Double suicide genes selectively kill human umbilical vein endothelial cells

<p>Abstract</p> <p>Background</p> <p>To construct a recombinant adenovirus containing CDglyTK double suicide genes and evaluate the killing effect of the double suicide genes driven by kinase domain insert containing receptor (KDR) promoter on human umbilical vein endothelial cells.</p> <p>Methods</p> <p>Human KDR promoter, <it>Escherichia coli </it>(<it>E. coli</it>) cytosine deaminase (CD) gene and the herpes simplex virus-thymidine kinase (TK) gene were cloned using polymerase chain reaction (PCR). Plasmid pKDR-CDglyTK was constructed with the KDR promoter and CDglyTK genes. A recombinant adenoviral plasmid AdKDR-CDglyTK was then constructed and transfected into 293 packaging cells to grow and harvest adenoviruses. KDR-expressing human umbilical vein endothelial cells (ECV304) and KDR-negative liver cancer cell line (HepG2) were infected with the recombinant adenoviruses at different multiplicity of infection (MOI). The infection rate was measured by green fluorescent protein (GFP) expression. The infected cells were cultured in culture media containing different concentrations of prodrugs ganciclovir (GCV) and/or 5-fluorocytosine (5-FC). The killing effects were measured using two different methods, i.e. annexin V-FITC staining and terminal transferase-mediated dUTP nick end-labeling (TUNEL) staining.</p> <p>Results</p> <p>Recombinant adenoviruses AdKDR-CDglyTK were successfully constructed and they infected ECV304 and HepG2 cells efficiently. The infection rate was dependent on MOI of recombinant adenoviruses. ECV304 cells infected with AdKDR-CDglyTK were highly sensitive to GCV and 5-FC. The cell survival rate was dependent on both the concentration of the prodrugs and the MOI of recombinant adenoviruses. In contrast, there were no killing effects in the HepG2 cells. The combination of two prodrugs was much more effective in killing ECV304 cells than GCV or 5-FC alone. The growth of transgenic ECV304 cells was suppressed in the presence of prodrugs.</p> <p>Conclusion</p> <p>AdKDR-CDglyTK/double prodrog system may be a useful method for suppressing tumor angiogenesis.</p

Pulse Diagnosis Signals Analysis of Fatty Liver Disease and Cirrhosis Patients by Using Machine Learning

Objective. To compare the signals of pulse diagnosis of fatty liver disease (FLD) patients and cirrhosis patients. Methods. After collecting the pulse waves of patients with fatty liver disease, cirrhosis patients, and healthy volunteers, we do pretreatment and parameters extracting based on harmonic fitting, modeling, and identification by unsupervised learning Principal Component Analysis (PCA) and supervised learning Least squares Regression (LS) and Least Absolute Shrinkage and Selection Operator (LASSO) with cross-validation step by step for analysis. Results. There is significant difference between the pulse diagnosis signals of healthy volunteers and patients with FLD and cirrhosis, and the result was confirmed by 3 analysis methods. The identification accuracy of the 1st principal component is about 75% without any classification formation by PCA, and supervised learning’s accuracy (LS and LASSO) was even more than 93% when 7 parameters were used and was 84% when only 2 parameters were used. Conclusion. The method we built in this study based on the combination of unsupervised learning PCA and supervised learning LS and LASSO might offer some confidence for the realization of computer-aided diagnosis by pulse diagnosis in TCM. In addition, this study might offer some important evidence for the science of pulse diagnosis in TCM clinical diagnosis

Putative carboxylesterase gene identification and their expression patterns in Hyphantria cunea (Drury)

The olfactory system of insects is important for behavioral activities as it recognizes internal and external volatile stimuli in the environment. Insect odorant degrading enzymes (ODEs), including antennal-specific carboxylesterases (CXEs), are known to degrade redundant odorant molecules or to hydrolyze important olfactory sex pheromone components and plant volatiles. Compared to many well-studied Type-I sex pheromone-producing lepidopteran species, the molecular mechanisms of the olfactory system of Type-II sex pheromone-producing Hyphantria cunea (Drury) remain poorly understood. In the current study, we first identified a total of ten CXE genes based on our previous H. unea antennal transcriptomic data. We constructed a phylogenetic tree to evaluate the relationship of HcunCXEs with other insects\u27 CXEs, and used quantitative PCR to investigate the gene expression of H. cunea CXEs (HcunCXEs). Our results indicate that HcunCXEs are highly expressed in antennae, legs and wings, suggesting a potential function in degrading sex pheromone components, host plant volatiles, and other xenobiotics. This study not only provides a theoretical basis for subsequent olfactory mechanism studies on H. cunea, but also offers some new insights into functions and evolutionary characteristics of CXEs in lepidopteran insects. From a practical point of view, these HcunCXEs might represent meaningful targets for developing behavioral interference control strategies against H. cunea

Serum anti-Mullerian hormone predicts ovarian response in (Macaca fascicularis) monkeys

AMH as a promising predictor of ovarian response has been studied extensively in women undergoing assisted reproductive technology treatment, but little is known about its prediction value in monkeys undergoing ovarian stimulation. In the current study, a total of 380 cynomolgus monkeys ranging from 5 to 12 years received 699 ovarian stimulation cycles. Serum samples were collected for AMH measure with enzyme-linked immunosorbent assay. It was found that serum AMH levels were positive correlated with the number of retrieved oocytes (P < 0.01) in the first, second and third stimulation cycles. In the first cycles, area under the curve (ROCAUC) of AMH is 0.688 for low response and 0.612 for high response respectively, indicating the significant prediction values (P = 0.000 and P = 0.005). The optimal AMH cutoff value was 9.68 ng/mL for low ovarian response and 15.88 ng/mL for high ovarian response prediction. In the second stimulation cycles, the significance of ROCAUC of AMH for high response rather than the low response was observed (P = 0.001 and P = 0.468). The optimal AMH cutoff value for high ovarian response was 15.61 ng/mL. In the third stimulation cycles, AMH lost the prediction value with no significant ROCAUC. Our data demonstrated that AMH, not age, is a cycle-dependent predictor for ovarian response in form of oocyte yields, which would promote the application of AMH in assisted reproductive treatment (ART) of female cynomolgus monkeys. AMH evaluation would optimize candidate selection for ART and individualize the ovarian stimulation strategies, and consequentially improve the efficiency in monkeys

Serial Monitoring of Circulating Tumor DNA in Patients With Metastatic Colorectal Cancer to Predict the Therapeutic Response

Early biomarkers of therapeutic responses can help optimize the treatment of metastatic colorectal cancers (mCRC). In this prospective exploratory study, we examined serial changes of plasma-circulating tumor DNA (ctDNA) in 41 mCRC patients receiving first-line chemotherapies and tested its association with treatment outcomes according to radiological assessments. Using next-generation sequencing technologies, we profiled somatic mutations in 50 cancer-related genes in ctDNA before each of the first four treatment cycles. We observed mutations in 95.7% of pre-treatment ctDNA samples. Using mutations of the maximal frequency in each pre-treatment plasma ctDNA sample as the candidate targets, we computed log2 fold changes of ctDNA levels between adjacent treatment cycles. We found that ctDNA reductions as early as prior to cycle 2 predicted responses after cycle 4. Log2 fold changes of ctDNA after cycle 1 (ctDNA log2 (C1/C0)) &gt; −0.126 predicted progressive disease, with an accuracy of 94.6%. These patients also showed significantly worse progression-free survival than those with ctDNA log2 (C1/C0) ≤ −0.126 (median 2.0 vs. 9.0 months; P = 0.007). Together, the present exploratory study suggests that early changes in ctDNA levels detected via targeted sequencing are potential biomarkers of future treatment responses in mCRCs

Identification and Characterization of Two Functionally Unknown Genes Involved in Butanol Tolerance of Clostridium acetobutylicum

Solvents toxicity is a major limiting factor hampering the cost-effective biotechnological production of chemicals. In Clostridium acetobutylicum, a functionally unknown protein (encoded by SMB_G1518) with a hypothetical alcohol interacting domain was identified. Disruption of SMB_G1518 and/or its downstream gene SMB_G1519 resulted in increased butanol tolerance, while overexpression of SMB_G1518-1519 decreased butanol tolerance. In addition, SMB_G1518-1519 also influences the production of pyruvate:ferredoxin oxidoreductase (PFOR) and flagellar protein hag, the maintenance of cell motility. We conclude that the system of SMB_G1518-1519 protein plays a role in the butanol sensitivity/tolerance phenotype of C. acetobutylicum, and can be considered as potential targets for engineering alcohol tolerance

Viral Aetiology in Adults with Acute Upper Respiratory Tract Infection in Jinan, Northern China

Our study investigated the epidemiology of respiratory viruses in adult patients with upper respiratory tract infection (URTI) between August 2009 and September 2010 in Jinan, northern China. Nasal and throat swabs (n=596) were collected from adult patients with URTIs. Nine respiratory-related viruses, including IFV, PIV, HRV, HMPV, HBoV, HCoV, ADV, RSV, and EV, were detected in all samples by conventional and reverse transcription polymerase chain reactions. Positive detection rate for respiratory virus was 38.76% and codetection rate was 4.70% in adults with acute respiratory tract infections. IFV (20.81%) was the dominant agent detected and IFVB had a higher incidence (12.58%) than IFVA (7.72%). Detection rates of 8.22%, 5.03%, 3.69%, and 2.52% were observed for HBoV, HRV, EV, and RSV, respectively. HCoV had the lowest detection rate of 0.50%. HBoV, HRV, EV, and ADV infection rates were higher in the 14–25-year-old group than in the 26–65-year-old group. Codetection rates were higher (7.52%) in the 14–25-year-old group than in the older age group (2.64%). The spectrum of respiratory virus infection in adult patients with URTIs was different in Jinan compared with other cities in China