MiR-125b and SATB1-AS1 might be shear stress-mediated therapeutic targets.

The aim of the study was to explore the potential molecular mechanism associated with shear stress on abdominal aortic aneurysm (AAA) progression. This study performed RNA sequencing on AAA patients (SQ), AAA patients after endovascular aneurysm repair (EVAR, SH), and normal controls (NC). Furthermore, we identified the differentially expressed microRNAs (miRNAs), long noncoding RNAs (lncRNAs), and circular RNA (cirRNAs) and constructed competing endogenous RNA (ceRNA) networks. Finally, 164 differentially expressed miRNAs, 179 co-differentially expressed lncRNAs, and 440 co-differentially expressed circRNAs among the three groups were obtained. The differentially expressed miRNAs mainly enriched in 325 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. Target genes associated with co-differentially expressed genes among the group of SH, SQ, and NC mainly enriched in 66 KEGG pathways. LncRNA-miRNA-mRNA interactions, including 15 lncRNAs, 63 miRNAs and 57 mRNAs, was constructed. CircRNA-miRNA-mRNA ceRNA network included 79 circRNAs, 21 miRNAs, and 49 mRNAs. Among them, KLRC2 and CSTF1, targeted by miR-125b, participated in cell-mediated immunity regulation. MiR-320-related circRNAs and SATB1-AS1 serving as the sponge of miRNAs, such as has-circ-0129245, has-circ-0138746, and has-circ-0139786, were hub genes in ceRNA network. In conclusion, AAA patients might be benefit from EVAR based on various pathways and some molecules, such as miR-125b and SATB1-AS1, related with shear stress.

lncRNA ANRIL accelerates wound healing in diabetic foot ulcers via modulating HIF1A/VEGFA signaling through interacting with FUS.

BACKGROUND: Diabetic foot ulcer (DFU) is a frequently diagnosed complication of diabetes, and remains a heathcare burden worldwide. However, the pathogenesis of DFU is still largely unclear. The objective of this study is to delineate the function and underlying mechanism of lncRNA antisense non coding RNA in the INK4 locus (ANRIL) in endothelial progenitor cells (EPCs) and DFU mice. METHODS: The DFU mouse model was established, and EPCs were subjected to high glucose (HG) treatment to mimic diabetes. qRT-PCR or western blot was employed to detected the expression of ANRIL, HIF1A, FUS and VEGFA. CCK-8 and Annexin V/PI staining were used to monitor cell proliferation and apoptosis. Wound healing, Transwell invasion and tube formation assays were conducted to assess cell migration, invasion and angiogenesis, respectively. The association between ANRIL and FUS was verified by RNA pull-down and RIP assays. Luciferase and ChIP assays were employed to investigate HIF1A-mediated transcriptional regulation of VEGFA and ANRIL. The histological alterations of DFU wound healing were observed by H&E and Masson staining. RESULTS: ANRIL was downregulated in peripheral blood samples of DFU patients, DFU mice and HG-treated EPCs. Mechanistically, ANRIL regulated HIFA mRNA stability via recruiting FUS. VEGFA and ANRIL were transcriptionally regulated by HIF1A. Functional experiments revealed that HG suppressed EPC proliferation, migration, invasion and tube formation, but promoted apoptosis via ANRIL/HIF1A axis. ANRIL accelerated DFU wound healing via modulating HIF1A expression in vivo. CONCLUSION: ANRIL accelerated wound healing in DFU via modulating HIF1A/VEGFA signaling in a FUS-dependent manner.

Enhanced Enzymatic Hydrolysis of Cellulose From Substrate and Indole-3-Acetic Acid Content-During the Fruiting Body Differentiation Stage by Sodium Acetate Addition.

Volvariella volvacea, with high commercial, nutritional and medicinal value, is widely cultivated in tropical and subtropical regions. The effects of supplementation on mushroom yield has been studied. We showed that the optimal application of sodium acetate (NaAc) was spray application of a 0.08% concentration during the substrate mixing stage which could increase yields by up to 89.16% and enhance the enzymatic hydrolysis of cellulose and hemicellulose from the substrate. For most enzymes tested maximum activity occurred during the fruiting body growth and development stage, which led to degradation of the substrate, increasing the available nutrients for mycelial propagation and fruiting body growth and development. Meanwhile, NaAc also significantly increased the indole-3-acetic acid (IAA) content in the early fruiting body development stage of V. volvacea, It was observed that IAA promotes not only plant primordium differentiation; but also the primordium differentiation of edible fungi. Furthermore, treatments with three acetate salts had an increase of yield by 30.22% on average. The mechanisms by which NaAc application may improve the yield of V. volvacea are discussed.

Identification of Cyt2Ba from a New Strain of Bacillus thuringiensis and Its Toxicity in Bradysia difformis.

Bradysia difformis is one of the most damaging pests in mushroom production in China. In this study, eight Bacillus thuringiensis strains were analyzed for insecticidal activity in B. difformis. The strain JW-1 showed the highest insecticidal activity against B. difformis larvae, but did not inhibit the mycelial growth of Pleurotus ostreatus and P. geesteranus. The 16S rRNA gene (1397 bp) and cyt2 gene (792 bp) were obtained from strain JW-1. The phylogenetic tree based on 16S rRNA gene and Cyt2 toxin showed that strain JW-1 was a member of B. thuringiensis and Cyt2 toxin belonged to Cyt2Ba toxin cluster. The Cyt2Ba toxin from strain JW-1 was overexpressed in E. coli as a fusion protein and the fusion protein (70 kDa) was purified by Ni-IDA affinity chromatography. The purified Cyt2Ba fusion protein was toxic to B. difformis larvae (LC50 was 2.25 ng/mL). The identification of Cyt2Ba from strain JW-1 and confirmation of the insecticidal activity of Cyt2Ba in B. difformis provided a new means of biological control of the important pest in mushroom production.

Dual Metabolomic Platforms Identified a Novel Urinary Metabolite Signature for Hepatitis B Virus-Infected Patients with Depression.

OBJECTIVE: Depression could make the treatment outcome worse. However, up to now, no objective methods were developed to diagnose depression in hepatitis B virus (HBV)-infected patients. Therefore, the dual metabolomic platforms were used here to identify potential biomarkers for diagnosing HBV-infected patients with depression (dHB). METHODS: Both gas chromatography-mass spectrometry-based and nuclear magnetic resonance-based metabolomic platforms were used to conduct urine metabolic profiling of dHB subjects and HBV-infected patients without depression (HB). Orthogonal partial least-squares discriminant analysis was used to identify the differential metabolites between dHB subjects and HB subjects, and the step-wise logistic regression analysis was used to identify potential biomarkers. RESULTS: In total, 21 important metabolites responsible for distinguishing dHB subjects from HB subjects were identified. Meanwhile, seven potential biomarkers (α-ydroxyisobutyric acid, hippuric acid, azelaic acid, isobutyric acid, malonic acid, levulinic acid, and phenylacetylglycine) were viewed as potential biomarkers. The simplified biomarker panel consisting of these seven metabolites had an excellent diagnostic performance in discriminating dHB subjects from HB subjects. Moreover, this panel could yield a higher accuracy in separating dHB subjects from HB subjects than our previous panels (identified by single metabolomic platform) did. CONCLUSION: These results suggested that the dual metabolomic platforms could yield a better urinary biomarker panel for dHB subjects than any single metabolomic platform did, and our results could be helpful for developing an objective method in future to diagnose depression in HBV-infected patients.

[Mechanism research of curcumin on cancer cells based on cell metabolic profiling].

In this study, gas chromatography coupled with mass spectrometry(GC-MS) was used to analyze the changes of 12 kinds of cancer cells treated by curcumin. The related differential metabolites were screened and the metabolic pathways were analyzed to explore the anti-tumor mechanism of curcumin. Methyl thiazol tetrazolium(MTT) assay was used to detect the 50% inhibiting concentration(IC_(50)) of curcumin on 12 human tumor cells. After treatment with curcumin for 48 h, the cells were collected and analyzed by GC-MS, followed by pathway analysis and multivariate data analysis including principal component analysis(PCA), orthogonal partial least squares discriminant analysis(OPLS-DA) and One-way analysis of variance(ANOVA),etc. Overall, 34 metabolites showed significant concentration changes after intervention for 48 h, mainly involving multiple metabolic pathways, including lysine degradation, glycine, serine and threonine metabolism, arginine and proline metabolism, cysteine and methionine metabolism, aminoacyl-tRNA biosynthesis, primary bile acid biosynthesis, lysine biosynthesis. In this study, the anti-tumor mechanisms of curcumin interfering with energy metabolism, amino acid metabolism, microtubule system, protein synthesis and oxidative stress response of tumor cells were analyzed from the perspective of metabolism, providing a new reference for further tumor pharmacology study.

Calorie restriction prevents the development of insulin resistance and impaired lipid metabolism in gestational diabetes offspring.

BACKGROUND: Gestational diabetes mellitus (GDM) has long-lasting influence on offspring, which is associated with increased risks of insulin resistance, obesity, and type II diabetes mellitus. Calorie restriction (CR) is one of the most common and available nutritional interventions to prevent obesity and diabetes. We are trying to explore the effect of CR on GDM offspring. METHODS: The streptozotocin was used to stimulate C57BL/6J mice to develop GDM, a number of metabolic characteristics and related protein expressions were determined in GDM offspring that were fed ad-libitum or treated with calorie restriction. RESULTS: CR reduced body weight and glucose levels in GDM offspring. CR modulated the lipid metabolism by decreasing triglyceride and cholesterol levels in plasma. We also found that the effect of CR on insulin sensitivity may involve in signaling pathway through the regulations of phosphatase and tensin homologue deleted on chromosome 10 (PTEN) and protein kinase B (Akt). CONCLUSION: GDM is a high risk factor for GDM offspring to develop insulin resistance, while CR could ameliorate this adverse outcome. Moreover, the specific decrease in PTEN activation and increase in Akt phosphorylation in livers of GDM offspring with CR improved insulin sensitivity and lipid metabolism.

The prevalence and determinants of drug-resistance-associated mutations in the HIV-1-infected MSM population of Henan Province in China.

To estimate the prevalence of human immunodeficiency virus (HIV) drug resistance (DR) in a population of men who have sex with men (MSM) from Henan Province of China and to identify the DR-associated HIV-1 mutations in these MSM. The HIV-positive status of the MSM subjects in this study was confirmed using ELISA and Western blotting. The MSM subjects were classified into non-treatment group (n = 106) and treatment group (n = 313). CD4(+) T-lymphocyte counts were obtained by flow cytometry, and viral load was measured by branched DNA (bDNA) signal amplification assay. HIV-1 genotypic resistance tests were performed by sequence analysis of the HIV-1 protease and reverse transcriptase genes. In the non-treatment group, 15 patients (14.2 %) displayed DR to non-nucleoside reverse transcriptase inhibitor (NNRTI). In the treatment group, the failure rate of viral suppression was 38.33 % and the DR rate was 33.2 %, which was higher than the rate observed in the non-treatment group (P < 0.05). The incidence of mutations corresponding to NNRTI resistance was significantly higher than the incidence of mutations corresponding to nucleoside reverse transcriptase inhibitor (NRTI) resistance (32.9 % vs. 26.5 %) in the cohort. After antiretroviral therapy (ART), the frequencies of K103N, G190A, Y181C, and V106A mutations were highly elevated. Logistic regression analysis results showed that duration of treatment, poor treatment compliance, drug abuse and homosexual orientation are the major risk factors for DR in this MSM population (all P < 0.05). Our results showed that DR-associated mutations in the HIV-1-infected MSM population increased significantly after ART. Furthermore, duration of treatment, poor treatment compliance, drug abuse and homosexual orientation were identified as the risk factors for DR in the MSM population from Henan Province in China.

Urinary metabonomics for diagnosis of depression in hepatitis B virus-infected patients.

BACKGROUND: Depression is concomitantly presented in Hepatitis B Virus (HBV)-infected patients and decreases these patients' quality of life. However, there are no laboratory-based methods to objectively diagnose this disorder. OBJECTIVES: The aim of this study was to investigate the alteration of urinary metabolites in depressed HBV-infected patients. PATIENTS AND METHODS: In this study, 81 depressed HBV-infected patients, 68 non-depressed HBV-infected patients and 64 Healthy Controls (HC) were recruited. A nuclear magnetic resonance (NMR)-based urinary metabonomic method was used to characterize the urinary metabolic profiling of depressed and non-depressed subjects. RESULTS: Seventeen differential urinary metabolites responsible for discriminating depressed HBV-infected patients from non-depressed HBV-infected patients and HC were identified. Among these metabolites, pyruvate, isobutyrate, N-methylnicotinamide, α-hydroxybutyrate, acetoacetate and malonate were identified as potential biomarkers for diagnosing depression in HBV-infected patients. A combined panel of these potential biomarkers could effectively discriminate depressed HBV-infected patients from non-depressed HBV-infected patients and HC, with an average accuracy of 89.6% in the training set and a predictive accuracy of 86.4% in the test set. CONCLUSIONS: These findings suggest that NMR-based urinary metabonomics approach might be a useful tool for the clinical diagnosis of depression in HBV-infected patients and the six potential biomarkers could be helpful for developing an objective diagnostic method. Limited by the number of recruited subjects, future studies are required to validate our conclusions.

An infectious molecular clone in early infection with HIV-1 subtype CRF01_AE strains: construction and biological properties.

Our aim was to construct infectious molecular clones of the CRF01_AE subtype in the primary infection phase of an acute HIV-1 infections in people screened from MSM populations, as well as continue preliminary research on this virus and its biological properties pertaining to deriving viruses. Walking sequencing was performed on a half-molecular clone with target fragment inserted. Western Blot was used to detect protein expression in HIV-1 infected 293T cells. Sequence analysis of HIV-1 genomic clones showed full-length HIV-1 genomic clones without frame shift mutation or termination codon. HIV-1 p24 antigens generated from 08-IMC were slightly greater than those from infectious molecular clones pNL4-3 3 and 93JP-NH1, but without statistical difference (all P > 0.05). The relative light units of 08-ISO was higher than those of 08-IMC, but no significant difference was observed (all P > 0.05). 08-IMC-driven virus was linked to lower replication kinetics. The replication levels of pNL4-3 and 08-ISO were significantly higher than the 08-IMC replication level but close to NH1 replication level (all P < 0.05). 08-IMC could infect the cells expressing CCR5 and be replicated in the CCR5-expressing cells with a positive percentage of 24.3 %, 08-ISO may use CCR5-using macrophage-tropic isolates as coreceptor, while pNL4-3 viruses with T cell tropisms utilize the CXCR4 co-receptor. Our study showed that the infectious molecular clones of viruses in the primary infection phase have a close relationship with the major prevalent CRF01_AE strains and have high homology with the viral RNA in plasma.

Aberrant expression profile of translationally controlled tumor protein and tumor-suppressive microRNAs in cervical cancer.

PURPOSE: Invasive and recurrent cervical cancer accounts for major mortality among women. The activity of biomarkers in cervical cancer varies with different pathological stages. The purpose of the present study was to evaluate the expression of 2 biomarkers in cervical cancer and their possible contribution to novel therapeutic strategies. METHODS: In this study, we assessed the expression of translationally controlled tumor protein (TCTP) using immunohistochemistry and Western blot analysis. The expression pattern of miR-143 was also evaluated using Northern blot analysis. RESULTS: HeLa cells and mice were used for tumor induction. A group of mice injected with HeLa cells and incubated for 6 weeks developed initial tumor, while a different group of mice injected with HeLa cells and incubated for about 10 weeks developed advanced stage cervical cancer. Histological analysis revealed higher proliferation of cells resulting in complex forms of tumor in advanced cervical cancer, whereas cell clustering was not found to be initiated in the initial stage. The results of immunohistochemistry and Western blot analysis indicated less variation in the expression of TCTP, but significant difference was observed in advanced stage. Expression of Bax apoptotic protein was higher in the initial stage of the tumor than in the advanced cervical cancer. Similar pattern of marginal downregulation of miR-143 was observed between control and initial tumor stages, but striking reduction in miR-143 expression was observed in advanced stages of tumor development. CONCLUSION: The results of this study reveal a new aspect of altered expression of biomarkers in different pathological stages that could help identify novel therapeutic strategies for cervical cancer treatment.

A study on the inhibitory effect of Radix Semiaquilegiae extract on human hepatoma HEPG-2 and SMMC-7721 cells.

The main objective of this paper was to investigate the extraction process of ethanol extract of Radix Semiaquilegiae, as well as its inhibitory activity on human hepatoma HepG-2 and SMMC-7721 cells, and to compare the inhibitory effects of different concentrations of ethanol extracts against these two hepatoma cells. Ethanol reflux extraction and ultrasound-assisted extraction with ethanol at room temperature were used in the extraction process, and MTT assay was mainly used in the activity experiment to perform in-vitro anti HepG-2 and SMMC-7721 cell activity screening of ethanol extract, and to calculate the cell inhibition rates of the extracts. The results showed that among the two types of extracts, ethanol reflux extract had more superior antitumour activity to that of the ultrasonic extract, but all of the extracts obtained had certain anti-cancer activities, and the anti-proliferative activity increased with the increase of concentration.

[Correlation of the expression of NF-κB p65 and hepatic fibrosis in hepatitis patients].

OBJECTIVE: To explore the relationship between the expression of NF-κB p65 and hepatic fibrosis in chronic hepatitis B (CHB) patients. METHODS: Sixty CHB patients with hepatic fibrosis underwent liver biopsy to determine the stages of liver fibrosis (S0-S4). The distribution and expression of collagens I, III and NF-κB p65 in different stages of fibrosis in liver tissue were observed by immunohistochemistry and the results analyzed statistically. RESULTS: The expression of NF-κB p65 was positively correlated with the stage of hepatic fibrosis. That was S4 > S3 > S2 > S1 (S0) (P < 0.01). And it was also positively correlated with the expression of collagens I and III (P < 0.01). CONCLUSION: The elevated expression of NF-κB p65 is closely associated with the occurrence and development of hepatic fibrosis. Its mechanism is probably related with the increased secretion of collagens I and III after the activation of hepatic stellate cell.

MicroRNA-372 is down-regulated and targets cyclin-dependent kinase 2 (CDK2) and cyclin A1 in human cervical cancer, which may contribute to tumorigenesis.

MicroRNAs are a class of noncoding RNAs that are ~22 nucleotides in length. MicroRNAs have been shown to play important roles in cell differentiation and in cancer. Recently, studies have shown that miR-372 is tumorigenic in human reproductive system cancers. However, we provide evidence that miR-372 acts as a tumor suppressor gene in cervical carcinoma. miR-372 was found down-regulated in cervical carcinoma tissues as compared with adjacent normal cervical tissues. Growth curve and FACS assays indicated that ectopic expression of miR-372 suppressed cell growth and induced arrest in the S/G2 phases of cell cycle in HeLa cells. We used bioinformatic predictions to determine that CDK2 and cyclin A1 were possible targets of miR-372 and confirmed this prediction using a fluorescent reporter assay. Taken together, these findings indicate that an anti-oncogenic role of miR-372 may be through control of cell growth and cell cycle progression by down-regulating the cell cycle genes CDK2 and cyclin A1.

[Effect of polymorphism of human intestinal fatty acid binding protein gene on the therapeutic efficacy of fenofibrate].

OBJECTIVE: To explore the effect of polymorphism in codon Ala54Thr of human intestinal fatty acid-binding protein gene (IFABP) on the therapeutic efficacy of fenofibrate. METHODS: Totally 147 patients with hyperlipidemia [72 men mean age: (56.2 +/- 8.63) years; 75 women mean age: (58.4 +/- 9.12) years] were enrolled. IFABP genotypes were detected by polymerase chain reaction, Hha I digestion, and sequencing. Four weeks before and after treatment, the levels of fasting serum total cholesterol (TC), triglyceride (TG), high density lipoprotein-cholesterol (HDL-C), low density lipoprotein-cholesterol (LDL-C), apolipoprotein A I (apoA I) and apolipoprotein B (apoB) were detected with biochemical techniques. RESULTS: The frequency of IFABP genotype was 0.47 for A/A, 0.37 for A/T, and 0.16 for T/T, and the allelic frequency was 0.65 for A and 0.35 for T. No significant different was found in lipid levels in every genotype before treatment (P > 0.05). After 4 weeks of treatment, the levels of TC, TG, LDL-C, and apoB significantly decreased (P < 00.01), and the levels of HDH-C and apoA I significantly increased (P < 0.01). The total therapeutic efficacy on A54A and A54T were 97% and 95%, respectively. In the patients with T54T genotype after treatment, no significant difference in lipids levels was found except TG (P < 0.05), and the total efficacy was only 38%. The total therapeutic efficacies of fenofibrate on A54A and A54T were higher than those of T54T, and there was significant different between A54A and T54T (P < 0.01). CONCLUSION: The polymorphism of human IFABP gene in hyperlipidemia is related with the therapeutic efficacy of fenofibrate, and the T54T IFABP genotype may have strong negative effect on such efficacy.

[Effect of antisense survivin RNA transfection on the growth of ovarian carcinoma SKOV3 cells in vitro and in vivo].

OBJECTIVE: To study the inhibition effect of survivin antisense RNA on the growth of ovarian carcinoma SKOV3 cells, and the tumorigenic ability of the transfected SKOV3 cells implanted subcutaneously in nude mice. METHODS: The recombinant vector pcDNA3-SVVas was constructed by directed cloning of fragments of survivin cDNA into the vector. The ovarian carcinoma SKOV3 cells were transfected with pcDNA3-SVVas by lipofectamine 2000 (SKOV3/SVVas cells), and blank vector pcDNA3 as control (SKOV3/neo cells). The effect of survivin antisense RNA on cell growth was assessed by growth curve. The inhibition of expression of endogenous survivin protein and mRNA was evaluated by immunohistochemical stain and RT-PCR. The apoptosis of cells was assessed by flow cytometry. Twenty-four nude mice were divided into three groups, then SKOV3/SVVas, SKOV3/neo and SKOV3 cells were implanted subcutaneously. The growth of tumor was observed and the tumor volume calculated. RESULTS: The expression of survivin significantly decreased in SKOV3/SVVas cells in comparison to that in SKOV3 and SKOV3/neo cells. The growth of the SKOV3/SVVas cells became significantly slower than those of SKOV3 cells and SKOV3/neo cells (P < 0.01). The expression of survivin protein and mRNA in the SKOV3/SVVas cells were (37.5 +/- 1.0)% and 0.407 +/- 0.022 respectively, compared with SKOV3 [(81.2 +/- 0.4)%, 0.793 +/- 0.042] and SKOV3/neo [(80.4 +/- 0.8)%, 0.734 +/- 0.039]. The difference was significant (P < 0.01). After transfected with pcDNA3-SVVas, cells were found apoptosis in early stage by flow cytometry. The apoptosis rate was (27.4 +/- 9.6)%. There was significant difference between SKOV3/SVVas cells and SKOV3 cells, as well as between SKOV3/SVVas and SKOV3/neo cells (P < 0.05). The tumorigenic ability of SKOV3/SVVas cells was reduced. The first time that tumor could be detected in SKOV3/SVVas group, (14.0 +/- 1.0) days was significantly prolonged compared to SKOV3/neo, (6.1 +/- 0.8) days and SKOV3, (5.8 +/- 0.9) days (P < 0.01). In SKOV3/SVVas group, 5 of 8 nude mice were found tumor, the growth rate of tumor was slower compared with the other two groups. When compared in volume, the difference was significant (P < 0.01). CONCLUSIONS: Stable expression of survivin antisense RNA can effectively inhibit the growth of SKOV3 cells and reduce the expression of endogenous survivin proteins and mRNA, induce the apoptosis of cells. Survivin antisense RNA can inhibit the tumorigenesis of SKOV3 cells in nude mice.