The aim of the present study was to investigate the key gene network in fracture healing. The dataset GSE45156 was downloaded from the Gene Expression Omnibus. Differentially expressed genes (DEGs) were identified using the linear models for microarray data package of Bioconductor. Subsequently, Gene Ontology (GO) functional and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were conducted for DEGs in day 2 and 6 fractured samples via the Database for Annotation, Visualization and Integrated Discovery. Furthermore, protein‑protein interactions (PPIs) of DEGs were analyzed and a PPI network was constructed. A total of 774 and 1,172 DEGs were identified in day 2 and 6 fractured samples, respectively, compared with unfractured controls. Of the DEGs in day 2 and 6 fractured samples, various upregulated DEGs, including protein kinase C α (Prkca) and B‑cell lymphoma antagonist/killer 1 were significantly enriched in GO terms associated with cell death, and certain downregulated DEGs, including fms‑related tyrosine kinase 1 (Flt1), nitric oxide synthase 3 (Nos3), bone morphogenetic protein 4 (Bmp4) and Notch1 were enriched in GO terms associated with angiogenesis. Furthermore, a series of downregulated DEGs were enriched in the Notch signaling pathway, including hes family bHLH transcription factor 1 and Notch1. Certain DEGs had a high degree and interacted with each other, including Flt1, Nos3, Bmp4 and Notch1, and Prkca and ras‑related C3 botulinum toxin substrate 3. The up and downregulated DEGs may exert critical functions by interactively regulating angiogenesis or apoptosis.

Enabled homolog (Enah), which is a member of the Ena/VASP family that also includes VASP (vasodilator-stimulated phosphoprotein) and Ena/VASP like, is a mammalian ortholog of Drosophila Enabled (Ena). An increasing number of studies demonstrated Enah overexpression is involved in human colorectal carcinomas, breast cancers and hepatocellular carcinoma. However, the significance of Enah expression in gastric cancer (GC) is poorly elucidated. Here, we demonstrate that Enah is upregulated in GC and associated with AJCC stage, depth of invasion and poor overall survival (OS). Knockdown of Enah inhibited GC cell proliferation and metastasis and vice versa. Further experiments suggested that p-Erk1/2, p-AKT, p-p65, Vimentin and Fibronectin were downregulated and E-cadherin was upregulated after Enah silencing, implicating altered functions in GC proliferation and metastasis. Thus, our study suggests that Enah is a harmful factor for GC and a novel target for GC treatment.

The expression of PD-L1 in tumor cells is one of the main causes of tumor immune escape. However, the exact mechanism for regulating PD-L1 expression in gastric cancer (GC) cells remains unclear. Our previous studies have shown that mesenchymal stem cells (MSCs) exert broad immunosuppressive potential, modulating the activity of cells either in innate or adaptive immune system to promote tumor progress. This study aims to investigate whether GCMSCs regulate the PD-L1 expression in GC cells and explore the specific molecular mechanism. The results have shown that GCMSCs enhanced PD-L1 expression in GC cells resulting in the resistance of GC cells to CD8+ T cells cytotoxicity. However, this resistance was attenuated with IL-8 inhibition. Further studies proved that IL-8 derived from GCMSCs induced PD-L1 expression in GC cells via c-Myc regulated by STAT3 and mTOR signaling pathways. Our data indicated that blocking IL-8 derived from GCMSCs may overcome the immune escape induced by PD-L1 in GC cells and provide a potential strategy to enhance the immunotherapy efficiency in GC.

The zinc finger E-box-binding homeobox 1 (ZEB1) induced the epithelial-mesenchymal transition (EMT) and altered ZEB1 expression could lead to aggressive and cancer stem cell (CSC) phenotypes in various cancers. Tissue specimens from 96 prostate cancer patients were collected for immunohistochemistry and CD34/periodic acid-Schiff double staining. Prostate cancer cells were subjected to ZEB1 knockdown or overexpression and assessment of the effects on vasculogenic mimicry formation in vitro and in vivo. The underlying molecular events of ZEB1-induced vasculogenic mimicry formation in prostate cancer were then explored. The data showed that the presence of VM and high ZEB1 expression was associated with higher Gleason score, TNM stage, and lymph node and distant metastases as well as with the expression of vimentin and CD133 in prostate cancer tissues. Furthermore, ZEB1 was required for VM formation and altered expression of EMT-related and CSC-associated proteins in prostate cancer cells in vitro and in vivo. ZEB1 also facilitated tumour cell migration, invasion and clonogenicity. In addition, the effects of ZEB1 in prostate cancer cells were mediated by Src signalling; that is PP2, a specific inhibitor of the Src signalling, dose dependently reduced the p-Src527 level but not p-Src416 level, while ZEB1 knockdown also down-regulated the level of p-Src527 in PC3 and DU-145 cells. PP2 treatment also significantly reduced the expression of VE-cadherin, vimentin and CD133 in these prostate cancer cells. Src signalling mediated the effects of ZEB1 on VM formation and gene expression.

The powerful Quantitative real-time PCR (RT-qPCR) was widely used to assess gene expression levels, which requires the optimal reference genes used for normalization. Oenococcus oeni (O. oeni), as the one of most important microorganisms in wine industry and the most resistant lactic acid bacteria (LAB) species to ethanol, has not been investigated regarding the selection of stable reference genes for RT-qPCR normalization under ethanol stress conditions. In this study, nine candidate reference genes (proC, dnaG, rpoA, ldhD, ddlA, rrs, gyrA, gyrB, and dpoIII) were analyzed to determine the most stable reference genes for RT-qPCR in O. oeni SD-2a under different ethanol stress conditions (8, 12, and 16% (v/v) ethanol). The transcript stabilities of these genes were evaluated using the algorithms geNorm, NormFinder, and BestKeeper. The results showed that dnaG and dpoIII were selected as the best reference genes across all experimental ethanol conditions. Considering single stress experimental modes, dpoIII and dnaG would be suitable to normalize expression level for 8% ethanol shock treatment, while the combination of gyrA, gyrB, and rrs would be suitable for 12% ethanol shock treatment. proC and gyrB revealed the most stable expression in 16% ethanol shock treatment. This study selected and validated for the first time the reference genes for RT-qPCR normalization in O. oeni SD-2a under ethanol stress conditions.

Lycorine has been revealed to inhibit the development of many kinds of malignant tumors, including glioblastoma multiforme (GBM). Although compelling evidences demonstrated Lycorine's inhibition on cancers through some peripheral mechanism, in-depth mechanism studies of Lycotine's anti-GBM effects still call for further exploration. Epidermal Growth Factor Receptor (EGFR) gene amplification and mutations are the most common oncogenic events in GBM. Targeting EGFR by small molecular inhibitors is a rational strategy for GBM treatment.

Imaging genetic studies typically focus on identifying single-nucleotide polymorphism (SNP) markers associated with imaging phenotypes. Few studies perform regression of SNP values on phenotypic measures for examining how the SNP values change when phenotypic measures are varied. This alternative approach may have a potential to help us discover important imaging genetic associations from a different perspective. In addition, the imaging markers are often measured over time, and this longitudinal profile may provide increased power for differentiating genotype groups. How to identify the longitudinal phenotypic markers associated to disease sensitive SNPs is an important and challenging research topic.

Although IGF-1 has been implicated in mediating hypersensitivity to pain, the underlying mechanisms remain unclear. We identified a novel functional of the IGF-1 receptor (IGF-1R) in regulating A-type K(+) currents (IA) as well as membrane excitability in small trigeminal ganglion neurons. Our results showed that IGF-1 reversibly decreased IA, whereas the sustained delayed rectifier K(+) current was unaffected. This IGF-1-induced IA decrease was associated with a hyperpolarizing shift in the voltage dependence of inactivation and was blocked by the IGF-1R antagonist PQ-401; an insulin receptor tyrosine kinase inhibitor had no such effect. An small interfering RNA targeting the IGF-1R, or pretreatment of neurons with specific phosphatidylinositol 3-kinase (PI3K) inhibitors abolished the IGF-1-induced IA decrease. Surprisingly, IGF-1-induced effects on IA were not regulated by Akt, a common downstream target of PI3K. The MAPK/ERK kinase inhibitor U0126, but not its inactive analog U0124, as well as the c-Raf-specific inhibitor GW5074, blocked the IGF-1-induced IA response. Analysis of phospho-ERK (p-ERK) showed that IGF-1 significantly activated ERK1/2 whereas p-JNK and p-p38 were unaffected. Moreover, the IGF-1-induced p-ERK1/2 increase was attenuated by PI3K and c-Raf inhibition, but not by Akt blockade. Functionally, we observed a significantly increased action potential firing rate induced by IGF-1; pretreatment with 4-aminopyridine abolished this effect. Taken together, our results indicate that IGF-1 attenuates IA through sequential activation of the PI3K- and c-Raf-dependent ERK1/2 signaling cascade. This occurred via the activation of IGF-1R and might contribute to neuronal hyperexcitability in small trigeminal ganglion neurons.

Most recently a novel avian-origin influenza A (H7N9) virus emerged in China and has been associated with lots of human infection and fatal cases. Genetic analysis of the viral genome revealed that this reassortant virus might be better adapted to humans than other avian influenza viruses. Molecular diagnostic methods are thus urgently needed in public health laboratories. In this study, a SYBR green-based one-step real time reverse transcription-PCR (RT-PCR) was developed to detect the novel H7N9 virus. The primer pairs on the basis of the hemagglutinin and neuraminidase gene sequences of H7N9 viruses amplified subtype-specific fragments with Tm values of 80.77±0.06°C for H7 and 81.20±0.17°C for N9 respectively. The standard curves showed a dynamic linear range across 6 log units of RNA copy number (10(6) to 10(1) copies/ µl) with a detection limit of 10 copies per reaction for both H7 and N9 assays by using serial ten-fold diluted in-vitro transcribed viral RNA. In addition, no cross-reactivity was observed with seasonal H1N1, H1N1 pdm09, H3N2, H5N1 and H9N2 viruses as well as other human respiratory viruses. When the assay was further evaluated in H7N9 virus infected clinical samples, positive amplification signals were obtained in all of the specimens with the accordance between H7 and N9 assays. Therefore, the established SYBR green-based real time RT-PCR assay could provide a rapid, sensitive, specific and reliable alternative approach with lower costs for high throughput screening of suspected samples from humans, animals and environments in first line public health laboratories.

Several reports indicate that melatonin alleviates bleomycin (BLM)-induced pulmonary fibrosis in rodent animals. Nevertheless, the exact mechanism remains obscure. The present study investigated the effects of melatonin on endoplasmic reticulum (ER) stress and epithelial-mesenchymal transition (EMT) during BLM-induced lung fibrosis. For the induction of pulmonary fibrosis, mice were intratracheally injected with a single dose of BLM (5.0 mg/kg). Some mice were intraperitoneally injected with melatonin (5 mg/kg) daily for a period of 3 wk. Twenty-one days after BLM injection, lung fibrosis was evaluated. As expected, melatonin significantly alleviated BLM-induced pulmonary fibrosis, as evidenced by Sirius red staining. Moreover, melatonin significantly attenuated BLM-induced EMT to myofibroblasts, as determined by its repression of α-SMA expression. Further analysis showed that melatonin markedly attenuated BLM-induced GRP78 up-regulation and elevation of the cleaved ATF6 in the lungs. Moreover, melatonin obviously attenuated BLM-induced activation of pulmonary eIF2α, a downstream target of the PERK pathway. Finally, melatonin repressed BLM-induced pulmonary IRE1α phosphorylation. Correspondingly, melatonin inhibited BLM-induced activation of XBP-1 and JNK, two downstream targets of the IRE1 pathway. Taken together, these results suggest that melatonin alleviates ER stress and ER stress-mediated EMT in the process of BLM-induced pulmonary fibrosis.

Mammalian target of rapamycin (mTOR) represents a key downstream intermediate for a myriad of oncogenic receptor tyrosine kinases. In the case of the insulin-like growth factor (IGF) pathway, the mTOR complex (mTORC1) mediates IGF-1 receptor (IGF-1R)-induced estrogen receptor alpha (ERα) phosphorylation/activation and leads to increased proliferation and growth in breast cancer cells. As a result, the prevalence of mTOR inhibitors combined with hormonal therapy has increased in recent years. Conversely, activated mTORC1 provides negative feedback regulation of IGF signaling via insulin receptor substrate (IRS)-1/2 serine phosphorylation and subsequent proteasomal degradation. Thus, the IGF pathway may provide escape (e.g. de novo or acquired resistance) from mTORC1 inhibitors. It is therefore plausible that combined inhibition of mTORC1 and IGF-1R for select subsets of ER-positive breast cancer patients presents as a viable therapeutic option.

The purpose of the present study was to investigate the molecular mechanisms of myotonic dystrophy (DM) 1 and 2 cataracts using bioinformatics methods. A microarray dataset (E‑MEXP‑3365) downloaded from the Array Express database included lens epithelial samples of DM1 and DM2 cataract patients (n=3/group) and non‑DM lens epithelial samples as a control (n=4). Differentially expressed genes (DEGs) were identified between DM1 and control samples, and between DM2 and control samples. Pathway enrichment analyses were performed for the DEGs. Potential micro (mi)RNAs regulating these DEGs were predicted. An miRNA‑target gene network was constructed for DM1 and DM2. The study identified 223 DEGs in DM1, and 303 DEGs in DM2. DM1 and DM2 shared 172 DEGs. The DEGs in DM1 were enriched with calcium, Wnt and axon guidance signaling pathways. The DEGs in DM2 were linked by adherens junction signaling pathways. miRNA (miR)‑197, miR‑29b and miR‑29c were included in the network modules of DM1. miR‑197, miR‑29c and miR‑29a were involved in the network modules of DM2. It is therefore hypothesized that these signaling pathways and miRNAs underlie DM1 and DM2 cataracts, and may represent potential therapeutic targets for the treatment of this disorder.

This study aims to investigate the prognostic value of pretreatment C-reactive protein/albumin ratio (CAR) in human malignancies by an updated meta-analysis.

Dilated cardiomyopathy (DCM) is characterized by left ventricular dilation, and is associated with systolic dysfunction and increased action potential duration. Approximately 50% of DCM cases are caused by inherited gene mutations with genetic and phenotypic heterogeneity. Next generation sequencing may be useful in screening unknown mutations in such cases.A family was identified with DCM, in which the affected family members developed heart failure, arrhythmia, and sudden death. Probands and 4 affected family members underwent whole exome sequencing (WES), bioinformatics methods, and gene annotation to identify potentially causative variants. The Sanger sequencing method was used to verify the candidate mutation.WES yielded 2,238,831 variations. KCNJ12 (p.Glu334del) was identified as a candidate mutation, and the heterozygous mutation was verified by Sanger sequencing.Our study emphasizes the application of WES in identifying causative mutations in DCM. This report is the first to describe the KCNJ12 gene as a cause of DCM in patients.

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.

Atopic dermatitis (AD) is characterized by a defective skin barrier, which increases the penetration of allergens and pathogens through the skin. The role of interleukin (IL)‑17, a pro‑inflammatory cytokine, in the pathogenesis of AD remains to be elucidated. The present study aimed to examine the effects of IL‑17 on skin barrier proteins in the HaCaT cell line. The expression levels of filaggrin (FLG) and involucrin (IVL) were evaluated by reverse transcription‑quantitative polymerase chain reaction and western blot analyses of the HaCaT cells following IL‑17 simulation. The role of IL‑17 was further examined by using small molecule inhibitors of extracellular signal‑regulated kinase (ERK) and P38. Treatment of the HaCaT cells with IL‑17 resulted in reduced expression levels of FLG and IVL at the mRNA and protein levels. In addition, the gene expression levels of FLG and IVL were significantly reduced in the HaCaT cells by IL‑4. Treatment with the mitogen‑activated protein kinase (MAPK) inhibitors, SB203580 and PD98059, significantly inhibited the effects of IL‑17 on the gene and protein expression levels of FLG and IVL. Finally, the protein levels of phosphorylated ERK and P38 were significantly increased following IL‑17 stimulation. Taken together, the results revealed that IL‑17 reduced the expression of FLG and IVL in HaCaT cells, and this effect involved the P38/ERK MAPK signaling pathways.

The present study aimed to explore the underlying molecular mechanisms of hepatocellular carcinoma (HCC). RNA‑sequencing profiles GSM629264 and GSM629265, from the GSE25599 data set, were downloaded from the Gene Expression Omnibus database and processed by quality evaluation. GSM629264 and GSM629265 were from HCC and adjacent non‑cancerous tissues, respectively. TopHat software was used for alignment analysis, followed by the detection of novel splicing sites. In addition, the Cufflinks software package was used to analyze gene expressions, and the Cuffdiff program was used to screen for differently expressed genes (DEGs) and differentially expressed splicing variants. Gene ontology functional enrichment and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses of DEGs were also performed. Transcription factors (TFs) and microRNAs (miRNAs) that regulate DEGs were identified, and a protein‑protein interaction (PPI) network was constructed. The hub node in the PPI network was obtained, and the TFs and miRNAs that regulated the hub node were further predicted. The quality of the sequencing data met the standards for analysis, and the clean reads were ~65%. Most sequencing reads mapped into coding sequence exons (CDS_exons), whereas other reads mapped into exon 3' untranslated regions (UTR_Exons), 5'UTR_Exons and Introns. Upregulated and downregulated DEGs between HCC and adjacent non‑cancerous tissues were screened. Genes of differentially expressed splicing variants were identified, including vesicle‑associated membrane protein 4, phosphatidylinositol glycan anchor biosynthesis class C, protein disulfide isomerase family A member 4 and growth arrest specific 5. Screened DEGs were enriched in the complement pathway. In the PPI network, ubiquitin C (UBC) was the hub node. UBC was predicted to be regulated by several TFs, including specificity protein 1 (SP1), FBJ murine osteosarcoma viral oncogene homolog (FOS), proto‑oncogene c‑JUN (JUN), FOS‑like antigen 2 (FOSL2) and SWI/SNF‑related, matrix‑associated, actin‑dependent regulator of chromatin, subfamily A, member 4 (SMARCA4), and several miRNAs, including miR‑30 and miR‑181. Results from the present study demonstrated that UBC, SP1, FOS, JUN, FOSL2, SMARCA4, miR‑30 and miR‑181 may participate in the development of HCC.

Pulmonary tuberculosis (TB) is a highly lethal infectious disease and early diagnosis of TB is critical for the control of disease progression. The objective of this study was to profile a panel of serum microRNAs (miRNAs) as potential biomarkers for the early diagnosis of pulmonary TB infection.

Altered circulating microRNA (miRNA) profiles have been noted in patients with microbial infections. We compared host serum miRNA levels in patients with hand-foot-and-mouth disease (HFMD) caused by enterovirus 71 (EV71) and coxsackievirus 16 (CVA16) as well as in other microbial infections and in healthy individuals. Among 664 different miRNAs analyzed using a miRNA array, 102 were up-regulated and 26 were down-regulated in sera of patients with enteroviral infections. Expression levels of ten candidate miRNAs were further evaluated by quantitative real-time PCR assays. A receiver operating characteristic (ROC) curve analysis revealed that six miRNAs (miR-148a, miR-143, miR-324-3p, miR-628-3p, miR-140-5p, and miR-362-3p) were able to discriminate patients with enterovirus infections from healthy controls with area under curve (AUC) values ranged from 0.828 to 0.934. The combined six miRNA using multiple logistic regression analysis provided not only a sensitivity of 97.1% and a specificity of 92.7% but also a unique profile that differentiated enterovirial infections from other microbial infections. Expression levels of five miRNAs (miR-148a, miR-143, miR-324-3p, miR-545, and miR-140-5p) were significantly increased in patients with CVA16 versus those with EV71 (p<0.05). Combination of miR-545, miR-324-3p, and miR-143 possessed a moderate ability to discrimination between CVA16 and EV71 with an AUC value of 0.761. These data indicate that sera from patients with different subtypes of enteroviral infection express unique miRNA profiles. Serum miRNA expression profiles may provide supplemental biomarkers for diagnosing and subtyping enteroviral HFMD infections.

Previous studies have shown that inflammation process involves pathogenesis of Alzheimer's disease (AD). But, the natural AD model of inflammation has not been obtained yet. In the present study, CD-1 mothers intraperitoneally received a 50 μg/kg lipopolysaccharide (LPS) or normal saline daily during gestational days 15-17. Body weight of the offspring was recorded at ages of 4-33 weeks. A different battery of behavioral tasks was, respectively, completed at ages of 35, 290 and 400 days. The results showed that there was no significant difference in body weight between LPS-treated and control mice during ages of 4-33 weeks. LPS-treated offspring had similar anxiety and locomotor behaviors, and spatial ability of learning and memory at the age of 35 days compared to the controls. At an age of 290 days, the LPS-treated offspring had similar sensorimotor ability, locomotor activity and anxiety, species-typical behaviors, and spatial ability of learning and memory. At an age of 400 days, there were similar sensorimotor ability, locomotor activity and anxiety between the LPS-treated offspring and controls. However, there were impaired species-typical behaviors, and spatial and non-spatial abilities of learning and memory in the LPS-treated offspring. Our results suggested that maternal exposure to LPS in adequate dose in late gestation can deliver term offspring which experience a normal duration of development and maturation, and an accelerated aged-related impairment in memory (spatial and non-spatial) and species-typical behaviors in middle-aged. These meet with the criteria of AD model in behaviors.