Notch signaling in thyrocytes is essential for adult thyroid function and mammalian homeostasis.

The thyroid functions as an apex endocrine organ that controls growth, differentiation and metabolism1, and thyroid diseases comprise the most common endocrine disorders2. Nevertheless, high-resolution views of the cellular composition and signals that govern the thyroid have been lacking3,4. Here, we show that Notch signalling controls homeostasis and thermoregulation in adult mammals through a mitochondria-based mechanism in a subset of thyrocytes. We discover two thyrocyte subtypes in mouse and human thyroids, identified in single-cell analyses by different levels of metabolic activity and Notch signalling. Therapeutic antibody blockade of Notch in adult mice inhibits a thyrocyte-specific transcriptional program and induces thyrocyte defects due to decreased mitochondrial activity and ROS production. Thus, disrupting Notch signalling in adult mice causes hypothyroidism, characterized by reduced levels of circulating thyroid hormone and dysregulation of whole-body thermoregulation. Inducible genetic deletion of Notch1 and 2 in thyrocytes phenocopies this antibody-induced hypothyroidism, establishing a direct role for Notch in adult murine thyrocytes. We confirm that hypothyroidism is enriched in children with Alagille syndrome, a genetic disorder marked by Notch mutations, suggesting that these findings translate to humans.

Single cell transcriptomic analysis of Graves' disease thyroid glands reveals the broad immunoregulatory potential of thyroid follicular and stromal cells and implies a major re-interpretation of the role of aberrant HLA class II expression in autoimmunity.

The study of the immune response in thyroid autoimmunity has been mostly focused on the autoantibodies and lymphocytes, but there are indications that intrinsic features of thyroid tissue cells may play a role in disrupting tolerance that needs further investigation. The overexpression of HLA and adhesion molecules by thyroid follicular cells (TFC) and our recent demonstration that PD-L1 is also moderately expressed by TFCs in autoimmune thyroid indicates that TFCs they may activate but also inhibit the autoimmune response. Intriguingly, we have recently found that in vitro cultured TFCs are able to suppress the proliferation of autologous lymphocyte T in a contact-dependent manner which is independent of the PD-1/PD-L1 signaling pathway. To get a more comprehensive picture of TFC activating and inhibitory molecules/pathways driving the autoimmune response in the thyroid glands, preparations of TFCs and stromal cells from five Graves' disease (GD) and four control thyroid glands were compared by scRNA-seq. The results confirmed the previously described interferon type I and type II signatures in GD TFCs and showed unequivocally that they express the full array of genes that intervene in the processing and presentation of endogenous and exogeneous antigens. GD TFCs lack however expression of costimulatory molecules CD80 and CD86 required for priming T cells. A moderate overexpression of CD40 by TFCs was confirmed. GD Fibroblasts showed widespread upregulation of cytokine genes. The results from this first single transcriptomic profiling of TFC and thyroid stromal cells provides a more granular view of the events occurring in GD. The new data point at an important contribution of stromal cells and prompt a major re-interpretation of the role of MHC over-expression by TFC, from deleterious to protective. Most importantly this re-interpretation could also apply to other tissues, like pancreatic beta cells, where MHC over-expression has been detected in diabetic pancreas.

Accuracy of an intraoral scanner based on sleeve type, decontamination, and calibration.

The purpose of this study was to evaluate the accuracy of an intraoral scanner based on camera sleeve type, decontamination protocol, and calibration status. Five extracted human teeth were set into a gypsum stone model and prepared for various indirect restorations. An optical impression was completed with a benchtop scanner to serve as a reference standard. A total of 160 optical impressions were completed using a sterilizable sleeve, an autoclavable sleeve with a single-use plastic window, or a single-use disposable plastic sleeve attached to a calibrated or an uncalibrated intraoral scanner. For the sterilizable sleeves, 2 decontamination protocols were used--high-level disinfection (HLD) or dry heat sterilization (DHS)--and scans were performed at baseline and after 25 and 50 cycles for each protocol. For the autoclavable (AS) and disposable single-use (SU) sleeves, scans were performed at baseline only. Thus, there were 10 optical impressions per test condition: sleeve type (HLD, DHS, AS, or SU) × decontamination status (baseline, 25 cycles [HLD or DHS], or 50 cycles [HLD or DHS]) × calibration status (calibrated or uncalibrated scanner). The individual optical impressions were compared to the reference standard impression by using 3-dimensional best-fit superimposition with the prepared tooth surfaces as reference points, and 3-dimensional linear differences were calculated for each superimposition. The median positive and absolute value median negative distance measurements were averaged for each impression to generate an average median discrepancy from baseline. The data were analyzed with Kruskal-Wallis and Mann-Whitney U tests (α = 0.05). No statistically significant differences in the median linear distance were found, regardless of sleeve type, decontamination protocol, or calibration status (P > 0.05). All groups demonstrated statistically similar linear disparities, ranging from 11.78 to 14.00 µm. The most precise sleeves were the single-use plastic sleeves, although their results were not significantly different from those of the multiuse sleeve. The results indicated that any of the currently available camera sleeves can provide similar accuracy in a clinical setting and that single-use disposable sleeves are a viable alternative to the currently accepted multiuse sleeves.

Case report: Accelerated regression of giant cardiac rhabdomyomas in neonates with low dose everolimus.

Cardiac rhabdomyoma (CRHM) is the principal cardiac tumor in children and is most often associated with tuberous sclerosis complex (TSC). Mutations in the TSC1 and TSC2 genes cause the overactivation of the mammalian Target of Rapamycin (mTOR). This protein family is responsible for abnormal cell proliferation leading to the formation of CRHMs and hamartomas in other organs. Despite the tendency for spontaneous regression, some CRHMs can cause heart failure and intractable arrhythmias, requiring surgical resection. In recent years, the use of everolimus and sirolimus (mTOR inhibitors) in the treatment of CRHMs has been reported. We report two cases of neonates with giant rhabdomyomas, with hemodynamic repercussions treated with low-dose everolimus (4.5 mg/m2/week). In both cases, we obtained an approximate decrease of 50% in the total area of the mass after three weeks of treatment. Despite rebound growth after stopping the drug, we were able to evidence that the use of low doses of everolimus immediately after birth is effective and safe in the treatment of giant CRHMs, avoiding surgical resection of the tumor and associated morbidity and mortality.

Thyroid cells from normal and autoimmune thyroid glands suppress T lymphocytes proliferation upon contact revealing a new regulatory inhibitory type of interaction independent of PD1/PDL1.

Immune Checkpoint Receptors include a number of inhibitory receptors that limit tissue damage during immune responses; blocking PD-1/PD-L1 checkpoint receptor axis led to a paradigm shift in cancer immunotherapy but also to autoimmune adverse effects, prominently thyroid autoimmunity. Although PD-L1 is known to be expressed on thyroid follicular cells (TFCs) of autoimmune glands the role on PD-1/PD-L1 in the interaction between T cells and thyroid cells in the tissue has not been investigated. Here we report that autologous primary TFCs, but not transformed TFCs, inhibit CD4 and CD8 T cell proliferation but no cytokine production. This effect is not, however, mediated by PD-1/PD-L1 nor locally produced cytokines. Beta galactosidase analysis excluded culture-induced senescence as an explanation. High resolution flow cytometry demonstrated that autologous TFC/T cells co-culture induced the expansion of several clusters of double negative (DN) T cells characterized by high expression of activation markers and negative immune checkpoints. Single cell transcriptomic profiling demonstrated that dissociated TFC express numerous candidate molecules for mediating this suppressive activity, including CD40, E-Cadherin and TIGIT ligands. These ligands directly or through the generation of a suppressor population of DN T cells, and not the PD-1/PD-L1 axis, are most likely the responsible of TFC immunosuppressive activity. These results contribute to reveal the complex network of inhibitory mechanism that operate at the tissue level to restrain autoimmunity but also point to pathways, other that PD-1/PD-L1, that can contribute to tumor evasion.

[Origins and development of a national scientific policy in Chile: Conicyt, 1967-1981]. / Orígenes y desarrollo de una política científica nacional en Chile: Conicyt, 1967-1981.

In 1967, the founding of the Comisión Nacional de Investigación Científica y Tecnológica (Conicyt) constituted a milestone by creating a national and public body, different from the university, which coordinated and financed scientific-technological activity, linked to the productive development in Chile, and the history of Latin American research councils. Conicyt led a demanding process by debating the contents of a scientific policy in a period in which Chile experienced opposing political-economic development models that conditioned its formulation and objectives. This article characterizes the origins and development of this commission from 1967 to 1981 when the Fondo Nacional de Ciencia y Tecnología was created and a new institutional stage began.

En 1967, la fundación de la Comisión Nacional de Investigación Científica y Tecnológica constituyó un hito al crearse un organismo nacional y público, distinto al universitario, que coordinó y financió la actividad científica-tecnológica, vinculándola al desarrollo productivo en Chile, y a la historia de los consejos de investigación latinoamericanos. La Comisión lideró un exigente proceso al debatir los contenidos de una política científica en un periodo en que Chile experimentó modelos de desarrollo político-económico opuestos que condicionaron su formulación y objetivos. Este artículo caracteriza los orígenes y desarrollo de esta comisión desde 1967 hasta 1981, cuando al crearse el Fondo Nacional de Ciencia y Tecnología se inicia una nueva etapa institucional.

Orígenes y desarrollo de una política científica nacional en Chile: Conicyt, 1967-1981 / Origins and development of a national scientific policy in Chile: Conicyt, 1967-1981

Resumen En 1967, la fundación de la Comisión Nacional de Investigación Científica y Tecnológica constituyó un hito al crearse un organismo nacional y público, distinto al universitario, que coordinó y financió la actividad científica-tecnológica, vinculándola al desarrollo productivo en Chile, y a la historia de los consejos de investigación latinoamericanos. La Comisión lideró un exigente proceso al debatir los contenidos de una política científica en un periodo en que Chile experimentó modelos de desarrollo político-económico opuestos que condicionaron su formulación y objetivos. Este artículo caracteriza los orígenes y desarrollo de esta comisión desde 1967 hasta 1981, cuando al crearse el Fondo Nacional de Ciencia y Tecnología se inicia una nueva etapa institucional.

Abstract In 1967, the founding of the Comisión Nacional de Investigación Científica y Tecnológica (Conicyt) constituted a milestone by creating a national and public body, different from the university, which coordinated and financed scientific-technological activity, linked to the productive development in Chile, and the history of Latin American research councils. Conicyt led a demanding process by debating the contents of a scientific policy in a period in which Chile experienced opposing political-economic development models that conditioned its formulation and objectives. This article characterizes the origins and development of this commission from 1967 to 1981 when the Fondo Nacional de Ciencia y Tecnología was created and a new institutional stage began.

Exposing and Overcoming Limitations of Clinical Laboratory Tests in COVID-19 by Adding Immunological Parameters; A Retrospective Cohort Analysis and Pilot Study.

Background: Two years since the onset of the COVID-19 pandemic no predictive algorithm has been generally adopted for clinical management and in most algorithms the contribution of laboratory variables is limited. Objectives: To measure the predictive performance of currently used clinical laboratory tests alone or combined with clinical variables and explore the predictive power of immunological tests adequate for clinical laboratories. Methods: Data from 2,600 COVID-19 patients of the first wave of the pandemic in the Barcelona area (exploratory cohort of 1,579, validation cohorts of 598 and 423 patients) including clinical parameters and laboratory tests were retrospectively collected. 28-day survival and maximal severity were the main outcomes considered in the multiparametric classical and machine learning statistical analysis. A pilot study was conducted in two subgroups (n=74 and n=41) measuring 17 cytokines and 27 lymphocyte phenotypes respectively. Findings: 1) Despite a strong association of clinical and laboratory variables with the outcomes in classical pairwise analysis, the contribution of laboratory tests to the combined prediction power was limited by redundancy. Laboratory variables reflected only two types of processes: inflammation and organ damage but none reflected the immune response, one major determinant of prognosis. 2) Eight of the thirty variables: age, comorbidity index, oxygen saturation to fraction of inspired oxygen ratio, neutrophil-lymphocyte ratio, C-reactive protein, aspartate aminotransferase/alanine aminotransferase ratio, fibrinogen, and glomerular filtration rate captured most of the combined statistical predictive power. 3) The interpretation of clinical and laboratory variables was moderately improved by grouping them in two categories i.e., inflammation related biomarkers and organ damage related biomarkers; Age and organ damage-related biomarker tests were the best predictors of survival, and inflammatory-related ones were the best predictors of severity. 4) The pilot study identified immunological tests (CXCL10, IL-6, IL-1RA and CCL2), that performed better than most currently used laboratory tests. Conclusions: Laboratory tests for clinical management of COVID 19 patients are valuable but limited predictors due to redundancy; this limitation could be overcome by adding immunological tests with independent predictive power. Understanding the limitations of tests in use would improve their interpretation and simplify clinical management but a systematic search for better immunological biomarkers is urgent and feasible.

Lymphocytic Thyroiditis Transcriptomic Profiles Support the Role of Checkpoint Pathways and B Cells in Pathogenesis.

Background: Autoimmune thyroid diseases are the most common types of autoimmune diseases, but their physiopathology is still relatively unexplored. Genotype-tissue expression (GTEx) is a publicly available repository containing RNAseq data, including profiles from thyroid. Approximately 14.8% of these glands were affected by focal lymphocytic thyroiditis and 6.3% were annotated as Hashimoto. We interrogated these data to improve the characterization of infiltrating cells and to identify new molecular pathways active in autoimmune thyroiditis. Materials and Methods: Histological GTEx images of 336 thyroid samples were classified into three categories, that is, non-infiltrated thyroid, small focal infiltrated thyroid, and extensive lymphoid infiltrated thyroid. Differentially expressed genes among these categories were identified and subjected to in silico pathway enrichment analysis accordingly. CIBERSORTx deconvolution was used to characterize infiltrating cells. Results: As expected, most of the transcriptional changes were dependent on tissue infiltration. Upregulated genes in tissues include-in addition to lineage-specific B and T cell genes-a broad representation of inhibitory immune checkpoint receptors expressed by B and T lymphocytes. CIBERSORTx analysis identified 22 types of infiltrating cells showed that T cells predominate 3:1 over B cells in glands with small infiltrates, only by 1.7:1 in those with large infiltrates. Follicular helper and memory CD4 T cells were significantly more abundant in glands with large infiltrates (p < 0.0001), but the most prominent finding in these glands was an almost sixfold increase in the number of naive B cells (p < 0.0001). A predominance of M2 macrophages over M1 and M0 macrophages was observed in the three gland categories (p < 0.001). Conclusions: Analysis of transcriptomic RNA-seq profiles constitutes a rich source of information for the analysis of autoimmune tissues. High-resolution transcriptomic data analysis of thyroid glands indicates the following: (a) in all infiltrated glands, active autoimmune response coexists with suppressor counteracting mechanisms involving several inhibitory checkpoint receptor pairs, (b) glands with small infiltrates contain an unexpected relatively high proportion of B lymphocytes, and (c) in highly infiltrated glands, there is a distinct transcriptomic signature of active tertiary lymphoid organs. These results support the concept that the autoimmune response is amplified in the thyroid tissue.

Stratification of hospitalized COVID-19 patients into clinical severity progression groups by immuno-phenotyping and machine learning.

Quantitative or qualitative differences in immunity may drive clinical severity in COVID-19. Although longitudinal studies to record the course of immunological changes are ample, they do not necessarily predict clinical progression at the time of hospital admission. Here we show, by a machine learning approach using serum pro-inflammatory, anti-inflammatory and anti-viral cytokine and anti-SARS-CoV-2 antibody measurements as input data, that COVID-19 patients cluster into three distinct immune phenotype groups. These immune-types, determined by unsupervised hierarchical clustering that is agnostic to severity, predict clinical course. The identified immune-types do not associate with disease duration at hospital admittance, but rather reflect variations in the nature and kinetics of individual patient's immune response. Thus, our work provides an immune-type based scheme to stratify COVID-19 patients at hospital admittance into high and low risk clinical categories with distinct cytokine and antibody profiles that may guide personalized therapy.

Effects of feeds, supplemented with humic substances and calcium carbonate, on performance, egg quality and heart rate variability in laying hens.

This study aimed to determine the effects of in-feed inclusion of humic substances (HS) and/or calcium carbonate (CaCO3) on the performance and welfare of laying hens. A total number of 144 Hy-Line Brown laying hens (55 weeks old) were randomly assigned to four treatments, T1-T4 (36 birds per treatment). T1 hens were fed on a control diet without HS or CaCO3, T2 hens were fed on the control diet + 2.00 g per bird per day CaCO3, T3 hens were fed on the control diet + 0.20% HS and T4 hens were fed on the control diet + 0.20% HS + 2.00 g/bird/day CaCO3. The experiment started after 15 days of adaptation and lasted 8 weeks. The parameters evaluated were percentage of hen-day egg production, food consumption, mortality, egg quality parameters and heart rate variability (welfare indicator). Hens in the T3 group showed a significantly lower feed intake than those in the other three groups, however, significantly higher daily egg production was recorded in groups T3 and T4 compared to T1 and T2. Eggshell quality characteristics were significantly improved by HS supplementation and eggs laid by hens in groups T1, and T4 presented paler yolk and shell color than those in the other groups. In conclusion, these results indicated that in-feed inclusion of HS had a beneficial effect on laying hens' productive performance including egg production and eggshells quality.

Peripheral and lung resident memory T cell responses against SARS-CoV-2.

Resident memory T cells (TRM) positioned within the respiratory tract are probably required to limit SARS-CoV-2 spread and COVID-19. Importantly, TRM are mostly non-recirculating, which reduces the window of opportunity to examine these cells in the blood as they move to the lung parenchyma. Here, we identify circulating virus-specific T cell responses during acute infection with functional, migratory and apoptotic patterns modulated by viral proteins and associated with clinical outcome. Disease severity is associated predominantly with IFNγ and IL-4 responses, increased responses against S peptides and apoptosis, whereas non-hospitalized patients have increased IL-12p70 levels, degranulation in response to N peptides and SARS-CoV-2-specific CCR7+ T cells secreting IL-10. In convalescent patients, lung-TRM are frequently detected even 10 months after initial infection, in which contemporaneous blood does not reflect tissue-resident profiles. Our study highlights a balanced anti-inflammatory antiviral response associated with a better outcome and persisting TRM cells as important for future protection against SARS-CoV-2 infection.

Cor Triatriatum Dexter: an unusual cause of neonatal cyanosis / Cor Triatriatum Dexter: una causa infrecuente de cianosis neonatal

We present the case of a term newborn, with no significant perinatal history, who was taken to the emergency room at 18 days old for intermittent episodes of cyanosis, with no signs of respiratory distress, oxygensaturation of 85%, arterial gases with moderate hypoxemia, and chest X-ray.

Subclinical cardiac alterations detected by echocardiography in Mexican schoolchildren with overweight and obesity.

Objective: The purpose of this study was to evaluate early changes in myocardial function in overweight and obese children without hypertension. Methods: Cross-sectional study involving 150 participants of both sexes between 6 and 15 years old. Anthropometric and biochemical evaluations were performed. Ventricular function was assessed by conventional echocardiographic methods and myocardial deformation analysis by two-dimensional speckle tracking echocardiography. One-way analysis of variance was employed for the global comparison of study variables between groups (children with normal weight, overweight and obesity), and post hoc analysis with Bonferroni correction was used for multiple comparison, considering normal-weight children as the reference category. Results: Overall, 142 participants were included, 50 (35%) with normal weight, 39 (28%) overweight and 53 (37%) obesity. Diastolic diameter of the left ventricular (LV) and interventricular septum, diameter of the left atrium and LV mass were significantly higher in children with obesity compared to those with normal weight. No significant differences in the conventional indicators of LV systolic and diastolic function were found between groups. Significant differences in the regional myocardial deformation between the three groups were observed. Mean global longitudinal myocardial deformation was smaller in patients with obesity (-20.9% vs. -23.5%, p < 0.05) compared to children with normal weight. Conclusions: The childhood obesity was associated with altered myocardial deformation, even in the presence of normal ejection fraction. Myocardial deformation evaluation is relevant in the assessment of pediatric patients with obesity.

Objetivo: El propósito de este estudio fue evaluar los cambios tempranos en la función miocárdica en niños con sobrepeso y obesidad, sin hipertensión arterial. Métodos: Estudio transversal en el que se incluyeron 150 participantes de ambos sexos entre 6 y 15 años. Se realizaron evaluaciones antropométricas, bioquímicas y de función ventricular mediante métodos ecocardiográficos convencionales y análisis de deformación miocárdica con ecocardiografía bidimensional speckle tracking. La comparación global entre los grupos de estudio (niños con peso normal, sobrepeso y obesidad) se llevó a cabo con la prueba de análisis de varianza (ANOVA) de una vía y análisis post hoc con corrección de Bonferroni para las comparaciones múltiples, y se consideró a los niños con peso normal como grupo de referencia. Resultados: La muestra final fue de 142 participantes, 50 (35%) con peso normal, 39 (28%) con sobrepeso y 53 (37%) con obesidad. El diámetro diastólico del ventrículo izquierdo (VI) y el septum interventricular, y el diámetro de la aurícula izquierda (AI) y la masa del VI fueron significativamente más altos en el grupo con obesidad en comparación con el grupo con peso normal. No se observaron diferencias significativas en los indicadores convencionales de la función sistólica y diastólica ventricular izquierda. Se observaron diferencias significativas en la deformación miocárdica regional entre los tres grupos. La media de deformación miocárdica longitudinal global fue más baja en los pacientes con obesidad (−20.9% vs. −23.5%; p < 0.05) en comparación con los niños con peso normal. Conclusiones: La obesidad infantil se asoció a alteraciones en la deformación miocárdica, incluso en presencia de fracción de expulsión normal. La evaluación de la deformación miocárdica es relevante en los pacientes pediátricos con obesidad.

Alteraciones cardíacas subclínicas detectadas por ecocardiografía en escolares Mexicanos con sobrepeso y obesidad / Subclinical cardiac alterations detected by echocardiography in Mexican schoolchildren with overweight and obesity

Resumen Objetivo: El propósito de este estudio fue evaluar los cambios tempranos en la función miocárdica en niños con sobrepeso y obesidad, sin hipertensión arterial. Métodos: Estudio transversal en el que se incluyeron 150 participantes de ambos sexos entre 6 y 15 años. Se realizaron evaluaciones antropométricas, bioquímicas y de función ventricular mediante métodos ecocardiográficos convencionales y análisis de deformación miocárdica con ecocardiografía bidimensional speckle tracking. La comparación global entre los grupos de estudio (niños con peso normal, sobrepeso y obesidad) se llevó a cabo con la prueba de análisis de varianza (ANOVA) de una vía y análisis post hoc con corrección de Bonferroni para las comparaciones múltiples, y se consideró a los niños con peso normal como grupo de referencia. Resultados: La muestra final fue de 142 participantes, 50 (35%) con peso normal, 39 (28%) con sobrepeso y 53 (37%) con obesidad. El diámetro diastólico del ventrículo izquierdo (VI) y el septum interventricular, y el diámetro de la aurícula izquierda (AI) y la masa del VI fueron significativamente más altos en el grupo con obesidad en comparación con el grupo con peso normal. No se observaron diferencias significativas en los indicadores convencionales de la función sistólica y diastólica ventricular izquierda. Se observaron diferencias significativas en la deformación miocárdica regional entre los tres grupos. La media de deformación miocárdica longitudinal global fue más baja en los pacientes con obesidad (−20.9% vs. −23.5%; p menor 0.05) en comparación con los niños con peso normal. Conclusiones: La obesidad infantil se asoció a alteraciones en la deformación miocárdica, incluso en presencia de fracción de expulsión normal. La evaluación de la deformación miocárdica es relevante en los pacientes pediátricos con obesidad.

Abstract Objective: The purpose of this study was to evaluate early changes in myocardial function in overweight and obese children without hypertension. Methods: Cross-sectional study involving 150 participants of both sexes between 6 and 15 years old. Anthropometric and biochemical evaluations were performed. Ventricular function was assessed by conventional echocardiographic methods and myocardial deformation analysis by two-dimensional speckle tracking echocardiography. One-way analysis of variance was employed for the global comparison of study variables between groups (children with normal weight, overweight and obesity), and post hoc analysis with Bonferroni correction was used for multiple comparison, considering normal-weight children as the reference category. Results: Overall, 142 participants were included, 50 (35%) with normal weight, 39 (28%) overweight and 53 (37%) obesity. Diastolic diameter of the left ventricular (LV) and interventricular septum, diameter of the left atrium and LV mass were significantly higher in children with obesity compared to those with normal weight. No significant differences in the conventional indicators of LV systolic and diastolic function were found between groups. Significant differences in the regional myocardial deformation between the three groups were observed. Mean global longitudinal myocardial deformation was smaller in patients with obesity (−20.9% vs. −23.5%, p less 0.05) compared to children with normal weight. Conclusions: The childhood obesity was associated with altered myocardial deformation, even in the presence of normal ejection fraction. Myocardial deformation evaluation is relevant in the assessment of pediatric patients with obesity.

Regulation of TSHR Expression in the Thyroid and Thymus May Contribute to TSHR Tolerance Failure in Graves' Disease Patients via Two Distinct Mechanisms.

Graves' disease (GD) involves the presence of agonistic auto-antibodies against the thyrotropin receptor (TSHR), which are responsible for the clinical symptoms. While failure of TSHR tolerance is central to GD pathogenesis, the process leading to this failure remains poorly understood. Two mechanisms intimately linked to tolerance have been proposed to explain the association of SNPs located in TSHR intron 1 to GD: (1) differential alternative splicing in the thyroid; and (2) modulation of expression in the thymus. To elucidate the relative contribution to these two mechanisms to GD pathogenesis, we analyzed the level of full-length and ST4 and ST5 isoform expression in the thyroid (n = 49) and thymus (n = 39) glands, and the influence of intron 1-associated SNPs on such expression. The results show that: (1) the level of flTSHR and ST4 expression in the thymus was unexpectedly high (20% that of the thyroid); (2) while flTSHR is the predominant isoform, the levels are similar to ST4 (ratio flTSHR/ST4 = 1.34 in the thyroid and ratio flTSHR/ST4 in the thymus = 1.93); (3) next-generation sequencing confirmed the effect of the TSHR intron 1 polymorphism on TSHR expression in the thymus with a bias of 1.5 ± 0.2 overexpression of the protective allele in the thymus compared to the thyroid; (4) GD-associated intron 1 SNPs did not influence TSHR alternative splicing of ST4 and ST5 in the thyroid and thymus; and (5) three-color confocal imaging showed that TSHR is associated with both thymocytes, macrophages, and dendritic cells in the thymus. Our findings confirm the effect of intron 1 polymorphisms on thymic TSHR expression and we present evidence against an effect on the relative expression of isoforms. The high level of ST4 expression in the thymus and its distribution within the tissue suggest that this would most likely be the isoform that induces central tolerance to TSHR thus omitting most of the hinge and transmembrane portion. The lack of central tolerance to a large portion of TSHR may explain the relatively high frequency of autoimmunity related to TSHR and its clinical consequence, GD.

Analysis of the PD-1/PD-L1 axis in human autoimmune thyroid disease: Insights into pathogenesis and clues to immunotherapy associated thyroid autoimmunity.

Autoimmune thyroid diseases (AITDs), i.e., Graves' disease (GD) and Hashimoto thyroiditis (HT), are the most prevalent organ-specific autoimmune diseases, but their pathogenesis is still incompletely understood. The PD-1/PD-L1 pathway is an important mechanism of peripheral tolerance that has not been investigated in AITDs. Here, we report the analysis of the expression of PD-1, PD-L1 and PD-L2 in PBMCs, infiltrating thyroid lymphocytes (ITLs) and in thyroid follicular cells (TFCs) in GD, HT and multinodular goiter (MNG) patients and healthy controls PBMCs (HC). By combining flow cytometry, tissue immunofluorescence and induction experiments on primary and thyroid cell line cultures, we show that: 1) while PD-1+ T cells are moderately expanded in PBMCs from GD vs HC, approximately half of T cells in the infiltrate are PD-1+ including some PD-1hi; 2) PD-L1, but not PD-L2, is expressed by 81% of GD glands and in 25% of non-autoimmune glands; 3) PD-L1, was expressed by TFCs in areas that also contain abundant PD-1 positive T cells but; 4) co-localization in TFCs indicated only partial overlap between the smaller areas of the PD-L1+ and the larger areas of HLA class II+ expression; 5) IFNγ is capable of inducing PD-L1 in >90% of TFCs in primary cultures and cell lines. Collectively these results indicate that the PD-1/PD-L1 axis is operative in AITD glands and may restrain the autoimmune response. Yet the discrepancy between easy induction in vitro and the limited expression in vivo (compared to HLA) suggests that PD-L1 expression in vivo is partially inhibited in GD and HT glands. In conclusions 1) the PD-1/PD-L1 pathway is activated in AITD glands but probably not to the extent to inhibit disease progression and 2) Thyroid autoimmunity arising after PD-1/PD-L1 blocking therapies in cancer patients may result from interfering PD-1/PD-L1 tolerance mechanism in thyroid with minimal (focal) thyroiditis. Finally acting on the PD-1/PD-L1 pathway could be a new approach to treat AITD and other organ-specific autoimmunity in the future.

Extended immunophenotyping reference values in a healthy pediatric population.

BACKGROUND: For the accurate diagnosis of immunodeficiencies is crucial to compare patients' immunology laboratory values with age-sex matched controls, yet there is a paucity of normal values for most populations. OBJECTIVES: To define appropriate reference values of extended lymphocyte subpopulations and T-cell receptor excision circle (TRECs) levels in healthy pediatric donors between 1 month and 18 years of age. METHODS: Extended immunophenotyping values were obtained by analysis of multiparameter flow cytometry panels for the following subpopulations: CD4+ and CD8+ Naive, Effector, Effector Memory and Central Memory, T helper subpopulations and their degrees of activation, T Regulatory cells, Recent Thymic Emigrants (RTE), B Lymphocyte subpopulations (Transitional, Naive, Preswitch-Memory, Switch-Memory, Plasmablasts, CD21low, and Exhausted), and subpopulations for Monocytes, NK cells and Dendritic Cells. RESULTS: Median values and the 10th and 90th percentiles were obtained for 32 lymphocyte and monocyte subpopulations, and for TRECs levels in each age group of children. Naive CD4+ and CD8+ T-cell populations tended to decrease with age, with significant difference between the groups, in parallel with the reduction in thymic function assessed by TRECs counts and the recent thymic emigrant population. Relative numbers of Th cell populations tended to increase with age. The percentage of class-switched B cell populations showed a significant increase between the youngest group and the others. CONCLUSION: This study provides essential data for interpreting extended immunophenotyping profiles in the pediatric and young adult populations, which could be of value for the diagnosis of PIDs and immune-mediated diseases, particularly those associated with subtle immunological abnormalities. © 2018 International Clinical Cytometry Society.

Subclinical cardiac alterations detected by echocardiography in Mexican schoolchildren with overweight and obesity.

Objective: The purpose of this study was to evaluate early changes in myocardial function in overweight and obese children without hypertension. Methods: This was a cross-sectional study involving 150 participants of both sexes between 6 and 15 years old. Anthropometric and biochemical evaluations were performed. Ventricular function was assessed by conventional echocardiographic methods and myocardial deformation analysis by two-dimensional speckle tracking echocardiography. One-way analysis of variance was employed for the global comparison of study variables between groups (children with normal weight, overweight, and obesity), and post hoc analysis with Bonferroni correction was used for multiple comparisons, considering normal weight children as the reference category. Results: Overall, 142 participants were included, 50 (35%) with normal weight, 39 (28%) overweight, and 53 (37%) obesity. The diastolic diameter of the left ventricular and interventricular septum, and diameter of the left atrium and LV mass were significantly higher in children with obesity compared to those with normal weight. No significant differences in the conventional indicators of LV systolic and diastolic function were found between groups. Significant differences in the regional myocardial deformation between the three groups were observed. Mean global longitudinal myocardial deformation was smaller in patients with obesity (-20.9% vs. -23.5%, p < 0.05) compared to children with normal weight. Conclusions: The childhood obesity was associated with altered myocardial deformation, even in the presence of normal ejection fraction. Myocardial deformation evaluation is relevant in the assessment of pediatric patients with obesity.

Objetivo: El propósito de este estudio fue evaluar los cambios tempranos en la función miocárdica en niños con sobrepeso y obesidad, sin hipertensión arterial. Métodos: Estudio transversal en el que se incluyeron 150 participantes de ambos sexos entre 6 y 15 años. Se realizaron evaluaciones antropométricas, bioquímicas y de función ventricular mediante métodos ecocardiográficos convencionales y análisis de deformación miocárdica con ecocardiografía bidimensional speckle tracking. La comparación global entre los grupos de estudio (niños con peso normal, sobrepeso y obesidad) se llevó a cabo con la prueba de análisis de varianza (ANOVA) de una vía y análisis post hoc con corrección de Bonferroni para las comparaciones múltiples, y se consideró a los niños con peso normal como grupo de referencia. Resultados: La muestra final fue de 142 participantes, 50 (35%) con peso normal, 39 (28%) con sobrepeso y 53 (37%) con obesidad. El diámetro diastólico del ventrículo izquierdo (VI) y el septum interventricular, y el diámetro de la aurícula izquierda (AI) y la masa del VI fueron significativamente más altos en el grupo con obesidad en comparación con el grupo con peso normal. No se observaron diferencias significativas en los indicadores convencionales de la función sistólica y diastólica ventricular izquierda. Se observaron diferencias significativas en la deformación miocárdica regional entre los tres grupos. La media de deformación miocárdica longitudinal global fue más baja en los pacientes con obesidad (−20.9% vs. −23.5%; p < 0.05) en comparación con los niños con peso normal. Conclusiones: La obesidad infantil se asoció a alteraciones en la deformación miocárdica, incluso en presencia de fracción de expulsión normal. La evaluación de la deformación miocárdica es relevante en los pacientes pediátricos con obesidad.