Complete transposition of the great arteries (TGA) is the most frequent cyanotic heart defect diagnosed in neonates. However, the exact etiology of TGA is unknown. The aim of the present study was to assess the association of TGA pathogenesis with single nucleotide polymorphisms (SNPs) in DNA methyltransferases (DNMTs)-1 and 3a- in Chinese children. We genotyped 5 SNPs (rs16999593, rs16999358, and rs2228611 in DNMT1; and rs2276599 and rs2276598 in DNMT3A) in 206 patients with complete TGA and 252 healthy children. Statistical analysis was performed to explore the association of the 5 SNPs with complete TGA susceptibility. Compared with the T/T and C/C genotypes, the heterozygous genotype C/T of rs16999593 correlated with a decreased risk for complete TGA under codominant (OR=0.46; 95% CI=0.29-0.72), dominant (OR=0.58; 95% CI=0.38-0.88), and overdominant (OR=0.44; 95% CI=0.28-0.68) models. In contrast, the genotype C/C of rs16999593 correlated with a higher risk for TGA under a recessive model (OR=3.15; 95% CI=1.14-8.68) compared with the T/T and C/T genotypes. Furthermore, the TGC, TGT, CGC, and CGT haplotypes of DNMT1 did not differ significantly between the two groups, whereas the frequency of the TAC haplotype was lower in the case group (OR<1; P=0.002). No significant differences in the frequencies of the TC, CC, TT, and CT haplotypes of DNMT3A were found between the two groups. Furthermore, logistic regression showed that sex and the rs16999358 SNP were two independent risk factors for complete TGA. Overall, the C/T genotype of the rs16999593 SNP in DNMT1 might decrease the risk of complete TGA pathogenesis in the Southern Chinese population.

Thyroid hormones are important for normal reproductive function. Although 3,5,3'-triiodothyronine (T3) enhances follicle-stimulating hormone (FSH)-induced preantral follicle growth and granulosa cells development in vitro, little is known about the molecular mechanisms regulating ovarian development via glucose. In this study, we investigated whether and how T3 combines with FSH to regulate glucose transporter protein (GLUT) expression and glucose uptake in granulosa cells. In this study, we present evidence that T3 and FSH cotreatment significantly increased GLUT-1/GLUT-4 expression, and translocation in cells, as well as glucose uptake. These changes were accompanied by upregulation of nitric oxide (NO) synthase (NOS)3 expression, total NOS and NOS3 activity, and NO content in granulosa cells. Furthermore, we found that activation of the mammalian target of rapamycin (mTOR) and phosphoinositide 3-kinase (PI3K)/Akt pathway is required for the regulation of GLUT expression, translocation, and glucose uptake by hormones. We also found that l-arginine upregulated GLUT-1/GLUT-4 expression and translocation, which were related to increased glucose uptake; however, these responses were significantly blocked by N(G)-nitro-l-arginine methylester. In addition, inhibiting NO production attenuated T3- and FSH-induced GLUT expression, translocation, and glucose uptake in granulosa cells. Our data demonstrate that T3 and FSH cotreatment potentiates cellular glucose uptake via GLUT upregulation and translocation, which are mediated through the activation of the mTOR/PI3K/Akt pathway. Meanwhile, NOS3/NO are also involved in this regulatory system. These findings suggest that GLUT is a mediator of T3- and FSH-induced follicular development.

Interest in preclinical drug development for ovarian cancer has stimulated development of patient-derived xenograft (PDX) or tumorgraft models. However, the unintended formation of human lymphoma in severe combined immunodeficiency (SCID) mice from Epstein-Barr virus (EBV)-infected human lymphocytes can be problematic. In this study, we have characterized ovarian cancer PDXs which developed human lymphomas and explore methods to suppress lymphoproliferative growth. Fresh human ovarian tumors from 568 patients were transplanted intraperitoneally in SCID mice. A subset of PDX models demonstrated atypical patterns of dissemination with mediastinal masses, hepatosplenomegaly, and CD45-positive lymphoblastic atypia without ovarian tumor engraftment. Expression of human CD20 but not CD3 supported a B-cell lineage, and EBV genomes were detected in all lymphoproliferative tumors. Immunophenotyping confirmed monoclonal gene rearrangements consistent with B-cell lymphoma, and global gene expression patterns correlated well with other human lymphomas. The ability of rituximab, an anti-CD20 antibody, to suppress human lymphoproliferation from a patient's ovarian tumor in SCID mice and prevent growth of an established lymphoma led to a practice change with a goal to reduce the incidence of lymphomas. A single dose of rituximab during the primary tumor heterotransplantation process reduced the incidence of CD45-positive cells in subsequent PDX lines from 86.3% (n = 117 without rituximab) to 5.6% (n = 160 with rituximab), and the lymphoma rate declined from 11.1% to 1.88%. Taken together, investigators utilizing PDX models for research should routinely monitor for lymphoproliferative tumors and consider implementing methods to suppress their growth.

Grapevine is an important and extensively grown fruit crop, which is severely hampered by drought worldwide. So, comprehending the impact of drought on grapevine genetic resources is necessary. In the present study, RNA-sequencing was executed using cDNA libraries constructed from both drought-stress and control plants. Results generated 12,451 differentially expressed genes (DEGs), out of which 8,021 genes were up-regulated, and 4,430 were down-regulated. Further physiological and biochemical investigations were also performed to validate the biological processes associated with the development of grapevine in response to drought stress. Results also revealed that decline in the rate of stomatal conductance, in turn, decrease the photosynthetic activity and CO2 assimilation in the grapevine leaves. Reactive oxygen species, including stress enzymes and their related proteins, and secondary metabolites were also activated in the present study. Likewise, various hormones also induced in response to drought stress. Overall, the present study concludes that these DEGs play both positive and negative roles in drought tolerance by regulating various biological pathways of grapevine. Nevertheless, our findings have provided valuable gene information for future studies of abiotic stress in grapevine and various other fruit crops.

Reactive oxygen species (ROS) have emerged as important signaling molecules that play crucial roles in carcinogenesis and cytotoxic responses. Nrf2 is the master regulator of ROS balance. Thus, uncovering mechanisms of Nrf2 regulation is important for the development of alternative treatment strategies for cancers. Here, we demonstrate that iASPP, a known p53 inhibitor, lowers ROS independently of p53. Mechanistically, iASPP competes with Nrf2 for Keap1 binding via a DLT motif, leading to decreased Nrf2 ubiquitination and increased Nrf2 accumulation, nuclear translocation, and antioxidative transactivation. This iASPP-Keap1-Nrf2 axis promotes cancer growth and drug resistance both in vitro and in vivo. Thus, iASPP is an antioxidative factor and represents a promising target to improve cancer treatment, regardless of p53 status.

Statins are well known to have muscle toxicity problem. Herba Cistanches (HC) is a Chinese herb traditionally used for pain in the loins and knees. Our previous in vitro study suggested that it could protect against statin-induced muscle toxicity. However, its in vivo protective effect has never been investigated. The objective of this study was to determine if the aqueous extract of HC (HCE) could prevent simvastatin-induced muscle toxicity in rats, and whether HCE could also exert beneficial effects on reducing high-fat diet-induced hypercholesterolemia and elevated liver cholesterol, thereby reducing the dose of simvastatin when used in combined therapy. From our results, HCE significantly restored simvastatin-induced reduction in muscle weights and reduced elevated plasma creatine kinase in rats. HCE also improved simvastatin-induced reduction in muscle glutathione levels, muscle mitochondrial membrane potential, and reduced simvastatin-induced muscle inflammation. Furthermore, HCE could exert reduction on liver weight, total liver lipid levels and plasma lipid levels in high-fat-fed mice. In conclusion, our study provided in vivo evidence that HCE has potential protective effect on simvastatin-induced toxicity in muscles, and also beneficial effects on diet-induced non-alcoholic fatty liver and hyperlipidemia when being used alone or in combination with simvastatin at a reduced dose.

Proteus vulgaris L-amino acid deaminase (pvLAAD) belongs to a class of bacterial membrane-bound LAADs mainly express in genus Proteus, Providencia and Morganella. These LAADs employ a non-cleavable N-terminal twin-arginine translocation (Tat) peptide to transport across membrane and bind to bacterial surface. Recent studies revealed that a hydrophobic insertion sequence (INS) in these LAADs also interacts with bacterial membrane. However, the functional significance of INS-membrane interaction is not clear. In this study, we made site-directed mutagenesis on the surface-exposed hydrophobic residues of pvLAAD INS, and we found that these mutations impaired the INS-membrane interaction but did not affect pvLAAD activity in the solution. We further found that when cell membrane is present, the catalytic activity can be increased by 8~10 folds for wild-type but not INS-mutated pvLAAD, indicating that the INS-membrane interaction is necessary for increasing activity of pvLAAD. Molecular dynamic (MD) simulations suggested that INS is flexible in the solution, and its conformational dynamics could lead to substrate channel distortion. Circular dichroism (CD) spectroscopy experiments indicated that bacterial membrane was able to maintain the conformation of INS. Our study suggests the function of the membrane binding of INS is to stabilize pvLAAD structure and increase its catalytic activity.

Long noncoding RNAs (lncRNAs) play a functional role in the initiation and progression of prostate cancer (PCa). This study aimed to determine differentially expressed lncRNA through high-throughput sequencing technology and investigate its expression, biological function and clinical correlation with PCa.

Protease-activated receptor 1 (PAR1) is the prototypical member of a family of G-protein-coupled receptors that mediate cellular responses to thrombin and related proteases. Thrombin irreversibly activates PAR1 by cleaving the amino-terminal exodomain of the receptor, which exposes a tethered peptide ligand that binds the heptahelical bundle of the receptor to affect G-protein activation. Here we report the 2.2 Å resolution crystal structure of human PAR1 bound to vorapaxar, a PAR1 antagonist. The structure reveals an unusual mode of drug binding that explains how a small molecule binds virtually irreversibly to inhibit receptor activation by the tethered ligand of PAR1. In contrast to deep, solvent-exposed binding pockets observed in other peptide-activated G-protein-coupled receptors, the vorapaxar-binding pocket is superficial but has little surface exposed to the aqueous solvent. Protease-activated receptors are important targets for drug development. The structure reported here will aid the development of improved PAR1 antagonists and the discovery of antagonists to other members of this receptor family.

The Pax gene Pox meso (Poxm) was the first and so far only gene whose initial expression was shown to occur specifically in the anlage of the somatic mesoderm, yet its role in somatic myogenesis remained unknown. Here we show that it is one of the crucial genes regulating the development of the larval body wall muscles in Drosophila. It has two distinct functions expressed during different phases of myogenesis. The early function, partially redundant with the function of lethal of scute [l(1)sc], demarcates the ;Poxm competence domain', a domain of competence for ventral and lateral muscle development and for the determination of at least some adult muscle precursor cells. The late function is a muscle identity function, required for the specification of muscles DT1, VA1, VA2 and VA3. Our results led us to reinterpret the roles of l(1)sc and twist in myogenesis and to propose a solution of the 'l(1)sc conundrum'.

To identify multiple microRNAs (miRNAs) for predicting the prognosis of gastric cancer (GC) patients by bioinformatics analysis.

Diabetic cardiomyopathy, a major cardiac complication, contributes to heart remodelling and heart failure. Our previous study discovered that CCAAT/enhancer-binding protein β (C/EBPβ), a transcription factor that belongs to a family of basic leucine zipper transcription factors, interacts with the angiotensin-converting enzyme 2 (ACE2) promoter sequence in other disease models. Here, we aimed to determine the role of C/EBPβ in diabetes and whether ACE2 expression is regulated by C/EBPβ. A type 1 diabetic mouse model was generated by an intraperitoneal injection of streptozotocin. Diabetic mice were injected with a lentivirus expressing either C/EBPβ or sh-C/EBPβ or treated with valsartan after 12 weeks to observe the effects of C/EBPβ. In vitro, cardiac fibroblasts and cardiomyocytes were treated with high glucose (HG) to investigate the anti-fibrosis, anti-apoptosis and regulatory mechanisms of C/EBPβ. C/EBPβ expression was down-regulated in diabetic mice and HG-induced cardiac neonatal cells. C/EBPβ overexpression significantly attenuated collagen deposition and cardiomyocyte apoptosis by up-regulating ACE2 expression. The molecular mechanism involved the binding of C/EBPβ to the ACE2 promoter sequence. Although valsartan, a classic angiotensin receptor blocker, relieved diabetic complications, the up-regulation of ACE2 expression by C/EBPβ overexpression may exert greater beneficial effects on patients with diabetic cardiomyopathy.

Background: Vein graft failure due to neointimal hyperplasia remains an important and unresolved complication of cardiovascular surgery. microRNA-21 (miR-21) plays a major role in regulating vascular smooth muscle cell (VSMC) proliferation and phenotype transformation. Thus, the purpose of this study was to determine whether adenovirus-mediated miR-21 sponge gene therapy was able to inhibit neointimal hyperplasia in rat vein grafts. Methods: Adenovirus-mediated miR-21 sponge was used to inhibit VSMC proliferation in vitro and neointimal formation in vivo. To improve efficiency of delivery gene transfer to the vein grafts, 20% poloxamer F-127 gel was used to increase virus contact time and 0.25% trypsin to increase virus penetration. Morphometric analyses and cellular proliferation were assessed for neointimal hyperplasia and VSMC proliferation. Results: miR-21 sponge can significantly decrease the expression of miR-21 and proliferation in cultured VSMCs. Cellular proliferation rates were significantly reduced in miR-21 sponge-treated grafts compared with controls at 28 days after bypass surgery (14.6±9.4 vs 34.9±10.8%, P=0.0032). miR-21 sponge gene transfer therapy reduced the intimal/media area ratio in vein grafts compared with the controls (1.38±0.08 vs. 0.6±0.10, P<0.0001). miR-21 sponge treatment also improved vein graft hemodynamics. We further identified that phosphatase and tensin homolog (PTEN) is a potential target gene that was involved in the miR-21-mediated effect on neointimal hyperplasia in vein grafts. Conclusions: Adenovirus-mediated miR-21 sponge gene therapy effectively reduced neointimal formation in vein grafts. These results suggest that there is potential for miR-21 sponge to be used to prevent vein graft failure.

Nitric oxide (NO) is a multifunctional gaseous molecule that plays important roles in mammalian reproductive functions, including follicular growth and development. Although our previous study showed that NO mediated 3,5,3'-triiodothyronine and follicle-stimulating hormone-induced granulosa cell development via upregulation of glucose transporter protein (GLUT)1 and GLUT4 in granulosa cells, little is known about the precise mechanisms regulating ovarian development via glucose. The objective of the present study was to determine the cellular and molecular mechanism by which NO regulates GLUT expression and glucose uptake in granulosa cells. Our results indicated that NO increased GLUT1/GLUT4 expression and translocation in cells, as well as glucose uptake. These changes were accompanied by upregulation of cyclic guanosine monophosphate (cGMP) level and cGMP-dependent protein kinase (PKG)-I protein content. The results of small interfering RNA (siRNA) analysis showed that knockdown of PKG-I significantly attenuated gene expression, translocation, and glucose uptake. Moreover, the PKG-I inhibitor also blocked the above processes. Furthermore, NO induced cyclic adenosine monophosphate response element binding factor (CREB) phosphorylation, and CREB siRNA attenuated NO-induced GLUT expression, translocation, and glucose uptake in granulosa cells. These findings suggest that NO increases cellular glucose uptake via GLUT upregulation and translocation, which are mediated through the activation of the cGMP/PKG pathway. Meanwhile, the activated CREB is also involved in the regulation. These findings indicate that NO has an important influence on the glucose uptake of granulosa cells.

To elucidate the molecular mechanisms underlying non-alcoholic fatty liver disease (NAFLD), we recruited 86 subjects with varying degrees of hepatic steatosis (HS). We obtained experimental data on lipoprotein fluxes and used these individual measurements as personalized constraints of a hepatocyte genome-scale metabolic model to investigate metabolic differences in liver, taking into account its interactions with other tissues. Our systems level analysis predicted an altered demand for NAD+ and glutathione (GSH) in subjects with high HS Our analysis and metabolomic measurements showed that plasma levels of glycine, serine, and associated metabolites are negatively correlated with HS, suggesting that these GSH metabolism precursors might be limiting. Quantification of the hepatic expression levels of the associated enzymes further pointed to altered de novo GSH synthesis. To assess the effect of GSH and NAD+ repletion on the development of NAFLD, we added precursors for GSH and NAD+ biosynthesis to the Western diet and demonstrated that supplementation prevents HS in mice. In a proof-of-concept human study, we found improved liver function and decreased HS after supplementation with serine (a precursor to glycine) and hereby propose a strategy for NAFLD treatment.

The essential pigment chlorophyll (Chl) plays important roles in light harvesting and energy transfer during photosynthesis. Here we present the data from a comparative proteomic analysis of chlorophyll-deficient Brassica napus mutant cde1 and its corresponding wild-type using the iTRAQ approach (Pu Chu et al., 2014 [1]). The distribution of length and number of peptides, mass and sequence coverage of proteins identified was calculated, and the repeatability of the replicates was analyzed. A total of 443 differentially expressed proteins were identified in B. napus leaves, including 228 down-accumulated proteins mainly involved in photosynthesis, porphyrin and chlorophyll metabolism, biosynthesis of secondary metabolites, carbon fixation and 215 up-accumulated proteins that enriched in the spliceosome, mRNA surveillance and RNA degradation.

Long noncoding RNAs (lncRNAs) have emerged as important regulators and biomarkers of tumor development and progression. This study investigated the clinical significance, biological functions, and underlying mechanisms of long intergenic non-protein-coding RNA 1296 (LINC01296) in prostate cancer.

CRISPR-Cas endonucleases directed against foreign nucleic acids mediate prokaryotic adaptive immunity and have been tailored for broad genetic engineering applications. Type VI-D CRISPR systems contain the smallest known family of single effector Cas enzymes, and their signature Cas13d ribonuclease employs guide RNAs to cleave matching target RNAs. To understand the molecular basis for Cas13d function and explain its compact molecular architecture, we resolved cryoelectron microscopy structures of Cas13d-guide RNA binary complex and Cas13d-guide-target RNA ternary complex to 3.4 and 3.3 Å resolution, respectively. Furthermore, a 6.5 Å reconstruction of apo Cas13d combined with hydrogen-deuterium exchange revealed conformational dynamics that have implications for RNA scanning. These structures, together with biochemical and cellular characterization, provide insights into its RNA-guided, RNA-targeting mechanism and delineate a blueprint for the rational design of improved transcriptome engineering technologies.

Purpose: Circular RNAs (circRNAs), are a large class of RNAs that from a covalently closed continuous loop and have recently showed huge capabilities as gene regulators in mammals. Although Hsa_circ_0001451 has been investigated in colorectal cancer, it remains unclear about the relationship between Hsa_circ_0001451 and clear cell renal cell carcinoma (ccRCC). Our research aims to reveal the function of Hsa_circ_0001451 in the proliferation and development in ccRCC cells. Methods: The expression of Hsa_circ_0001451 in 52 pairs of ccRCC tissues and paraneoplastic tissues was detected by quantitative real-time polymerase chain reaction (qRT-PCR). The correlation between Hsa_circ_0001451 and the clinicopathological features was evaluated using the chi-sequare test. Receiver operating characteristic (ROC) curve was built by SPSS to evaluate the diagnostic values. The effects of Hsa_circ_0001451 on ccRCC cells were determined via a MTT assay, clone formation assay, flow cytometry and Western blot analysis. Results: The expression of Hsa_circ_0001451 was significantly correlated with differentiation (P<0.05). The area under ROC curve of Hsa_circ_0001451 was 0.704 (P<0.05). Knockdown of Hsa_circ_0001451 significantly promoted tumor growth in vitro. Bioinformatics results also displayed that Hsa_circ_0001451 might be involved in the regulation of tumor progression. Conclusion: Taken together, our finding showed that Hsa_circ_0001451 might become a novel potential biomarker in the diagnosis of ccRCC and a potential novel target for the treatment of ccRCC.

Aberrant activation of Wnt/β-catenin signaling is a common event in the development of colorectal cancer (CRC). It is important to identify new molecules and mechanisms that can negatively regulate Wnt/β-catenin signaling. MicroRNAs are considered as promising candidates for cancer diagnosis and therapy. In our study, we found that miR-377-3p was significantly decreased in CRC samples compared to the normal mucosa tissues, especially in the patients at stage III/IV. Functional studies showed that overexpression of miR-377-3p suppressed and silence of miR-377-3p enhanced the proliferation, migration and chemoresistance of CRC cells. Molecularly, miR-377-3p inhibited Wnt/β-catenin signaling by directly targeting ZEB2 and XIAP, which were the positive regulators of Wnt/β-catenin signaling. Overexpression of ZEB2/XIAP could counteract the tumor suppressing phenotypes induced by miR-377-3p. Therefore, we uncovered the anti-cancer role and the relevant mechanisms of miR-377-3p in CRC, which might provide novel targets for designing new anti-tumor strategies.