Luiz Carlos Conti de Freitas

Background: Endoplasmic reticulum stress (ERS) and the unfolded protein response (UPR) are adaptive mechanisms for conditions of high protein demand, marked by an accumulation of misfolded proteins in the endoplasmic reticulum (ER). Rheumatic autoimmune diseases (RAD) are known to be associated with chronic inflammation and an ERS state. However, the activation of UPR signaling pathways is not completely understood in Sjögren’s disease (SD). This study evaluated the expression of ERS-related genes in glandular tissue of patients with primary SD (pSD), associated SD (aSD) with other autoimmune diseases, and non-Sjögren sicca syndrome (NSS). Methods: In a cross-sectional study, minor salivary gland biopsies were obtained from 44 patients with suspected SD and 13 healthy controls (HC). Patients were classified as pSD, aSD, or NSS based on clinical, serological, and histological assessment. Histopathological analysis and mRNA expression analysis of genes associated with ERS and UPR (PERK, XBP1, ATF-6, ATF-4, CANX, CALR, CHOP, and BIP) were performed on the samples. Differences between groups (pSD, aSD, NSS, and HC) were assessed. The influence of chloroquine (CQ) on the ER was also investigated. Results: Twenty-eight SD patients showed increased expression of PERK ( p = 0.0117) and XBP1 ( p = 0.0346), and reduced expression of ATF-6 ( p = 0.0003) and CHOP ( p = 0.0003), compared to the HC group. Increased expression of BIP ( p < 0.0001), PERK ( p = 0.0003), CALR ( p < 0.0001), and CANX ( p = 0.0111) was also observed in the SD group compared to the NSS group ( n = 16). Patients receiving CQ ( n = 16) showed a significant increase in ATF-6 ( p = 0.0317) compared to patients not taking the medication ( n = 29). Conclusions: Altogether, the results suggest a greater activation of the ERS and UPR genes in patients with SD, especially in the pSD group. Antimalarial drugs, like CQ, used to treat RAD, may affect the ER function in exocrine glands.

Human leukocyte antigen (HLA)-G is an immune checkpoint molecule that is highly expressed in papillary thyroid carcinoma (PTC). The HLA-G gene presents several functional polymorphisms distributed across the coding and regulatory regions (5′URR: 5′ upstream regulatory region and 3′UTR: 3′ untranslated region) and some of them may impact HLA-G expression and human malignancy. To understand the contribution of the HLA-G genetic background in PTC, we studied the HLA-G gene variability in PTC patients in association with tumor morbidity, HLA-G tissue expression, and plasma soluble (sHLA-G) levels. We evaluated 185 PTC patients and 154 healthy controls. Polymorphic sites defining coding, regulatory and extended haplotypes were characterized by sequencing analyses. HLA-G tissue expression and plasma soluble HLA-G levels were evaluated by immunohistochemistry and ELISA, respectively. Compared to the controls, the G0104a_(5′URR)G*01:04:04_(coding)UTR-03_(3’UTR) extended haplotype was underrepresented in the PTC patients, while G0104a_(5′URR)G*01:04:01_(coding)UTR-03_(3′UTR) was less frequent in patients with metastatic and multifocal tumors. Decreased HLA-G tissue expression and undetectable plasma sHLA-G were associated with the G010102a_(5′URR)G*01:01:02:01_(coding)UTR-02_(3′UTR) extended haplotype. We concluded that the HLA-G variability was associated with PTC development and morbidity, as well as the magnitude of the encoded protein expression at local and systemic levels.

As BRAF, TERT, HLA-G, and microRNAs have been individually associated with papillary thyroid carcinoma (PTC), we aimed to evaluate the individual and collaborative role of these markers in PTC in the same patient cohort. HLA-G and BRAF tumor expression was evaluated by immunohistochemistry. Using molecular methods, BRAF^V600E and TERT promoter mutations were evaluated in thyroid fine needle aspirates. MicroRNA tumor profiling was investigated using massively parallel sequencing. We observed strong HLA-G (67.96%) while BRAF (62.43%) staining was observed in PTC specimens. BRAF overexpression was associated with poor response to therapy. The BRAF^V600E (52.9%) and TERT^C228T (13%) mutations were associated with extrathyroidal extension, advanced-age, and advanced-stage cancer. The TERT rs2853669 CC+TC genotypes (38%) were overrepresented in metastatic tumors. Nine modulated microRNAs targeting the BRAF, TERT, and/or HLA-G genes were observed in PTC and involved with cancer-related signaling pathways. The markers were individually associated with PTC features, emphasizing the synergistic effect of BRAF^V600E and TERT^C228T; however, their collaborative role on PTC outcome was not fully demonstrated. The differentially expressed miRNAs targeting the BRAF and/or HLA-G genes may explain their increased expression in the tumor milieu.

Dietary restriction (DR) reduces adiposity and improves metabolism in patients with one or more symptoms of metabolic syndrome. Nonetheless, it remains elusive whether the benefits of DR in humans are mediated by calorie or nutrient restriction. This study was conducted to determine whether isocaloric dietary protein restriction is sufficient to confer the beneficial effects of dietary restriction in patients with metabolic syndrome. We performed a prospective, randomized controlled dietary intervention under constant nutritional and medical supervision. Twenty-one individuals diagnosed with metabolic syndrome were randomly assigned for caloric restriction (CR; n = 11, diet of 5941 ± 686 KJ per day) or isocaloric dietary protein restriction (PR; n = 10, diet of 8409 ± 2360 KJ per day) and followed for 27 days. Like CR, PR promoted weight loss due to a reduction in adiposity, which was associated with reductions in blood glucose, lipid levels, and blood pressure. More strikingly, both CR and PR improved insulin sensitivity by 62.3% and 93.2%, respectively, after treatment. Fecal microbiome diversity was not affected by the interventions. Adipose tissue bulk RNA-Seq data revealed minor changes elicited by the interventions. After PR, terms related to leukocyte proliferation were enriched among the upregulated genes. Protein restriction is sufficient to confer almost the same clinical outcomes as calorie restriction without the need for a reduction in calorie intake. The isocaloric characteristic of the PR intervention makes this approach a more attractive and less drastic dietary strategy in clinical settings and has more significant potential to be used as adjuvant therapy for people with metabolic syndrome.

Parathyroid tumors are very prevalent conditions among endocrine tumors, being the second most common behind thyroid tumors. Secondary hyperplasia can occur beyond benign and malignant neoplasia in parathyroid glands. Adenomas are the leading cause of hyperparathyroidism, while carcinomas represent less than 1% of the cases. Tumor suppressor gene mutations such as MEN1 and CDC73 were demonstrated to be involved in tumor development in both familiar and sporadic types; however, the epigenetic features of the parathyroid tumors are still a little-explored subject. We present a review of epigenetic mechanisms related to parathyroid tumors, emphasizing advances in histone modification and its perspective of becoming a promising area in parathyroid tumor research.

Background Increasing evidence shows that chronic inflammation plays an important role in thyroid tumorigenesis. Cytokines as central mediators in inflammatory microenvironment can present both pro‐tumour and anti‐tumour effects and cytokine release may be influenced by soluble HLA‐G (sHLA‐G), an immune checkpoint molecule whose expression can also be induced by certain cytokines. Aim To understand the role of these soluble factors in papillary thyroid cancer (PTC). Methods We evaluated plasma levels of sHLA‐G and of 13 cytokines using ELISA and flow cytometry, respectively, in PTC patients at two time points: pre‐ and post‐thyroidectomy; and control subjects. Results Compared with controls, IL‐6 levels were increased, while IL‐1β, IFN‐α and TGF‐β1 levels were decreased in pre‐thyroidectomy PTC patients. IFN‐α and TGF‐β1 efficiently discriminated patients from controls and were associated with extrathyroidal extension and lymph node metastasis, respectively. In addition, TNF and IL‐13 were associated with male gender, lymph node metastasis and Hashimoto thyroiditis, and sHLA‐G with tumour invasion. Compared with pre‐thyroidectomy, IL‐4, IL‐10, TNF, IFN‐α and TGF‐β1 levels were increased in post‐thyroidectomy. Conclusion There are significant changes in the cytokine profile after surgical removal of the thyroid tumour, and IFN‐α e TGF‐β1 showed to be promising cytokines for discriminating PTC patients from controls. We also found that different cytokines are associated with clinicohistopathological characteristics of PTC related to poor prognosis, suggesting that cytokines seem to play an important role in PTC development and management.

Objetivo: O objetivo desse estudo foi avaliar a eficácia do diagnóstico e tratamento de lesões orais entre diferentes níveis de saúde. Material e Métodos: Todas as informações foram obtidas dos prontuários médicos de um Hospital Universitário e de fichas do Sistema Público de Regulação, considerando as características clínico-demográficas tal como, doenças orais diagnosticadas, sistema de referência e terapia empregada. A avaliação foi baseada na relação entre diagnóstico final, hipótese diagnóstica e prioridade para agendamento da consulta, também foi considerado o tempo decorrido entre o encaminhamento e o primeiro atendimento, data do diagnóstico final e início da terapia. Depois, a hipótese diagnóstica preenchida por profissionais de saúde do nível primário e terciário foram comparadas e a concordância entre ambos foi determinada. Resultados: Cerca de 51% dos encaminhamentos preenchidos pelos profissionais da atenção primária estavam incompletos, ou a hipótese diagnóstica estava descrita de forma inadequada. No nível terciário, o tempo médio entre o encaminhamento e a primeira consulta foi de 23 dias, a média entre o diagnóstico final e o início do tratamento foi de 22 dias. O tratamento cirúrgico foi o mais rápido (31 dias) quando comparado a radioterapia para lesões malignas (81 a 100 dias). Conclusão: É necessário melhorar o treinamento de profissionais de saúde da Atenção Básica no reconhecimento de lesões orais malignas. Claramente, o preenchimento adequado das hipóteses de diagnóstico, com o grau de prioridade correspondente acelera o diagnóstico e tratamento do paciente.

Background: Familial partial lipodystrophy of the Dunnigan type is one of the most common inherited lipodystrophies variables. These individuals have important metabolic disorders that cause predisposition to various diseases. In this study we aimed to demonstrate the relation between the metabolic abnormalities, inflammatory profile and the expression of genes involved in the activation of the endoplasmic reticulum stress (ERS) in subjects with FPLD. Methods: We evaluated 14 female FPLD patients and compared with 13 female healthy individuals. The subjects were paired with their respective BMI and age and categorized into two groups: Familial partial lipodystrophy of the Dunnigan type (FPLD) and control. Patients were fasted for 12 h before blood collection for measurement of HbA1c, glucose, insulin, lipids and inflammatory markers. Subcutâneous adipose tissue was collected by puncture aspiration of submental region during ambulatorial surgical aesthetic procedure. Results: We demonstrate that patients with FPLD show increased HbA1c (p < 0.01), fasting glucose (p < 0.002) and triglycerides (p < 0.005) while HDL/cholesterol (p < 0.001) was lower when compared to healthy individuals. We found that 64.2% FPLD patients had metabolic syndrome according to International Diabetes Federation definition. We also observe increased AUC of glucose (p < 0.001) and insulin during oGTT, featuring a frame of hyperglycemia and hyperinsulinemia, suggesting insulin resistance. Also we found hyperactivation of several genes responsible for ERS such as ATF-4 (p < 0.01), ATF-6 (p < 0.01), EIF2α3K (p < 0.005), CCT4 (p < 0.001), CHOP (p < 0.01), CALR (p < 0.001) and CANX (p < 0.005), that corroborate the idea that diabetes mellitus and metabolic syndrome are associated with direct damage to the endoplasmic reticulum homeostasis. Ultimately, we note that individuals with lipodystrophy have an increase in serum interleukins, keys of the inflammatory process, as IL-1β, TNF-α and IL-6 (p < 0.05 all), compared with healthy individuals, which can be the trigger to insulin resistance in this population. Conclusion: Individuals with FPLD besides having typical dysfunctions of metabolic syndrome, show a hyperactivation of ERS associated with increased systemic inflammatory profile, which together may explain the complex clinical aspect of this diseases. Trial registration HCRP no 6711/2012