[Short-term prognostic predictive value of the neutrophil/lymphocyte ratio combined with prognostic nutritional index in hepatitis B virus-related acute-on-chronic liver failure].

Objective: To explore the prognostic predictive value of neutrophil/lymphocyte ratio (NLR) combined with prognostic nutritional index (PNI) in patients with hepatitis B virus-related acute-on-chronic liver failure (HBV-ACLF). Methods: Clinical data from 149 HBV-ACLF patients admitted to the infectious diseases Department of the General Hospital of Ningxia Medical University were retrospectively analyzed. Demographic data of the enrolled patients and the initial clinical-related data after admission were collected. Patients were divided into survival (93 cases) and death groups (56 cases) according to their prognostic condition 90 days after discharge. Demographic and clinical differences were compared between the two groups data. Receiver operating characteristic (ROC) curves were plotted to determine the optimal cutoff values for NLR and PNI in predicting the 90-day mortality rate of HBV-ACLF patients. The COX regression model was used to conduct univariate and multivariate analyses to investigate the correlation between NLR and PNI and the prognosis of HBV-ACLF patients. Kaplan-Meier survival analysis was used to explore the effects of NLR and PNI on the survival of HBV-ACLF patients. Results: The death group NLR was higher than that of the survival group, while the PNI was lower than that of the survival group, with a statistically significant difference. The area under the receiver operating characteristic curve (0.842, 95% CI: 0.779-0.906) showed patients with adverse prognosis assessed by NLR combined with PNI had a superior prognosis than that of the Model for End-Stage Liver Disease (MELD) and its combined serum sodium (MELD-Na) and Child-Turcotte-Pugh (CTP) scores. COX regression analysis showed that NLR≥3.03 and MELD score were independent risk factors affecting the prognosis of HBV-ACLF patients. PNI > 36.13 was a protective factor for evaluating the prognosis of HBV-ACLF patients. Conclusion: NLR combined with PNI can enhance the prognostic predictive value of HBV-ACLF.

[Quantitative hepatitis B core antibody levels can be used as a predictive index of HBsAg clearance].

Objective: To explore the possibility of hepatitis B core antibody (anti-HBc) in predicting hepatitis B virus surface antigen (HBsAg) clearance. Methods: Sixty cases with chronic hepatitis B who were previously treated with peginterferon α-2a combined with nucleos(t)ide analogues (NAs) antiviral therapy were divided according to the HBsAg clearance or non-clearance; 41 cases in the clearance group and 19 cases in the non-clearance group. Double antigen sandwich method was used to detect patients anti-HBc quantitative levels during the course of treatment and at baseline, 24, 48, 72 and 96 weeks. Logistic regression analysis and receiver operating characteristic curve (ROC) were used to evaluate the predictive ability of related influencing factors for HBsAg clearance. Results: With antiviral treatment prolongation, anti-HBC quantitative levels in the overall population showed a progressive downward trend in the clearance group and the non-clearance group, but the anti-HBC level in the clearance group was significantly higher than non-clearance group at the baseline and successive detection time points during the antiviral treatment (P < 0.05). Multivariate logistic regression showed that baseline quantitative anti-HBC level, HBsAg decline at week 24 (log10 IU / ml), and alanine aminotransferase (ALT) > 1.5 times the upper limit of normal value (ULN) were all influencing factors for HBsAg clearance during the treatment (OR = 0.156, P = 0.026; OR = 0.134, P = 0.023; OR = 0.239, P = 0.028). Among them, the baseline quantitative anti-HBc level was the best independent predictor for HBsAg clearance (OR = 0.235; P = 0.004), and the sensitivity and specificity for predicting HBsAg clearance at > 3.40 log10 IU/ mL were 56.1% and 89.5%, respectively. Logistic regression model was used as a reference to construct combined predictors in order to improve the prediction accuracy. Among them, the combined factor 3 had the highest predictive value (the area under the ROC curve had reached up to 0.870; 95%CI was 0.781 ~ 0.960; P < 0.001). The cut-off value of combined factor 3 was > 0.386, and the sensitivity and specificity were 80.5% and 78.9%, respectively. In addition, the combined index had further improved the predictive value, which is the combination of any two or more indexes based on the baseline quantitative anti-HBC level, and HBsAg clearance predictive rate had reached 94.12% ~ 100%. Conclusion: The baseline quantitative anti-HBC level has the highest predictive value for HBsAg clearance. The combination of ALT > 1.5×ULN and HBsAg decline at 24 weeks during the treatment can more precisely predict HBsAg clearance. Therefore, it is a reliable non-invasive biomarker.

MicroRNA-362 inhibits cell growth and metastasis in glioblastoma by targeting MAPK1.

OBJECTIVE: Glioblastoma (GBM) is a deadly brain cancer that seriously threatens the lives of patients. Moreover, various microRNAs (miRNAs) have been found to be involved in the progression of GBM. The purpose of this study is to preliminarily elucidate the regulatory mechanism of miR-362 in GBM. PATIENTS AND METHODS: The abnormal expression of miR-362 and MAPK1 was detected by RT-qPCR or Western blot analysis in GBM tissues and cells. CCK-8 and transwell assays were performed to measure cell proliferation, migration and invasion. The relationship between miR-362 and MAPK1 was confirmed by luciferase reporter assay. RESULTS: MiR-362 expression was reduced in GBM tissues and cells. The decreased expression of miR-362 predicted poor prognosis in GBM patients. Functionally, overexpression of miR-362 inhibited the proliferation and metastasis of GBM cells. In addition, miR-362 directly targets MAPK1. MAPK1 was negatively correlated with miR-362 expression in GBM. Moreover, MAPK1 was upregulated and served as a tumor promoter in GBM. More importantly, the upregulation of MAPK1 weakened the inhibitory effect of miR-362 on cell proliferation and metastasis in GBM. CONCLUSIONS: MiR-362 restrains cell proliferation and metastasis in GBM by targeting MAPK1, indicating that miR-362 functions as a tumor suppressor in GBM.

MiRNA-802 inhibits the metastasis of colorectal cancer by targeting FOXE1.

OBJECTIVE: The aim of this study was to explore the role of microRNA-802 (miRNA-802) in the progression of colorectal cancer (CRC) and the underlying mechanism. PATIENTS AND METHODS: The relative expression levels of miRNA-802 and FOXE1 in 40 paired CRC tissues and adjacent normal tissues were detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). The correlation between miRNA-802 expression and the pathological indexes of CRC patients was assessed. Meanwhile, the prognostic potentials of miRNA-802 and FOXE1 in CRC patients were identified through the Kaplan-Meier method. After overexpression of miRNA-802, the changes in the proliferative, migratory, and invasive capacities of HT29 and HCT-8 cells were evaluated in vitro. The Dual-Luciferase Reporter Gene Assay was applied to investigate the binding relationship between miRNA-802 and FOXE1. Finally, the rescue experiments were carried out to uncover the role of the miRNA-802/FOXE1 axis in regulating the cellular behaviors of CRC. RESULTS: MiRNA-802 was significantly downregulated in CRC tissues and cell lines. CRC patients with a low level of miRNA-802 had significantly higher rates of lymphatic metastasis and distant metastasis, as well as worse overall survival. The transfection of miRNA-802 mimics remarkably attenuated the proliferation, migration, and invasion of HT29 and HCT-8 cells. FOXE1 expression was significantly upregulated in CRC tissues and cell lines. Meanwhile, the expression of FOXE1 was negatively correlated with miRNA-802 in CRC tissues. A higher level of FOXE1 indicated the worse prognosis of CRC patients. The Dual-Luciferase Reporter Gene Assay further verified the binding relationship between FOXE1 and miRNA-802. Importantly, the overexpression of FOXE1 could reverse the regulatory effects of miRNA-802 on the cellular behaviors of CRC. CONCLUSIONS: MiRNA-802 is significantly downregulated in CRC, and is closely related to lymphatic and distant metastasis of CRC. Furthermore, miRNA-802 alleviates the malignant progression of CRC via negatively regulating FOXE1.

[Strategy of nursing care on the face skin injuries caused by wearing medical-grade protective equipment].

For effective resistance to virus attack and infection, reducing virus transmission chance, it is extremely important for the medical staff and related workers to have their own safe protection. This paper summarizes the development causes, common locations, and prevention ways about the device related pressure injuries on the face resulted from wearing medical-grade protective equipment for a long working time. The paper proposes the nursing strategy for device related pressure injuries and other nursing strategy is proposed to take care efficiently the device related pressure injuries. Meantime, a corresponding nursing strategy is also suggested to deal with the correlative skin diseases during the application of medical-grade protective equipment. These paper aims to provide reference for the prevention of device related pressure injuries and the care of skin-related diseases for clinical working staff, especially to the respectable personnel in front line of fighting against Corona virus disease 2019.

[The role of pulmonary arterial pressure in chronic obstructive pulmonary disease phenotypes based on cluster analysis and its prognostic value].

Objective: To explore the role of pulmonary arterial pressure in chronic obstructive pulmonary disease (COPD) phenotypes based on cluster analysis and its prognostic value. Methods: Three hundred and nineteen patients admitted to Beijing Chaoyang Hospital and Xuanwu Hospital from April 2013 to April 2016 were recruited in the study. All the patients were older than 40 years old and in stable COPD. One-year follow-up was performed and the endpoint was acute exacerbation of COPD or all-cause mortality. Age, body mass index (BMI), smoking index, history of exacerbation, modified British medical research council (mMRC), forced expiratory volume in first second (FEV(1)), pulmonary arterial pressure and right ventricular transverse diameter measured by echocardiography were selected as cluster indicators to classify patients, survival analysis was performed. Results: Eight cluster indexes were converted into four independent principal components by principal component analysis (PCA), with a cumulative contribution rate of 70.1%. The extracted principal components were used for cluster analysis. Patients were divided into four categories, each contained different GOLD grades and had statistically significant differences in age, symptoms, degree of pulmonary function impairment and pulmonary arterial pressure (all P<0.001). The four categories were: class 1: young, pulmonary function damage was medium, lower pulmonary arterial pressure, good prognosis; class 2: elderly, pulmonary function damage was mild, higher pulmonary arterial pressure, poor prognosis; class 3: young, pulmonary function damage was serious, normal pulmonary arterial pressure, the best prognosis; class 4: elderly, pulmonary function damage was medium, pulmonary arterial pressure increased significantly, the worst prognosis. Conclusion: Cluster analysis based on pulmonary artery pressure can be used to identify COPD patients with different risk of acute exacerbation or death, suggesting that pulmonary hypertension as a COPD phenotype plays a role in prognostic assessment.

[A preliminary study of serum marker alpha-enolase in the diagnosis of hepatocellular carcinoma].

Objective: To investigate the diagnostic value of serum α-enolase (ENO1) in the primary hepatocellular carcinoma. Methods: From May 2012 to March 2017, 163 cases with liver diseases who met the inclusion and exclusion criteria were admitted to the Infectious Diseases Department of the General Hospital of Ningxia Medical University. Among them, 28 cases were of chronic hepatitis B (CHB), 31 cases with liver cirrhosis (LC), 104 cases with hepatocellular carcinoma (HCC), and 18 healthy volunteers (NC). Patient data and serum samples were collected and liver disease related indicators were measured to detect ENO1 levels with enzyme-linked immunosorbent assay (ELISA). The measured indicators were expressed in median. Mann-Whitney U nonparametric test was used to analyze the differences between the data. A Spearman's correlation analysis was used for bivariate correlation analysis. The sensitivity and specificity of ENO1 and alpha-fetoprotein in the diagnosis of liver cancer were analyzed by ROC curve. Results: Serum level of ENO1 in CHB group, LC group and HCC group was significantly higher than normal group. Serum level of ENO1 in HCC group was higher than CHB group (P = 0.001) and LC group (P < 0.01). Area under the curve (AUC) for serum ENO1 and alpha-fetoprotein were 0.782 (cut-off value 75.96, P = 0.000 1) and 0.800 (cut-off value 27.02, P = 0.000 1), respectively. There was a positive correlation between ENO1 and AFP (P = 0.001). The combined detection had significantly improved the detection efficiency (AUC = 0.835). Serum ENO1 was statistically significant (P < 0.05) in HCC tumor size (AUC = 0.663), tumor metastasis (AUC = 0.681), TNM stage (AUC = 0.710, stage I vs. II), and Edmondson grade (AUC = 0.685) (P < 0.05) and the elevated levels of ENO1 had significantly reduced (P < 0.05) the survival time. Conclusion: ENO1 can be a new candidate marker for the diagnosis of early stage HCC and its progression.

[Clinical study on liver function, virology, serological changes and the safety of drug withdrawal in pregnant women who are chronic HBV carriers during pregnancy and postpartum].

Objective: To observe the changes of liver function, virology and serology and the safety of drug withdrawal in pregnant women who are chronic hepatitis B virus (HBV) carriers. Methods: A prospective clinical cohort was established to enroll pregnant women who are chronic HBV carriers and they were divided into the nucleoside/nucleotide analogs (NAs) intervention group and the non-NAs intervention group according to patients' wishes. Liver function, HBV DNA and HBV serological markers were detected at gestation, postpartum 6 weeks, 12 weeks, 24 weeks, 36 weeks and 48 weeks. Results: 351 patients were enrolled, 320 in the NAs intervention group and 31 in the non-NAs intervention group. The proportion of postpartum hepatitis flares in both groups was higher than that in pregnancy (39.4% vs 12.5%, P < 0.001; 38.7% vs 3.2%, P = 0.001). Six weeks postpartum was the peak period of hepatitis flares, and 96.0% (121/126) of the hepatitis flares occurred within 24 weeks postpartum. At 6 weeks postpartum, there were 6 cases of alanine aminotransferase (ALT) ≥ 10 times upper limit of normal (ULN) in the NAs intervention group. The rate of the hepatitis flare after drug withdrawal was 16.7% (34/203). Conclusion: Regardless of the presence or absence of NAs intervention, pregnant women who are chronic HBV carriers have a certain proportion of hepatitis flares during pregnancy and postpartum, and the hepatitis flare even have a tendency to be severe. Therefore, drug withdrawal after delivery is not always safe, which requires close observation and classification. At 6 weeks postpartum, the incidence of hepatitis flares was high, and those who meet the treatment indications can get better therapeutic effects if given appropriate treatment. The vast majority (96%) of postpartum hepatitis flares occur within 24 weeks, so it is recommended to follow up to at least 24 weeks postpartum after discontinuation.

[Low-levels of HBsAg quantification at 48-week in HBeAg-negative chronic hepatitis B patients are the advantageous population for HBsAg clearance].

Objective: To analyze the therapeutic effect on HBeAg-negative chronic hepatitis B patients treated with Peg-IFNα-2a combined with NAs to obtain the influencing factors for predicting HBsAg clearance. Methods: A retrospective study was conducted to investigate the effect of pegylated interferon alpha-2a combined with nucleoside analogues (lamivudine/adefovir dipivoxil) on HBeAg-negative chronic hepatitis B. The treatment course was 96 weeks. Patients were followed up 120 weeks after the treatment. HBsAg clearance at 120 weeks was taken as the objective of the study. Logistic regression and receiver operating characteristic curve analysis screened the related factors affecting HBsAg clearance. χ (2) test was used to compare count data. Results: 111 patients were treated with pegylated interferon alpha-2a combined with nucleoside analogues, and 107 patients completed the scheduled course of treatment and follow-up. HBsAg clearance rate at120 week was 29.0% (31/107). The influencing factors for analysis were: (1) gender had no effect on HBsAg clearance rate; age and baseline levels of HBV DNA and alanine aminotransferase had no significant effect on HBsAg clearance; low baseline level of HBsAg (< 3.023 lgIU/ml) was beneficial to HBsAg clearance. The area under the working characteristic curve of the subjects was 0.746, the positive predictive value was 44.4%, and the negative predictive value was 86.8%. (2) HBsAg quantification or decline in 24 weeks and 48 weeks of treatment had a good predictive effect on HBsAg clearance, and the 48 weeks predicted value was higher than 24 weeks. When the HBsAg quantification was≤2.070 lgIU/ml at 48 weeks, the area under the receiver operating characteristic curve was 0.931, the positive predictive value was 52.8%, and the negative predictive value was 94.4%. When HBsAg decreased from baseline to≥0.991 lgIU/ml, the area under the receiver operating characteristic curve was 0.888, the positive predictive value was 50.8%, and the negative predictive value was 97.9%. (3) The analysis of HBsAg subgroup levels at 48 weeks suggested that the "interval analysis" can forecast HBsAg clearance more exactly than "nodal analysis" .The final HBsAg clearance rate of 100 IU/ml < HBsAg≤1 000 IU/ml, 10 IU/ml < HBsAg≤100 IU/ml and HBsAg≤10 IU/ml groups reached 6.7%, 31.8% and 67.7%, respectively. (4) The ALT abnormal group in the course of treatment obtained a higher HBsAg clearance rate (48.0%, 12/25). Conclusion: 96-weeks long-term treatment with pegylated interferon-alpha -alpha-2a combined with nucleoside analogues for HBeAg-negative chronic hepatitis B has a good predictive value for HBsAg clearance at baseline and during treatment. The "interval level" of HBsAg at 48-weeks is more accurate in predicting HBsAg clearance, suggesting that HBeAg-negative chronic hepatitis B patients with low HBsAg levels at 48-weeks are the advantageous populations with HBsAg clearance. These patients are worthy of prolonged treatment to pursue "clinical cure".

Long non-coding RNA CPS1-IT1 is a positive prognostic factor and inhibits epithelial ovarian cancer tumorigenesis.

OBJECTIVE: The aim of the present study was to explore the prognostic value of long non-coding RNA CPS1-IT1 (CPS1-IT1) expression in epithelial ovarian cancer (EOC) patients and identify the effect of CPS1-IT1 on cell proliferation and apoptosis of EOC cells. PATIENTS AND METHODS: Expression levels of CPS1-IT1 in tissues and cells were detected by the Real-time quantitative RT-PCR assay. The χ2-test was used to analyze the relationship between CPS1-IT1 expression and the clinicopathological characteristics. Survival analysis was performed using the Kaplan-Meier method and Cox's proportional hazards model. The capacity for cellular proliferation was measured with cell counting Kit-8. Cell apoptosis assays were performed using flow cytometry. Western blot was used to detect the expression levels of cell apoptosis-related proteins. RESULTS: We observed that CPS1-IT1 was significantly downregulated in EOC cell lines and tissue samples. The expression of CPS1-IT1 was significantly associated with FIGO stage and lymph node metastases. In addition, EOC patients in the low tissue CPS1-IT1 expression group had significantly shorter 5-year overall survival time than those in the high tissue CPS1-IT1 expression group. Furthermore, univariate and multivariable Cox regression analysis identified low CPS1-IT1 expression in EOC tissues as an independent poor prognostic marker of overall survival. It was also found that over-expression of CPS1-IT1 markedly promoted proliferation of EOC cells. Further studies revealed that over-expression of CPS1-IT1 induced cell apoptosis by through regulating apoptosis-related proteins. CONCLUSIONS: CPS1-IT1 may be a functional tumor suppressor in EOC. It may also serve as an independent prognostic factor for patients with EOC.

[Role of enolase 1 in hepatocellular carcinoma and possible mechanism].

Objective: To investigate the role of enolase 1 (ENO1) in hepatocellular carcinoma (HCC) and possible mechanism. Methods: Real-time PCR and Western blot were used to measure the expression of ENO1 in HCC tissue, adjacent tissue, hepatoma cells, and normal hepatocytes. The siRNA interference technique was used for ENO1 knockout in HepG2 cells, and then CCK-8, colony formation assay, and transwell assay were used to measure the proliferation, migration, and invasion abilities of HepG2 cells. Real-time PCR and Western blot were used to measure the expression of proteins and genes involved in the activation of the Notch signaling pathway. The two-independent-samples t test and a one-way analysis of variance were used for comparison. Results: HCC tissue and HepG2 cells had significantly higher expression of ENO1 than adjacent tissue and normal hepatocytes (P < 0.05). There were significant reductions in the proliferation, migration, and invasion abilities of HepG2 cells after siRNA interference (P < 0.05). There were also significant reductions in the expression of N1ICD, snail, slug, HEY1, HES1, and HES5 (P < 0.05). Conclusion: ENO1 may promote the development of HCC, possibly by participating in the regulation of the Notch signaling pathway.

Quantifying global soil carbon losses in response to warming.

The majority of the Earth's terrestrial carbon is stored in the soil. If anthropogenic warming stimulates the loss of this carbon to the atmosphere, it could drive further planetary warming. Despite evidence that warming enhances carbon fluxes to and from the soil, the net global balance between these responses remains uncertain. Here we present a comprehensive analysis of warming-induced changes in soil carbon stocks by assembling data from 49 field experiments located across North America, Europe and Asia. We find that the effects of warming are contingent on the size of the initial soil carbon stock, with considerable losses occurring in high-latitude areas. By extrapolating this empirical relationship to the global scale, we provide estimates of soil carbon sensitivity to warming that may help to constrain Earth system model projections. Our empirical relationship suggests that global soil carbon stocks in the upper soil horizons will fall by 30 ± 30 petagrams of carbon to 203 ± 161 petagrams of carbon under one degree of warming, depending on the rate at which the effects of warming are realized. Under the conservative assumption that the response of soil carbon to warming occurs within a year, a business-as-usual climate scenario would drive the loss of 55 ± 50 petagrams of carbon from the upper soil horizons by 2050. This value is around 12-17 per cent of the expected anthropogenic emissions over this period. Despite the considerable uncertainty in our estimates, the direction of the global soil carbon response is consistent across all scenarios. This provides strong empirical support for the idea that rising temperatures will stimulate the net loss of soil carbon to the atmosphere, driving a positive land carbon-climate feedback that could accelerate climate change.

[Role of glucose-6-phosphate dehydrogenase in hepatitis B virus replication and its possible mechanism of action].

OBJECTIVE: To investigate the role of glucose-6-phosphate dehydrogenase (G6PD) in hepatitis B virus (HBV) replication and its possible mechanism of action. METHODS: Tissue microarray, quantitative real-time PCR, and Western blot were performed to analyze the differences in G6PD expression levels in the HBV-positive and HBV-negative liver tissues, HepG2.2.15 cells, and HepG2 cells. The siRNA transfection technique was used to knock down G6PD gene in HepG2.2.15 cells for 48 hours. Chemiluminescence was used for HBsAg and HBeAg quantification in supernatant, and quantitative real-time PCR was used to measure HBV DNA, type I interferon (IFN), and downstream IFN-stimulated genes. The t-test was used for comparison between groups. RESULTS: G6PD expression was significantly upregulated in the HBV-positive liver tissues and cells compared with HBV-negative liver tissues and cells, and the stain intensity and immunohistochemical scores were 89.69±54.92 and 31.90±18.62, respectively (P < 0.05). After G6PD expression in HepG2.2.15 cells was interfered by siRNA, the quantitative levels of HBV DNA, HBsAg, and HBeAg in supernatant were reduced significantly, and the mRNA expression levels of IFNα1, IFNß1, and five downstream IFN-stimulated genes (OAS1, ISG15, OAS3, EIF2α, and PKR) increased significantly (all P < 0.05). CONCLUSION: G6PD plays a vital role in HBV replication, and its mechanism of action in regulating HBV replication may be related to type I IFN signaling pathway.

Effects of nucleotide usage on the synonymous codon usage patterns of biofilm-associated genes in Haemophilus parasuis.

To provide a new perspective on the evolutionary characteristics shaping the genetic diversity of Haemophilus parasuis biofilms, the relative synonymous codon usage values, codon usage bias values, effective number of codons (ENC) values, codon adaptation index (CAI) values, and the base components were calculated. Our objective was to implement a comparative analysis to evaluate the dynamic evolution of biofilm-associated genes in H. parasuis. The analysis of genetic diversity provides evidence that some biofilm-associated genes have similar genetic features. However, other genes show some variations in genetic direction. Furthermore, preferential selection of the synonymous codons and amino acids is apparent in biofilm-associated genes. Additionally, the ENC and CAI data from this study all strongly suggested that biofilm-associated genes may depend on deoptimization to adapt to environmental changes, and the mutation effect of biofilm-associated genes in H. parasuis plays an important role in shaping the genetic features. Our results reveal that the mutations of biofilm-associated genes form a set of sophisticated strategies for combating the environmental changes arising from the host cell in the evolution of H. parasuis.

Oncogenic miR-20a and miR-106a enhance the invasiveness of human glioma stem cells by directly targeting TIMP-2.

Emerging evidence has shown that cancer stem cells (CSCs) are the cellular determinants to promote cancer invasion and metastasis. However, the mechanism underlying CSC invasion remains unknown. MicroRNAs are evolutionally conserved small noncoding RNAs that are critical for the regulation of gene expression, and their expressions are often dysregulated in cancers. In the present study, we demonstrated that two functionally related microRNAs, miR-20a and -106a (miR-20a/106a), were capable of enhancing the invasiveness of CD133(+) glioma stem cells (GSCs) isolated from both glioblastoma cell line U87 and primary human glioma specimens. We found that the level of miR-20a/106a in GSCs was significantly higher than that in the committed CD133(-) glioma cells, and correlated with the invasive capability of GSCs. By bioinformatic analysis, we identified tissue inhibitor of metalloproteinases-2 (TIMP-2) as one of the miR-20a/106a-targeted genes. TIMP-2 level correlated inversely with miR-20/106 expression. Directly targeting by miR-20a/106a on 3'-untranslation region (3'-UTR) of TIMP-2 mRNA was confirmed by 3'-UTR dual-luciferase reporter assay. Knockdown of miR-20a/106a in GSCs increased endogenous TIMP-2 protein abundance, thereby inhibiting GSC invasion. We also found that Nordy, a synthetic lipoxygenase inhibitor, inhibited GSC invasiveness by elevating the expression of TIMP-2 via downregulation of miR-20a/106a. Our results indicate that miR-20a/106a has a key role in GSC invasion and may serve as targets for treatment of glioblastoma.

Pentamethylquercetin generates beneficial effects in monosodium glutamate-induced obese mice and C2C12 myotubes by activating AMP-activated protein kinase.

AIMS/HYPOTHESIS: Pentamethylquercetin (PMQ) has recently been shown to have glucose-lowering properties. Here, we aimed to characterise the effectiveness and underlying mechanisms of PMQ for ameliorating metabolic disorders in vivo and vitro. METHODS: We generated a mouse model of obesity by neonatal administration of monosodium glutamate (MSG) and used it to assess the properties of PMQ as a treatment for metabolic disorders. We also investigated the possible underlying mechanisms of PMQ in the prevention of metabolic disorders. RESULTS: Compared with normal mice, MSG mice had metabolic disorders, including central obesity, hyperinsulinaemia, insulin resistance, hyperglycaemia, hyperlipidaemia, decreased phosphorylation of AMP-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC), and downregulated levels of GLUT4 in gastrocnemius muscles. In MSG mice, PMQ treatment (5, 10, 20 mg/kg daily) reduced body weight gain, waist circumference, adipose tissue mass, serum glucose, triacylglycerol and total cholesterol, while improving insulin resistance, activating AMPK and increasing ACC phosphorylation and GLUT4 abundance. In C2C12 myotubes, PMQ (10 µmol/l) increased glucose consumption by ∼65%. PMQ treatment (1-10 µmol/l) also activated AMPK, increased ACC phosphorylation and GLUT4 abundance, and upregulated the expression of some key genes involved in fatty acid oxidation. CONCLUSIONS/INTERPRETATION: These findings suggest that PMQ can ameliorate metabolic disorders at least in part via stimulation of AMPK activity.

Calcium overload is associated with lipofuscin formation in human retinal pigment epithelial cells fed with photoreceptor outer segments.

PURPOSE: To investigate the role of Ca²(+) in lipofuscin formation in human retinal pigment epithelial (RPE) cells that phagocytize bovine photoreceptor outer segments (POSs). METHODS: Cultured human RPE cells fed with 2 × 107per l bovine POS were treated with flunarizine, an antagonist of Ca²(+) channel, or/and centrophenoxine, a lipofuscin scavenger. The Ca²(+) changes and lipofuscin formation were measured with fluoresence dye Fluo-3/AM ester, laser scanning confocal microscopy (LSCM) and flow cytometry (FCM). The activity of RPE cells was measured by methyl thiazolyl tetrazolium (MTT) assay and argyrophilic nucleolar organizer regions (AgNORs) assay. RESULTS: The Ca²(+) fluorescence intensity (CFI) of RPE cells fed with POS was significantly increased compared with the controls (165.36 ± 29.92 U). It reached a peak with 777.33 ± 63.86 U (P<0.01) at 12 h, and then decreased but still maintained a high level of 316.90 ± 36.07 U (P<0.01) for 4 days. Flunarizine and centrophenoxine significantly decreased the Ca²(+) overload to 227.18 ± 14.00 U at 12 h and 211.06 ± 20.45 U at 4 days. FCM confirmed these changes. The drugs also showed an inhibitory effect on the lipofuscin formation. The proliferation rate of the cells fed with POS increased significantly. Both drugs had inhibitory effects on the activity of the cultured cells. This tendency was confirmed by AgNORs assay. CONCLUSIONS: The Ca²(+) inflow initiated lipofuscin accumulation in RPE cells fed with POS. Flunarizine and centrophenoxine can decrease Ca²(+) overload and lipofuscin formation in RPE cells, accompanied by maintaining cellular vitality.