N-Substituted Benzyl Matrinic Acid Derivatives Inhibit Hepatitis C Virus (HCV) Replication through Down-Regulating Host Heat-Stress Cognate 70 (Hsc70) Expression

Heat-stress cognate 70 (Hsc70) is a host factor that helps hepatitis C virus (HCV) to complete its life cycle in infected hepatocytes. Using Hsc70 as a target for HCV inhibition, a series of novel N-substituted benzyl matrinic/sophoridinic acid derivatives was synthesized and evaluated for their anti-HCV activity in vitro. Among these analogues, compound 7c possessing N-p-methylbenzyl afforded an appealing ability to inhibit HCV replication with SI value over 53. Furthermore, it showed a good oral pharmacokinetic profile with area-under-curve (AUC) of 13.4 µM·h, and a considerably good safety in oral administration in mice (LD₅₀>1000 mg/kg). As 7c suppresses HCV replication via an action mode distinctly different from that of the marketed anti-HCV drugs, it has been selected as a new mechanism anti-HCV candidate for further investigation, with an advantage of no or decreased chance to induce drug-resistant mutations.     

Alcohol Use and Subsequent Sex among HIV-Infected Patients in an Ethnic Minority Area of Yunnan Province,China

Objective

To examine alcohol use and subsequent HIV risky behaviors among a sample of predominately ethnic minority people living with HIV/AIDS (PLWHA) in a rural community in Yunnan Province, China.

Method

A cross-sectional study with a face-to-face questionnaire interview was conducted among eligible participants.

Results

In total, 455 (94.4%) out of 482 eligible HIV patients participated in the study. Of them, 82.6% were ethnic minorities; 15.4% were never married; 96.5% were sexually experienced; 55.4% had used drugs, 67% were receiving antiretroviral therapy (ART). Over 65% were ever drinkers; of whom 61.5% were current drinkers. Among current drinkers, 32.4% drank daily and 41.2% were hazardous drinkers. Chinese white wine was the preferred choice. Higher level of alcohol use among drinkers in the preceding month was positively associated with being males (OR = 2.76, 95%CI: 1.03–7.43), ethnic minorities (OR _Jingpo = 2.21, 95%CI: 1.06–4.59; OR _{other minorities} = 3.20, 95%CI: 1.34–7.62), higher education (OR_1–6 = 1.98, 95%CI: 0.99–3.96; OR_≥7 = 2.35, 95%CI: 1.09–5.06) and being ART-naive (OR = 2.69, 95%CI: 1.67–4.32). About 39% of ever drinkers reported having engaged in sex after drinking since HIV diagnosis. Those who were younger than 46 years (OR_16–25 = 7.77, 95%CI: 1.22–49.60, OR_26–35 = 2.79, 95%CI: 1.06–7.35, OR_36–45 = 2.96, 95%CI: 1.57–7.58), hazardous drinkers (OR = 1.99, 95%CI: 1.00–3.97) and drug users (OR = 3.01, 95%CI: 1.19–7.58) were more likely to have had sex after drinking. Approximately 56% of drug users had used drugs after drinking.

Conclusions

High prevalence of alcohol use and subsequent risky behaviors including sexual engagement and drug use among HIV patients in rural Yunnan require tremendous and integrated efforts for prevention and control of alcohol and drug abuse and HIV spreading.     

Proteomic Evaluation of Acquired Enamel Pellicle during In Vivo Formation

Acquired enamel pellicle (AEP) is a protein film that forms on the enamel surface of teeth by selective adsorption of proteins and peptides present in the mouth. This protein film forms the interface between enamel and the damage oral biofilm, which modulates the attachment of bacteria found in oral biofilm. The overall goal of this study was to gain insight into the biological formation of the human in vivo AEP. This study hypothesized that AEP is created by the formation of successive protein layers, which consist of initial binding to enamel and subsequent protein-protein interactions. This hypothesis was examined by observing quantitative and qualitative changes in pellicle composition during the first two hours of AEP formation in the oral cavity. Quantitative mass spectrometry approaches were used to generate an AEP protein profile for each time-point studied. Relative proteomic quantification was carried out for the 50 proteins observed in all four time-points. Notably, the abundance of important salivary proteins, such as histatin 1, decrease with increasing of the AEP formation, while other essential proteins such as statherin showed constant relative abundance in all time-points. In summary, this is the first study that investigates the dynamic process to the AEP formation by using proteomic approaches. Our data demonstrated that there are significant qualitative and quantitative proteome changes during the AEP formation, which in turn will likely impact the development of oral biofilms.     

Reduced CTGF Expression Promotes Cell Growth,Migration, and Invasion in Nasopharyngeal Carcinoma

Background

The role of CTGF varies in different types of cancer. The purpose of this study is to investigate the involvement of CTGF in tumor progression and prognosis of human nasopharyngeal carcinoma (NPC).

Experimental design

CTGF expression levels were examined in NPC tissues and cells, nasopharynx (NP) tissues, and NP69 cells. The effects and molecular mechanisms of CTGF expression on cell proliferation, migration, invasion, and cell cycle were also explored.

Results

NPC cells exhibited decreased mRNA expression of CTGF compared to immortalized human nasopharyngeal epithelial cell line NP69. Similarly, CTGF was observed to be downregulated in NPC compared to normal tissues at mRNA and protein levels. Furthermore, reduced CTGF was negatively associated with the progression of NPC. Knocking down CTGF expression enhanced the colony formation, cell migration, invasion, and G1/S cell cycle transition. Mechanistic analysis revealed that CTGF suppression activated FAK/PI3K/AKT and its downstream signals regulating the cell cycle, epithelial-mesenchymal transition (EMT) and MMPs. Finally, DNA methylation microarray revealed a lack of hypermethylation at the CTGF promoter, suggesting other mechanisms are associated with suppression of CTGF in NPC.

Conclusion

Our study demonstrates that reduced expression of CTGF promoted cell proliferation, migration, invasion and cell cycle progression through FAK/PI3K/AKT, EMT and MMP pathways in NPC.     

A Dual-Mode Surface Display System for the Maturation and Production of Monoclonal Antibodies in Glyco-Engineered Pichia pastoris

State-of-the-art monoclonal antibody (mAb) discovery methods that utilize surface display techniques in prokaryotic and eukaryotic cells require multiple steps of reformatting and switching of hosts to transition from display to expression. This results in a separation between antibody affinity maturation and full-length mAb production platforms. Here, we report for the first time, a method in Glyco-engineered Pichia pastoris that enables simultaneous surface display and secretion of full-length mAb molecules with human-like N-glycans using the same yeast cell. This paradigm takes advantage of homo-dimerization of the Fc portion of an IgG molecule to a surface-anchored "bait" Fc, which results in targeting functional “half” IgGs to the cell wall of Pichia pastoris without interfering with the secretion of full length mAb. We show the utility of this method in isolating high affinity, well-expressed anti-PCSK9 leads from a designed library that was created by mating yeasts containing either light chain or heavy chain IgG libraries. Coupled with Glyco-engineered Pichia pastoris , this method provides a powerful tool for the discovery and production of therapeutic human mAbs in the same host thus improving drug developability and potentially shortening the discovery time cycle.     

Bmi-1 Promotes Glioma Angiogenesis by Activating NF-κB Signaling

Angiogenesis in glioma is associated with the poor prognosis of the disease and closely correlates with the highly invasive phenotype of glioma cells, which represents the most challenging impediment against the currently glioma treatments. Bmi-1, an onco-protein, has been implicated in the progression of various human cancers, including gliomas, whereas its role in glioma angiogenesis remains unclear. Our current study examined the effects of Bmi-1 on glioma angiogenesis in vitro as well as in vivo. We found that overexpression of Bmi-1 enhanced, whereas knockdown of Bmi-1 diminished, the capability of glioma cells to induce tubule formation and migration of endothelial cells and neovascularization in chicken chorioallantoic membrane. In vivo, Bmi-1 overexpression and knockdown, respectively, promoted and inhibited angiogenesis in orthotopically transplanted human gliomas. Furthermore, NF-κB activity and VEGF-C expression was induced by Bmi-1 overexpression, whereas Bmi-1 knockdown attenuated NF-κB signaling and decreased VEGF-C expression. Additionally suppression of NF-κB activity using a specific chemical inhibitor abrogated the NF-κB activation and the pro-angiogenic activities of glioma cells. Together, our data suggest that Bmi-1 plays an important role in glioma angiogenesis and therefore could represent a potential target for anti-angiogenic therapy against the disease.     

Function and Expression of Cystic Fibrosis Transmembrane Conductance Regulator after Small Intestinal Transplantation in Mice

The secretion function of intestinal graft is one of the most important factors for successful intestinal transplantation. Cystic fibrosis transmembrane conductance regulator (CFTR) mediates HCO₃ ^- and Cl^- secretions in intestinal epithelial cells. In this study, we made investigation on the expression and function of CFTR in an experimental model of murine small intestinal transplantation. Heterotopic intestinal transplantations were performed in syngeneic mice. The mRNA and protein expressions of CFTR were analyzed by real time PCR and western blot. Murine intestinal mucosal HCO₃ ^- and Cl^- secretions were examined in vitro in Ussing chambers by the pH stat and short circuit current (I_sc) techniques. The results showed that forskolin, an activator of CFTR, stimulated jejunal mucosal epithelial HCO₃ ^- and Cl^- secretions in mice, but forskolin-stimulated HCO₃ ^- and Cl^- secretions in donor and recipient jejunal mucosae of mice after heterotopic jejunal transplantation were markedly decreased, compared with controls (P<0.001). The mRNA and protein expression levels of CFTR in donor and recipient jejunal mucosae of mice were also markedly lower than those in controls (P<0.001), and the mRNA and protein expression levels of tumor necrosis factor α (TNFα) were markedly increased in donor jejunal mucosae of mice (P<0.001), compared with controls. Further experiments showed that TNFα down-regulated the expression of CFTR mRNA in murine jejunal mucosa. In conclusion, after intestinal transplantation, the function of CFTR was impaired, and its mRNA and protein expressions were down-regulated, which may be induced by TNFα.     

Association between MMP1 -1607 1G>2G Polymorphism and Head and Neck Cancer Risk: A Meta-Analysis

Background

MMP1 is an important member of the MMP endopeptidase family that plays a critical role in the development of head and neck cancer (HNC). Several studies have investigated the association between the MMP1 -1607 1G>2G polymorphism and risk of HNC, but their results have been inconsistent. Here, we conducted a meta-analysis to further explore the role of the MMP1 -1607 1G>2G polymorphism in HNC development.

Methods

We identified all eligible studies in the electronic databases of PubMed, ISI Web of Knowledge, MEDLINE, Embase, and Google Scholar (from January 2000 to June 2012). A meta-analysis was performed to evaluate the association between the MMP1 -1607 1G>2G polymorphism and risk of HNC by calculating odds ratios (OR) and 95% confidence interval (CIs).

Results

Twelve studies were included in this meta-analysis. In overall comparison, significant associations were found using the recessive and allelic contrast models (OR, 1.38; 95% CI, 1.07–1.79 and OR, 1.27; 95% CI, 1.05–1.53, respectively), but no association was detected using the dominant model. In the stratified analyses by several variables, significant associations were observed using the recessive, dominant, and allelic contrast models in the Asian population (OR, 1.64; 95% CI, 1.29–2.08; OR, 1.39; 95% CI, 1.06–1.82; and OR, 1.41; 95% CI, 1.21–1.65, respectively), European population (OR, 0.58; 95% CI, 0.40–0.84; OR, 0.64; 95% CI, 0.44–0.92; and OR, 0.68; 95% CI, 0.54–0.85, respectively), and population-based subgroup (OR, 1.24; 95% CI,1.05–1.47; OR,1.48; 95% CI,1.04–2.12; and OR, 1.22; 95% CI, 1.07–1.38, respectively). Furthermore, significant associations were detected in oral cavity cancer and nasopharyngeal cancer under the recessive model.

Conclusion

Our results suggest that the MMP1 -1607 1G>2G polymorphism is associated with risk of HNC and that it plays different roles in Asian and European populations. Further studies with large sample size are needed to validate our findings.     

The Significance of Notch1 Compared with Notch3 in High Metastasis and Poor Overall Survival in Hepatocellular Carcinoma

Background

The prognosis for patients with hepatocellular carcinoma (HCC) is poor, and the mechanisms underlying the development of HCC remain unclear. Notch1 and Notch3 may be involved in malignant transformation, although their roles remain unknown.

Materials and Methods

HCC tissues were stained with anti-Notch1 or -Notch3 antibody. The migration and invasion capacities of the cells were measured with transwell cell culture chambers. RT-PCR was used to measure the expression of Notch1 and Notch3 mRNA. Additionally, western blot analysis was used to assess the protein expression of Notch1, Notch3, CD44v6, E-cadherin, matrix metalloproteinase-2 (MMP-2), MMP-9, and urokinase-type plasminogen activator (uPA). RNA interference was used to down-regulate the expression of Notch1 and Notch3. Cell viability was assessed using MTT.

Results

Based on immunohistochemistry, high Notch1 expression was correlated with tumor size, tumor grade, metastasis, venous invasion and AJCC TNM stage. High Notch3 expression was only strongly correlated with metastasis, venous invasion and satellite lesions. Kaplan-Meier curves demonstrated that patients with high Notch1 or Notch3 expression were at a significantly increased risk for shortened survival time. In vitro, the down-regulation of Notch1 decreased the migration and invasion capacities of HCC cells by regulating CD44v6, E-cadherin, MMP-2, MMP-9, and uPA via the COX-2 and ERK1/2 pathways. Down-regulation of Notch3 only decreased the invasion capacity of HCC cells by regulating MMP-2 and MMP-9 via the ERK1/2 pathway.

Conclusions

Based on the migration and invasion of HCC, we hypothesize that targeting Notch1 may be more useful than Notch3 for designing novel preventive and therapeutic strategies for HCC in the near future.     

Predicting National Suicide Numbers with Social Media Data

Suicide is not only an individual phenomenon, but it is also influenced by social and environmental factors. With the high suicide rate and the abundance of social media data in South Korea, we have studied the potential of this new medium for predicting completed suicide at the population level. We tested two social media variables (suicide-related and dysphoria-related weblog entries) along with classical social, economic and meteorological variables as predictors of suicide over 3 years (2008 through 2010). Both social media variables were powerfully associated with suicide frequency. The suicide variable displayed high variability and was reactive to celebrity suicide events, while the dysphoria variable showed longer secular trends, with lower variability. We interpret these as reflections of social affect and social mood, respectively. In the final multivariate model, the two social media variables, especially the dysphoria variable, displaced two classical economic predictors – consumer price index and unemployment rate. The prediction model developed with the 2-year training data set (2008 through 2009) was validated in the data for 2010 and was robust in a sensitivity analysis controlling for celebrity suicide effects. These results indicate that social media data may be of value in national suicide forecasting and prevention.