 75). Using diaries, we collected data on duration of sleep, time awake in bed, and sleep efficiency. Follicle-stimulating hormone, 17ß-estradiol, and cortisol levels were quantified by radioimmunoassay, and serum anti-Müllerian hormone levels were measured using enzyme-linked immunosorbent assay. Correlations between sleep measurements and symptoms were assessed using a generalized linear model. RESULTS: The reported duration of sleep was similar for both groups of women (close to 6.9 h), and sleep efficiency was 88%. We did not find any factor that was associated with duration of sleep. Sleep efficiency was negatively associated with age, perimenopause/postmenopause status, loss of sexual interest, hot flashes, and depressed mood. Time awake in bed was positively associated with depressed mood (P < 0.000001), cigarette smoking (P < 0.000041), menopause status (P < 0.00009), and age (P < 0.0009). These associations remained after controlling for exercise, alcohol consumption, and caffeine consumption as confounding variables. Finally, morning salivary cortisol was reduced in postmenopausal women. CONCLUSIONS: Time awake in bed shows the most significant associations. Depressed mood, age, and menopause status are the main factors associated with sleep disturbances.

Plasmacytoid dendritic cells in patients with autoimmune thyroid disease.

BACKGROUND: Patients with autoimmune thyroid diseases (AITD) show defects in immunoregulatory mechanisms. Herein we assessed the expression of different regulatory receptors in circulating and thyroid dendritic cells (DCs). DESIGN: Peripheral blood samples from 49 patients with Hashimoto's thyroiditis, 35 with Graves' disease, and 34 healthy subjects were studied. Clinical parameters included grades of goiter and ophthalmopathy, thyroid function, and antibody tests. Thyroid tissue samples from 10 AITD patients were also analyzed. Levels of DCs and their expression of different regulatory molecules (IDO, ILT2, ILT3, PSGL-1, PD-L1) were studied. In vitro interferon-α response by plasmacytoid DCs (pDCs) and tryptophan (Trp) metabolites were determined. RESULTS: Significant low levels of pDCs, but not conventional DCs, were detected in the peripheral blood from AITD patients, mainly in those with severe disease. Furthermore, a diminished expression of ILT3, PSGL-1, and CD69 by peripheral blood pDCs from AITD patients was observed. An increased number of pDCs was found in thyroid tissue, showing a diminished expression of ILT3 and PSGL-1. A lower proportion of IDO+ pDCs, a significant increase in Trp levels, a decrease in the kyneruine/Trp ratio, and an increased in vitro interferon-α response were present in AITD patients. Finally, a significant correlation was found between the in vitro synthesis of IL-10 by stimulated T cells and expression of IDO by pDCs. CONCLUSIONS: The diminished number of pDCs in the peripheral blood from AITD patients as well as their abnormal phenotype could contribute significantly to the pathogenesis.

The association of the angiopoietin/Tie-2 system with the development of metastasis and leukocyte migration in neuroendocrine tumors.

The aim of this study was to explore the possible involvement of the angiopoietin (Ang)-1, -2/Tie-2 system in the development, growth, and metastases evolution of gastroenteropancreatic-neuroendocrine tumors (GEP-NETs). We prospectively examined the serum levels of Tie-2, Ang-1, and Ang-2 by ELISA in 42 patients with proven GEP-NETs and 27 controls. We also determined the expression of the Ang/Tie-2 system in freshly isolated peripheral blood monocytes and in tumor cells from malignant primary tumors and/or liver metastases samples from GEP-NET patients by flow cytometry and/or RT-PCR. Furthermore, the function of the Ang/Tie-2 system in monocytes from controls and patients was assessed by a chemotaxis assay. GEP-NET patients showed enhanced serum levels of soluble form of Tie-2 (sTie-2), Ang-1, and Ang-2 (P<0.05 in all cases), compared to controls. sTie-2 and Ang-2 levels were significantly higher in GEP-NETs with metastases compared to those with no metastases. In addition, a significant correlation was detected between Ang-2 levels and chromogranin A or sTie-2 concentrations or 5-hydroxy-indole acetic acid excretion (r=0.71, r=0.60, and r=0.81 respectively, P<0.01 in all cases). Furthermore, we observed an enhanced expression of Ang-1, Ang-2, and Tie-2 in freshly isolated tumor cells from GEP-NET both by immunohistochemistry and by RT-PCR. Interestingly, an enhanced expression and function of Tie-2 was detected in monocytes from GEP-NET patients. Our data suggest that the Ang/Tie-2 system is involved in the growth and development of metastases of GEP-NETs, and that favors the recruitment of Tie-2(+) monocytes to the tumor site, where they can promote inflammation and angiogenesis.

Increased circulating pro-inflammatory cytokines and Th17 lymphocytes in Hashimoto's thyroiditis.

CONTEXT: Th17 lymphocytes play an important role in different chronic inflammatory and autoimmune conditions. AIM: The aim of the study was to explore the status of Th17 cells in patients with autoimmune thyroid diseases (AITD). DESIGN: We assessed the serum levels and in vitro synthesis of IL-17 and IL-22 and of different cytokines (IL-6, IL-15, and IL-23) involved in the differentiation of Th17 cells in the peripheral blood and thyroid glands of 26 patients with AITD, eight with Graves' disease, and 18 with Hashimoto's thyroiditis (HT) as well as 10 healthy controls. RESULTS: We found enhanced levels of T cells synthesizing IL-17 and IL-22 in the peripheral blood from AITD patients, mainly in those with HT. In addition, a stronger expression of IL-17 and IL-22 and an enhanced number of IL-23R(+) cells was detected in thyroid glands from HT patients compared with Graves' disease or controls. Furthermore, increased concentrations of IL-6 and IL-15 were detected in sera from HT patients, whereas serum levels of IL-23 tended to be higher in these patients. Finally, an enhanced in vitro differentiation of T lymphocytes into Th17 cells induced by IL-23/IL-6 was observed in AITD patients. Accordingly, a strong induction of RORC2 gene was detected in lymphocytes from HT patients when stimulated with IL-23. CONCLUSION: Our results indicate that there is an increased differentiation of Th17 lymphocytes and an enhanced synthesis of Th17 cytokines in AITD, mainly in HT. These phenomena may have an important role in the pathogenesis of thyroid autoimmunity.

Tie-2 is overexpressed by monocytes in autoimmune thyroid disorders and participates in their recruitment to the thyroid gland.

CONTEXT: The angiopoietin/Tie system seems to have an important role in the pathogenesis of inflammatory diseases. Although Tie-2 is mainly expressed by endothelium, it is also detected in monocytes, which participate in the development of angiogenic and inflammatory phenomena. AIM: The aim was to study the expression and function of Tie-2 and their ligands, angiopoietin-1 (Ang-1) and Ang-2, in thyroid glands and monocytes from patients with autoimmune thyroid disease (AITD). DESIGN: We studied the expression of Tie-2, Ang-1, and Ang-2 by immunohistochemical techniques in surgical thyroid tissues from 17 patients with Graves' disease, 8 with Hashimoto's thyroiditis, and 3 healthy glands. In addition, we explored the expression and function of Tie-2 in peripheral blood monocytes from 17 patients with Graves' disease, 11 with Hashimoto's thyroiditis, and 14 healthy controls. RESULTS: We found that the expression of Ang-1, Ang-2, and Tie-2 was up-regulated in thyroid glands from AITD patients. Flow cytometry, immunofluorescence, ELISA, and RT-PCR analyses confirmed the synthesis and release of Ang-1, Ang-2, and Tie-2 by thyroid follicular cells (TFC) from AITD patients. In addition, these patients showed increased levels of Tie-2(+) monocytes in the peripheral blood, which exhibited an enhanced chemotactic response to Ang-2 or autologous TFC. CONCLUSIONS: Our data suggest that the Ang/Tie-2 system, through the participation of blood vessels, inflammatory cells, and TFC, may have an important role in the recruitment of monocytes to the thyroid gland and the pathogenesis of the tissue damage seen in AITD.

Serum levels of angiogenic molecules in autoimmune thyroid diseases and their correlation with laboratory and clinical features.

CONTEXT: Because angiogenesis has a role in the pathogenesis of inflammatory conditions, we studied angiogenesis soluble markers in autoimmune thyroid diseases. OBJECTIVE: The aim of the study was to measure concentrations of angiopoietins, Tie-2, and vascular endothelial growth factor in sera from autoimmune thyroid disease patients. DESIGN: Soluble Tie-2 (sTie-2), angiopoietin-1, angiopoietin-2, and vascular endothelial growth factor were quantified by ELISA in sera from 44 untreated Graves' disease (GD) patients, 25 untreated Hashimoto's thyroiditis (HT) patients, 13 non-GD hyperthyroid patients, and 22 age-matched controls. Subgroups of patients with active and non-active Graves' ophthalmopathy (GO) were analyzed. Correlations among these markers and clinical parameters were assessed by bivariate and multivariate analyses. RESULTS: STIE-2 levels were higher in GD or HT patients compared to controls (P < 0.01). In addition, serum Ang-2 concentrations were higher in untreated GD patients compared to controls, HT patients, or non-GD hyperthyroid patients (P < 0.01), and no difference was observed between HT patients and controls. Significant correlations were found between free T(4)/sTie-2 and free T(4)/Ang-2 levels (r = 0.464, P < 0.01; and r = 0.463, P < 0.01, respectively) as well as between sTie-2/anti-TSH receptor antibody (r = 0.527; P < 0.01) and sTie-2/Ang-2 (r = 0.563; P = 0.001). Furthermore, sTie-2 levels were significantly higher in patients with active GO when compared to those with inactive GO (P < 0.05). Interestingly, Ang-2 levels decreased significantly after treatment with antithyroid drugs (P < 0.01). CONCLUSIONS: Ang-2 and sTie-2 could participate in the pathogenesis of GD and potentially be used as markers of GO activity. Antithyroid drugs affect the angiogenic pattern in GD.

Expresión de la enzima óxido nítrico sintetasa inducible en las enfermedades tiroideas autoinmunitarias / Inducible nitric oxide synthase (iNOS) expression in autoimmune thyroid disorders (AITD)

Introducción: Diferentes tipos de células producen óxido nítricomediante la acción de la enzima óxido nítrico sintetasa (NOS). Existen cuatro isoformas de la enzima NOS, la neuronal, la endotelial, la mitocondrial y la inducible (iNOS). La producción de NO puede tener un papel importante en la patogenia de la inflamación y el daño de tejidosq ue no ha sido explorado adecuadamente en la enfermedad tiroideaautoinmunitaria. Objetivo: Estudiar la expresión de iNOS, tanto en proteína como en el ARNm, en las enfermedades tiroideas autoinmunitarias. Pacientes y método: Se evaluó la expresión de iNOS en los tiroides de10 pacientes con enfermedad de Graves (EG), 5 con tiroiditis de Hashimoto (TH) y 10 controles sanos, mediante inmunohistoquímica y transcripción inversa-reacción en cadena de la polimerasa (RT-PCR).Resultados: Se encontró un incremento en la expresión de iNOS en lag lándula tiroides de pacientes con EG y con TH tanto por PCR como mediante inmunohistoquímica. Esta expresión fue mayor en pacientes con EG. Por el contrario, en la glándula tiroides de sujetos sanos no se observó expresión de iNOS. La expresión de iNOS, tanto en EG como en TH, se detectó en células foliculares tiroideas, principalmente en las próximas al infiltrado inflamatorio. Asimismo, se observó expresión de iNOS en células endoteliales y células mononucleares del infiltrado inflamatorio en EG y TH. Conclusiones: La expresión prominente de iNOS en la enfermedad tiroidea autoinmunitaria indica que el NO puede tener un papel importante en la patogenia del fenómeno inflamatorio y el daño del tejido que ocurre en esta afección (AU)

Introduction: Nitric oxide is synthesized by different cell types through the action of the enzyme nitric oxide synthase (NOS).There are four is forms of this enzyme: the neuronal, the endothelial, the mitochondrial, and the inducible (iNOS) forms. Although NO production may play an important role in the pathogenesis of inflammation and tissue damage, its possible role in autoimmune thyroiditis has not been adequately explored. Objective: To study protein and mRNA expression of iNOS in human autoimmune thyroid disorders (AITD).Patients and method: We evaluated the expression of iNOS in thyroid glandspecimens from 10 patients with Graves’ disease (GD), from five patients with Hashimoto’s thyroiditis (HT) and from 10controls by immunohistochemical and reverse transcription-polymerase chain reaction (RT-PCR).Results: Both immunohistochemistry and PCR techniques showed up-regulated expression of iNOS in the thyroid glands of patients with GD and HT. iNOS expression was higher in GD than in HT. In contrast,no iNOS expression was detected in normal thyroid tissue. In both GD and HT, iNOS was detected in thyrocytes, mainly int hose localized in areas close to the inflammatory cell infiltrate. In addition, upregulated expression of iNOS was observed in endothelial cells and thyroidinfiltrating mononuclear cells in both GD and HT. Conclusions: The enhanced expression of iNOS in autoimmune thyroiditis suggests that NO synthesis may play an important role in the inflammatory phenomena and tissue damage observed in this disease (AU)