BACKGROUND: Chitosan is one of the most significant non-viral vector materials with the advantages of outstanding biocompatibility. Quarternary chitosan derivatives can improve transfection efficiency and solubility of chitosan in a broader range of pH values. OBJECTIVE: To synthesize a new vector of quarternary chitosan and to study its complex conditions with plasmid and transfection efficiency compared with chitosan. DESIGN, TIME AND SETTING: A contrast observational study was performed in Second Affiliated Hospital of School of Medicine of Zhejiang University between August and October 2008. MATERIALS: Quarternary chitosan was synthesized in Polymer Materials Lab of Beijing Institute of Technology. Plasmid pEGFP-C1 was presented friendly by Mr. Zheng of School of Medicine of Zhejiang University. Hela cells were provided by Miss. Cheng of School of Medicine of Zhejiang University. METHODS: Quarternary chitosan was prepared according to mass concentration of 0.2 g/L, pH value 5.5 (or 6.9, 7.6) and sodium acetate concentration of 50 mmol/L, and rapidly mixed with pEGFP-C1. The mixture was swirled for 15-30 second and stood at room temperature for 30 minutes at least. MAIN OUTCOME MEASURES: The impacts of pH values and time on complex ability of quarternary chitosan and plasmid were studied by gel retardation test. Transfection efficiency of quarternary chitosan on Hela cells was observed by inversed fluorescence microscope and also compared with chitosan. RESULTS: Quarternaty chitosan could form complex with plasmid in acidic, neutral and basic conditions. It could be used in a broader range of pH values. In an acidic condition, the combination of quarternary chitosan with plasmid was superior to chitosan. A stable complex was formed via a combination of quarternary chitosan or chitosan with plasmid within 30 minutes, and the stability lasted for 12 hours. Transfection efficiency of quarternary chitosan on Hela cells demonstrated that transfection efficiency of quarternary chitosan was superior to chitosan. CONCLUSION: Quarternary chitosan has a broader range in use and higher transfection efficiency than chitosan; however, there is no significant difference in stability between quarternary chitosan and chitosan. Additionally, transfection efficiency of quarternary chitosan on Hela cells is superior to chitosan, which needs a further research.

Adolescent ; Adult ; Antigens, Nuclear ; metabolism ; Brain Diseases ; complications ; diagnosis ; pathology ; surgery ; Brain Neoplasms ; pathology ; Child ; Child, Preschool ; Diagnosis, Differential ; Epilepsy ; etiology ; Female ; Ganglioglioma ; pathology ; Glial Fibrillary Acidic Protein ; metabolism ; Humans ; Infant ; Magnetic Resonance Imaging ; Male ; Malformations of Cortical Development ; classification ; complications ; diagnosis ; pathology ; surgery ; Malformations of Cortical Development, Group I ; Microtubule-Associated Proteins ; metabolism ; Neoplasms, Neuroepithelial ; pathology ; Nerve Tissue Proteins ; metabolism ; Neurofilament Proteins ; metabolism ; Retrospective Studies ; Vimentin ; metabolism ; Young Adult

OBJECTIVETo investigate the effects and mechanisms of high glucose on the phenotype transformation of rat vascular smooth muscle cells (VSMCs).

METHODSVSMCs ere isolated from rat thoracic aorta and the 3rd-5th VSMCs were incubated with normal glucose (5.5 mmol/L), high glucose (25 mmol/L), or high glucose (25 mmol/L) + P38 inhibitor (25 mmol/L +SB203580) for another 24 hours. Then the gene expression of osteopontin (OPN), alpha smooth-actin (alpha-SMA), matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9(MMP-9) were assayed by real time RT-PCR, the protein expression of P38 were assayed by Western blot.

RESULTS(1) High glucose promoted the phenotype transformation of VSMCs and up-regulated the expression of MMP-2 and MMP-9. (2) High glucose promoted the phosphorylation of P38. (3) SB203580, the inhibitor of P38/MAPK signal pathway, inhibited the effects of high glucose on phenotype transformation and expression of MMP-2 and MMP-9.

CONCLUSIONHigh glucose may promote phenotype transformation of VSMCs via the signal pathway of P38/MAPK.

Actins ; metabolism ; Animals ; Aorta, Thoracic ; cytology ; Blotting, Western ; Cells, Cultured ; Glucose ; pharmacology ; Imidazoles ; pharmacology ; MAP Kinase Signaling System ; Matrix Metalloproteinase 2 ; Matrix Metalloproteinase 9 ; metabolism ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; cytology ; drug effects ; Osteopontin ; metabolism ; Phenotype ; Pyridines ; pharmacology ; Rats ; p38 Mitogen-Activated Protein Kinases ; metabolism

OBJECTIVETo observe therapeutic effects of bird-pecking moxibustion in children of hand, foot and mouth disease, and to analyze the mechanism.

METHODSSeventy-five children of hand, foot and mouth disease were randomly divided into 3 groups, a combined moxibustion and medicine group (n = 22), a Chinese medicine group (n = 29), and a western medicine group (n = 24). The combined moxibustion and medicine group was treated with bird-pecking moxibustion combined with routine western medicine, the Chinese medicine group with oral administration of Chinese medicine and routine western medical therapy, and the western medicine group with routine western medicine. After treatment of 7 days, the therapeutic effects on skin rash, oral herpes, constipation or loose stool, dyspepsia and anorexia, etc. were comprehensively assessed, and their therapeutic effects were compared.

RESULTSThe total effective rate was 95.5% in the combined moxibustion and medicine group, 86.2% in the Chinese medicine group, and 83.3% in the western medicine group, the former being significantly better than those of the other two groups (both P < 0.05). The combined moxibustion and medicine group was significantly better than the other two groups in the relieving time of skin rash, oral herpes, constipation or loose stool, dyspepsia and anorexia, etc. (all P < 0.05)

CONCLUSIONThe combined moxibustion and medication can effectively improve symptoms of the digestive tract, shorten duration of disease, reduce pain in the patient with hand, foot and mouth disease.

Child ; Child, Preschool ; Combined Modality Therapy ; Drug Therapy ; methods ; Drugs, Chinese Herbal ; administration & dosage ; therapeutic use ; Female ; Hand, Foot and Mouth Disease ; pathology ; therapy ; Humans ; Infant ; Male ; Moxibustion ; methods ; Treatment Outcome

OBJECTIVEA novel multiplex real-time RT-PCR kit was developed to detect EV71, CoxA16 and other human enteroviruses simultaneously with an internal amplification control to avoids false negatives, which used for hand, foot and mouth disease in the clinical diagnosis and epidemiological surveillance.

METHODSDesign specific primers and probes of EV71, CA16, other intestinal virus and internal amplification control, improve the extraction method of virus nucleic acid. Optimization the detection system of real-time quantitative PCR. Research the products of the accuracy, stability, precision, amplification efficiency and detection of linear range.

RESULTSThe primers and probes had high spicificity. The Viral RNA extraction effect of this Kit is as same as that of QIAamp Viral RNA mini Kit (QIAGEN company), but less reagent cost. The optimal concentrations of primers and probes are 0.2 micromol/L for all the upstream and downstream primers, 0.06 micromol/L for probes of other human enteroviruse, 0.08 micromol/L for probes of EV71 and CA16 respectively. The kit has good stability, accuracy and precision. The amplification efficiencies of EV71, CoxA16 and other human enteroviruses are 106% ,101% and 105% and the detection of linear range is from 10(9) copies/microl-10(2) copies/microl.

CONCLUSIONThe novel multiplex real-time RT-PCR kit for detecting EV71, CoxA16 and other human enteroviruses simultaneously with an internal amplification control has good stability, accuracy, precision and amplification efficiencies. So it has great value in clinical application.

Enterovirus ; isolation & purification ; Humans ; Reagent Kits, Diagnostic ; Real-Time Polymerase Chain Reaction ; methods

BACKGROUNDIt has been reported that the nicotinic acetylcholine receptor subunit α4 gene (CHRNA4) might be associated with smoking behaviors in the previous studies. Up to now, there are few reports on the relationship between CHRNA4 and smoking initiation. In this study, we tried to explore the role of two polymorphisms in CHRNA4 (rs1044396 and rs1044397) in smoking initiation and nicotine dependence in Chinese male smokers.

METHODSNine hundred and sixty-six Chinese male lifetime nonsmokers and smokers were assessed by the Fagerström test for nicotine dependence (FTND), smoking quantity (SQ) and the heaviness of smoking index (HSI). All subjects were divided into four groups based on their tobacco use history and the FTND scores. The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was performed to find two polymorphisms of CHRNA4 in these subjects.

RESULTSThe χ(2) test showed that rs1044396 was significantly associated with smoking initiation (χ(2) = 4.65, P = 0.031), while both rs1044396 and rs1044397 were significantly associated with nicotine dependence (χ(2) = 5.42, P = 0.020; χ(2) = 7.58, P = 0.005). Furthermore, the T-G (3.9%) haplotype of rs1044396-rs1044397 showed significant association with smoking initiation (χ(2) = 6.30, P = 0.012) and the C-G haplotype (58.9%) remained positive association with nicotine dependence (χ(2) = 8.64, P = 0.003) after Bonferroni correction. The C-G haplotype also significantly increased the HSI (P = 0.002) and FTND scores (P = 0.001) after Bonferroni correction.

CONCLUSIONThese findings suggest that CHRNA4 may be associated with smoking initiation and the C-G haplotype of rs1044396-rs1044397 might increase the vulnerability to nicotine dependence in Chinese male smokers.

Adult ; Aged ; Asian Continental Ancestry Group ; genetics ; Genetic Predisposition to Disease ; genetics ; Humans ; Male ; Middle Aged ; Polymorphism, Genetic ; genetics ; Receptors, Nicotinic ; genetics ; Smoking ; adverse effects

AIMTo approach the relationship between lung injury induced by shock/reperfusion and nitric oxide as well as the beneficial effect of taurine.

METHODSTwenty four rabbits were divided randomly into 3 groups (n = 8): control group, shock group, taurine group. The model of lung injury induced by shock/reperfusion was used. The activities of nitric oxide synthase (NOS), superoxide dismutase (SOD), the contents of malondialdehyde (MDA), nitric oxide products (NO2-/NO3-) in plasma and lung homogenate, lung wet/dry weight, lung water content, lung permeability index, and protein content in the pulmonary alveolar lavage fluid were measured. Meanwhile, pathologic samples treated routinely.

RESULTS(1) At 3 hours after reperfusion, the activities of SOD in plasma and lung homogenate decreased markedly, but the other indexes above mentioned were increased significantly compared with the control group (P < 0.01). (2) A close correlation was shown between MDA content and NO2-/NO3- content in plasma and lung. Furthermore, the content of NO2-/NQ3- in lung homogenate showed strong positive correlation with the lung injury parameters. (3) Taurine (40 mg x kg(-1) i.v.) could attenuate all the changes above mentioned at the same time points of reperfusion.

CONCLUSIONNO may play an important role in lung injury induced by shock/reperfusion. Taurine can ameliorate the lung injury, mechanism of which may be related to decreasing the generation of NO and anti-lipoperoxidation.

Animals ; Lung ; drug effects ; metabolism ; Lung Injury ; etiology ; prevention & control ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase ; metabolism ; Rabbits ; Reperfusion Injury ; complications ; drug therapy ; Shock, Hemorrhagic ; complications ; drug therapy ; Superoxide Dismutase ; metabolism ; Taurine ; pharmacology ; therapeutic use

AIMTo investigate the effect of taurine on nitric oxide synthase (NOS) activity and nitric oxide products (NO2 /NO3 ) content in myocardium and plasma during shock resuscitation.

METHODSTwenty-four rabbits were divided randomly into 3 groups (n=8): control group, shock group, taurine group. The model of hemorrhagic shock resuscitation was used. The activities of nitric oxide synthase (NOS), lactate dehydrogenase (LDH) and the contents of nitric oxide products (NO2- /NO3-) in plasma were observed before shock and shock 1.5 hours, after resuscitation 1 hour, 2 hours and 3 hours. The activities of NOS and the contents of NO2-/NO3- in myocardium homogenate were measured after resuscitation 3 hours. Meanwhile, pathologic samples treated routinely.

RESULTS(1) During resuscitation, the activities of NOS, LDH and the contents of NO2- /NO3- in plasma of shock group were significantly higher than that of before shock and shock 1.5 hours (P < 0.01). (2) After resuscitation 3 hours, the activity of NOS and the contents of NO2- / NO3 in myocardium of shock group were significantly higher than that of control group (P < 0.01). The cardiac myocyte appeared edema, fatty degeneration. (3) All the changes of above mentioned could be attenuated by intravenous injection taurine (40 mg/kg) (P < 0.01).

CONCLUSIONThese results suggest that the NOS activation and NO release may mediated myocardium injury induced by shock resuscitation, taurine can ameliorate the myocardium injury, which may be related to decreasing the generation of NO.

Animals ; Myocardium ; metabolism ; pathology ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase ; metabolism ; Plasma ; metabolism ; Rabbits ; Resuscitation ; Shock, Hemorrhagic ; blood ; metabolism ; Taurine ; pharmacology

OBJECTIVE: To study the clinical effect of calsurf, a domestic exogenous pulmonary surfactant, in the treatment of severe neonatal infectious pneumonia. METHODS: A total of 208 neonates with severe infectious pneumonia who hospitalized in 5 hospitals of China were enrolled. According to their parents' wishes on admission, these neonates were administered with conventional treatment (control group; n=81) and calsurf  treatment + conventional treatment (calsurf treatment group, n=127). The two groups were compared in terms of the degree of oxygen dependence on admission, blood gas parameters before and after treatment, lung ultrasound results, duration of mechanical ventilation, length of hospital stay, hospital costs, complications and prognosis. RESULTS: Compared with the control group on admission, the calsurf treatment group had significantly higher inhaled oxygen concentration and partial pressure of carbon dioxide and significantly lower arterial partial pressure of oxygen and oxygenation index (P<0.01). After 1 hour of treatment, both groups had significant improvements in the above indices (P<0.05), and the improvements were more significant in the calsurf treatment group (P<0.05). After 4-6 hours of calsurf administration, there was a significant reduction in the degree of pulmonary consolidation. The calsurf treatment group had significantly shorter duration of mechanical ventilation and length of hospital stay than the control group, while there was no significant difference in the incidence rate of complications between the two groups. The neonates of both groups had a good prognosis. CONCLUSIONS In neonates with severe infectious pneumonia, calsurf treatment can significantly improve oxygenation, reduce the degree of pulmonary consolidation, and shorten the duration of mechanical ventilation and length of hospital stay. Therefore, it should be considered in neonates with severe infectious pneumonia.
China ; Humans ; Infant, Newborn ; Pneumonia ; Prospective Studies ; Pulmonary Surfactants ; Respiration, Artificial

OBJECTIVETo investigate the effects of endothelia progenitor cells conditioned medium (EPC-CM) on the migration, adhesion and proliferation of vascular smooth muscle cells (VSMCs).

METHODSMononuclear cells were isolated from rat bone marrow by density gradient centrifugation,plated on dishes precoated with 5% fibronectin, and then cultured with complete M199 medium (including 15% fetal calf serum, 10 microg/L VEGF and 5 microg/L bFGF). EPC-CM was collected and used to incubate VSMCs isolated from rat arteriae aorta. After 24 h, VSMCs proliferation, adhesion and migration were assayed with CCK-8, adhesion test and modified Boyden chamber assay, respectively.

RESULTSThe proliferation, adhesion and migration of VSMCs were obviously decreased when the cells were cultured with EPC-CM.

CONCLUSIONEPC-CM could inhibit VSMC functions, which would be one of the mechanisms against atherosclerosis by EPCs.

Animals ; Cell Adhesion ; Cell Movement ; Cell Proliferation ; Cells, Cultured ; Culture Media, Conditioned ; chemistry ; Endothelial Cells ; cytology ; Male ; Muscle, Smooth, Vascular ; cytology ; Rats ; Rats, Sprague-Dawley ; Stem Cells ; cytology