Periodontitis is an inflammatory disease that is characterized by progressive destruction of the periodontium and causes tooth loss in adults. Periodontitis is known to be associated with dysbiosis of the oral microflora, which is often linked to various diseases. However, the complexity of plaque microbial communities of periodontitis, antibiotic resistance, and enhanced virulence make this disease difficult to treat. In this study, using metagenomic shotgun sequencing, we investigated the etiology, antibiotic resistance genes (ARGs), and virulence genes (VirGs) of periodontitis. We revealed a significant shift in the composition of oral microbiota as well as several functional pathways that were represented significantly more abundantly in periodontitis patients than in controls. In addition, we observed several positively selected ARGs and VirGs with the Ka/Ks ratio > 1 by analyzing our data and a previous periodontitis dataset, indicating that ARGs and VirGs in oral microbiota may be subjected to positive selection. Moreover, 5 of 12 positively selected ARGs and VirGs in periodontitis patients were found in the genomes of respiratory tract pathogens. Of note, 91.8% of the background VirGs with at least one non-synonymous single-nucleotide polymorphism for natural selection were also from respiratory tract pathogens. These observations suggest a potential association between periodontitis and respiratory infection at the gene level. Our study enriches the knowledge of pathogens and functional pathways as well as the positive selection of antibiotic resistance and pathogen virulence in periodontitis patients, and provides evidence at the gene level for an association between periodontitis and respiratory infection.

OBJECTIVE:To determine the upregulation of IL-21-inducible genes in minor salivary glands (MSGs) in 28 primary SS (pSS) patients and 12 non-pSS subjects and correlate it with disease characteristics.METHODS:RNA sequencing was utilized to compare IL-21-inducible genes expression in the MSGs between pSS and non-pSS subjects. The subgroups were characterized according to the IL-21 score calculated by seven IL-21-inducible genes. Furthermore, the disease characteristics and transcripts implicated in hypoxia and interferon signalling were assessed in two pSS subgroups.RESULTS:We observed that the expression of the IL-21-inducible genes (IL-21, IL-21R, JAK3, STAT1, HLA-B, CCR7 and CXCL10), the so-called IL-21 signature genes, was significantly increased in pSS patients. The upregulation of JAK3 expression may be induced by hypomethylation of the JAK3 promoter in pSS patients and putatively associated with POU2F2. The patients with increased IL-21 signature gene expression showed an increased EULAR Sjögren's Syndrome Disease Activity Index score and increased enrichment of B cells, memory B cells, CD4+ T cells and CD8+ T cells. Furthermore, the IL-21 scores in the anti-SSA+, SSB+, ANA+ and high IgG samples were higher than those in the respective antibody-negative samples and normal IgG. In addition, we found both hypoxia and IFN-relevant genes showed strong correlation with IL-21 signature gene expression, indicating their interaction in pSS.CONCLUSION:IL-21 signature gene was associated with typical disease characteristics in pSS, which provides insight into the contribution of the IL-21 signalling pathway to the pathogenesis of the disease and might provide a novel treatment strategy for this subtype of pSS.

Background Urinary retention (UR) is a common urinary system disease can be caused by urinary tract obstruction with numerous reasons, however, the role of urine microbes in these disorders is still poorly understood. The aim of this study was to identify the urine microbial features of two common types of obstructive UR, caused by urinary stones or urinary tract tumors, with comparison to healthy controls. Methods Urine samples were collected from a cohort of 32 individuals with stone UR, 25 subjects with tumor UR and 25 healthy controls. The urine microbiome of all samples was analyzed using high-throughput 16S rRNA (16S ribosomal RNA) gene sequencing. Results We observed dramatically increased urine microbial richness and diversity in both obstructive UR groups compared to healthy controls. Despite different origins of UR, bacteria such as Pseudomonas, Acinetobacter and Sphingomonas were enriched, while Lactobacillus , Streptococcus , Gardnerella , Prevotella and Atopobium were decreased in both UR groups in comparison with healthy controls, exhibited an approximate urine microbial community and functional characteristics of two types of obstructive UR. Furthermore, disease classifiers were constructed using specific enriched genera in UR, which can distinguish stone UR or tumor UR patients from healthy controls with an accuracy of 92.29% and 97.96%, respectively. Conclusion We presented comprehensive microbial landscapes of two common types of obstructive urinary retention and demonstrated that urine microbial features of these patients are significantly different from that of healthy people. The urine microbial signatures would shed light on the pathogenesis of these types of urinary retention and might be used as potential classification tools in the future.

The novel coronavirus SARS-CoV-2 (causing the disease COVID-19) has caused a highly transmissible and ongoing pandemic worldwide. Due to its rapid development, next-generation sequencing plays vital roles in many aspects. Here, we summarize the current knowledge on the origin and human transmission of SARS-CoV-2 based on NGS analysis. The ACE2 expression levels in various human tissues and relevant cells were compared to provide insights into the mechanism of SAS-CoV-2 infection. Gut microbiota dysbiosis observed by metagenome sequencing and the immunogenetics of COVID-19 patients according to single-cell sequencing analysis were also highlighted. Overall, the application of these sequencing techniques could be meaningful for finding novel intermediate SARS-CoV-2 hosts to block interspecies transmission. This information will further benefit SARS-CoV-2 diagnostic development and new therapeutic target discovery. The extensive application of NGS will provide powerful support for our fight against future public health emergencies.

Neurodevelopmental disorders, including autism spectrum disorder (ASD), intellectual disability (ID), developmental disorders (DD) and epileptic encephalopathy (EE), have a strong clinical comorbidity, which indicates a common genetic etiology across various disorders. However, the underlying genetic mechanisms of comorbidity and specificity remain unknown across neurodevelopmental disorders. Based on de novo mutations, we compared systematically the functional characteristics between shared and unique genes under these disorders, as well as the spatiotemporal trajectory of development in brain and common molecular pathways of all shared genes. We observed that shared genes present more constrained against functional rare genetic variation, and harbor more pathogenic rare variants than do unique genes in each disorder. Furthermore, 71 shared genes formed two clusters related to synaptic transmission, transcription regulation and chromatin regulator. Particularly, we also found that two core genes STXBP1 and SCN2A , that were shared by the four neurodevelopmental disorders showed prominent pleiotropy. Our findings shed light on the shared and specific patterns across neurodevelopmental disorders and will enable us to further comprehend the etiology and provide valuable information for the diagnosis of neurodevelopmental disorders.

Recent whole-exome sequencing (WES) studies have demonstrated the contribution of de novo mutations (DNMs) to epileptic encephalopathies (EEs). Here, we performed WES on four trios with West syndrome and identified three loss-of-function DNMs in both CSNK1E (c.885+1G>A) and STXBP1 (splicing, c.1111-2A>G; nonsense, p.(Y519X)). The splicing mutation in CSNK1E creates insertion of 116 new amino acids at position 246 followed by a premature stop codon. Both CSNK1E and STXBP1 showed a closer coexpression relationship with epilepsy candidate genes beyond that expected by chance. In addition, genes coexpressed with CSNK1E were enriched in early prenatal stages across multiple brain regions. We also found that 60 CSNK1E-interacting genes share an association with multiple neuropsychiatric disorders, and these genes formed a significant interconnected interaction network with roles in the midbrain development. Our study supported the potential role of CSNK1E variants in EE susceptibility and expanded the phenotypic spectrum associated with CSNK1E variation.

Schimke immuno-osseous dysplasia (SIOD) is an extremely rare autosomal recessive pleiotropic disease. Although biallelic mutations in SMARCAL1 gene have been reported to be the genetic etiology of SIOD, its molecular diagnosis has been challenging in a relatively proportion of cases due to the extreme rarity. Here, we made a definitive SIOD diagnosis of a 5-year-old girl with an extremely mild phenotype by applying whole exome sequencing (WES). As a result, a novel maternal mutation (c.2141+5G > A) confirmed to create a novel splice donor site combined with a known paternal mutation (c.1933C > T; p.Arg645Cys) were detected. In addition, previous reported SIOD cases showed excessive enrichment for mutations in the helicase ATP-binding and C-terminal domains of SMARCAL1. Similarly, the novel mutation we identified caused a mutant protein truncated in the SMARCAL1 C-terminus. Interestingly, based on the phenotypic profile, compared to reported cases, the patient in our study exhibited milder symptoms with renal dysfunctions limited to asymptomatic proteinuria, but no neurological signs or recurrent infections. Moreover, we identified 73 SMARCAL1-interacting genes, which formed a significant interconnected interaction network with roles in disease-related pathways such as double-strand break repair via homologous recombination, DNA repair, and replication fork processing. Notably, the top 15 SMARCAL1-interacting genes all showed a similar renal temporal expression pattern. Altogether, to our knowledge, the case in this study is the first case diagnosed originally based on a genetic test via WES rather than a characteristic phenotype. The identification of the novel allelic mutation (c.2141+5G > A) extends the phenotypic spectrum of SMARCAL1 mutations and the following bioinformatics analysis presents additional genetic evidence to illustrate the role of SMARCAL1 in SIOD.

Pak choi (Brassica rapa subsp. chinensis) is a leafy vegetable that is widely available in Asia and consumed in rising quantities in Europe. Pak choi contains high levels of secondary plant metabolites, such as carotenoids, chlorophylls, glucosinolates, phenolic compounds, and vitamin K, which are beneficial for humans if consumed on a regular basis. The evaluation of the genotype-induced variation of secondary plant metabolites revealed that the cultivar 'Amur' contained the highest concentration of secondary plant metabolites. Furthermore, steaming retained more chlorophylls, glucosinolates, phenolic acids and flavonoid compounds than boiling. In contrast, both domestic cooking methods - boiling, and steaming - reduced the formation of glucosinolate breakdown products, especially the undesired epithionitriles and nitriles but less of the health-beneficial isothiocyanates.

Leydig cells reside in the interstitium of the testis and make testosterone and insulin-like 3 for reproductive health. Leydig cells contain a series of androgen biosynthetic and metabolizing enzymes. The luteinizing hormone/cAMP/PKA signaling is critical for the stimulation of steroidogenesis. Environmental toxicants on the Leydig cell function act via disrupting its development, blocking LH/cAMP/PKA signaling, downregulating the expressions of steroidogenesis-related proteins and insulin-like 3 or directly inhibiting one or more enzyme activities for testosterone biosynthesis and metabolic activation. These environmental toxicants include industrial materials and natural resources. This chapter reviews these environmental toxicants targeting Leydig cells.

Abstract The neurotoxicity of ziram is largely unknown. In this study, we investigated the direct inhibitions of ziram on rat neurosteroid synthetic and metabolizing enzymes, 5α-reductase 1 (SRD5A1), 3α-hydroxysteroid dehydrogenase (AKR1C14), and retinol dehydrogenase 2 (RDH2). Rat SRD5A1, AKR1C14, and RDH2 were cloned and transiently expressed in COS1 cells, and the effects of ziram on these enzymes were measured. Ziram inhibited rat SRD5A1 and AKR1C14 with IC50 values of 1.556 ± 0.078 and 1.017 ± 0.072 μM, respectively, when 1000 nM steroid substrates were used. Ziram weakly inhibited RDH2 at 100 μM, when androstanediol (1000 nM) was used. Ziram competitively inhibited SRD5A1 and non-competitively inhibited AKR1C14 when steroid substrates were used. Docking study showed that ziram bound to NADPH-binding pocket of AKR1C14. In conclusion, our results demonstrated that ziram inhibited SRD5A1 and AKR1C14 activities, thus possibly interfering with neurosteroid production in rats.

Citral is a widely used penetration enhancer that has been used to assist the delivery of drugs through the skin. In this study we aimed to investigate the effectiveness of combination treatments of citral with hyperthermic intraperitoneal chemotherapy (HIPEC) for colorectal cancer and to unravel the underlying mechanism by which citral increased the efficacy of HIPEC. In vitro experiments indicated that citral increased cytoplasmic absorption of pirarubicin and potentiated the effects of pirarubicin on colorectal cancer cells to induce apoptosis. Intracellular reactive oxygen species (ROS) activity was elevated after single or combo treatments with pirarubicin, leading to compromised NF-κB signaling. Therefore, the results suggested that the effects of citral were mediated by increasing cell permeability and ROS productions. Furthermore, the colorectal xenograft model was used to evaluate the efficacy of the combo treatment at the histological and molecular levels, which showed that the cotreatment with citral for colorectal cancer increased the efficacy of HIPEC with pirarubicin with respect to both ascite control and tumor load. The results indicated that citral was an effective additive for HIPEC with pirarubicin for colorectal cancer, which warrant further effort to explore the translational application of this new treatment regimen.

Cadmium is an endocrine disruptor, impairing male reproduction. The objective of this study is to investigate whether cadmium affects rat Leydig cell regeneration and to dissect the underlying mechanism. Adult male Sprague-Dawley rats received a single intraperitoneal injection (i.p.) of 0, 0.5 or 1.0 mg/kg of cadmium chloride, followed by ethane dimethane sulfonate (EDS) treatment to eliminate adult Leydig cells 20 days later. Compared to control (0 dose), cadmium treatment reduced serum testosterone levels by days 21, 35, and 56 after EDS treatment. Serum luteinizing hormone (LH) levels were also affected by day 56, the only time point examined. There were fewer regenerated Leydig cells in the cadmium-treated testis on days 35 and 56 after EDS treatment. Further studies demonstrated that the mRNA or protein levels of Leydig ( Lhcgr , Scarb1 , Star , Cyp11a1 , Hsd3b1 , Cyp17a1 , Hsd17b3 , and Hsd11b1 ), non-Leydig ( Fshr and Dhh ), and gonadotroph ( Lhb ) cells were also significantly lower in cadmium-treated animals. Since LH and desert hedgehog (DHH) are critical factors for Leydig cell differentiation, our result demonstrated that the lower doses of cadmium exposure, even briefly, may permanently damage Leydig cell regeneration.

Fetal exposure to di-n-butyl phthalate (DBP) causes the adult disease such as lower testosterone production and infertility. However, the mechanism is still unknown. The objective of the present study is to determine how DBP affects the involution of fetal Leydig cells during the neonatal period and how this event causes the delayed development of the adult Leydig cells during puberty. The pregnant Sprague Dawley dams were randomly divided into 3 groups and were gavaged with 0 (corn oil, the vehicle control), 100 or 500mg/kg DBP from gestational day 12 (G12) to G21. The blood and testes were collected from male pups on postnatal day 4 (P4), P7, P14, P21, P28, and P56. Serum testosterone concentrations were assessed and the mRNA levels of Leydig cell- or gonadotroph cell-specific genes were measured. Prenatal exposure to DBP caused the aggregation of fetal Leydig cells, which slowly disappeared when compared to the control. This effect was associated with the reduction of testicular testosterone secretion and down-regulation of the mRNA levels of Leydig cell biomarkers including Scarb1, Star, Cyp11a1, Hsd3b1, Hsd11b1, and Hsd17b3 as well as the gonadotroph biomarkers including Lhb and Gnrhr. In conclusion, we demonstrated that the increased aggregation of fetal Leydig cells by DBP delayed fetal Leydig cell involution, thus leading to the disrupted development of the adult Leydig cells.

In the testis, KIT ligand (KITL, also called stem cell factor) is expressed by Sertoli cells and its receptor (c-kit, KIT) is expressed by spermatogonia and Leydig cells. Although KITL-KIT signaling is critical for the spermatogenesis, its roles in Leydig cell development during puberty are not clear. In the present study, we investigated effects of KITL on stem Leydig cell proliferation and differentiation. Using an in vitro culture system of seminiferous tubules from Leydig cell-depleted testis, we found that KITL increased the proliferation activity of putative stem Leydig cells at higher concentration (10 and 100 ng/ml). Low concentration (1 ng/ml) of KITL significantly induced the differentiation of stem Leydig cells via increasing the expression level of steroidogenic acute regulatory protein (Star). In contrast, higher concentration (100 ng/ml) of KITL inhibited the differentiation of stem Leydig cells via inhibiting the steroidogenic enzyme (Cyp11a1, Cyp17a1, and Hsd17b3) expression levels. We cultured rat progenitor Leydig cells with KITL for 48 h and did not find any influence of KITL on the proliferation and androgen production of these cells. In conclusion, KITL is a growth factor that regulates the development of the stem Leydig cell.

Placenta produces progesterone and estradiol for maintaining pregnancy. Two critical enzymes are responsible for their production: 3β-hydroxysteroid dehydrogenase 1 (HSD3B1) that catalyzes the formation of progesterone from pregnenolone and aromatase that catalyzes the production of estradiol from testosterone. Fungicide ziram may disrupt the placental steroid production. In the present study, we investigated the effects of ziram on steroid formation in human placental cell line JEG-3 cells and on HSD3B1 and aromatase in the human placenta. Ziram did not inhibit progesterone production in JEG-3 cells and HSD3B1 activity at 100μM. Ziram was a potent aromatase inhibitor with the half maximal inhibitory concentration (IC50) value of 333.8nM. When testosterone was used to determine the mode of action, ziram was found to be a competitive inhibitor. Docking study showed that ziram binds to the testosterone binding pocket of the aromatase. In conclusion, ziram is a potent inhibitor of human aromatase.

Ziram [zinc, bis (dimethyldithiocarbamate)] is an agricultural dithiocarbamate fungicide. By virtual screening, we have identified that ziram is a potential endocrine disruptor. To investigate its effects on pubertal development of Leydig cells, 35-day-old male Sprague Dawley rats orally received ziram (2 or 4 mg/kg/d) for 4 weeks and immature Leydig cells isolated from 35-day-old rat testes were treated with ziram (0.5-50 μM in vitro). Serum hormones, Leydig cell number and specific gene or protein expression levels after in vivo treatment were determined and medium androgen levels were measured as well as apoptosis of Leydig cells after in vitro treatment were determined. In vivo exposure to ziram lowered testosterone and follicle-stimulating hormone levels, and reduced Leydig cell number, and downregulated Leydig cell specific gene or protein expression levels. Ziram exposure in vitro inhibited androgen production and steroidogenic enzyme activities in Leydig cells by downregulating expression levels of P450 cholesterol side cleavage enzyme (Cyp11a1), 3β-hydroxysteroid dehydrogenase 1 (Hsd3b1), 17α-hydroxylase/17,20-lyase (Cyp17a1), and 17β-hydroxysteroid dehydrogenase 3 (Hsd17b3) via downregulating the steroidogenic factor 1 (Nr5a1) at a concentration as low as 5 μM. In conclusion, ziram exposure disrupts Leydig cell development during puberty possibly via downregulating Nr5a1.

Gastric cancer is one of the leading causes of cancer mortality in the world. Curcumin is a natural product with multiple pharmacological activities, while its clinical application has been limited by the poor chemical stability. We have previously designed a series of curcumin derivatives with high stability and anticancer potentials. The present study aims to identify the anti‐cancer effects and mechanisms of WZ26, an analog of curcumin, in gastric cancer cells. In vitro, WZ26 showed higher chemical stability and much stronger anti‐proliferative effects than curcumin, accompanied by dose‐dependent induction of cell cycle arrest and apoptosis in gastric cancer cells. Mechanistically, the novel compound WZ26 induced ROS production, resulting in the activation of JNK‐mitochondrial and ER stress apoptotic pathways. Blockage of ROS production totally reversed WZ26‐induced JNK activation, Bcl‐2/Bax decrease, ER stress activation, and final cell apoptosis in SGC‐7901 cells. WZ26 also exhibited potent anti‐tumor effects in human gastric cancer cell xenograft models. WZ26 could be considered as a potential chemotherapeutic agent for the treatment of advanced gastric cancer. In addition, this study also demonstrated that ROS production could be act as a vital candidate pathway for inducing tumor cell apoptosis by targeting mitochondrial and ER stress‐related death pathway. © 2015 Wiley Periodicals, Inc.

BACKGROUND. The limited treatment option for recurrent prostate cancer and eventual resistant to conventional chemotherapy drugs has fueled continued interest in finding new anti-neoplastic agents. WZ35, a chemical analog of curcumin, had been demonstrated to have high chemical stability and potential anticancer effects in gastric cancer cells. The present study aimed to investigate the anti-prostate cancer effects of WZ35 in vitro and in vivo as well as the underlying mechanism.METHODS. Two prostate cancer cell lines RM-1 and DU145 were utilized to test the anticancer effects of WZ35 and the underlying mechanism. MTT assay was used to assess the cytotoxic effect of WZ35. Cell cycle distribution, apoptosis, alteration of ROS, and [Ca2+](i) level were evaluated using flow cytometry. Western blotting assay was applied to measure the levels of proteins associated with apoptosis and cell cycle. Immunofluorescence staining and Electron micrographs were used to evaluate activation of mitochondrial apoptosis pathway. Tumor models in nude mice were induced by injection of RM-1 prostate cancer cells to test the in vivo anticancer action of WZ35.RESULTS. Our results showed that WZ35 treatment induced loss of cell viability, cell apoptosis, and G2/M cycle arrest in both RM-1 and DU145 cells, coupled with ROS overproduction, intracellular calcium surge, and activation of mitochondrial apoptosis pathway in RM-1 cells. Interestingly, all above changes induced by WZ35 were completely reversed by ROS blockage. In addition, prevention of [Ca2+](i) elevation by BAPTA/AM also inhibited activation of mitochondrial apoptosis pathway induced by WZ35. In vivo studies, WZ35 treatment significantly inhibited RM-1 homograft tumor growth along with increased ROS accumulation, mitochondrial disruption, and cell apoptosis in tumor tissues.CONCLUSIONS. In conclusion, this work provides a novel anticancer candidate for the treatment of prostate cancer and demonstrated that increased ROS mediate the anti-cancer effects of WZ35 via activating mitochondrial apoptosis pathway. Importantly, this work also reveals that targeting ROS generation might be an effective strategy in human androgenresistant prostate cancer treatment. (C) 2016 Wiley Periodicals, Inc.

Referring to the comments of Svingen [1] on our latest publication about Effects of in utero Exposure to Dicyclohexyl Phthalate on Rat Fetal Leydig Cells [2], we would like to give some comments.[...].

Polybrominated diphenyl ethers (PBDEs) are a class of brominated flame retardants. PBDEs have been widely used in textiles, flexible polyurethane foams, electronic components, electrical components, and plastics. 11β-Hydroxysteroid dehydrogenases, isoform 1 (HSD11B1) and isoform 2 (HSD11B2), have been demonstrated to be the regulators of local glucocorticoid levels. In this study, the potencies of 4 different PBDEs (BDE-3, BDE-47, BDE-100, and BDE-153) with 1-6 bromine atoms attached in inhibition of rat and human HSD11B1 and HSD11B2 activities were compared to 4-bromobiphenyl (BBP), a structurally similar compound. All 4 PBDEs and BBP did not inhibit rat and human HSD11B1. BDE-3 and BDE-47 potently inhibited rat HSD11B2, and BDE-47 and BDE-153 potently inhibited human HSD11B2, with the half maximal inhibitory concentration values of 12.42, 5.95, 11.97, and 4.41 µmol/l, respectively. All PBDEs noncompetitively inhibited HSD11B2 when a steroid substrate was used. However, PBDEs exerted uncompetitive inhibition when the cofactor NAD+ was used. In conclusion, some PBDEs are selective inhibitors of HSD11B2, possibly causing excessive glucocorticoid action in local tissues.