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The vascular endothelium plays a key role in the pathobiology of atherosclerotic cardiovascular disease. Endothelial cell Piezo1 mediates blood vessel formation, angiogenesis and regulation of blood pressure. However, changes of Piezo1 expression in atherosclerosis (AS) and the role of Piezo1 in the progression of atherosclerotic diseases remains obscure. Thus, the current study is to elucidate the role and mechanism of which Piezo1 mediates vascular inflammation in atherosclerotic mice and vascular endothelial inflammation induced by oxidized low density lipoprotein (ox-LDL) in vitro. Here, we have shown that the expression of Piezo1 was significantly increased in the stenotic carotid artery of ApoE-/- mice fed by high-fat diet (HFD). Pharmacological inhibition of Piezo1 (GsMTx-4) attenuated plaque formation, decreased the level of inflammation related factors (JNK, TNF-α, NF-κB, VCAM-1) of carotid plaque in atherosclerotic mice. Meanwhile, ox-LDL also upregulates Piezo1 and inflammation proteins (NF-κB, JNK and TNF-α) in endothelium cells (ECs). YAP/TAZ is activated accompanied by the enhanced Piezo1 activity in ECs induced by ox-LDL. Interference by siRNA of Piezo1 abolished the expression of YAP/TAZ and inflammation proteins (JNK, NF-κB and TNF-α). In addition, Ca 2+ influx in ECs induced by ox-LDL was increased than control group, Piezo1 siRNA can reduce the calcium content. Piezo1 agonist Yoda1 increased Ca 2+ influx and promote YAP nucleus translocation in ECs, genetic deletion of Piezo1 reversed it. Our results indicate that Piezo1 could mediate endothelial atherogenic inflammatory responses via regulation of YAP/TAZ activation and nuclear localization. Piezo1 may be a potential therapeutic target for atherosclerotic diseases in the future., (© 2021. Japan Human Cell Society.)

Background: Dihydroartemisinin (DHA) possesses an inhibitory effect on ovarian cancer and promotes reversion-inducing cysteine-rich protein with Kazal motifs (RECK) expression in glioma cells. This study explored the role of DHA and RECK on ovarian cancer., Methods: The RECK level in ovarian cancer was analyzed under GEPIA 2 database and proved by RT-qPCR. After being treated with DHA or infected with siRECK lentivirus, the viability, apoptosis, migration, and invasion of ovarian cancer cells were evaluated by CCK-8, flow cytometry, wound healing, and transwell assays. Also, the expressions of factors related to apoptosis and epithelial-mesenchymal transition were measured by Western blot or RT-qPCR., Results: DHA-treatment weakened the viability, migration, invasion, and enhanced apoptosis of ovarian cancer cells. DHA also down-regulated the levels of Bcl-2, N-cadherin, and Vimentin, and up-regulated the levels of Bax, C-caspase-3 and E-cadherin in ovarian cancer cells. RECK was lowly expressed in both ovarian cancer tissues and cells. siRECK not only had an effect opposite to DHA on the viability, apoptosis, migration, invasion, and related-factors of ovarian cancer cells but also offset the effect of DHA on ovarian cancer cells., Conclusion: DHA regulated apoptosis, migration, and invasion of ovarian cancer cells via mediating RECK., Competing Interests: Declaration of competing interest The authors indicated no potential conflicts of interest., (Copyright © 2021 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Background: It has been reported that the dysregulation of microRNAs (miRNAs) is implicated in the biological processes of diverse diseases, including the tumorigenesis of human cancers. MicroRNA-628-5p (miR-628-5p) is differentially expressed and plays a critical role in several cancers, but the role of miR-628-5p in cervical cancer has not been well studied., Methods: The TCGA database and RT-qPCR were used to evaluate the expression profile of miR-628-5p in cervical cancer tissues. Transfection efficiency of synthetic miRNAs was detected using RT-qPCR. The biological effects of miR-628-5p on cervical cancer cells were assessed by the CCK-8 assay, flow cytometry, western blot analysis, and the tube formation assay. The expression levels of key proteins involved in cell apoptosis, the cell cycle and the PI3K pathway were analyzed by western blot analysis. Bioinformatic analysis and the luciferase reporter assay were performed to investigate the targeted relationship between miR-628-5p and vascular endothelial growth factor (VEGF)., Results: MiR-628-5p was downregulated and negatively correlated with Ki-67 expression in cervical cancer tissues, and its low level predicted poor survival of patients. Functional assays indicated that miR-628-5p inhibited cell proliferation and promoted cell apoptosis. Mechanically, VEGF was verified to be a downstream target of miR-628-5p. Moreover, overexpression of VEGF could reverse the effects of miR-628-5p on VEGF/PI3K/AKT signaling, cell proliferation, apoptosis, the cell cycle and angiogenesis in cervical cancer., Conclusions: MiR-628-5p inhibited cervical cancer cell proliferation and promoted apoptosis by targeting VEGF., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.)

The aging brain is associated with reduced cell surface expression of N-methyl-d-aspartate receptors (NMDARs), but the mechanism remains poorly understood. In the present study, we showed that in the striatum and hippocampus but not the cerebellum and parietal cortex, levels of α-synuclein monomers and oligomers increased with age, which correlated negatively with the expression of GluN1, and positively with the expression of total Rab5B. The oligomer-α-synuclein exhibited a stronger correlation with the expression of surface GluN1 and total Rab5B. In MES23.5 cells, the monomer- or oligomer-α-synuclein were shown to increase in a manner dependent on the concentrations of the added monomers and oligomers. Again, the oligomer-α-synuclein showed more potent effects than the monomer-α-synuclein on surface GluN1 and total Rab5B expression. Accordingly, the oligomer-treated cells showed a greater reduction in NMDA-evoked Ca 2+ influx than the monomer-treated cells, which was largely inhibited by pistop2, a clathrin inhibitor. These results suggest that the age-dependent accumulation of α-synuclein monomers and oligomers differentially contributes to the reduction in surface NMDAR expression in selective brain regions., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Inulin is considered an efficient prebiotic and is beneficial for metabolic diseases via promoting intestinal probiotic enrichment and the metabolites of short-chain fatty acids (SCFAs). However, the effect of inulin on patients with InR deficiencies has seldom been reported. In this study, the lifespan, related gene expression, and gut microbiota of InR p5545 under inulin treatment were investigated. The results showed that the lifespan was extended in only males and not in females. Furthermore, distinctly different patterns of gene expression were found between males and females, especially in the insulin/insulin-like growth factor (IGF)-like signalling (IIS) and target of rapamycin (TOR) pathways. Additionally, as a link between inulin and lifespan responses, the gut microbiota was distinctly separated by gender in both the standard diet group and the inulin treatment group, and the relationship between lifespan and the gut microbiota community was stronger in male flies than in females. This study provides preliminary evidence for the gender-dependent lifespan responses to inulin in insulin signalling-deficient Drosophila melanogaster under inulin treatment were investigated. The results showed that the lifespan was extended in only males and not in females. Furthermore, distinctly different patterns of gene expression were found between males and females, especially in the insulin/insulin-like growth factor (IGF)-like signalling (IIS) and target of rapamycin (TOR) pathways. Additionally, as a link between inulin and lifespan responses, the gut microbiota was distinctly separated by gender in both the standard diet group and the inulin treatment group, and the relationship between lifespan and the gut microbiota community was stronger in male flies than in females. This study provides preliminary evidence for the gender-dependent lifespan responses to inulin in insulin signalling-deficient Drosophila . However, controls such as wild-type and TM3 flies, and more InR mutant strains with different genetic backgrounds need to be further investigated to elucidate the mechanisms underlying the phenomenon.

The present study evaluated the distribution of sympathetic and parasympathetic nerves and the expression of the α9 nicotinic acetylcholine receptor (α9nAChR) and investigated their potential association with colorectal cancer (CRC) development. The distribution of autonomic nerves and α9nAChR in CRC was detected by immunohistochemistry, which was then used to analyze their association with clinicopathological parameters and prognosis. Sympathetic fibers were primarily observed in the stroma adjacent to cancer cells, whereas parasympathetic fibers were primarily observed in the stroma away from cancer cells. Patients with samples positive for sympathetic nerve fibers had less lymph node invasion and a better prognosis compared with patients with samples negative for sympathetic nerve fibers. The expression of parasympathetic nerves in patients >60 years old was increased compared with patients ≤60 years old. The expression of parasympathetic nerves in patients with lymph node invasion was increased compared with patients without lymph node invasion. The detection of parasympathetic nerves gradually increased as CRC (T stage) advanced. Patients with parasympathetic negative samples had better prognoses compared with patients with parasympathetic positive samples. The expression of α9nAChR was principally localized in cellular membranes and the cytoplasm of CRC tissues and it was revealed to have a positive association with the number of parasympathetic nerves. Increased α9nAChR expression was observed in patients >60 years old compared with patients <60 years old. The detection rate of α9nAChR in tissues from patients with lymph node invasion was increased compared with patients without lymph node invasion. The detection of α9nAChR gradually increased as the CRC stage advanced. The prognoses for patients with α9nAChR negative tissue were improved compared with the prognoses for patients with α9nAChR positive tissue. Sympathetic nerves were primarily detected in the early phases of CRC and indicated a good prognosis. Parasympathetic nerves and α9nAChR were principally observed in the late phases of cancer and indicated a poor prognosis. The present study revealed that parasympathetic nerves may promote the progression of CRC through α9nAChR.

Understanding the responses of plant growth and secondary metabolites to differential light conditions is very important to optimize cultivation conditions of medicinal woody plants. As a highly valued and multiple function tree species, Cyclocarya paliurus is planted and managed for timber production and medical use. In this study, LED-based light including white light (WL), blue light (BL), red light (RL), and green light (GL) were used to affect leaf biomass production, flavonoid accumulation and related gene expression of one-year C. paliurus seedlings in controlled environments. After the treatments of 60 days, the highest leaf biomass appeared in the treatment of WL, while the lowest leaf biomass was found under GL. Compared to WL, the total flavonoid contents of C. paliurus leaves were significantly higher in BL, RL, and GL, but the highest values of selected flavonoids (kaempferol, isoquercitrin and quercetin) were observed under BL. Furthermore, the greatest yields of total and selected flavonoids in C. paliurus leaves per seedling were also achieved under BL, indicating that blue light was effective for inducing the production of flavonoids in C. paliurus leaves. Pearson's correlation analysis showed that there were significantly positive correlations between leaf flavonoid content and relative gene expression of key enzymes (phenylalanine ammonia lyase, PAL; 4-coumaroyl CoA-ligase, 4CL; and chalcone synthase, CHS) in the upstream, which converting phenylalanine into the flavonoid skeleton of tetrahydroxy chalcone. It is concluded that manipulating light quality may be potential mean to achieve the highest yields of flavonoids in C. paliurus cultivation, however this needs to be further verified by more field trials., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Objectives: (1) To investigate the expression patterns of MΦ1 and MΦ2 phenotype markers of peripheral blood monocyte (PBMC)-derived macrophages in atherosclerosis patients and healthy controls, as well as the expression correlation among these genes. (2) To elucidate whether a high level of liver X receptor α (LXRα) expression is associated with anti-inflammatory MΦ2-type polarization., Design: Peripheral blood monocytes (PBMCs) were obtained from 28 patients with carotid artery plaques and 10 normal persons, who did not have carotid artery plaques. M1 and M2 phenotype markers were analyzed after cellular differentiation into macrophages. Human macrophages derived from healthy donors were transfected with plasmid DNA encoding LXRα and control null-plasmids. Gene expression levels were quantified after further differentiation., Results: Three genes (LXRα, CD68, and CD36) were expressed at a significantly lower rate in the atherosclerotic group than normal patients. There were correlations between the expression of LXRα, CD68, and peroxisome proliferator-activated receptor (PPARγ), and between CD163, CD36 and scavenger receptor class A (SRA1). Macrophages over-expressing LXRα exhibited enhanced expression level of MΦ2-type genes and decreased expression level of MΦ1-type genes., Conclusions: PBMCs from healthy persons were predisposed to the MΦ2 differentiation phenotype, which exhibits elevated cholesterol uptake and anti-inflammatory properties. LXRα over-expression polarizes macrophages towards the anti-inflammatory MΦ2 phenotype.

Background: Sex is an important but understudied factor in the genetics of human diseases. Analyses using a combination of gene expression data, ENCODE data, and evolutionary data of sex-biased gene expression in human tissues can give insight into the regulatory and evolutionary forces acting on sex-biased genes., Methods: In this study, we analyzed the differentially expressed genes between males and females. On the X chromosome, we used a novel method and investigated the status of genes that escape X-chromosome inactivation (escape genes), taking into account the clonality of lymphoblastoid cell lines (LCLs). To investigate the regulation of sex-biased differentially expressed genes (sDEG), we conducted pathway and transcription factor enrichment analyses on the sDEGs, as well as analyses on the genomic distribution of sDEGs. Evolutionary analyses were also conducted on both sDEGs and escape genes., Results: Genome-wide, we characterized differential gene expression between sexes in 462 RNA-seq samples and identified 587 sex-biased genes, or 3.2% of the genes surveyed. On the X chromosome, sDEGs were distributed in evolutionary strata in a similar pattern as escape genes. We found a trend of negative correlation between the gene expression breadth and nonsynonymous over synonymous mutation (dN/dS) ratios, showing a possible pleiotropic constraint on evolution of genes. Genome-wide, nine transcription factors were found enriched in binding to the regions surrounding the transcription start sites of female-biased genes. Many pathways and protein domains were enriched in sex-biased genes, some of which hint at sex-biased physiological processes., Conclusions: These findings lend insight into the regulatory and evolutionary forces shaping sex-biased gene expression and their involvement in the physiological and pathological processes in human health and diseases.

In this study, we investigated the expression profiles of Toll-like receptors(TLRs) in eutopic endometrium(EU) and ectopic endometrium(EC) and its implication in the inflammatory pathogenesis of adenomyosis. Thirty adenomyosis patients who underwent laparoscopy were recruited in this study. We tested the mRNA and protein expression of TLRs, and the mRNA expression of IL-6 and IL-8 in EU and EC of adenomyosis patients, and control endometrium without adenomyosis(CE). We found that the mRNA expression of IL-6 and IL-8 in EU was significantly higher than that in CE, and was the highest in EC (P < 0.01). The mRNA and protein expression of TLRs were higher in EU, with the expression of TLR1-6, 8 and 9 being significantly higher in EU than in CE, and were the highest in EC (except TLR6) (P < 0.05 or P < 0.01). Pearson correlation analysis showed that the expression of TLR1, 2, 4, 5 and 9 in EU and EC was positively correlated with that of IL-6 and IL-8 (P < 0.00139). This study suggested that adenomyosis was a state of inflammatory pathology. High expression of TLRs in EU and EC were positively correlated with IL-6 and IL-8, which may be involved in the inflammatory pathogenesis of adenomyosis.

Background: Cancer stem cell (CSC) epithelial cell adhesion molecule (Ep-CAM) is frequently expressed in colorectal cancer (CRC). However, the clinical significance of Ep-CAM expression in CRC is not clear. This study evaluated whether Ep-CAM provided valuable insight as a molecular biomarker for CRC diagnosis and prognosis and the potential of Ep-CAM as a novel therapeutic target in CRC., Methods: Publications were selected online using electronic databases. The pooled odds ratios (ORs) or hazard ratios (HRs) with their 95% confidence intervals (95% CIs), and the combined sensitivity, specificity, and area under the curve (AUC) were calculated and summarized., Results: Eleven eligible articles published in English involving 4561 cases were analyzed in this study. Ep-CAM expression was significantly higher in CRC compared with normal controls, and its overexpression was negatively linked to tumor differentiation, tumor stage, vascular invasion, depth of tumor invasion, lymph node metastasis, distant metastasis, and tumor budding in CRC patients. The loss of Ep-CAM expression positively correlated with these characteristics. Multivariate analysis of loss of Ep-CAM expression correlated with a poor prognosis in disease-free survival (DFS), disease-specific survival (DSS), and overall survival (OS). The pooled sensitivity, specificity and AUC values of Ep-CAM expression in patients with CRC vs. normal controls were 0.93, 0.90, and 0.94, respectively., Conclusions: The present findings suggest that Ep-CAM expression may be associated with CRC carcinogenesis, while the loss of Ep-CAM expression is correlated with the progression, metastasis, and poor prognosis of CRC. Ep-CAM expression may be a useful biomarker for the clinical diagnosis of CRC., (Copyright © 2017. Published by Elsevier B.V.)

Background & Aims: Intestinal tissues from patients with inflammatory bowel disease (IBD) and colorectal cancer have increased expression of microRNA-301a (MIR301A) compared with tissues from patients without IBD. We studied the mechanisms of MIR301A in the progression of IBD in human tissues and mice., Methods: We isolated intestinal epithelial cells (IECs) from biopsy samples of the colon from 153 patients with different stages of IBD activity, 6 patients with colitis-associated cancer (CAC), and 35 healthy individuals (controls), enrolled in the study in Shanghai, China. We measured expression of MIR301A and BTG anti-proliferation factor 1 (BTG1) by IECs using quantitative reverse-transcription polymerase chain reaction. Human colon cancer cell lines (HCT-116 and SW480) were transfected with a lentivirus that expresses MIR301A; expression of cytokines and tight junction proteins were measured by quantitative reverse transcription polymerase chain reaction, flow cytometry, and immunofluorescence staining. We generated mice with disruption of the microRNA-301A gene (MIR301A-knockout mice), and also studied mice that express a transgene-encoding BTG1. Colitis was induced in knockout, transgenic, and control (C57BL/B6) mice by administration of dextran sulfate sodium (DSS), and mice were given azoxymethane to induce colorectal carcinogenesis. Colons were collected and analyzed histologically and by immunohistochemistry; tumor nodules were counted and tumor size was measured. SW480 cells expressing the MIR301A transgene were grown as xenograft tumors in nude mice., Results: Expression of MIR301A increased in IECs from patients with IBD and CAC compared with controls. MIR301A-knockout mice were resistant to the development of colitis following administration of DSS; their colon tissues expressed lower levels of interleukin 1β (IL1β), IL6, IL8, and tumor necrosis factor than colons of control mice. Colon tissues from MIR301A-knockout mice had increased epithelial barrier integrity and formed fewer tumors following administration of azoxymethane than control mice. Human IECs expressing transgenic MIR301A down-regulated expression of cadherin 1 (also called E-cadherin or CDH1). We identified BTG1 mRNA as a target of MIR301A; levels of BTG1 mRNA were reduced in inflamed mucosa from patients with active IBD compared with controls. There was an inverse correlation between levels of BTG1 mRNA and levels of MIR301A in inflamed mucosal tissues from patients with active IBD. Human colon cancer cell lines that expressed a MIR301A transgene increased proliferation; they had increased permeability and decreased expression of CDH1 compared with cells transfected with a control vector, indicating reduced intestinal barrier function. BTG1 transgenic mice developed less severe colitis than control mice following administration of DSS. SW480 cells expressing anti-MIR301A formed fewer xenograft tumors in nude mice than cells expressing a control vector., Conclusions: Levels of MIR301A are increased in IECs from patients with active IBD. MIR301A reduces expression of BTG1 to reduce epithelial integrity and promote inflammation in mouse colon and promotes tumorigenesis. Strategies to decrease levels of MIR301A in colon tissues might be developed to treat patients with IBD and CAC., (Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Hemophilia B occurs due to a deficiency in human blood coagulation factor IX (hFIX). Currently, no effective treatment for hemophilia B has been identified, and gene therapy has been considered the most appropriate treatment. Mesenchymal stem cells (MSCs) have homing abilities and low immunogenicity, and therefore they may be potential cell carriers for targeted drug delivery to lesional tissues. The present study constructed an adeno‑associated virus integration site 1 (AAVS1)‑targeted vector termed AAVS1‑green fluorescent protein (GFP)‑hFIX and a zinc finger nuclease (ZFN) expression vector. Nucleofection was used to co‑transfect the targeting vector and the ZFN expression vector into human MSCs. The GFP‑positive cells were selected using flow cytometry. Site‑specific integration clones were obtained following the monoclonal culture, subsequent detections were performed using polymerase chain reaction and Southern blotting. Following the confirmation of stem cell traits of the site‑specific integration MSCs, the in vivo and in vitro expression levels of hFIX were detected. The results demonstrated that the hFIX gene was successfully transfected into the AAVS1 locus in human MSCs. The clones with the site‑specific integration retained stem cell traits of the MSCs. In addition, hFIX was effectively expressed in vivo and in vitro. No significant differences in expression levels were identified among the individual clones. In conclusion, the present study demonstrated that the exogenous gene hFIX was effectively expressed following site‑specific targeting into the AAVS1 locus in MSCs; therefore, MSCs may be used as potential cell carriers for gene therapy of hemophilia B.

Human GRP78 has been shown to promote cancer progression and is regarded as a novel target for anticancer drugs. However, generation of recombinant full-length GRP78 remains challenging. This report demonstrates that E. coli autoinduction is an excellent method for the preparation of active recombinant GRP78 protein. The final yield was approximately 50 mg/liter of autoinduction culture. Gel-filtration experiments confirmed that the chaperone is a monomer. The purified human GRP78 catalyzed the conversion of ATP to ADP without requiring metal ions as cofactors. Three mutants, T38A, T229A, and S300A, exhibited much lower activity than wild-type GRP78, indicating that the active sites of the ATPase are located at the negatively charged cavity. Three mutants in the negatively charged cavity region dramatically reduced GRP78 activity, further confirming the region as the site of ATPase activity.

Background: MicroRNAs (miRNAs) are small, noncoding RNAs that regulate gene expression post-transcriptionally. Dysregulation of miRNA has been implicated in the development and progression of prostate cancer. Through next generation miRNA sequencing, we recently identified a panel of five miRNAs associated with prostate cancer recurrence and metastasis. Of the five miRNAs, miR-301a had the strongest association with prostate cancer recurrence. Overexpression of miR-301a in prostate cancer cells, PC3, and LNCaP resulted in increased growth both in vitro and in xenografted tumors. We therefore sought to examine its role in prostate carcinogenesis in greater detail., Methods: We examined the effect of miR-301a expression on biochemical recurrence and metastasis among 585 men treated with radical prostatectomy for prostate cancer. We examined the mechanism of growth deregulation by miR-301a in prostate cancer cells using analysis of the miRome of prostate cancer cell lines, quantitative PCR, and Western blotting., Results: High levels of miR-301a (above the median) were associated with an increased risk of biochemical recurrence (adjusted hazard ratio [aHR] 1.42, 95% confidence interval (CI) 1.06-1.90, P = 0.002) but not of metastasis (aHR 0.84, 95%CI 0.41-1.70, P = 0.6) after adjustment for known prognostic factors. RNA transcriptome sequencing analysis of miR-301a overexpressing prostate cancer cell lines identified the tumor suppressor p63 as a potential direct miR-301a target. Transcriptome sequencing, qPCR and Western blotting showed that miR-301a induced epithelial-mesenchymal transition (EMT) in prostate cancer cells through a pathway initiated by p63 inhibition. Luciferase assay verified p63 as a direct target of miR-301a. Loss of p63 resulted in miR-205 downregulation, releasing Zeb1 and Zeb2 from inhibition, culminating in Zeb1/Zeb2 suppression of E-cadherin. This pathway of growth alteration mediated by miR-301a upregulation was shown to be valid in prostate cancer cell lines and patient-derived tumors., Conclusions: These data indicate that miR-301a functions as an oncogene in prostate cancer by directly targeting the p63 tumor suppressor leading to loss of E-cadherin and EMT. Hence, miR-301a may serve as a novel biomarker in prostate cancer as well as a therapeutic target for prostate cancer management. Prostate 76:869-884, 2016. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Deeper understanding of the anatomical intermediaries for disease and other complex genetic traits is essential to understanding mechanisms and developing new interventions. Existing ontology tools provide functional, curated annotations for many genes and can be used to develop mechanistic hypotheses; yet information about the spatial expression of genes may be equally useful in interpreting results and forming novel hypotheses for a trait. Therefore, we developed an approach for statistically testing the relationship between gene expression across the body and sets of candidate genes from across the genome. We validated this tool and tested its utility on three applications. First, we show that the expression of genes in associated loci from GWA studies implicates specific tissues for 57 out of 98 traits. Second, we tested the ability of the tool to identify novel relationships between gene expression and phenotypes. Specifically, we experimentally confirmed an underappreciated prediction highlighted by our tool: that white blood cell count--a quantitative trait of the immune system--is genetically modulated by genes expressed in the skin. Finally, using gene lists derived from exome sequencing data, we show that human genes under selective constraint are disproportionately expressed in nervous system tissues., (© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The small peptides representation of the original proteins are a valuable source of information that can be used as biomarkers involved in toxicity mechanism for chemical exposure. The aim of this study is to investigate serum peptide biomarkers of benzene exposure. C57BL/6 mice were enrolled into control group and benzene groups of 150 and 300 mg/kg/d Serum peptides were identified by mass spectrometry using an assisted laser desorption ionization/time of flight mass spectrometry (MS). Differential peptide spectra were obtained by tandem mass spectrometry and analyzed by searching the International Protein Index using the Sequest program. Forty-one peptide peaks were found in the range of 1000-10,000 Da molecular weight. Among them, seven peaks showed significantly different expression between exposure groups and control group. Two peptide peaks (1231.2 and 1241.8), which showed a two-fold increase in expression, were sequenced and confirmed as glucose 6-phosphate dehydrogenase (G6PD) and heat shock protein 90 Beta (HSP90 Beta), respectively. Furthermore, the expression of the two proteins in liver cells showed the same trend as in serum. In conclusion, G6PD and HSP90 beta might be the candidate serum biomarkers of benzene exposure. It also provided possible clues for the molecular mechanism of benzene-induced oxidative stress.

The Gax gene has been implicated in a variety of cell-developmental and biological processes, and aberrant Gax expression is linked to many diseases. In this study, to provide important insights for Gax-based gene therapy in vein graft restenosis and its anti-restenotic mechanism, we used rabbit vascular smooth muscle cells (VSMCs) to investigate the effects of Gax overexpression on proliferation, migration, cell cycle, and apoptosis in a serum-stimulated culture. Rabbit VSMC lines that stably overexpressed Gax were established by transfection with recombinant adenoviral vector Ad5-Gax. The effect of Gax overexpression on in vitro serum-induced VSMCs proliferation, migration, cell cycle, and apoptosis was assessed by MTT, wound healing, and flow cytometry assays, respectively. To investigate the effect of Gax overexpression on PCNA and MMP-2 in serum-induced VSMCs, immunocytochemistry, RT-PCR, and gelatin zymography were performed. The results clearly showed that Gax overexpression decreases PCNA expression in serum-induced VSMCs. Gax overexpression also significantly inhibited cell proliferation by blocking entry into the S-phase of the cell cycle, promoted cell apoptosis, and reduced cell migration activity by downregulating MMP-2 release and activity. These findings indicate that Gax would be an optimal target gene for gene therapy to treat vein graft restenosis.

Background: Abdominal aortic aneurysm (AAA) is a multi-factorial disease and its underlying pathogenesis remains poorly understood., Aim: We aim to search for the underlying etiology of AAA using whole exome sequencing and gene expression analysis., Materials and Methods: We performed whole exome sequencing for AAA and adjacent normal abdominal aorta tissue from one male AAA patient. Further gene expression analysis using downloaded dataset from the GEO database was also carried out to explore the underlying molecular mechanisms., Results: A total of 5.97 Gb clean data were generated for the two samples, achieving a mean depth of coverage of 31.96 and 32.88 for the AAA and normal samples, respectively. We identified 203 somatic variants and confirmed 34 protein-altering somatic mutations in 25 genes. Among the confirmed variants, 11 mutations were not reported in the dbSNP database before. According to the literature review, none of these 25 genes were reportedly associated with AAA., Conclusions: Our findings here may provide potential targets for effective prevention of human AAA development and progression.

Thrombolytic therapy using tissue plasminogen activator (TPA) is an effective method for treating acute myocardial infarction. However, the systemic administration of TPA is associated with the risk of hemorrhage. Mesenchymal stem cells (MSCs) from bone marrow are characterized by low immunogenicity and homing toward damaged tissues and are therefore ideal cell carriers to achieve lesion-targeting medication. In this article, TPA gene was integrated into the AAVS1 of mesenchymal stem cells, which has been confirmed to be a safe chromosomal locus. The targeting efficiency was 83%. The clones with the site-specific integration retained the stem cell traits of MSCs, displayed a normal karyotype and could persistently and effectively express TPA, as demonstrated by an average expression activity of 1.5 units/mL (3.4-fold that of the control group). After subculture and subsequent growth for two weeks, the clones showed an average TPA activity of 1.43 units/mL and exhibited no significant differences among the individual clones. In summary, the foreign TPA gene can be specifically introduced to the AAVS1 locus, whereby it can be stably and effectively expressed. MSCs can serve as cell carriers for the targeted treatment of a thrombus using TPA., (Crown Copyright © 2013. Published by Elsevier Inc. All rights reserved.)

Although it has been known for many years that T. spiralis muscle larvae (ML) can not invade intestinal epithelial cells unless they are exposed to the intestinal milieu and activated into intestinal infective larvae (IIL), which genes in IIL are involved in the process of invasion is still unknown. In this study, suppression subtractive hybridization (SSH) was performed to identify differentially expressed genes between IIL and ML. SSH library was constructed using cDNA generated from IIL as the 'tester'. About 110 positive clones were randomly selected from the library and sequenced, of which 33 T. spiralis genes were identified. Thirty encoded proteins were annotated according to Gene Ontology Annotation in terms of molecular function, biological process, and cellular localization. Out of 30 annotated proteins, 16 proteins (53.3%) had binding activity and 12 proteins (40.0%) had catalytic activity. The results of real-time PCR showed that the expression of nine genes (Ts7, Ndr family protein; Ts8, serine/threonine-protein kinase polo; Ts11, proteasome subunit beta type-7; Ts17, nudix hydrolase; Ts19, ovochymase-1; Ts22, fibronectin type III domain protein; Ts23, muscle cell intermediate filament protein OV71; Ts26, neutral and basic amino acid transport protein rBAT and Ts33, FACT complex subunit SPT16) from 33 T. spiralis genes in IIL were up-regulated compared with that of ML. The present study provide a group of the potential invasion-related candidate genes and will be helpful for further studies of mechanisms by which T. spiralis infective larvae recognize and invade the intestinal epithelial cells.

Aire plays an important role in central immune tolerance by regulating the transcription of thousands of genes. However, the role of Aire in the peripheral immune system is poorly understood. Regulatory T (Treg) cells are considered essential for the maintenance of peripheral tolerance, but the effect of Aire on Treg cells in the peripheral immune system is currently unknown. In this study, we investigated the effects of macrophages overexpressing Aire on CD4+Foxp3+ Treg cells by co-culturing Aire-overexpressing RAW264.7 cells or their supernatant with splenocytes. The results show that macrophages overexpressing Aire enhanced the expression of Foxp3 mRNA and induced different subsets of Treg cells in splenocytes through cell-cell contact or a co-culture supernatants. TGF-β is a key molecule in the increases of CD4+CD45RA+Foxp3hi T cell and activating Treg (aTreg) levels observed following cell‑supernatant co-culturing. Subsets of Treg cells were induced by Aire-overexpressing macrophages, and the manipulation of Treg cells by the targeting of Aire may provide a method for the treatment of inflammatory or autoimmune diseases.

In previous studies, HSPC016 was identified as a differentially expressed gene in dermal papilla cells (DPC) with aggregative behaviour. To clarify its role in the regulation of DPC, the recombinant HSPC016 protein was expressed using the Pichia pastoris expression system, and three small interfering ribonucleic acid duplexes (siRNA) were designed and transfected into DPC. Results showed that DPC with the HSPC016 gene inhibited, exhibit non-aggregative behaviour and grow much slower than normal DPC, and that the recombinant HSPC016 protein could promote the proliferation of high-passage DPC and induce its aggregative behaviour., (© 2010 The Authors. Australasian Journal of Dermatology © 2010 The Australasian College of Dermatologists.)

Genetic transformation is useful for basic research and applied biotechnology. However, genetic transformation of microalgae is usually quite difficult due to the technical limitations of existing methods. We cloned the promoter and terminator of the nitrate reductase gene from the microalga Phaeodactylum tricornutum and used them for optimization of a transformation system of the microalga Chlorella vulgaris. This species has been used for food production and is a promising candidate as a bioreactor for large-scale production of value-added proteins. A construct was made containing the CAT (chloramphenicol acetyltransferase) reporter gene driven by the nitrate reductase promoter. This construct was transferred into the C. vulgaris genome by electroporation. Expression of CAT in transgenic Chlorella conferred resistance to the antibiotic chloramphenicol and enabled growth in selective media. Overall efficiency for the transformation was estimated to be approximately 0.03%, which is relatively high compared with other available Chlorella transformation systems. Expression of CAT was induced in the presence of nitrate and inhibited in the presence of ammonium as a sole nitrogen source. This study presented an inducible recombinant gene expression system, also providing more gene regulation elements with potential for biotechnological applications.

Four non-magnetic mutants of Magnetospirillum gryphiswaldense strain MSR-1 were isolated by transposon mutagenesis and found to contain interruption of either the mamO or mamE gene within the mamAB operon. Studies indicated that mamO and mamE genes are essential for magnetosome crystal biomineralization in MSR-1. This is the first report of a single gene (mamO or mamE) whose mutation affects crystal biomineralization in MSR-1., (Copyright © 2010 Elsevier Masson SAS. All rights reserved.)

High mammalian gene expression was obtained for more than twenty different proteins in different cell types by just a few laboratory scale stable gene transfections for each protein. The stable expression vectors were constructed by inserting a naturally-occurring 1.006 kb or a synthetic 0.733 kb DNA fragment (including intron) of extremely GC-rich at the 5' or/and 3' flanking regions of these protein genes or their gene promoters. This experiment is the first experimental evidence showing that a non-coding extremely GC-rich DNA fragment is a super "chromatin opening element" and plays an important role in mammalian gene expression. This experiment has further indicated that chromatin-based regulation of mammalian gene expression is at least partially embedded in DNA primary structure, namely DNA GC-content.

Avian pathogenic Escherichia coli (APEC) and uropathogenic E. coli (UPEC) establish infections in extraintestinal habitats of different hosts. As the diversity, epidemiological sources and evolutionary origins of extraintestinal pathogenic E. coli (ExPEC) are so far only partially defined, in the present study,100 APEC isolates and 202 UPEC isolates were compared by their content of virulence genes and phylogenetic groups. The two groups showed substantial overlap in terms of their serogroups, phylogenetic groups and virulence genotypes, including their possession of certain genes associated with large transmissible plasmids of APEC. In a chicken challenge model, both UPEC U17 and APEC E058 had similar LD(50), demonstrating that UPEC U17 had the potential to cause significant disease in poultry. To gain further information about the similarities between UPEC and APEC, the in vivo expression of 152 specific genes of UPEC U17 and APEC E058 in both a murine urinary tract infection (UTI) model and a chicken challenge model was compared with that of these strains grown statically to exponential phase in rich medium. It was found that in the same model (murine UTI or chicken challenge), various genes of UPEC U17 and APEC E058 showed a similar tendency of expression. Several iron-related genes were upregulated in the UTI model and/or chicken challenge model, indicating that iron acquisition is important for E. coli to survive in blood or the urinary tract. Based on these results, the potential for APEC to act as human UPEC or as a reservoir of virulence genes for UPEC should be considered. Further, this study compared the transcriptional profile of virulence genes among APEC and UPEC in vivo.

We consider the problem of predicting alternative splicing patterns from a set of expressed sequences (cDNAs and ESTs). Some of these expressed sequences may be errorous, thus forming incorrect exons/introns. These incorrect exons/introns may cause a lot of false positives. For example, we examined a popular alternative splicing database, ECgene, which predicts alternate splicing patterns from expressed sequences. The result shows that about 81.3%-81.6% (sensitivity) of known patterns are found, but the specificity can be as low as 5.9%. Based on the idea that errorous sequences are usually not consistent with other sequences, in this paper we provide an alternative approach for finding alternative splicing patterns which ensures that individual exons/introns of the reported patterns have enough support from the expressed sequences. On the same dataset, our approach can achieve a much higher specificity and a slight increase in sensitivity (38.9% and 84.9%, respectively). Our approach also gives better results compared with popular alternative splicing databases (ASD, ECgene, SpliceNest) and the software ClusterMerge.

To determine whether the neuronal activity of the spindle cord in muscle spindle afferent pathways is altered after a period of hindlimb unloading (Hu), and after recovery, we focused on c-Fos-immunoreactivity in the spinal cord evoked by excitation of the muscle spindle in normal gravity and after 3, 7, and 14 days of Hu, and after 14 days of Hu with an additional 2, 5, and 9 days of unrestricted cage activity. High frequency sinusoidal vibration (HFV) applied to the Achilles tendon was used for activation of the muscle spindle. Results showed that c-Fos-immunoreactive neurons evoked by HFV were mainly concentrated in lamina IV-VII of the ipsilateral spinal cord. After 14 days of Hu, the total number of labeled neurons in the spinal cord was significantly increased (P<0.05). The number of c-Fos-immunoreactive neurons in lamina IV and VI-VII were also significantly higher than that in normal rats (P<0.05). After 9 days of reloading, the total number of c-Fos-immunoreactive neurons and the pattern of their lamellar distribution were both recovered to pre-experimental levels. Our findings suggest that simulated weightlessness induces a large increase in neuronal excitation of the spinal cord in muscle spindle afferent pathways. Weight bearing immediately after Hu could induce completed recovery of excitability of the spinal cord in muscle spindle afferent pathways.

Although cure rate of childhood acute lymphoblastic leukemia (ALL) has surpassed 80%, drug resistance remains a major cause of treatment failure. We previously identified a panel of 33 genes differentially expressed in prednisolone sensitive versus resistant ALL cells from newly diagnosed children. Here we used bioinformatics to identify resistance genes most likely to contain single nucleotide polymorphisms (SNPs) in their promoter region. The highest priority gene was SMARCB1, a core member of the SWI/SNF complex which promotes glucocorticoid effects through nucleosome remodeling. We identified several SNPs in the SMARCB1 promoter in lymphoblastoid cells from 90 individuals in the Centre d'Etude du Polymorphisme Humain (CEPH) panel. Among these SNPs, the -228G>T SNP (allele frequency 9.4%) was the only one that significantly increased reporter activity in human ALL cell lines. Furthermore, we identified nuclear protein poly (ADP-ribose) polymerase family, member 1 (PARP1) as a nuclear protein binding to the SMARCB1 promoter and showed that the -228 SNP significantly altered PARP1 binding affinity. The -228G>T SNP altered SMARCB1 mRNA and protein levels and a positive association was found between the SMARCB1 mRNA level and both the -228 genotype and prednisolone sensitivity in CEPH cell lines. Finally, knockdown experiments performed in human ALL cell lines confirmed that lower SMARCB1 expression increased prednisolone resistance. In summary, we provide functional evidence that SMARCB1 is involved in prednisolone resistance and identified a promoter SNP that alters the level of SMARCB1 mRNA and protein expression and the binding of PARP1 to the SMARCB1 promoter.

Mouse genetic models that probe important pathways in intestinal cell maturation, such as cell-cycle regulation, apoptosis, and, especially, lineage specific differentiation, have provided profound insight into the underlying mechanisms of intestinal tumor formation and progression. However, a wealth of epidemiological and experimental data indicates that environment, especially the diet, is a principal determinant of relative risk for tumor development. We have demonstrated that even in mouse models in which tumor incidence is strongly initiated by genetic manipulation of genes, such as Apc, p21(WAF1/cip1), and p27(Kip1), a Western-style diet that is high in fat and low in calcium and vitamin D can dramatically increase and accelerate tumor formation. Moreover, experiments show that modulation of calcium and vitamin D levels can substantially influence tumor formation in both the mouse genetic models, as well as in a new dietary model that appears to mimic the development of sporadic colon cancer. Finally, analysis of gene expression profiles provides important insights into how diets may alter metabolic profiles and regulatory pathways that influence probability of tumor formation in the histologically and physiologically normal intestinal mucosa.

Background: Childhood acute lymphoblastic leukemia (ALL) is curable with chemotherapy in approximately 80 percent of patients. However, the cause of treatment failure in the remaining 20 percent of patients is largely unknown., Methods: We tested leukemia cells from 173 children for sensitivity in vitro to prednisolone, vincristine, asparaginase, and daunorubicin. The cells were then subjected to an assessment of gene expression with the use of 14,500 probe sets to identify differentially expressed genes in drug-sensitive and drug-resistant ALL. Gene-expression patterns that differed according to sensitivity or resistance to the four drugs were compared with treatment outcome in the original 173 patients and an independent cohort of 98 children treated with the same drugs at another institution., Results: We identified sets of differentially expressed genes in B-lineage ALL that were sensitive or resistant to prednisolone (33 genes), vincristine (40 genes), asparaginase (35 genes), or daunorubicin (20 genes). A combined gene-expression score of resistance to the four drugs, as compared with sensitivity to the four, was significantly and independently related to treatment outcome in a multivariate analysis (hazard ratio for relapse, 3.0; P=0.027). Results were confirmed in an independent population of patients treated with the same medications (hazard ratio for relapse, 11.85; P=0.019). Of the 124 genes identified, 121 have not previously been associated with resistance to the four drugs we tested., Conclusions: Differential expression of a relatively small number of genes is associated with drug resistance and treatment outcome in childhood ALL., (Copyright 2004 Massachusetts Medical Society)

Jellyfish, Cyanea capillata, has an important position in head patterning and ion channel evolution, in addition to containing a rich source of toxins. In the present study, 2153 expressed sequence tags (ESTs) from the tentacle cDNA library of C. capillata were analyzed. The initial ESTs consisted of 198 clusters and 818 singletons, which revealed approximately 1016 unique genes in the data set. Among these sequences, we identified several genes related to head and foot patterning, voltage-dependent anion channel gene and genes related to biological activities of venom. Five kinds of proteinase inhibitor genes were found in jellyfish for the first time, and some of them were highly expressed with unknown functions.

A series of fluorescent diboronic acids was synthesized in nine steps as potential sensors for sialyl Lewis X (sLex). The fluorescent binding studies of these sensors with sLex were carried out in a mixed aqueous solution. Compound 7e was found to show the strongest fluorescence enhancement upon binding with sLex. Using cell cultures, 7e was shown to label sLex-expressing HEPG2 cells at 1 microM, while non-sLex-expressing cells were not labeled.

After the pig platelet-activating factor receptor (PAFr) gene was cloned and sequenced, the chromosomal location of this gene was studied using a pig/rodent somatic cell hybrid panel containing 27 cell lines. The results indicated that the pig PAFr gene is located on SSC6q22-23. Platelet-activating factor (PAF, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) is thought to be very important in the animal reproductive processes. Its function is mediated through a membrane-bound receptor. Pig PAFr mRNA distribution in different tissues was tested using reverse transcription and PCR (RT-PCR) reactions. All tissues examined expressed PAFr. Using a pig PAFr gene DNA competitor, PAFr expression was quantificated. The pig PAFr mRNA expression level was estimated to be from 1 x 10(2) to 1.2 x 10(4) copies of complementary DNA (cDNA) per 50 ng of total RNA. The highest level was found in lung, and the lowest in the skeletal muscle. These results demonstrated that PAFr was differentially expressed in pig tissues.

Background: Human macrophage metalloelastase (HME), also referred as matrix metalloproteinase 12, has been shown to convert plasminogen into angiostatin, an essential inhibitor of tumor angiogenesis. The current study was designed to investigate the association of HME mRNA expression with disease progression of in patients with colorectal carcinoma., Methods: The expression of HME mRNA was quantified by Northern blot analysis in tumorous (T) and nontumorous (N) tissues obtained from 54 patients with primary colorectal carcinoma, and the ratio of these two tissue types was defined as the HME T/N ratio. The prognostic significance of this ratio after surgery, along with its correlation with disease progression and metastatic potential, was evaluated. The tissues also were subjected to in situ hybridization analysis for HME. The microvessel count after immunohistochemical staining of factor VIII was used to assess angiogenesis., Results: The HME T/N ratio showed an inverse correlation with the depth of the intestinal wall invasion, the lymphatic invasion, and the vascular invasion. The patients with overexpression of HME mRNA (HME T/N ratio > 1.191) significantly demonstrated better survival compared with those patients who did not show overexpression of HME mRNA. In situ hybridization identified the colorectal carcinoma cells as mainly responsible for the signal shown in Northern blot analysis. A considerable but not statistically significant lower microvessel density (MVD) was observed in the patients with HME overexpression., Conclusions: These findings demonstrate that the HME gene plays an important role in the inhibition of tumor progression in patients with colorectal carcinoma and that its overexpression is correlated closely with a better prognosis., (Copyright 2001 American Cancer Society.)

Objective: To express recombinant human platelet factor 4 (PF4) in methanol yeast., Methods: PF4 cDNA amplified by PCR was cloned into the mating factor alpha (MFalpha) signal sequence, downstream the alcohol oxidase 1 (AOX 1) promoter of the pPIC9 vector. The reconstructed vector was subsequently integrated into the methanol yeast P. pastoris strain GS115. The highly expressing strain was selected and the protein sequences and their biological activities were assayed., Results: The amino acid sequence of rhPF4 was the same as that of natural one. The rhPF4 can neutralize the anticoagulation effect of heparin in a dose-dependent manner., Conclusion: The amount of rhPF4 expressed in methanol yeast meets well the scale for manufacturing. The biological activities of the rhPF4 were the same as that of natural PF4.

The cyclin D1 (PRAD1) oncogene is rearranged with the PTH gene and is transcriptionally activated in a subset of parathyroid adenomas. Because of heterogeneity in rearrangement breakpoints, the true percentage of adenomas with cyclin D1 deregulation is unknown. Overexpression of the cyclin D1 protein in parathyroid adenomas appears to be a unifying consequence of all cyclin D1 gene rearrangements and can, therefore, be examined to more comprehensively identify adenomas in which cyclin D1 is pathogenetically important. We studied cyclin D1 expression in 65 parathyroid adenomas (from 64 patients), 51 normal parathyroid glands (from the same patients), and 4 parathyroid carcinoma specimens (from 3 patients) using a microwave-enhanced immunohistochemical method and affinity-purified cyclin D1 polyclonal antiserum. When available, data on adenoma mass, intact PTH level, and concurrent serum calcium level were also collected. Twelve of the 65 adenomas (18%) showed diffuse nuclear staining of approximately 30-70% of the tumor cells. All 51 normal glands were negative, except 1 gland that showed scattered cells ( < 10%) with positive nuclear staining. In addition, scattered positive cells were seen in the compressed rim of histologically normal parathyroid tissue surrounding 2 adenomas that were cyclin D1 negative. No significant differences in adenoma mass, intact PTH levels, or concurrent calcium levels were found between positive and negative tumors. Two of 4 parathyroid carcinoma specimens from 2 of 3 patients showed strong nuclear staining for cyclin D1. Overexpression of the cyclin D1 oncogene in 18% of our cases, due to the cyclin D1/PTH translocation and/or other mechanisms, suggests that overexpressed cyclin D1 plays a role in the pathogenesis of a much larger proportion of parathyroid adenomas than previously appreciated. Cyclin D1 overexpression is a feature of typical parathyroid adenomas and is not confined to unusually large, symptom-causing adenomas as had been suggested by early DNA studies. Although only three patients with parathyroid carcinoma were studied, two of the patients' tumors stained for cyclin D1, raising the possibility that the frequency of cyclin D1 overexpression may be even greater in carcinomas. Cyclin D1 overexpression appears to highlight a central pathway in parathyroid neoplasia.

The inhibitory effect of antisense phosphorothioate oligodeoxyribonucleotides (S-oligos) on the production of hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) was studied using Hep G2 cells transfected with hepatitis B virus (HBV). Synthetic antisense oligos (15-mers) directed against the cap site of mRNA transcribed from the SP II promoter and regions of the translational initiation site of the S gene showed a sequence-specific, dose-dependent inhibitory effect on HBV gene expression between concentrations of 1.0 microM-5.0 microM. Oligos directed against the middle of the S gene had little effect on HBsAg and HBeAg expression, as did non-complementary random-sequence control. The cells remained viable throughout the experiments and no morphological abnormalities were observed with antisense S-oligos at concentrations below 20.0 microM. These results suggest a therapeutic potential for antisense oligonucleotides in the treatment of patients who are chronically infected with HBV.

Asthma is a respiratory disease with a clinically high incidence, and repeated attacks of asthma severely affect the quality of life and even pose a threat to health, leading to severe burdens on families and even the society. A thorough understanding of the pathogenesis of asthma is essential for the prevention and treatment of asthma. This study aimed to examine the effect of the microRNA miR-27a on asthma and its relationship with mitogen activated protein kinase 4 (MAP2K4). Patients with asthma admitted to our hospital from August 2016 to August 2018 and healthy participants in the same period were included in this prospective analysis. The mRNA expression levels of miR-27a and MAP2K4 in peripheral blood were determined. Airway smooth muscle cells (ASMCs) were used to study the effects of miR-27a and MAP2K4 on cell biological behavior. The relationship between miR-27a and MAP2K4 was verified using dual-luciferase reporter assay. miR-27a expression was increased and MAP2K4 mRNA expression was decreased in asthma (P < 0.05). Increasing miR-27a expression and inhibiting MAP2K4 expression could enhance the activity of ASMCs, whereas inhibiting miR-27a expression and increasing MAP2K4 expression had the opposite effect (P < 0.05). Dual-luciferase reporter assay results showed that the fluorescence activity of MAP2K4-wild type was inhibited by increased miR-27a expression (P < 0.05). miR-27a promotes the proliferation and invasion of ASMCs by targeting MAP2K4 and is involved in the occurrence of asthma. [ABSTRACT FROM AUTHOR]

Summary: Egg‐type ducks and meat‐type ducks are predominantly commercial or indigenous and have been subjected to artificial directional selection. These two duck types differ substantially in body shape, production performance and reproductivity. However, the genetic changes associated with phenotypic differences remain unclear. Here, we compared the two duck types at the genomic and transcriptomic levels. We identified a large number of SNPs and genes in genomic divergent regions in terms of FST and θπ values. The corresponding genes were mainly enriched in embryonic development function and metabolic pathway. RNA‐seq analysis also revealed differential gene expression in the liver and gonads. The differentially expressed genes were functionally associated with signal transmission and substance metabolism respectively. Furthermore, we found that seven genes were related to differentiation between the two types by both g genome and transcriptome analysis and were plausible candidate genes. These genes were annotated to GO categories of cell development and disease immunity. These findings will enable a better understanding of the artificial selection history of meat and egg ducks and provide a valuable resource for future research on the breeding of these two lineages. [ABSTRACT FROM AUTHOR]

Immunotherapy has been found to be efficient in a variety of cancers and, therefore, may be a promising strategy for breast cancer (BC), particularly due to the limited therapeutic options currently available for triple-negative BC (TNBC). However, heterogeneity of tumor mutation burden (TMB), immune gene expression and mismatch repair (MMR) gene activity across BC subtypes has not been well characterized. In the present study, a comprehensive analysis of TMB, expression levels of immune cell type marker genes, and expression levels of MMR-associated genes was performed. A total of 5 MMR-associated genes, including MLH1, MLH3, MSH2, MSH6 and PMS2, were analyzed. Patients that harbored at least two MMR genes with expression levels in the lower 10% percentile across the cohort were considered as potentially aberrant (lost expression). Hormone receptor (HR)-negative BC is associated with a higher TMB and immune gene expression compared with HR-positive BC [TMB, estrogen receptor (ER)-negative vs. ER-negative, 55 vs. 32, respectively; P=4.1×10−13; progesterone receptor (PR)-negative vs. PR-positive, 53 vs. 31, respectively; P<2.2×10−16]. By contrast, human epidermal growth factor receptor 2 (HER2)-negative BC tended to have a lower TMB and decreased immune gene expression compared with HER2-positive BC (TMB, HER2-negative vs. HER2-positive, 36 vs. 48, respectively; P=0.02). Furthermore, aberrant expression of MMR genes was found to be more common in HR-negative compared with HR-positive BC (P<0.001). Significantly higher expression levels of each immune marker gene of four major immune cell types were found in patients who were HR-negative compared with patients who were HR-positive (P<0.001). The findings of the present study suggest that HR-negative or HER2-positive BC exhibits elevated TMB and immunogenic activity, and immunotherapeutic options are recommended. [ABSTRACT FROM AUTHOR]

Abstract: Aims: Available bacterins, commercial or autogenous, for Actinobacillus pleuropneumoniae disease control have, thus far, shown debatable protective efficacy and only in homologous challenges. Our study sought to determine whether the addition of reombinant protein ApxIV to the multicomponent vaccine could enhance protection against homologous and heterologous challenge of A. pleuropneumoniae. Methods and Results: The virulence of ApxI, ApxII, ApxIV and OMP were cloned and expressed using a prokaryotic system; these recombinant proteins were combined with inactivated A. pleuropneumoniae serovar 1 to formulate different multicomponent vaccines. Immune response and protective efficacy of the vaccines were evaluated in mice and pigs. A protection rate of 67% was observed against heterologous challenge in mice vaccinated with the rApxIV formulation. Piglets vaccinated with vaccine containing ApxIV produced significantly higher antibody titre and provided complete protection and reduced gross lesions by 67% when compared with the nonimmunized group after homologous challenge. Additionally, flow cytometry analysis showed significant cellular immune response. Conclusions: The results of our vaccination experiments revealed that a combination of inactivated bacteria and the recombinant antigens rApxI, rApxII, rApxIV and rOMP can provide effective protection against heterologous A. pleuropneumoniae challenge. Significance and Impact of the Study: The addition of ApxIV to the multicomponent vaccine could enhance homologous and heterologous protection in mice and pigs, respectively, against challenge by A. pleuropneumoniae. [ABSTRACT FROM AUTHOR]

Abstract: The genetic improvement of redlip mullet Liza haematocheila through breeding programmes is of interest for this important aquaculture species. Lipoprotein lipase (LPL) is a key enzyme in lipid deposition and metabolism. The purpose of this study was to investigate the nutritional regulation of LPL in redlip mullet. We cloned and identified the LPL gene, determined LPL gene expression in various tissues, and examined the effect of dietary lipid level on hepatic LPL gene expression. The LPL gene of redlip mullet Liza haematocheila (L.hLPL) was 2,395 bp in length and encoded 516 amino acids. Sequence analysis showed that L.hLPL shared 61%–90.3% identity with LPLs in other species. Expression patterns of hepatic L.hLPL were studied in redlip mullet fed diets containing 2.0, 4.8, 7.5, 9.8, 12.0 or 14.6 g/kg, crude fat for 60 days by real‐time quantitative PCR. The abundance of LPL mRNA in hepatic tissue increased with the increase in dietary fat. The expression L.hLPL mRNA was significantly higher in the groups fed diets with 14.6 and 12.0 g/kg fat than in the other groups (p < .05). Gene expression was significantly higher in the abdominal fat of redlip mullet (p < .05) compared with other tissues. In conclusion, a high fat diet (9.79–14.59 g/kg) induces L.hLPL expression in abdominal fat. [ABSTRACT FROM AUTHOR]

The present study evaluated the effects of isovalerate supplementation on the development of the small intestinal mucosa in dairy calves. Forty-eight Chinese Holstein bull calves at 15 days of age and 45.1 ± 0.36 kg of body weight were assigned randomly to four groups. The treatments were control, low-isovalerate, moderate-isovalerate and high-isovalerate with 0, 3, 6 and 9 g isovalerate per calf per day, respectively. The study comprised 75 days with a 15-day adaptation period followed by a 60-day sampling period. Calves were weaned at 60 days of age. Six calves were chosen from each treatment at random and slaughtered at 30 and 90 days of age. The small intestine morphology and activities of amylase and trypsin improved significantly with increasing age. No interaction between treatments and age was observed. The small intestine length, mucosa layer thickness, villus height and crypt depth increased linearly with increasing isovalerate supplementation. However, the ratio of villus height to crypt depth was not affected by treatment. Activities of amylase and trypsin increased linearly. The lactase activity increased linearly during the 75-day period and for pre-weaned calves but was unaltered for post-weaned calves. The relative mRNA expressions of growth hormone receptor, insulin-like growth factor-1 receptor and sodium-glucose co-transporter-1 in the small intestine mucosa increased linearly, and a similar pattern was observed for the expression of peptide transporter-1 in the duodenum and proximal jejunum. The results suggested that small intestine development was promoted by isovalerate in a dose-dependent manner. [ABSTRACT FROM AUTHOR]

Our present work showed that the expression of genes encoding PTOX (terminal oxidase of chlororespiration) and PGR5 (one essential component of cyclic electron transfer) were stimulated by red and blue light, but the stimulation under red light was soon reversed by subsequent far-red light. The expression levels of PTOX and PGR5 under simulated light quality conditions in line with maize-soybean relay strip intercropping (SRI) were obviously lower than those under simulated soybean monocropping (SM), since the lower red:far-red ratio under SRI. Measurements on photosynthetic and chlorophyll fluorescence parameters suggested a decline of assimilatory power supply and a lower nonphotochemical quenching under SRI as compared to SM. In this case, weaker PGR-dependent cyclic electron transfer and chlororespiration under SRI, suggested by lower expression levels of PGR5 and PTOX, could be considered as means of reducing excitation energy dissipation to allocate more power toward CO assimilation. [ABSTRACT FROM AUTHOR]