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The increasing incidence of prostate cancer (PCa) indicates an urgent need for the development of new effective drug therapy. There are limited options to treat the PCa, this study tried to determine a new therapy option for this acute cancer. Androgen-independent PCa cell lines PC3 and DU145 were treated with different melatonin concentrations (0.1~3.5 mM) for 1~3 days and assessed cell migration, cell invasion, cycle arrest in G0/G1 phase as well as apoptosis. We utilized RNA-seq technology to analyze the transcriptional misregulation pathways in DU145 prostate cancer cell line with melatonin (0.5 mM) treatment. Data revealed 20031 genes were up and down-regulated, there were 271 genes that differentially expressed: 97 up-regulated (P

This study aimed to investigate the mechanism of human umbilical cord blood stem cell (HUCBSC) transplantation on restenosis after percutaneous transluminal angioplasty (PTA) for diabetic hindlimb vascular disease in rabbits. After successfully preparing a rabbit model of diabetic hindlimb vascular disease, 16 rabbits were randomly assigned to two groups. Of these, 8 rabbits received PTA surgery alone (PTA group), and the other 8 rabbits received PTA and HUCBSC (PTA+HUCBSC group) treatments. Five more healthy rabbits were set as healthy control (HC group). Samples were collected after 4 weeks of treatment. The expressions of regulator of calcineurin 1 (RCAN1) and calcineurin A (CnA) in the diseased artery were detected by immunofluorescence staining. The distribution of HUCBSCs was observed by pathological examination in transplanted artery, distal artery, and liver. Cytology experiments were applied to assess the levels of JAK and STAT3, and the migration and proliferation of human aortic vascular smooth muscle cells (HA-VSMC). In the rabbit model of diabetic vascular lesions in the hindlimbs, we found the stenosis of the femoral artery became more and more serious with time, and the expression level of PCNA positive cells was also gradually increased. The expression levels of RCAN1 and CnA in the PTA+HUCBSC group were significantly lower than those in PTA group. HUCBSC inhibited the migration and proliferation of HA-VSMC via JAK/STAT3 pathway. After HUCBSC local transplantation, HUCBSC had no distal tissue distribution. HUCBSC transplantation may prevent restenosis after PTA of diabetic hindlimb vascular disease through JAK/STAT3 pathway.

Myotonia congenita and hypokalemic periodic paralysis type 2 are both rare genetic channelopathies caused by mutations in the CLCN1 gene encoding voltage-gated chloride channel CLC-1 and the SCN4A gene encoding voltage-gated sodium channel Nav1.4. The patients with concomitant mutations in both genes manifested different unique symptoms from mutations in these genes separately. Here, we describe a patient with myotonia and periodic paralysis in a consanguineous marriage pedigree. By using whole-exome sequencing, a novel F306S variant in the CLCN1 gene and a known R222W mutation in the SCN4A gene were identified in the pedigree. Patch clamp analysis revealed that the F306S mutant reduced the opening probability of CLC-1 and chloride conductance. Our study expanded the CLCN1 mutation database. We emphasized the value of whole-exome sequencing for differential diagnosis in atypical myotonic patients.

Conventional education results in unsatisfactory morphological understanding of acetabular fractures due to lack of three-dimensional (3D) details and tactile feedback of real fractures. Virtual reality (VR) and 3D printing (3DP) techniques are widely applied in teaching. The purpose of this study was to identify the effect of physical model (PM), VR and 3DP models in education of morphological understanding of acetabular fractures. 141 students were invited to participate in this study. Participants were equally and randomly assigned to the PM, VR and 3DP learning groups. Three-level objective tests were conducted to evaluate learning, including identifying anatomical landmarks, describing fracture lines, identifying classification, and inferring fracture mechanism. Four subjective questions were asked to evaluate the usability and value of instructional materials. Generally, the 3DP group showed a clear advantage over the PM and VR groups in objective tests, while there was no significant difference between the PM and VR groups. 3DP was considered to be the most valuable learning tool for understanding acetabular fractures. The findings demonstrate that 3DP modelling of real fractures is an effective learning instrument that can be used to understand the morphology of acetabular fractures and promote subjective interest.

Identification of genes involved in mangrove species' adaptation to salt stress can provide valuable information for developing salt-tolerant crops and understanding the molecular evolution of salt tolerance in halophiles. Ceriops tagal is a salt-tolerant mangrove tree growing in mudflats and marshes in tropical and subtropical areas, without any prior genome information. In this study, we assessed the biochemical and transcriptional responses of C. tagal to high salt treatment (500 mmol/L NaCl) by hydroponic experiments and RNA-seq. In C. tagal root tissues under salt stress, proline accumulated strongly from 3 to 12 h of treatment; meanwhile, malondialdehyde content progressively increased from 0 to 9 h, then dropped to lower than control levels by 24 h. These implied that C. tagal plants could survive salt stress through biochemical modification. Using the Illumina sequencing platform, approximately 27.39 million RNA-seq reads were obtained from three salt-treated and control (untreated) root samples. These reads were assembled into 47,111 transcripts with an average length of 514 bp and an N50 of 632 bp. Approximately 78% of the transcripts were annotated, and a total of 437 genes were putative transcription factors. Digital gene expression analysis was conducted by comparing transcripts from the untreated control to the three salt treated samples, and 7,330 differentially expressed transcripts were identified. Using k-means clustering, these transcripts were divided into six clusters that differed in their expression patterns across four treatment time points. The genes identified as being up- or downregulated are involved in salt stress responses, signal transduction, and DNA repair. Our study shows the main adaptive pathway of C. tagal in saline environments, under short-term and long-term treatments of salt stress. This provides vital clues as to which genes may be candidates for breeding salt-tolerant crops and clarifying molecular mechanisms of salt tolerance in C. tagal. The expression levels of 20 candidate genes measured by RNA-Seq were validated via qRT-PCR. Eighteen genes showed consistent expression patterns in RNA-Seq and qRT-PCR results, suggesting that the RNA-seq dataset was dependable for gene expression pattern analysis.

RNA replication of positive-strand (+)RNA viruses requires the protein-protein interactions among viral replicases and the association of viral replicases with intracellular membranes. Protein A from Wuhan nodavirus (WhNV), which closely associate with mitochondrial membranes, is the sole replicase required for viral RNA replication. Here, we studied the direct effects of mitochondrial membrane lipids (MMLs) on WhNV protein A activity in vitro. Our investigations revealed the self-interaction of WhNV protein A is accomplished via two different patterns (i.e., homotypic and heterotypic self-interactions via different interfaces). MMLs stimulated the protein A self-interaction, and this stimulation exhibited selectivity for specific phospholipids. Moreover, we found that specific phospholipids differently favor the two self-interaction patterns. Furthermore, manipulating specific phospholipid metabolism affected protein A self-interaction and the activity of protein A to replicate RNA in cells. Taken together, our findings reveal the direct effects of membrane lipids on a nodaviral RNA replicase.

Flock House virus (FHV) is a positive-stranded RNA virus with a bipartite genome of RNAs, RNA1 and RNA2, and belongs to the family Nodaviridae. As the most extensively studied nodavirus, FHV has become a well-recognized model for studying various aspects of RNA virology, particularly viral RNA replication and antiviral innate immunity. FHV RNA1 encodes protein A, which is an RNA-dependent RNA polymerase (RdRP) and functions as the sole viral replicase protein responsible for RNA replication. Although the RNA replication of FHV has been studied in considerable detail, the mechanism employed by FHV protein A to initiate RNA synthesis has not been determined. In this study, we characterized the RdRP activity of FHV protein A in detail and revealed that it can initiate RNA synthesis via a de novo (primer-independent) mechanism. Moreover, we found that FHV protein A also possesses a terminal nucleotidyl transferase (TNTase) activity, which was able to restore the nucleotide loss at the 3'-end initiation site of RNA template to rescue RNA synthesis initiation in vitro, and may function as a rescue and protection mechanism to protect the 3' initiation site, and ensure the efficiency and accuracy of viral RNA synthesis. Altogether, our study establishes the de novo initiation mechanism of RdRP and the terminal rescue mechanism of TNTase for FHV protein A, and represents an important advance toward understanding FHV RNA replication.

Insects and plants have coexisted for over 350 million years and their interactions have affected ecosystems and agricultural practices worldwide. Variation in herbivorous insects' virulence to circumvent host resistance has been extensively documented. However, despite decades of investigation, the genetic foundations of virulence are currently unknown. The brown planthopper (Nilaparvata lugens) is the most destructive rice (Oryza sativa) pest in the world. The identification of the resistance gene Bph1 and its introduction in commercial rice varieties prompted the emergence of a new virulent brown planthopper biotype that was able to break the resistance conferred by Bph1. In this study, we aimed to construct a high density linkage map for the brown planthopper and identify the loci responsible for its virulence in order to determine their genetic architecture. Based on genotyping data for hundreds of molecular markers in three mapping populations, we constructed the most comprehensive linkage map available for this species, covering 96.6% of its genome. Fifteen chromosomes were anchored with 124 gene-specific markers. Using genome-wide scanning and interval mapping, the Qhp7 locus that governs preference for Bph1 plants was mapped to a 0.1 cM region of chromosome 7. In addition, two major QTLs that govern the rate of insect growth on resistant rice plants were identified on chromosomes 5 (Qgr5) and 14 (Qgr14). This is the first study to successfully locate virulence in the genome of this important agricultural insect by marker-based genetic mapping. Our results show that the virulence which overcomes the resistance conferred by Bph1 is controlled by a few major genes and that the components of virulence originate from independent genetic characters. The isolation of these loci will enable the elucidation of the molecular mechanisms underpinning the rice-brown planthopper interaction and facilitate the development of durable approaches for controlling this most destructive agricultural insect.

The worldwide expansion of four serotypes of dengue virus (DENV) poses great risk to global public health. Several vaccine candidates are under development. However, none is yet available for humans. In the present study, a novel strategy to produce tetravalent DENV vaccine based on envelope protein domain III (EDIII) was proposed. Tandem EDIIIs of two serotypes (type 1-2 and type 3-4) of DENV connected by a Gly-Ser linker ((Gly4Ser)3) were expressed in E. coli, respectively. Then, the two bivalent recombinant EDIIIs were equally mixed to form the tetravalent vaccine candidate MixBiEDIII, and used to immunize BALB/c mice. The results showed that specific IgG and neutralizing antibodies against all four serotypes of DENV were successfully induced in the MixBiEDIII employing Freund adjuvant immunized mice. Furthermore, in the suckling mouse model, sera from mice immunized with MixBiEDIII provided significant protection against four serotypes of DENV challenge. Our data demonstrated that MixBiEDIII, as a novel form of subunit vaccine candidates, might have the potential to be further developed as a tetravalent dengue vaccine in the near future.

PurposeTo investigate the efficacy of leflunomide in experimental autoimmune uveitis (EAU) in rats.MethodsLewis rats were immunized with interphotoreceptor retinoid-binding peptide (IRBP) in order to generate EAU. Rats received three dose of leflunomide through intragastric administration (prevention or treatment protocols) after immunization at three separate doses (3 mg/kg/d; 6 mg/kg/d; 12 mg/kg/d). Cyclosporin A was administered as a positive) control. Rats were euthanized during peak disease activity (day 14 or 15). Treatment effectiveness was evaluated in vivo using clinical EAU scoring (d14) and histopathological evaluation of enucleated eyes after experimental termination. The expression levels of inflammatory cytokines in the serum were quantified by ELISA. Eyeball of rats were harvested and mRNA expression of interleukin 17 (IL17) and IFN-γ were quantified through RT-PCR. Intracellular expression of interleukin (IL)-17 in the activated CD4(+) T cells was assessed by flow cytometry. The effects of leflunomide inhibition on immune responses in rats were investigated in isolated lymphocytes.ResultsHistopathological and clinical data revealed severe intraocular inflammation in the immunized rat. Inflammation reached its peak on day 14 in this EAU model. Treatment with leflunomide significantly prevented and treated EAU-induced ocular inflammation and decreased clinical and pathological scores compared to vehicle-treated eyes. Gene expression of IL17 and IFN-γ was markedly reduced in leflunomide-treated eyes. Leflunomide significantly decreased the serum levels of IL17 and IFN-γ. The study of IL17+ T cells in peripheral blood and spleen by flow cytometry showed a decreased number of Th17 cell in rats of leflunomide prevented group. Lymphocytes from animals treated with leflunomide had decreased antigen-specific proliferation in vitro compared with lymphocytes from untreated animals.ConclusionsOral administration of leflunomide effectively suppressed IRBP-induced uveitis in rats. These results suggest that leflunomide may be potentially clinical application in uveitis.

OBJECTIVE: There are numerous reports about the leptin concentration (LC) in postmenopausal women (PW). Changes in LC can elicit different clinical outcomes. We systematically analyzed the LC in PW. METHODS: A search was conducted in original English-language studies published from 1994 to October 2012 in the following databases: Medline (78), Cochrane Center (123) Embase (505), Biological abstracts (108), Cochrane (53) and Science Finder Scholar (0). A meta-analysis was undertaken on the correction coefficient (r) between the serum LC and body mass index (BMI) for healthy PW across studies containing a dataset and sample size. Pre-analytical and analytical variations were examined. Pre-analytical variables included fasting status (FS) and sampling timing. Analytical variation comprised assay methodology, LC in those undertaking hormone replacement therapy (HRT) and those not having HRT as well as LC change according to age. RESULTS: Twenty-seven studies met the inclusion criteria. Eighteen studies detected LC in the morning in a FS, 15 studies denoted the r between leptin and the BMI. A combined r was counted for the 15 studies (r = 0.51 [95% confidence interval (CI), 0.46-0.54], P = 0.025), and if sampling collection was in the FSat morning, a combined r was form 10 studies (r = 0.54 [95% CI, 0.45-0.54], P = 0.299) and heterogeneity was diminished. LC did not change between HRT users and non-users in 7 studies. Five studies analyzed changes in LC according to age. CONCLUSION: Based on all studies that investigated both LC and BMI, LC was positively correlated with the BMI. No studies established reference ranges according to the Clinical and Laboratory Standards Institute (CLSI) in healthy PW, and there was a wide variation in LC values. These differences suggest that caution should be used in the interpretation and comparison between studies.