Significance of Furin Expression in Thyroid Neoplastic Transformation.

Angiotensin-Converting Enzyme 2 (ACE2), Transmembrane Serine Protease 2 (TMPRSS2), and Furin were known to be key players in the SARS-CoV-2 infection, and the thyroid gland was revealed to be one of the relevant targets of the virus. Regardless of the viral infection, the expression of these molecules in the thyroid gland and their putative role in the neoplastic transformation of the thyrocytes has not been thoroughly explored. In this work, we aimed to characterize the mRNA and protein expression pattern of ACE2, TMPRSS2, and Furin in a series of patients with thyroid lesions. Our main results revealed a significantly decreased expression of ACE2 mRNA in the thyroid neoplasms in comparison to normal adjacent tissue. Furin mRNA was significantly increased in thyroid neoplasms when compared to normal adjacent tissue. In addition, a higher Furin mRNA level in thyroid carcinomas was associated with the presence of lymph node metastasis. Furin mRNA expression revealed a high discriminatory power between adjacent tissue and neoplasms. Protein expression of these molecules did not correlate with mRNA expression. Our study shows the mRNA downregulation of ACE2 and overexpression of Furin in thyroid neoplasms. Further studies are required to clarify if Furin expression can be a potential diagnostic indicator in thyroid neoplasia.

Generation of an Obese Diabetic Mouse Model upon Conditional Atrx Disruption.

Atrx loss was recently ascertained as insufficient to drive pancreatic neuroendocrine tumour (PanNET) formation in mice islets. We have identified a preponderant role of Atrx in the endocrine dysfunction in a Rip-Cre;AtrxKO genetically engineered mouse model (GEMM). To validate the impact of a different Cre-driver line, we used similar methodologies and characterised the Pdx1-Cre;AtrxKO (P.AtrxKO) GEMM to search for PanNET formation and endocrine fitness disruption for a period of up to 24 months. Male and female mice presented different phenotypes. Compared to P.AtrxWT, P.AtrxHOM males were heavier during the entire study period, hyperglycaemic between 3 and 12 mo., and glucose intolerant only from 6 mo.; in contrast, P.AtrxHOM females started exhibiting increased weight gains later (after 6 mo.), but diabetes or glucose intolerance was detected by 3 mo. Overall, all studied mice were overweight or obese from early ages, which challenged the histopathological evaluation of the pancreas and liver, especially after 12 mo. Noteworthily, losing Atrx predisposed mice to an increase in intrapancreatic fatty infiltration (FI), peripancreatic fat deposition, and macrovesicular steatosis. As expected, no animal developed PanNETs. An obese diabetic GEMM of disrupted Atrx is presented as potentially useful for metabolic studies and as a putative candidate for inserting additional tumourigenic genetic events.

Endocrine-disrupting chemicals and endocrine neoplasia: A forty-year systematic review.

INTRODUCTION: Endocrine disrupting chemicals (EDCs) are exogenous substances recognised as relevant tumourigenic chemicals. Studies show that even EDCs which were long abolished are still contributing to the increasing incidence of neoplasia. AIM: To investigate the association between human exposure to EDCs and the risk of endocrine-related tumours: breast, prostate, thyroid, uterus, testis, and ovary. METHODS: A systematic review using PubMed, Scopus, and Embase was conducted, searching for original observational studies published between 1980 and 2020, approaching EDCs exposure and endocrine tumourigenic risk in humans. We comprised neoplasia of six endocrine organs. We included all the studies on EDCs reporting tumour odds ratio, risk ratio, or hazard ratio. Study levels of confidence and risk of bias were accessed applying accredited guidelines. Human-made accidents and natural EDCs were not considered in the present study. RESULTS: Our search returned 3271 papers. After duplicate removal and screening, only 237 papers were included (corresponding to 268 records). EDCs were grouped from the most frequently (pesticides) to the least frequently studied (salts). The most tumourigenic EDC groups were phthalates (63%), heavy metals (54%), particulate matter (47%), and pesticides (46%). Pesticides group comprised the highest number of retrieved studies (n = 133). Increased neoplasia risk was found in 43-67% of the studies, with a lower value for ovary (43%) and a higher value for thyroid (67%). CONCLUSIONS: The innovative nature of our review comes from including human studies of six endocrine-related neoplasia aiming to understand the contribution of specific EDCs groups to each organ's tumourigenesis. Thyroid was the organ presenting the highest cancer risk after EDC exposure which may explain the increasing thyroid cancer incidence. However, detailed and controlled works reporting the effects of EDCs are scarce, probably justifying conflicting results. Multinational and multicentric human studies with biochemical analysis are needed to achieve stronger and concordant evidence.

An update on genetically engineered mouse models of pancreatic neuroendocrine neoplasms.

Pancreatic neuroendocrine neoplasms (PanNENs) are rare and clinically challenging entities. At the molecular level, PanNENs' genetic profile is well characterized, but there is limited knowledge regarding the contribution of the newly identified genes to tumor initiation and progression. Genetically engineered mouse models (GEMMs) are the most versatile tool for studying the plethora of genetic variations influencing PanNENs' etiopathogenesis and behavior over time. In this review, we present the state of the art of the most relevant PanNEN GEMMs available and correlate their findings with the human neoplasms' counterparts. We discuss the historic GEMMs as the most used and with higher translational utility models. GEMMs with Men1 and glucagon receptor gene germline alterations stand out as the most faithful models in recapitulating human disease; RIP-Tag models are unique models of early-onset, highly vascularized, invasive carcinomas. We also include a section of the most recent GEMMs that evaluate pathways related to cell cycle and apoptosis, Pi3k/Akt/mTOR, and Atrx/Daxx. For the latter, their tumorigenic effect is heterogeneous. In particular, for Atrx/Daxx, we will require more in-depth studies to evaluate their contribution; even though they are prevalent genetic events in PanNENs, they have low/inexistent tumorigenic capacity per se in GEMMs. Researchers planning to use GEMMs can find a road map of the main clinical features in this review, presented as a guide that summarizes the chief milestones achieved. We identify pitfalls to overcome, concerning the novel designs and standardization of results, so that future models can replicate human disease more closely.

Characterisation of an Atrx Conditional Knockout Mouse Model: Atrx Loss Causes Endocrine Dysfunction Rather Than Pancreatic Neuroendocrine Tumour.

ATRX is a chromatin remodeller that maintains telomere homeostasis. Loss of ATRX is described in approximately 10% of pancreatic neuroendocrine tumours (PanNETs) and associated with poorer prognostic features. Here, we present a genetically engineered mouse model (GEMM) addressing the role of Atrx loss (AtrxKO) in pancreatic ß cells, evaluating a large cohort of ageing mice (for up to 24 months (mo.)). Atrx loss did not cause PanNET formation but rather resulted in worsening of ageing-related pancreatic inflammation and endocrine dysfunction in the first year of life. Histopathological evaluation highlighted an exacerbated prevalence and intensity of pancreatic inflammation, ageing features, and hepatic steatosis in AtrxKO mice. Homozygous floxed mice presented hyperglycaemia, increased weights, and glucose intolerance after 6 months, but alterations in insulinaemia were not detected. Floxed individuals presented an improper growth of their pancreatic endocrine fraction that may explain such an endocrine imbalance. A pilot study of BRACO-19 administration to AtrxKO mice resulted in telomere instability, reinforcing the involvement of Atrx in the maintenance of ß cell telomere homeostasis. Thereby, a non-obese dysglycaemic GEMM of disrupted Atrx is here presented as potentially useful for metabolic studies and putative candidate for inserting additional tumourigenic genetic events.

Exercise performed during pregnancy positively modulates liver metabolism and promotes mitochondrial biogenesis of female offspring in a rat model of diet-induced gestational diabetes.

Gestational diabetes mellitus (GDM) is associated with a high-risk for metabolic complications in offspring. However, exercise is recognized as a non-pharmacological strategy against metabolic disorders and is recommended in GDM treatment. This study aimed to investigate whether gestational exercise (GE) could modulate maternal high-fat high-sucrose (HFHS) diet-related hepatic metabolic and mitochondrial outcomes in female offspring of mothers with HFHS-induced GDM. Female Sprague-Dawley rats were fed with control or HFHS diet and kept sedentary or submitted to GE. Their female offspring were fed with control diet and kept sedentary. Hepatic lipid accumulation, lipid metabolism regulators, mitochondrial biogenesis and dynamics markers, and microRNAs associated to the regulation of these markers were evaluated. Female offspring of GDM mothers showed increased body weight at early age, whereas GE prevented this effect of maternal HFHS-feeding and reduced hepatic lipid accumulation. GE stimulated hepatic mRNA transcription and protein expression of mitochondrial biogenesis markers (peroxisome proliferator-activated receptor-gamma co-activator-1alpha and mitochondrial transcription factor A) and mRNA transcription of mitochondrial dynamics markers (mitofusin-1, mitofusin-2, and dynamin-related protein-1) that were altered by maternal GDM, while mitochondrial dynamics markers protein expression was not affected by maternal diet/GE except for optic atrophy-1. MicroRNAs associated with these processes (miR-122, miR-34a, miR-130b, miR-494), and the expression of auto/mitophagy- and apoptosis-related proteins were not substantially influenced by altered intrauterine environment. Our findings suggest that GE is an important regulator of the intrauterine environment positively affecting liver metabolism and promoting liver mitochondrial biogenesis in female offspring despite eventual effects of maternal HFHS-feeding and related GDM.

Maternal high-fat high-sucrose diet and gestational exercise modulate hepatic fat accumulation and liver mitochondrial respiratory capacity in mothers and male offspring.

BACKGROUND: Maternal high-caloric nutrition and related gestational diabetes mellitus (GDM) are associated with a high-risk for developing metabolic complications later in life and in their offspring. In contrast, exercise is recognized as a non-pharmacological strategy against metabolic dysfunctions associated to lifestyle disorders. Therefore, we investigated whether gestational exercise delays the development of metabolic alterations in GDM mothers later in life, but also protects 6-week-old male offspring from adverse effects of maternal diet. METHODS: Female Sprague-Dawley rats were fed with either control (C) or high-fat high-sucrose (HFHS) diet to induce GDM and submitted to gestational exercise during the 3 weeks of pregnancy. Male offspring were sedentary and fed with C-diet. RESULTS: Sedentary HFHS-fed dams exhibited increased gestational body weight gain (p < 0.01) and glucose intolerance (p < 0.01), characteristic of GDM. Their offspring had normal glucose metabolism, but increased early-age body weight, which was reverted by gestational exercise. Gestational exercise also reduced offspring hepatic triglycerides accumulation (p < 0.05) and improved liver mitochondrial respiration capacity (p < 0.05), contributing to the recovery of liver bioenergetics compromised by maternal HFHS diet. Interestingly, liver mitochondrial respiration remained increased by gestational exercise in HFHS-fed dams despite prolonged HFHS consumption and exercise cessation. CONCLUSIONS: Gestational exercise can result in liver mitochondrial adaptations in GDM animals, which can be preserved even after the exercise program cessation. Exposure to maternal GDM programs liver metabolic setting of male offspring, whereas gestational exercise appears as an important preventive tool against maternal diet-induced metabolic alterations.

Relevant dose of the environmental contaminant, tributyltin, promotes histomorphological changes in the thyroid gland of male rats.

Organotin compounds, such as tributyltin (TBT), are common environmental contaminants and suspected endocrine-disrupting chemicals. Tributyltin is found in antifouling paints, widely used in ships and other vessels. The present study evaluated whether a 15-day treatment with TBT at a dose of 100 ng/kg/day could induce histomorphological changes in the thyroid gland of rats. TBT promoted relevant alterations in the thyroid architecture, being the most relevant histological findings the presence of increased number of small-size follicles in the treated group. In qualitative analyses, colloid vacuolization, papillary budging structures, cystic degeneration and chronic thyroiditis, were observed. Moreover, histomorphometric analysis showed statistically significant changes in the follicular architecture of TBT-treated rats, mainly a decrease in the follicle area (colloid) and an increased epithelial height that resulted in an increased epithelial height/colloid ratio. Augmented collagen deposition was also seen in the thyroids of treated groups. In immunohistochemical (IHC) analyses, the localization of NIS protein was described and a significant increased proliferation index (evaluated by Ki67 positive cells) in the treated group was reported. As an indirect measurement of oxidative stress, mitochondrial protein SDHA was also analyzed by IHC analysis. Although the cytoplasmic expression of SDHA was observed in both groups, the staining intensity score was higher in TBT-treated group. Our results suggest that besides causing histomorphological changes, environmental relevant dose of TBT treatment can also induce oxidative alterations.

Understanding Resistance vs. Susceptibility in Visceral Leishmaniasis Using Mouse Models of Leishmania infantum Infection.

Every year, up to 90,000 new cases of Visceral Leishmaniasis and 30,000 resultant deaths are estimated to occur worldwide. Such numbers give relevance to the continuous study of this complex form of the disease: a zoonosis and an anthroponosis; two known etiological agents (Leishmania infantum and L. donovani, respectively); with an estimated average ratio of 1 symptomatic per 10 asymptomatic individuals; and sometimes associated with atypical clinical presentations. This complexity, which results from a long co-evolutionary process involving vector-host, host-pathogen, and pathogen-vector interactions, is still not completely understood. The determinants of visceralization are not fully defined and the dichotomy resistance vs. susceptibility remains unsolved, translating into obstacles that delay the progress of global disease control. Inbred mouse models, with different susceptibility patterns to Leishmania infection, have been very useful in exploring this dichotomy. BALB/c and C57BL/6 mice were described as susceptible strains to L. donovani visceral infection, while SV/129 was considered resistant. Here, we used these three mouse models, but in the context of L. infantum infection, the other Leishmania species that cause visceral disease in humans, and dynamically compared their local and systemic infection-induced immune responses in order to establish a parallel and to ultimately better understand susceptibility vs. resistance in visceral leishmaniasis. Overall, our results suggest that C57BL/6 mice develop an intermediate "infection-phenotype" in comparison to BALB/c and SV/129 mouse strains, considering both the splenic parasite burden and the determined target organs weights. However, the immune mechanisms associated with the control of infection seem to be different in each mouse strain. We observed that both BALB/c and SV/129, but not C57BL/6 mice, show an infection-induced increase of splenic T follicular helper cells. On the other hand, differences detected in terms of CD21 expression by B cells early after infection, together with the quantified anti-Leishmania specific antibodies, suggest that SV/129 are faster than BALB/c and C57BL/6 mice in the assembly of an efficient B-cell response. Additionally, we observed an infection-induced increase in polyfunctional CD4+ T cells in the resistant SV/129 model, opposing an infection-induced increase in CD4+IL-10+ cells in susceptible BALB/c mice. Our data aligns with the observations reported for L. donovani infection and suggest that not only a single mechanism, but an interaction of several could be necessary for the control of this parasitic disease.

Telomere Maintenance Mechanisms in Cancer.

Tumour cells can adopt telomere maintenance mechanisms (TMMs) to avoid telomere shortening, an inevitable process due to successive cell divisions. In most tumour cells, telomere length (TL) is maintained by reactivation of telomerase, while a small part acquires immortality through the telomerase-independent alternative lengthening of telomeres (ALT) mechanism. In the last years, a great amount of data was generated, and different TMMs were reported and explained in detail, benefiting from genome-scale studies of major importance. In this review, we address seven different TMMs in tumour cells: mutations of the TERT promoter (TERTp), amplification of the genes TERT and TERC, polymorphic variants of the TERT gene and of its promoter, rearrangements of the TERT gene, epigenetic changes, ALT, and non-defined TMM (NDTMM). We gathered information from over fifty thousand patients reported in 288 papers in the last years. This wide data collection enabled us to portray, by organ/system and histotypes, the prevalence of TERTp mutations, TERT and TERC amplifications, and ALT in human tumours. Based on this information, we discuss the putative future clinical impact of the aforementioned mechanisms on the malignant transformation process in different setups, and provide insights for screening, prognosis, and patient management stratification.