Raman spectroscopy for the preoperative diagnosis of thyroid cancer and its subtypes: An in vitro proof-of-concept study.

OBJECTIVE: In 2016, there were an estimated 56 870 new cases of thyroid cancer (TC) in the USA. Fine needle aspiration cytology (FNAC) is the most safe, accurate and cost-effective method for the initial investigation of thyroid nodules. FNAC is limited by the inability to diagnose malignancy in follicular-patterned lesions accurately and, as a result, 20%-30% of cases under investigation for TC are classified as cytologically indeterminate, illustrating a problem with current FNAC procedure. Raman spectroscopy has shown promising results for the detection of many cancers; however, to date there has been no report on the performance of Raman spectroscopy on thyroid cytological samples. The aim of this study was to examine whether Raman spectroscopy could be used to correctly classify cell lines representing benign thyroid cells and various subtypes of TC. METHODS: A benign thyroid cell line and seven TC cell lines were prepared as ThinPrep® cytology slides and analysed with Raman spectroscopy. Principal components analysis and linear discriminant analysis were implemented to develop effective diagnostic algorithms for classification of Raman spectra of different TC subtypes. RESULTS: The spectral differences separating benign and TC cell lines were assigned to differences in the composition of nucleic acids, lipids, carbohydrates and protein in the benign and cancer cells. Good sensitivities (74%-85%), specificities (65%-93%) and diagnostic accuracies (71%-88%) were achieved for the identification of TC. CONCLUSION: These findings suggest that Raman spectroscopy has potential for preoperative TC diagnosis on FNAC samples.

Transcriptomic profiling of the response to excess iodide in Keap1 hypomorphic mice reveals new gene-environment interactions in thyroid homeostasis.

Iodide plays a pivotal role in thyroid homeostasis due to its crucial involvement in thyroid hormone biosynthesis. Exposure to pharmacological doses of iodide elicits in the thyroid an autoregulatory response to preserve thyroid function, as well as an antioxidant response that is mediated by the Keap1/Nrf2 signaling pathway. The objective of the present study was to investigate the transcriptional response of the thyroid to excess iodide in a background of enhanced Nrf2 signaling. Keap1 knockdown (Keap1KD) mice that have activated Nrf2 signaling were exposed or not to excess iodide in their drinking water for seven days and compared to respective wild-type mice. RNA-sequencing of individual mouse thyroids identified distinct transcriptomic patterns in response to iodide, with Keap1KD mice showing an attenuated inflammatory response, altered thyroidal autoregulation, and enhanced cell growth/proliferative signaling, as confirmed also by Western blotting for key proteins involved in antioxidant, autoregulatory and proliferative responses. These findings underscore novel gene-environment interactions between the activation status of the Keap1/Nrf2 antioxidant response system and the dietary iodide intake, which may have implications not only for the goiter phenotype of Keap1KD mice but also for humans harboring genetic variations in KEAP1 or NFE2L2 or treated with Nrf2-modulating drugs.

Traditional Chinese Medicine for Hashimoto's Thyroiditis: Focus on Selenium and Antioxidant Phytochemicals.

Hashimoto's thyroiditis (HT) is not only the most frequent autoimmune thyroid disease (AITD), but it also has a significant impact on patients' health-related quality of life (HRQoL), and it has been variably associated with differentiated thyroid carcinoma. Even though its pathogenesis is still incompletely understood, oxidative stress is believed to play an important role. Hypothyroidism related to later stages of HT can be treated with levothyroxine substitution therapy; various approaches such as selenium supplementation and iodine-restricted diets have been proposed as disease-modifying treatments for earlier stages, and even thyroidectomy has been suggested for refractory cases of painful HT. Nevertheless, many patients still report suboptimal HRQoL, highlighting an unmet medical need in this area. The concepts and approaches of traditional Chinese medicine (TCM) in treating HT are not broadly known in the West. Here, we provide an overview of TCM for HT, including combinations of TCM with selenium. We encompass evidence from clinical trials and other studies related to complex TCM prescriptions, single herbs used in TCM, and phytochemicals; wherever possible, we delineate the probable underlying molecular mechanisms. The findings show that the main active components of TCM for HT have commonly known or presumed antioxidant and anti-inflammatory activities, which may account for their potential utility in HT. Further exploring the practices of TCM for HT and combining them with evidence- and mechanism-based approaches according to Western standards may help to identify new strategies to alter the clinical course of the disease and/or to treat patients' symptoms better and improve their HRQoL.

Rare types of rotaviruses isolated from children with acute gastroenteritis in Patras, Greece.

BACKGROUND: Acute diarrhea remains a major public health issue worldwide, with gastroenteritis agents associated with a high morbidity, especially in infants and young children. A 1-year study (2009-2010) of rotavirus (RV)-caused pediatric diarrhea was performed on hospitalized children admitted with symptoms of acute diarrhea to the University Hospital of Patras, Greece. METHODS: Twenty-nine fecal samples were investigated for RVs, adenoviruses (AdVs), and enteroviruses (EVs) in an attempt to characterize these enteric viruses, which have been implicated in hospitalized diarrhea. RESULTS: A 44.8% (13/29) incidence of viral infection was reported for the viral targets. Monoinfections accounted for 31% (9/29), while bi-infections accounted for 13.4% (4/29). Sequencing of positive samples allowed identification of RVs G4P[8] (2/29), G9P[8] (1/29), and interestingly of the rare type G12P[8] (2/29). AdV types 1, 2, and 6 were also identified in 4/29, 1/29, and 1/29 of the samples, respectively. Regarding the EVs, EV71 (2/29), coxsackievirus A4 (1/29), echovirus 11 (1/29), and EV96 (1/29) were typed. CONCLUSION: The results of the present study, and specifically the detection of rare RV G12 and EV71 strains, address the need for continuous epidemiological surveys to provide epidemiological pictures of pediatric viral infections circulating in the community.

ASIC1a affects hypothalamic signaling and regulates the daily rhythm of body temperature in mice.

The body temperature of mice is higher at night than during the day. We show here that global deletion of acid-sensing ion channel 1a (ASIC1a) results in lower body temperature during a part of the night. ASICs are pH sensors that modulate neuronal activity. The deletion of ASIC1a decreased the voluntary activity at night of mice that had access to a running wheel but did not affect their spontaneous activity. Daily rhythms of thyrotropin-releasing hormone mRNA in the hypothalamus and of thyroid-stimulating hormone ß mRNA in the pituitary, and of prolactin mRNA in the hypothalamus and pituitary were suppressed in ASIC1a-/- mice. The serum thyroid hormone levels were however not significantly changed by ASIC1a deletion. Our findings indicate that ASIC1a regulates activity and signaling in the hypothalamus and pituitary. This likely leads to the observed changes in body temperature by affecting the metabolism or energy expenditure.

Emerging roles of Keap1/Nrf2 signaling in the thyroid gland and perspectives for bench-to-bedside translation.

The signaling pathway centered on the transcription factor nuclear erythroid factor 2-like 2 (Nrf2) has emerged during the last 15 years as a target for the prevention and treatment of diseases broadly related with oxidative stress such as cancer, neurodegenerative and metabolic diseases. The roles of Nrf2 are expanding beyond general cytoprotection, and they encompass its crosstalk with other pathways as well as tissue-specific functions. The thyroid gland relies on reactive oxygen species for its main physiological function, the synthesis and secretion of thyroid hormones. A few years ago, Nrf2 was characterized as a central regulator of the antioxidant response in the thyroid, as well as of the transcription and processing of thyroglobulin, the major thyroidal protein that serves as the substrate for thyroid hormone synthesis. Herein, we summarize the current knowledge about the roles of Nrf2 in thyroid physiology, pathophysiology and disease. We focus specifically on the most recent publications in the field, and we discuss the implications for the preclinical and clinical use of Nrf2 modulators.

Thyroid Disorders in Patients Treated with Dimethyl Fumarate for Multiple Sclerosis: A Retrospective Observational Study.

BACKGROUND: Dimethyl fumarate (DMF), a drug used for the treatment of multiple sclerosis (MS) and psoriasis, has been shown to activate the Keap1/Nrf2 antioxidant response. Nrf2 exerts pleiotropic roles in the thyroid gland; among others, single nucleotide polymorphisms (SNPs) in the gene encoding Nrf2 modulate the risk of Hashimoto's thyroiditis (HT), suggesting that pharmacological activation of Nrf2 might also be protective. However, a patient with acute exacerbation of HT after starting DMF for MS was recently reported, raising questions about the thyroidal safety of Nrf2 activators. METHODS: In a retrospective observational study, we investigated the prevalence and incidence of thyroid disorders (TD) among 163 patients with MS treated with DMF. RESULTS: Only 7/163 patients (4.3%) were diagnosed with functional TD; most (5/163, 3.0%) were diagnosed before DMF treatment. Functional TD were diagnosed under or after DMF in only 2 patients (1.2%). Under DMF, one patient developed transient mild hypothyroidism with negative thyroid autoantibodies. After DMF discontinuation, another patient developed hyperthyroidism due to Graves' disease. No patient developed thyroid structural disease under or after DMF. CONCLUSIONS: The very low incidence of functional TD indicates an overall very good thyroid tolerance of DMF, arguing against screening for TD in MS patients considered for or treated with DMF, and supporting the further study of Nrf2 activators for the prevention and treatment of TD.

Brown adipose tissue responds to cold and adrenergic stimulation by induction of FGF21.

Fibroblast growth factor-21 (FGF21) is a pleiotropic protein involved in glucose, lipid metabolism and energy homeostasis, with main tissues of expression being the liver and adipose tissue. Brown adipose tissue (BAT) is responsible for cold-induced thermogenesis in rodents. The role of FGF21 in BAT biology has not been investigated. In the present study, wild-type C57BL/6J mice as well as a brown adipocyte cell line were used to explore the potential role of cold exposure and ß3-adrenergic stimulation in the expression of FGF21 in BAT. Our results demonstrate that short-term exposure to cold, as well as ß3-adrenergic stimulation, causes a significant induction of FGF21 mRNA levels in BAT, without a concomitant increase in FGF21 plasma levels. This finding opens new routes for the potential use of pharmaceuticals that could induce FGF21 and, hence, activate BAT thermogenesis.

Development of an optimized method for the detection of airborne viruses with real-time PCR analysis.

BACKGROUND: Airborne viruses remain one of the major public health issues worldwide. Detection and quantification of airborne viruses is essential in order to provide information regarding public health risk assessment. FINDINGS: In this study, an optimized new, simple, low cost method for sampling of airborne viruses using Low Melting Agarose (LMA) plates and a conventional microbial air sampling device has been developed. The use of LMA plates permits the direct nucleic acids extraction of the captured viruses without the need of any preliminary elution step. Molecular detection and quantification of airborne viruses is performed using real-time quantitative (RT-)PCR (Q(RT-)PCR) technique. The method has been tested using Adenoviruses (AdVs) and Noroviruses (NoVs) GII, as representative DNA and RNA viruses, respectively. Moreover, the method has been tested successfully in outdoor experiments, by detecting and quantifying human adenoviruses (HAdVs) in the airborne environment of a wastewater treatment plant. CONCLUSIONS: The great advantage of LMA is that nucleic acids extraction is performed directly on the LMA plates, while the eluted nucleic acids are totally free of inhibitory substances. Coupled with QPCR the whole procedure can be completed in less than three (3) hours.

Molecular detection of multiple viral targets in untreated urban sewage from Greece.

BACKGROUND: Urban sewage virological analysis may produce important information about the strains that cause clinical and subclinical infections in the population, thus supporting epidemiological studies. METHODS: In the present study, a twenty one-month survey (November 2007 to July 2009) was conducted in order to evaluate the presence of human adenoviruses (hAdV), hepatitis A viruses (HAV), hepatitis E viruses (HEV), Noroviruses (NoV), and human Polyomaviruses (hPyV) in untreated sewage samples collected from the inlet of Patras' municipal biological wastewater treatment plant, located in southwestern Greece. Nucleic acid amplification techniques were applied for viral nucleic acid detection. Positive samples were confirmed by sequencing and comparative phylogenetic analysis was performed on the isolated viral strains. RESULTS: In total, viruses were detected in 87.5% (42/48) of sewage samples. AdVs, PyVs, HAV, and NoVs were detected in 45.8% (22/48), 68.8% (33/48), 8.3% (4/48), and 6.3% (3/48) of the samples collected from the plant's inlet, while HEV was not detected at all. Adenovirus types 8 (Ad8), 40 (Ad40) and 41 (Ad41) were recognized, while JC and BK polyomaviruses were recorded. Noroviruses were identified as GII.4. HAV was typed as genotype IA. CONCLUSIONS: Our study demonstrates the advantages of environmental surveillance as a tool to elucidate the molecular epidemiology of community circulating viruses. We underline the need of environmental surveillance programs in countries such as Greece with inadequate and problematic epidemiological surveillance system and no environmental surveillance system currently in action.

Mice Hypomorphic for Keap1, a Negative Regulator of the Nrf2 Antioxidant Response, Show Age-Dependent Diffuse Goiter with Elevated Thyrotropin Levels.

Background: Familial nontoxic multinodular goiter (MNG) is a rare disease. One of the associated genes is Kelch-like ECH-associated protein 1 (KEAP1), which encodes the main inhibitor of nuclear factor erythroid 2-related transcription factor 2 (Nrf2), a central mediator of antioxidant responses. The association of KEAP1 with familial MNG is based on only two loss-of-function mutations identified in two families, only one of which included proper phenotyping and adequate demonstration of co-segregation of the phenotype and the mutation. There is no experimental evidence from model organisms to support that decreased Keap1 levels can, indeed, cause goiter. This study used mice hypomorphic for Keap1 to test whether decreased Keap1 expression can cause goiter, and to characterize the activation status of Nrf2 in their thyroid. Methods: C57BL/6J Keap1flox/flox (Keap1 knock-down [Keap1KD]) mice were studied at 3 and 12 months of age. Plasma and thyroid glands were harvested for evaluation of thyroid function tests and for gene and protein expression by real-time polymerase chain reaction and immunoblotting, respectively. Results: Keap1KD mice showed diffuse goiter that began to develop in early adult life and became highly prominent and penetrant with age. The goiter was characterized by a markedly increased size of thyroid follicles, most notably of the colloid compartment, and by absence of thyroid nodules or hyperplasia. Keap1KD mice also showed decreased T4 levels in early adult life that were eventually well compensated over time by increased thyrotropin (TSH) levels. Nrf2 was activated in the thyroid of Keap1KD mice. Despite a known stimulatory effect of Nrf2 on thyroglobulin (Tg) gene transcription and Tg protein abundance, the expression levels were decreased in the thyroid of Keap1KD mice. No clear patterns were observed in the expression profiles of other thyroid hormone synthesis-specific factors, with the exception of Tg-processing and Tg-degrading cathepsins, including an increase in mature forms of cathepsins D, L, and S. Conclusions: Keap1KD mice develop age-dependent diffuse goiter with elevated TSH levels. The precise mechanism accounting for the thyroidal phenotype remains to be elucidated, but it may involve enhanced Tg solubilization and excessive lysosomal Tg degradation.

Simvastatin lowers reactive oxygen species level by Nrf2 activation via PI3K/Akt pathway.

The beneficial effects of HMG-CoA (3-hydroxy-3-methyl-glutaryl-CoA) reductase inhibitors (statins) have been attributed not only to their cholesterol lowering effect but also to their pleiotropic actions and especially to their anti-oxidant activity. Nrf2 (NF-E2-related factor 2) is a transcription factor that orchestrates the transcriptional response of cells to oxidative stressors and electrophilic xenobiotics. In this study, primary mouse embryonic fibroblasts from wild type or Nrf2 knock out C57BL6J mice and ST-2 cells were used to investigate the implication of Nrf2 in the mediation of the anti-oxidant effects of statins and the possible involvement of PI3K/Akt pathway in this process. We show for the first time that simvastatin lowers reactive oxygen species (ROS) by activating Nrf2 through the PI3K/Akt pathway.

Patent Review (2017-2020) of the Keap1/Nrf2 Pathway Using PatSeer Pro: Focus on Autoimmune Diseases.

Research on the antioxidant pathway comprising the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) and its cytoplasmic inhibitor Kelch-like ECH-associated protein 1 (Keap1) is ever increasing. As modulators of this pathway have started to be used in clinical trials and clinical practice, Nrf2 has become the subject of several patents. To assess the patent landscape of the last three years on Nrf2 and evaluate the main fields they refer to, we used the web-based tool PatSeer Pro to identify patents mentioning the Nrf2 pathway between January 2017 and May 2020. This search resulted in 509 unique patents that focus on topics such as autoimmune, neurodegenerative, liver, kidney, and lung diseases and refer to modulators (mainly activators) of the Nrf2 pathway as potential treatments. Autoimmunity emerged as the main theme among the topics of Nrf2 patents, including a broad range of diseases, such as systemic sclerosis, systemic lupus erythematosus, multiple sclerosis, inflammatory bowel diseases, Hashimoto's thyroiditis, etc.; however, there was a dearth of experimental support for the respective patents' claims. Given that chronic inflammation is the main element of the pathophysiology of most autoimmune diseases, the majority of patents referring to activation of Nrf2 as a method to treat autoimmune diseases base their claims on the well-established anti-inflammatory role of Nrf2. In conclusion, there is strong interest in securing intellectual property rights relating to the potential use of Nrf2 pathway activators in a variety of diseases, and this trend parallels the rise in related research publications. However, in the case of autoimmunity, more research is warranted to support the potential beneficial effects of Nrf2 modulation in each disease.

The Keap1/Nrf2 Signaling Pathway in the Thyroid-2020 Update.

The thyroid gland has a special relationship with oxidative stress. On the one hand, like all other tissues, it must defend itself against reactive oxygen species (ROS). On the other hand, unlike most other tissues, it must also produce reactive oxygen species in order to synthesize its hormones that contribute to the homeostasis of other tissues. The thyroid must therefore also rely on antioxidant defense systems to maintain its own homeostasis in the face of continuous self-exposure to ROS. One of the main endogenous antioxidant systems is the pathway centered on the transcription factor Nuclear factor erythroid 2-related factor 2 (Nrf2) and its cytoplasmic inhibitor Kelch-like ECH-associated protein 1 (Keap1). Over the last few years, multiple links have emerged between the Keap1/Nrf2 pathway and thyroid physiology, as well as various thyroid pathologies, including autoimmunity, goiter, hypothyroidism, hyperthyroidism, and cancer. In the present mini-review, we summarize recent studies shedding new light into the roles of Keap1/Nrf2 signaling in the thyroid.

Rare and common genetic variations in the Keap1/Nrf2 antioxidant response pathway impact thyroglobulin gene expression and circulating levels, respectively.

Nuclear factor, erythroid 2-like 2 (Nrf2) is a transcription factor that has been gaining attention in the field of pharmacology and especially in the chemoprevention of diseases such as cancer, metabolic and neurodegenerative diseases, etc. This is because natural compounds such as sulforaphane, which is found in broccoli sprout extracts, can activate Nrf2. The repertoire of the roles of Nrf2 is ever increasing; besides its traditional antioxidant and cytoprotective effects, Nrf2 can have other functions as a transcription factor. We have recently shown that Nrf2 directly regulates the expression of thyroglobulin (Tg), which is the most abundant thyroidal protein and the precursor of thyroid hormones. Two functional binding sites for Nrf2 (antioxidant response elements, AREs) were identified in the regulatory region of the TG gene. Interestingly, we then observed that one of these AREs harbors a rare single-nucleotide polymorphism (SNP). Also recently, we performed the first genome-wide association study (GWAS) for common SNPs that impact the circulating levels of Tg. Based on these investigations, we were triggered (i) to investigate whether common SNPs in the Nrf2 pathway correlate with circulating Tg levels; and (ii) to examine whether the rare SNP in one of the TG regulatory AREs may affect gene expression. To address the first question, we analyzed GWAS data from a general population and its two subpopulations, one with thyroid disease and/or abnormal thyroid function tests and the other without, in which circulating Tg levels had been measured. Statistically significant associations with Tg levels were observed in the genes encoding Nrf2 and Keap1, including, notably, a known functional SNP in the promoter of the gene encoding Nrf2. Regarding the rare SNP (rs778940395) in the proximal ARE of the TG enhancer, luciferase reporter gene expression studies in PCCL3 rat thyroid follicular cells showed that this SNP abrogated the basal and sulforaphane- or TSH-induced luciferase activity, behaving as a complete loss-of-function mutation. Thus, both rare and common genetic variation in the Keap1/Nrf2 pathway can impact TG expression and Tg circulating levels, respectively.

The Transcriptomic Response of the Murine Thyroid Gland to Iodide Overload and the Role of the Nrf2 Antioxidant System.

BACKGROUND: Thyroid follicular cells have physiologically high levels of reactive oxygen species because oxidation of iodide is essential for the iodination of thyroglobulin (Tg) during thyroid hormone synthesis. Thyroid follicles (the functional units of the thyroid) also utilize incompletely understood autoregulatory mechanisms to defend against exposure to excess iodide. To date, no transcriptomic studies have investigated these phenomena in vivo. Nuclear erythroid factor 2 like 2 (Nrf2 or Nfe2l2) is a transcription factor that regulates the expression of numerous antioxidant and other cytoprotective genes. We showed previously that the Nrf2 pathway regulates the antioxidant defense of follicular cells, as well as Tg transcription and Tg iodination. We, thus, hypothesized that Nrf2 might be involved in the transcriptional response to iodide overload. METHODS: C57BL6/J wild-type (WT) or Nrf2 knockout (KO) male mice were administered regular water or water supplemented with 0.05% sodium iodide for seven days. RNA from their thyroids was prepared for next-generation RNA sequencing (RNA-Seq). Gene expression changes were assessed and pathway analyses were performed on the sets of differentially expressed genes. RESULTS: Analysis of differentially expressed messenger RNAs (mRNAs) indicated that iodide overload upregulates inflammatory-, immune-, fibrosis- and oxidative stress-related pathways, including the Nrf2 pathway. Nrf2 KO mice showed a more pronounced inflammatory-autoimmune transcriptional response to iodide than WT mice. Compared to previously published datasets, the response patterns observed in WT mice had strong similarities with the patterns typical of Graves' disease and papillary thyroid carcinoma (PTC). Long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) also responded to iodide overload, with the latter targeting mRNAs that participate mainly in inflammation pathways. CONCLUSIONS: Iodide overload induces the Nrf2 cytoprotective response and upregulates inflammatory, immune, and fibrosis pathways similar to autoimmune hyperthyroidism (Graves' disease) and PTC.

Disease-related anemia in chronic lymphocytic leukemia is not due to intrinsic defects of erythroid precursors: a possible pathogenetic role for tumor necrosis factor-alpha.

BACKGROUND/AIMS: Disease-related anemia in chronic lymphocytic leukemia (CLL) occurs when the obvious causes are excluded while its pathogenesis is still obscure. We investigated its underlying mechanisms in 56 untreated patients with CLL. METHODS: Bone marrow (BM) lymphocytic infiltration was estimated in trephine biopsies. Serum erythropoietin (EPO) and tumor necrosis factor-alpha (TNF-alpha) levels were measured by ELISA. The potential of BM CD34+ to differentiate into erythroid cells was evaluated by methylcellulose-based assays and in liquid cultures supplemented with EPO, SCF, IL-3 +/- TNF-alpha. The response of erythroid precursors to EPO +/- TNF-alpha was assessed by detecting activated key proteins of EPO-EPO receptor signalling pathway using Western Blot and EMSA. RESULTS: Bone marrow lymphocytic infiltration was not exclusively responsible for disease-related anemia and CD34+ cells were intrinsically capable of generating erythroid precursors. Also, no deficiency of serum erythropoietin (EPO) or defective intracellular response of erythroid precursors to EPO +/- TNF-alpha stimulation was observed. Serum TNF-alpha levels were found increased in anemic CLL patients and TNF-alpha appeared to directly inhibit the erythroid development in early stages of erythropoiesis. CONCLUSION: We concluded that CLL-related anemia was not due to intrinsic defects of erythroid precursors, but might result from the direct suppressive effect of TNF-alpha on the erythroid production.

Simvastatin activates Keap1/Nrf2 signaling in rat liver.

Some of the statins' pleiotropic actions have been attributed to their antioxidant activity. The Nrf2 transcription factor controls the expression of a number of protective genes in response to oxidative stress. In the present study, wistar rats, primary hepatocytes as well as ST2 cells, were employed to explore the potential role of Nrf2 in mediating the reported antioxidant effects of statins. Simvastatin triggered nuclear translocation of Nrf2 in rat liver and in primary rat hepatocytes in a mevalonate-dependent and cholesterol-independent way. In liver, nuclear extracts from simvastatin-treated rats, the DNA-binding activity of Nrf2, was significantly increased and the mRNA of two known targets of Nrf2 (HO-1 and GPX2) was induced. In ST2 cells stably transfected with constructs bearing Nrf2-binding site (antioxidant responsive element), simvastatin enhanced Nrf2-mediated transcriptional activity in a mevalonate-dependent and cholesterol-independent fashion. In conclusion, activation of Keap1/Nrf2 signaling pathway by simvastatin might provide effective protection of the cell from the deleterious effects of oxidative stress.

Keap1/Nrf2 Signaling: A New Player in Thyroid Pathophysiology and Thyroid Cancer.

The Keap1/Nrf2 pathway is a key mediator of general redox and tissue-specific homeostasis. It also exerts a dual role in cancer, by preventing cell transformation of normal cells but promoting aggressiveness, and drug resistance of malignant ones. Although Nrf2 is well-studied in other tissues, its roles in the thyroid gland are only recently emerging. This review focuses on the involvement of Keap1/Nrf2 signaling in thyroid physiology, and pathophysiology in general, and particularly in thyroid cancer. Studies in mice and cultured follicular cells have shown that, under physiological conditions, Nrf2 coordinates antioxidant defenses, directly increases thyroglobulin production and inhibits its iodination. Increased Nrf2 pathway activation has been reported in two independent families with multinodular goiters due to germline loss-of-function mutations in KEAP1. Nrf2 pathway activation has also been documented in papillary thyroid carcinoma (PTC), due to somatic mutations, or epigenetic modifications in KEAP1, or other pathway components. In PTC, such Nrf2-activating KEAP1 mutations have been associated with tumor aggressiveness. Furthermore, polymorphisms in the prototypical Nrf2 target genes NQO1 and NQO2 have been associated with extra-thyroidal extension and metastasis. More recently, mutations in the Nrf2 pathway have also been found in Hürthle-cell (oncocytic) thyroid carcinoma. Finally, in in vitro, and in vivo models of poorly-differentiated, and undifferentiated (anaplastic) thyroid carcinoma, Nrf2 activation has been associated with resistance to experimental molecularly-targeted therapy. Thus, Keap1/Nrf2 signaling is involved in both benign and malignant thyroid conditions, where it might serve as a prognostic marker or therapeutic target.

A Bibliometric Review of the Keap1/Nrf2 Pathway and its Related Antioxidant Compounds.

Nrf2 is a master transcriptional regulator of antioxidant and cytoprotective pathways. Currently in its third decade, research on Nrf2 has expanded to encompass not only basic but also clinical studies. In the present bibliometric review, we employed the VOSviewer tool to describe the existing Nrf2 literature landscape. As of July 2019, 11,931 papers on Nrf2 were listed in the "Web of Science" database, with more than 1000 new papers published each year. As expected, terms related to oxidative stress and antioxidant molecules occur very often in the Nrf2 literature throughout the years. Interestingly, there is also a gradual increase in the occurrence of terms related to diseases or to natural compounds, the most prominent being sulforaphane, curcumin, and resveratrol that modulate the Nrf2 pathway. Going beyond molecular biology/biochemistry and related fields, Nrf2 research has begun to spread into more clinical areas like endocrinology/metabolism, cardiology, and nephrology, likely reflecting an increased interest in clinical applications of Nrf2 pathway activators. China has become the most prolific producer of Nrf2 papers the last five years followed by the USA and Japan, a reverse pattern compared to the past. In conclusion, Nrf2 is the subject of a globally active research field that keeps growing and extends from bench to bedside.