[Construction and identification of a novel adeno-integrase hybrid system for hemophilia B].

This study was aimed to construct an adenovirus hybrid system with high transduction efficiency and site-specific integration. By a series of DNA manipulation, a hybrid system of two adenovirus vectors was constructed. One vector contains loxP-flanked transgene expression cassette, in which there are hFIX and DsRed coding sequences and attB for phiC31 recolonization. The other vector carries Cre and phiC31 gene. Vectors only expressing Cre or phiC31 were used as controls. 293A cells were constructed and transfected with the adenoviral vectors by Lipofectamine 2000, and the expression of target genes was identified by fluorescence microscopy and RT-PCR. The results showed that after being identified by PCR, restriction analysis and sequencing, an adeno-integrase hybrid system was successfully constructed. The system expressed RFP, GFP, hFIX, Cre and phiC31 in 293A cells in vitro. It is concluded that the adeno-integrase hybrid system is successfully constructed, which lays a good foundation for further investigation of its therapeutic application.

Construction and identification of a novel adeno-integrase hybrid system for hemophilia B / 中国实验血液学杂志

This study was aimed to construct an adenovirus hybrid system with high transduction efficiency and site-specific integration. By a series of DNA manipulation, a hybrid system of two adenovirus vectors was constructed. One vector contains loxP-flanked transgene expression cassette, in which there are hFIX and DsRed coding sequences and attB for phiC31 recolonization. The other vector carries Cre and phiC31 gene. Vectors only expressing Cre or phiC31 were used as controls. 293A cells were constructed and transfected with the adenoviral vectors by Lipofectamine 2000, and the expression of target genes was identified by fluorescence microscopy and RT-PCR. The results showed that after being identified by PCR, restriction analysis and sequencing, an adeno-integrase hybrid system was successfully constructed. The system expressed RFP, GFP, hFIX, Cre and phiC31 in 293A cells in vitro. It is concluded that the adeno-integrase hybrid system is successfully constructed, which lays a good foundation for further investigation of its therapeutic application.

Serum enzyme profile characteristics of victims following the Wenchuan earthquake in China.

BACKGROUND: Earthquakes are major causes of morbidity and mortality. The Wenchuan region of China was devastated by a catastrophic earthquake on May 12, 2008, at 02:28 p.m. (Beijing time), registering magnitude 8.0 on the Richter scale and causing more than 69,181 deaths. As a first-line general hospital in the disaster area, Mianyang Central Hospital admitted a large number of the victims. METHODS: A total of 534 victims (246 males, 288 females) were categorized as non-crush injury patients (n=239), simple crush injury patients (n=136), and crush syndrome patients (n=69) according to their traumatic conditions. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyltransferase (GGT), lactate dehydrogenase (LDH), alkaline phosphatase (ALP), cholinesterase (CHS), and creatine kinase (CK) levels were measured. RESULTS: ALT, AST, LDH, CHS, and CK levels showed significant differences among the three groups by one-way analysis of variance (ANOVA). Pearson correlation analysis showed that correlative changes between any two of the following: ALT, AST, GGT, ALP, LDH, and CHS were similar among three groups, with the following exceptions. The correlation coefficients of ALT-GGT, AST-GGT, and ALP-CHS changed from positive to negative values, and ALP-LDH changed from a negative value to a positive value. Receiver-operating characteristic (ROC) curve analysis showed the highest diagnostic effectiveness of 99.4% for CK, with 100% specificity [positive predictive value (PPV)=100%] and 99.4% sensitivity [negative predictive value (NPV)=99.0%] in distinguishing crush injury (including crush syndrome) from non-crush injury. AST had the best diagnostic effectiveness in distinguishing crush syndrome from crush injury; 53.8%, with 85.5% specificity (PPV=64.4%) and 77.9% sensitivity (NPV=90.7%). Multivariate logistic analysis revealed that CK was best at distinguishing crush injury (including crush syndrome) from non-crush injury (OR 409.636, 95% CI 382.96-438.17), and AST was best for distinguishing crush syndrome from crush injury (OR 50.08, 95% CI 46.84-53.55). CONCLUSIONS: Crush injury and crush syndrome are severe in victims following accidents or natural catastrophes. Serum CK, LDH, AST, ALT, GGT, and ALP activities were all helpful biochemical parameters in estimating the severity of crush injury and/or crush syndrome and preventing the development of further complications.

Pharmacokinetics of recombinant human endostatin in rats.

The pharmacokinetics of recombinant human endostatin (rh-Endo) has not been established in the rat, although this species of animal is commonly used in the pharmacological studies of rh-Endo. This study aimed to investigate the pharmacokinetics, tissue distribution, and excretion of rh-Endo in rats. 125I-radiolabeled rh-Endo was administered to healthy rats by intravenous (i.v) bolus injection at 1.5, 4.5 and 13.5 mg/kg. The maximum plasma concentration (Cmax) and area under the plasma concentration versus time curve (AUC) of rh-Endo increased proportionally with the increase of the dosage. There were no significant differences in total body clearance (CL) and elimination half-life (t1/2beta) of rh-Endo among the three dosages used. A 93.5% and 2.2% of the radioactivity was recovered in the urine and feces, respectively, in bile-duct intact rats; whereas only 0.1% of the total radioactivity was excreted into the bile in bile-duct cannulated rats. rh-Endo was rapidly and widely distributed in the liver, kidneys, spleen and lungs. Furthermore, a significant allometric relationship between CL, but not volume of distribution (Vd) and t1/2beta of rh-Endo, and the body weight was observed across mouse, rat and monkey, with the predicted values in humans significantly lower than those observed in cancer patients. rh-Endo exhibited a linear pharmacokinetics in rats and it is mainly excreted through the urine.