Epigallocatechin gallate modulates ferroptosis through downregulation of tsRNA-13502 in non-small cell lung cancer.

Ferroptosis, an iron-dependent regulated cell death mechanism, holds significant promise as a therapeutic strategy in oncology. In the current study, we explored the regulatory effects of epigallocatechin gallate (EGCG), a prominent polyphenol in green tea, on ferroptosis and its potential therapeutic implications for non-small cell lung cancer (NSCLC). Treatment of NSCLC cell lines with varying concentrations of EGCG resulted in a notable suppression of cell proliferation, as evidenced by a reduction in Ki67 immunofluorescence staining. Western blot analyses demonstrated that EGCG treatment led to a decrease in the expression of glutathione peroxidase 4 (GPX4) and solute carrier family 7 member 11 (SLC7A11) while increasing the levels of acyl-CoA synthetase long-chain family member 4 (ACSL4). These molecular changes were accompanied by an increase in intracellular iron, malondialdehyde (MDA), and reactive oxygen species (ROS), alongside ultrastructural alterations characteristic of ferroptosis. Through small RNA sequencing and RT-qPCR, transfer RNA-derived small RNA 13502 (tsRNA-13502) was identified as a significant target of EGCG action, with its expression being upregulated in NSCLC tissues compared to adjacent non-tumorous tissues. EGCG was found to modulate the ferroptosis pathway by downregulating tsRNA-13502 and altering the expression of key ferroptosis regulators (GPX4/SLC7A11 and ACSL4), thereby promoting the accumulation of iron, MDA, and ROS, and ultimately inducing ferroptosis in NSCLC cells. This study elucidates EGCG's multifaceted mechanisms of action, underscoring the modulation of ferroptosis as a viable therapeutic approach for enhancing NSCLC treatment outcomes.

Association between dietary fatty acids and urinary incontinence.

Background: Dietary nutrient intake contributes to urination; however, the association between dietary nutrient intake, especially that of fat, and urinary incontinence (UI) is not well understood. The most common types of UI include stress UI (SUI) and urgency UI (UUI). Objective: To investigate the potential effect(s) of dietary fat intake on UI and explore its mechanism of action in relation to body mass index (BMI). Methods: A cross-sectional survey of data from 15,121 individuals (20-85 years of age) from the National Health and Nutrition Examination Survey (2001-2008), a random population-based sample, was performed. Data regarding dietary nutrient intake were collected through 24 h dietary recall interviews. UI and covariate data were collected through in-person interviews. UI was assessed according to the American Urological Association Symptom Index. The odds ratio (OR) for SUI and UUI were calculated using multivariate logistic regression analysis. The mediation effect was estimated using observational mediation analysis. Results: Higher total fat intake was positively associated with increased odds for developing UI (OR 1.44 [95% confidence interval (CI) 1.08-1.93]). Females who consumed more saturated fatty acids (SFA), monounsaturated fatty acids (MUFA), and polyunsaturated fatty acids (PUFA) were more likely to develop SUI. BMI partially explained the association between total fat, SFA, MUFA, and PUFA and SUI; the proportions of the mediation effect of BMI were 14.7%, 13.0%, 18.7%, and 16.3%, respectively. Conclusions: Results of this study emphasize the key role of dietary fat intake in the prevalence of UI. Higher fat intake was positively associated with UI and BMI partially mediated the effect of fat intake on SUI.

Deficiency of the HGF/Met pathway leads to thyroid dysgenesis by impeding late thyroid expansion.

The mechanisms of bifurcation, a key step in thyroid development, are largely unknown. Here we find three zebrafish lines from a forward genetic screening with similar thyroid dysgenesis phenotypes and identify a stop-gain mutation in hgfa and two missense mutations in met by positional cloning from these zebrafish lines. The elongation of the thyroid primordium along the pharyngeal midline was dramatically disrupted in these zebrafish lines carrying a mutation in hgfa or met. Further studies show that MAPK inhibitor U0126 could mimic thyroid dysgenesis in zebrafish, and the phenotypes are rescued by overexpression of constitutively active MEK or Snail, downstream molecules of the HGF/Met pathway, in thyrocytes. Moreover, HGF promotes thyrocyte migration, which is probably mediated by downregulation of E-cadherin expression. The delayed bifurcation of the thyroid primordium is also observed in thyroid-specific Met knockout mice. Together, our findings reveal that HGF/Met is indispensable for the bifurcation of the thyroid primordium during thyroid development mediated by downregulation of E-cadherin in thyrocytes via MAPK-snail pathway.

TAF1 is needed for the proliferation and maturation of thyroid follicle cells via Notch signaling.

Thyroid dysgenesis (TD) is the common pathogenic mechanism of congenital hypothyroidism (CH). In addition, known pathogenic genes are limited to those that are directly involved in thyroid development. To identify additional candidate pathogenetic genes, we performed forward genetic screening for TD in zebrafish, followed by positional cloning. The candidate gene was confirmed in vitro using the Nthy-ori 3.1 cell line and in vivo using a zebrafish model organism. We obtained a novel zebrafish line with thyroid dysgenesis and identified the candidate pathogenetic mutation TATA-box binding protein associated Factor 1 (taf1) by positional cloning. Further molecular studies revealed that taf1 was needed for the proliferation of thyroid follicular cells by binding to the NOTCH1 promoter region. Knockdown of TAF1 impaired the proliferation and maturation of thyroid cells, thereby leading to thyroid dysplasia. This study showed that TAF1 promoted Notch signaling and that this association played a pivotal role in thyroid development.NEW & NOTEWORTHY In our study, we obtained a novel zebrafish line with thyroid dysgenesis (TD) and identified the candidate pathogenetic mutation TATA-box binding protein associated Factor 1 (taf1). Further researches revealed that taf1 was required for thyroid follicular cells by binding to the NOTCH1 promoter region. Our findings revealed a novel role of TAF1 in thyroid morphogenesis.

TSHR Variant Screening and Phenotype Analysis in 367 Chinese Patients With Congenital Hypothyroidism.

Background: Genetic defects in the human thyroid-stimulating hormone (TSH) receptor (TSHR) gene can cause congenital hypothyroidism (CH). However, the biological functions and comprehensive genotype-phenotype relationships for most TSHR variants associated with CH remain unexplored. We aimed to identify TSHR variants in Chinese patients with CH, analyze the functions of the variants, and explore the relationships between TSHR genotypes and clinical phenotypes. Methods: In total, 367 patients with CH were recruited for TSHR variant screening using whole-exome sequencing. The effects of the variants were evaluated by in-silico programs such as SIFT and polyphen2. Furthermore, these variants were transfected into 293T cells to detect their Gs/cyclic AMP and Gq/11 signaling activity. Results: Among the 367 patients with CH, 17 TSHR variants, including three novel variants, were identified in 45 patients, and 18 patients carried biallelic TSHR variants. In vitro experiments showed that 10 variants were associated with Gs/cyclic AMP and Gq/11 signaling pathway impairment to varying degrees. Patients with TSHR biallelic variants had lower serum TSH levels and higher free triiodothyronine and thyroxine levels at diagnosis than those with DUOX2 biallelic variants. Conclusions: We found a high frequency of TSHR variants in Chinese patients with CH (12.3%), and 4.9% of cases were caused by TSHR biallelic variants. Ten variants were identified as loss-of-function variants. The data suggest that the clinical phenotype of CH patients caused by TSHR biallelic variants is relatively mild. Our study expands the TSHR variant spectrum and provides further evidence for the elucidation of the genetic etiology of CH.

Machine learning reveals diverse cell death patterns in lung adenocarcinoma prognosis and therapy.

Cancer cell growth, metastasis, and drug resistance pose significant challenges in the management of lung adenocarcinoma (LUAD). However, there is a deficiency in optimal predictive models capable of accurately forecasting patient prognoses and guiding the selection of targeted treatments. Programmed cell death (PCD) pathways play a pivotal role in the development and progression of various cancers, offering potential as prognostic indicators and drug sensitivity markers for LUAD patients. The development and validation of predictive models were conducted by integrating 13 PCD patterns with comprehensive analysis of bulk RNA, single-cell RNA transcriptomics, and pertinent clinicopathological details derived from TCGA-LUAD and six GEO datasets. Utilizing the machine learning algorithms, we identified ten critical differentially expressed genes associated with PCD in LUAD, namely CHEK2, KRT18, RRM2, GAPDH, MMP1, CHRNA5, TMPRSS4, ITGB4, CD79A, and CTLA4. Subsequently, we conducted a programmed cell death index (PCDI) based on these genes across the aforementioned cohorts and integrated this index with relevant clinical features to develop several prognostic nomograms. Furthermore, we observed a significant correlation between the PCDI and immune features in LUAD, including immune cell infiltration and the expression of immune checkpoint molecules. Additionally, we found that patients with a high PCDI score may exhibit resistance to immunotherapy and standard adjuvant chemotherapy regimens; however, they may benefit from other FDA-supported drugs such as docetaxel and dasatinib. In conclusion, the PCDI holds potential as a prognostic signature and can facilitate personalized treatment for LUAD patients.

Low-dose polystyrene microplastics exposure impairs fertility in male mice with high-fat diet-induced obesity by affecting prostate function.

The harmful effects of microplastics (MPs) on male fertility are receiving more and more attention. However, the impact of low-dose MPs exposure on the reproductive function of male mice is still unclear. In this study, we exposed male mice to low-dose MPs (25-30 µg/kg body weight/day) or low-dose MPs combined with high-fat diet (HFD) feeding. Our results showed that low-dose MPs exposure or HFD feeding significantly reduced sperm quality and the number of offspring born, while low-dose MPs exposure combined with HFD feeding further enhanced the above effects. The combination of low-dose MPs exposure and HFD feeding resulted in a notable elevation of inflammatory level within the prostate of mice and induced apoptosis of prostate epithelium and a decrease in nutrients (zinc, citrate) in seminal plasma fluid. Our findings in this study could provide valuable clues for better understanding the influence of low-dose MPs exposure on the reproductive system under metabolic disorders and facilitate the development of the prevention of reproductive toxicity caused by MPs exposure.

Contactin 6, A Novel Causative Gene for Congenital Hypothyroidism, Mediates Thyroid Hormone Biosynthesis Through Notch Signaling.

Background: Congenital hypothyroidism (CH) is the most common neonatal metabolic disorder. In patients with CH in China, thyroid dyshormonogenesis is more common than thyroid dysgenesis; however, the genetic causes of CH due to thyroid dyshormonogenesis remain largely unknown. Therefore, we aimed at identifying novel candidate causative genes for CH. Methods: To identify novel CH candidate genes, a total of 599 patients with CH were enrolled and next-generation sequencing was performed. The functions of the identified variants were confirmed using HEK293T and FTC-133 cell lines in vitro and in a mouse model organism in vivo. Results: Three pathogenic contactin 6 (CNTN6) variants were identified in two patients with CH. Pedigree analysis showed that CH caused by CNTN6 variants was inherited in an autosomal recessive pattern. The CNTN6 gene was highly expressed in the thyroid in humans and mice. Cntn6 knockout mice presented with thyroid dyshormonogenesis and CH due to the decreased expression of crucial genes for thyroid hormone biosynthesis (Slc5a5, Tpo, and Duox2). All three CNTN6 variants resulted in the blocking of the release of the Notch intracellular domain, which could not translocate into the nucleus, impaired NOTCH1 transcriptional activity, and decreased expression of SLC5A5, TPO, and DUOX2. Further, we found that DTX1 was required for CNTN6 to promote thyroid hormone biosynthesis through Notch signaling. Conclusions: This study demonstrated that CNTN6 is a novel causative gene for CH through the mediation of thyroid hormone biosynthesis via Notch signaling, which provides new insights into the genetic background and mechanisms involved in CH and thyroid dyshormonogenesis.

Genetic screening and functional analysis of TPO variants in Chinese patients with congenital hypothyroidism.

INTRODUCTION: Congenital hypothyroidism (CH), the most common neonatal endocrine disorder world-wide, can be caused by variants in the thyroid peroxidase (TPO) gene. This study aimed to identify TPO variants in Chinese patients with CH, analyze their impact on TPO function, and establish relationships between TPO genotypes and clinical characteristics. METHODS: A total of 328 patients with CH were screened for TPO variants by performing whole exome sequencing. The function of the detected TPO variants was investigated via transfection assays in vitro. The pathogenic effect of five novel variants was further assessed in silico. RESULTS: Among 328 patients with CH, 19 TPO variants, including six novel ones, were identified in 43 patients. Eighteen patients (5.5%) carried biallelic TPO variants. In vitro experiments showed that TPO activity was impaired to varying degrees in 17 variants. Furthermore, we determined that a residual TPO enzyme activity threshold of 15% may serve as a criterion for differentiating CH severity. CONCLUSIONS: According to our study, the prevalence of TPO variants among Chinese patients with CH was 13.1 %. Five novel variants led to impaired TPO function by altering its structure or by affecting its expression or cellular localization, which should result in impaired thyroid hormone synthesis.

Influence of two anti-tumor drugs, pazopanib, and axitinib, on the development and thyroid-axis of zebrafish (Danio rerio) embryos/larvae.

Introduction: In recent years, the potential toxicities of different pharmaceuticals toward the thyroid system have received increasing attention. In this study, we aim to evaluate the toxic effects of pazopanib and axitinib, two anti-tumor drugs with widespread clinical use, on thyroid function in the zebrafish model. Methods: We measured levels of thyroid-related hormones using the commercial Enzyme-Linked Immunosorbent Assay (ELISA) kit. Whole-mount in situ hybridization (WISH) analysis was employed to detect target gene expression changes. Morphology of the thyroid were evaluated by using transgenic Tg (tg: EGFP) fish line under a confocal microscope. The relative mRNA expression of key genes was verified through quantitative real-time polymerase chain reaction (RT‒qPCR). The size and number of the follicles was quantified whereby Hematoxylin-Eosin (H & E) staining under a light microscope. Results: The results revealed that fertilized zebrafish embryos were incubated in pazopanib or axitinib for 96 hours, development and survival were significantly affected, which was accompanied by significant disturbances in thyroid endocrine system (e.g., increased thyroid-stimulating hormone (TSH) content and decreased triiodothyronine (T3) and thyroxine (T4) content, as well as transcription changes of genes associated with the hypothalamus-pituitary-thyroid (HPT) axis. Moreover, based on whole-mount in situ hybridization staining of tg and histopathological examination of zebrafish embryos treated with pazopanib and axitinib, we observed a significantly abnormal development of thyroid follicles in the Tg (tg: EGFP) zebrafish transgenic line. Conclusion: Collectively, these findings indicate that pazopanib and axitinib may have toxic effects on thyroid development and function, at least partially, by influencing the regulation of the HPT axis. Thus, we believe that the potential thyroid toxicities of pazopanib and axitinib in their clinical applications should receive greater attention.

Pathogenic variations in MAML2 and MAMLD1 contribute to congenital hypothyroidism due to dyshormonogenesis by regulating the Notch signalling pathway.

BACKGROUND: In several countries, thyroid dyshormonogenesis is more common than thyroid dysgenesis in patients with congenital hypothyroidism (CH). However, known pathogenic genes are limited to those directly involved in hormone biosynthesis. The aetiology and pathogenesis of thyroid dyshormonogenesis remain unknown in many patients. METHODS: To identify additional candidate pathogenetic genes, we performed next-generation sequencing in 538 patients with CH and then confirmed the functions of the identified genes in vitro using HEK293T and Nthy-ori 3.1 cells, and in vivo using zebrafish and mouse model organisms. RESULTS: We identified one pathogenic MAML2 variant and two pathogenic MAMLD1 variants that downregulated canonical Notch signalling in three patients with CH. Zebrafish and mice treated with N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butylester, a γ-secretase inhibitor exhibited clinical manifestations of hypothyroidism and thyroid dyshormonogenesis. Through organoid culture of primary mouse thyroid cells and transcriptome sequencing, we demonstrated that Notch signalling within thyroid cells directly affects thyroid hormone biosynthesis rather than follicular formation. Additionally, these three variants blocked the expression of genes associated with thyroid hormone biosynthesis, which was restored by HES1 expression. The MAML2 variant exerted a dominant-negative effect on both the canonical pathway and thyroid hormone biosynthesis. MAMLD1 also regulated hormone biosynthesis through the expression of HES3, the target gene of the non-canonical pathway. CONCLUSIONS: This study identified three mastermind-like family gene variants in CH and revealed that both canonical and non-canonical Notch signalling affected thyroid hormone biosynthesis.

The isl2a transcription factor regulates pituitary development in zebrafish.

Background: ISL LIM homeobox 2, also known as insulin gene enhancer protein ISL-2 (ISL2), is a transcription factor gene that participates in a wide range of developmental events. However, the role of ISL2 in the hypothalamus-pituitary-thyroid axis is largely unknown. In the present study, we characterized the expression patterns of ISL2 and revealed its regulative role during embryogenesis using zebrafish. Methods: We used the CRISPR/Cas9 system to successfully establish homozygous ISL2-orthologue (isl2a and isl2b) knockout zebrafish. Moreover, we utilized these knockout zebrafish to analyze the pituitary and thyroid phenotypes in vivo. For further molecular characterization, in situ hybridization and immunofluorescence were performed. Results: The isl2a mutant zebrafish presented with thyroid hypoplasia, reduced whole-body levels of thyroid hormones, increased early mortality, gender imbalance, and morphological retardation during maturity. Additionally, thyrotropes, a pituitary cell type, was notably decreased during development. Importantly, the transcriptional levels of pituitary-thyroid axis hormones-encoding genes, such as tshba, cga, and tg, were significantly decreased in isl2a mutants. Finally, the thyroid dysplasia in isl2a mutant larvae may be attributed to a reduction in proliferation rather than changes in apoptosis. Conclusions: In summary, isl2a regulates the transcriptional levels of marker genes in hypothalamus-pituitary-thyroid axis, and isl2a knockout causing low thyroid hormone levels in zebrafish. Thus, isl2a identified by the present study, is a novel regulator for pituitary cell differentiation in zebrafish, resulting in thyroid gland hypoplasia and phenotypes of hypothyroidism.

Large-scale forward genetic screening of zebrafish affecting thyroid development.

The thyroid follicular cells originate from the foregut endoderm and elucidating which genes and signaling pathways regulate their development is crucial for understanding developmental disorders as well as diseases in adulthood. We exploited unique advantages of the zebrafish model to carry an ENU-based forward mutagenesis screen aiming at identifying genes involved in the development and function of the thyroid follicular cells. ENU is an excellent chemical mutagen due to its high mutation efficiency and an indiscriminate selection of genes. A total of 1606 F2 families from 36 ENU treated founders was raised and embryos from F3 generation were collected at 5dpf to perform the whole embryo in situ hybridization with a cocktail probe of thyroid marker thyroglobulin(tg), pituitary marker thyroid stimulating hormone (tshba) to determine the mutagenic phenotype. Among the 1606 F2 families, 112 F2 mutant families with normal development stages except for thyroid dysfunction were identified and divided into three different groups according to their phenotypic characteristics. Further studies of the mutants are likely to shed more insights into the molecular basis of both the thyroid development and function in the zebrafish and vertebrate.

The association between serum vitamin A and NAFLD among US adults varied in different BMI groups: a cross-sectional study.

Background and aims: The association between serum vitamin A (VA) and non-alcoholic fatty liver disease (NAFLD) has not been adequately studied. This study aimed to evaluate the association between them in different BMI groups among US adults. Methods: A cross-sectional study was performed using the 2017-2018 National Health and Nutrition Examination Survey (NHANES) datasets (N = 4.723). Linear/logistic regression, interaction effect and mediation analyses were adopted to analyze the association. Results: NAFLD tended to be more prevalent in adults in the middle and high tertiles of serum VA than in those in the low tertile of serum VA (OR [95% CI], 1.17 [0.94, 1.45] and 1.43 [1.16, 1.75]). In the sensitivity analysis, subjects in the middle or high tertile of serum VA had 10% (OR, 1.10 [0.88, 1.39] and 31% (OR, 1.31 [1.09, 1.58]) higher odds of NAFLD than those in the low tertile of serum VA. In the normal weight group, higher serum VA was associated with 125% and 333% higher odds of NAFLD in the middle and high tertiles, respectively, (OR, 2.25 [1.46, 3.48] and 4.33 [2.43, 7.69]) compared with the low tertile serum VA group. However, serum VA and NAFLD were not significantly associated with the obese group. Among different BMI groups (<30 compared with ≥30), serum triglycerides and insulin resistance mediated the association between VA and NAFLD in adults to varying degrees. Conclusions: In the weighted survey, serum VA was positively associated with the degree of NAFLD, especially in the non-obese population.

Corilagin prevents non-alcoholic fatty liver disease via improving lipid metabolism and glucose homeostasis in high fat diet-fed mice.

Non-alcoholic fatty liver disease (NAFLD) has been considered to be one of the most common chronic liver diseases. However, no validated pharmacological therapies have been officially proved in clinic due to its complex pathogenesis. The purpose of this study was to examine the protective effects of Corilagin (referred to Cori) against NAFLD in mice under a high fat diet (HFD) condition. Mice were fed either a normal control diet (NCD) or HFD with or without Cori (5 or 10 mg/kg body weight) for 15 weeks. In our results, Cori treatment significantly attenuated HFD-induced hepatic steatosis, high NAFLD activity score (NAD) and liver injury. Consistently, Cori treatment remarkably alleviated HFD-induced hepatic lipid accumulation (e.g., triglycerides (TG) and total cholesterol (TC) contents in liver), and improved plasma lipid concentrations (e.g., plasma TG, TC, low-density lipoprotein cholesterol (LDL-c), high-density lipoprotein cholesterol (HDL-c)). Moreover, Cori treatment ameliorated NAFLD associated metabolic disorders such as glucose intolerance and insulin resistance in HFD-fed mice. Additionally, Cori treatment dramatically changed HFD-induced liver gene expression profiles, and identified overlapped differentially expressed genes (DEGs) between NCD vs. HFD group and HFD vs. HCR (high fat diet plus treatment with Cori) group. With these DEGs, we observed a marked enrichment of Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, which were closely associated with the metabolic balance in liver. Particularly, we found several potential hub proteins against NAFLD development with analyses of protein-protein interaction (PPI) network and qPCR assays. Collectively, our results revealed the important protective effects of Cori against the progress of NAFLD, which was probably mediated through improving dysregulated lipid metabolism and insulin resistance in HFD-fed mice. Additionally, Cori-dependent overlapped DEGs might serve as a featured NAFLD-associated gene expression signature for the diagnosis, treatment, as well as drug discovery and development of NAFLD in the near future.

The influence of sunitinib and sorafenib, two tyrosine kinase inhibitors, on development and thyroid system in zebrafish larvae.

Recently, the potential toxic effects of various pharmaceuticals on the thyroid endocrine system have raised considerable concerns. In this study, we evaluated the adverse effects of sorafenib and sunitinib, two widely used anti-tumor drugs, on the developmental toxicities and thyroid endocrine disruption by using zebrafish (Danio rerio) model. Zebrafish embryos/larvae were exposed to different contentions (0, 10, 50 and 100 nM) of sorafenib and sunitinib for 96 hpf. The results revealed that waterborne exposure to sorafenib and sunitinib exhibited remarkable toxic effects on the survival and development in zebrafish embryos/larvae, which was accompanied by obvious disturbances of thyroid endocrine system (e.g., decreased T3 and T4 content, increased TSH content) and genes' transcription changes within the hypothalamus-pituitary-thyroid (HPT) axis. In addition, we verified a strikingly abnormal thyroid gland organogenesis in zebrafish larvae in response to sorafenib and sunitinib, by assessing the development of thyroid follicles using the WISH staining of tg, the Tg (tg:GFP) zebrafish transgenic line, and histopathological analysis. Taken together, our results indicated sorafenib and sunitinib exposure could induce obvious developmental toxicities and thyroid function disruption in zebrafish embryos/larvae, which might involve a regulatory mechanism, at least in part, by destroying the thyroid follicle structure, and by disturbing the balance of the HPT axis.

miR-185-5p Regulates Inflammation and Phagocytosis through CDC42/JNK Pathway in Macrophages.

Macrophage activation is an essential component of systemic chronic inflammation and chronic inflammatory diseases. Emerging evidence implicates miR-185-5p in chronic inflammation diseases. However, the regulatory role of miR-185-5p in macrophage pro-inflammatory activation has not been studied previously. Here, we identified that miR-185-5p was one of the top genes and effectively downregulated in two macrophage miRNA expression datasets from GEO. Under LPS stress, miR-185-5p overexpression reduced pro-inflammatory cytokine expression, suppressed phagocytosis in RAW264.7 macrophage. miR-185-5p inhibitors augmented pro-inflammatory effects of LPS in macrophage. Mechanically, miR-185-5p sponged and negatively regulated the protein expression of CDC42. Ablation of CDC42 with selective CDC42 inhibitor CASIN reversed the pro-inflammatory effect of miR-185-5p inhibitors through inhibiting MAPK/JNK pathways. Collectively, these data demonstrate that miR-185-5p exhibited anti-inflammatory functions in LPS-induced RAW264.7 macrophages at least partially through CDC42/JNK pathways. Our findings yield insights into the understanding of miR-185-5p-regulated network in macrophages inflammation, which is beneficial for exploring miRNA-protein interaction in atherosclerotic inflammation.

E2F transcription factor 2-activated DLEU2 contributes to prostate tumorigenesis by upregulating serum and glucocorticoid-induced protein kinase 1.

Long noncoding RNAs (lncRNAs) participate in biological processes in multiple types of tumors. However, the regulatory patterns of lncRNAs in prostate cancer remain largely unclear. Here, we evaluated the expression and roles of the lncRNA DLEU2 in prostate cancer. Our results showed that DLEU2 was upregulated in advanced prostate cancer tissues. Patients with prostate cancer displaying high expression of DLEU2 had a poor prognosis. Moreover, we demonstrated that overexpression of DLEU2 facilitated the proliferation, migration, and invasion of prostate cancer in vitro. Mechanistically, DLEU2 promoted serum and glucocorticoid-induced protein kinase 1 (SGK1) expression by acting as an miR-582-5p sponge, and the transcription of DLEU2 was activated by the dysregulation of E2F transcription factor 2 (E2F2) expression in prostate cancer. Furthermore, knockdown of DLEU2 attenuated prostate cancer tumorigenesis in vivo. Notably, these findings suggested that E2F2-activated DLEU2 may function as a competing endogenous RNA to facilitate prostate cancer progression by targeting the miR-582-5p/SGK1 axis.

Mutation Screening and Functional Study of SLC26A4 in Chinese Patients with Congenital Hypothyroidism

Objective: Defects in the human solute carrier family 26 member 4 (SLC26A4) gene are reported to be one of the causes of congenital hypothyroidism (CH). We aimed to identify SLC26A4 mutations in Chinese patients with CH and analyze the function of the mutations. Methods: Patients with primary CH were screened for 21 CH candidate genes mutations by targeted next-generation sequencing. All the exons and exon-intron boundaries of SLC26A4 were identified and analyzed. The function of six missense mutation in SLC26A4 were further investigated in vitro. Results: Among 273 patients with CH, seven distinct SLC26A4 heterozygous mutations (p.S49R, p.I363L, p.R409H, p.T485M, p.D661E, p.H723R, c.919-2A>G) were identified in 10 patients (3.66%, 10/273). In vitro experiments showed that mutation p.I363L, p.R409H, p.H723R affect the membrane location and ion transport of SLC26A4, while p.S49R did not. Mutation p.T485M and p.D661E only affected ion transport, but had no effect on the membrane location. Conclusion: The prevalence of SLC26A4 mutations was 3.66% in Chinese patients with CH. Five mutations (p.I363L, p.R409H, p.T485M, p.D661E and p.H723R) impaired the membrane location or ion transport function of SLC26A4, suggesting important roles for Ile363, Arg409, Thr485, Asp661, and His723 residues in SLC26A4 function. As all variants identified were heterozygous, the pathogenesis of these patients cannot be explained, and the pathogenesis of these patients needs further study.

The mutation screening in candidate genes related to thyroid dysgenesis by targeted next-generation sequencing panel in the Chinese congenital hypothyroidism.

OBJECTIVE: Congenital hypothyroidism (CH) is known to be due to thyroid dyshormonogenesis (DH), which is mostly inherited in an autosomal recessive inheritance pattern or thyroid dysgenesis (TD), whose inheritance pattern is controversial and whose molecular etiology remains poorly understood. DESIGN AND METHODS: The variants in 37 candidate genes of CH, including 25 genes related to TD, were screened by targeted exon sequencing in 205 Chinese patients whose CH cannot be explained by biallelic variants in genes related to DH. The inheritance pattern of the genes was analyzed in family trios or quartets. RESULTS: Of the 205 patients, 83 patients carried at least one variant in 19 genes related to TD, and 59 of those 83 patients harbored more than two variants in distinct candidate genes for CH. Biallelic or de novo variants in the genes related to TD in Chinese patients are rare. We also found nine probands carried only one heterozygous variant in the genes related to TD that were inherited from a euthyroid either paternal or maternal parent. These findings did not support the monogenic inheritance pattern of the genes related to TD in CH patients. Notably, in family trio or quartet analysis, of 36 patients carrying more than two variants in distinct genes, 24 patients carried these variants inherited from both their parents, which indicated that the oligogenic inheritance pattern of the genes related to TD should be considered in CH. CONCLUSIONS: Our study expanded the variant spectrum of the genes related to TD in Chinese CH patients. It is rare that CH in Chinese patients could be explained by monogenic germline variants in genes related to TD. The hypothesis of an oligogenic origin of the CH should be considered.