Objective To isolate the microsatellite DNA of Phlebotomus chinensis and screen the polymorphic loci.Methods Genomic DNA fragments were hybridized with biotinylated oligonucleotide probes(AAT)17,(GA)25,(CCT)17,and(TG)18.The hybridized fragments were captured with Vectrex Avidin D and enriched by centrifugal ultrafiltration with ultra-4-column ultrafiltrate.The target fragments were amplified,cloned and sequenced.The suitable microsatellite loci were chosen in Ph.chinensis's library to establish PCR amplification assay.The polymorphism screening was conducted by PAGE with Ph.chinensis field specimens.Results The enrichment protocol used in this study was efficient,with a percentage of recombinant clone as 78.6%.There were 118 microsatellite sequences in library,the GenBank accession numbers were from FJ919812 to FJ919932(except GenBank accession numbers FJ919833,FJ919836,and FJ919869).There were 72 typical microsatellite sequences occupying 61.0% and the rest were 46 nontypical microsatellite sequences in the library.Twenty-two loci were chosen to polymorphism screening and PAGE showed that 14 loci were polymorphic.The loci of dinucleotide repeat were more polymorphic than those of trinucleotide and polynucleotide repeat.Conclusion The microsatellite-containing library of Ph.chinensis has been constructed with 118 sequences,and 14 new polymorphic microsatellite loci are reported.

To study the induced condition and characteristics of T cell anergy in vitro. Methods: Anergic Tcell was induced by combination of B7-1 mAb and cyclosporin A (CsA) in vitro, cytokine gene of anergic T cells was detected by RT-PCR. Results: T cell anergy was antigen-specific. The state of T cell anergy can be reversed by PHA, CD3 mAb and PMA plus A23187. IL-2 can prevent the induction of T cell anergy, but it can not reverse the state of un-responsiveness. IL-2 and IFN mRNA can not express in anergic T cells. In contrast, IL-4 and IL-10 mRNA were detectable. Conclusion: T cell anergy can be induced in vitro , cytokine profile of anergic T cells deviated to Th2-like phe-norype.

To compare the chemical change of Paeoniae Radix Alba (PRA) after vinegar-baking processing, as well as the effect of vinegar types exerted on the processing, 1H NMR-based metabolomic approach combined with multivariate statistical analysis was used to investigate the different metabolites between the raw and two vinegar-baked PRA. More than thirty metabolites were identified in the 1H NMR spectrum of PRA, and the multivariate statistical analysis showed that raw and two vinegar-baked PRA could be separated obviously. After vinegar-baking, the contents of isoleucine, lactate, alanine, arginine, albiflorin, and 5-hydroxymethyl furfural (5-HMF) elevated, while those of sucrose, paeoniflorin and its analogues (calculated by benzoate) decreased. The chemical compositions of two vinegar-baked PRA were also different. Shanxi vinegar- baked PRA showed higher levels of leucine, isoleucine, valine, and albiflorin, while rice vinegar-baked PRA contained more sucrose and paeoniflorin's analogues (calculated by benzoate). And the chemical changes in Shanxi vinegar-baked PRA were greater than those of rice vinegar-baked PRA. The results revealed the chemical differences between raw and vinegar-baked PRA, as well as the influence of vinegar type on processing, in a holistic manner, the results obtained suggested that the correlations between the chemical change and the drug action after processing, as well as the vinegar type used in processing, should be further studied.
Acetic Acid ; Benzoates ; Bridged-Ring Compounds ; Cooking ; Drugs, Chinese Herbal ; Furaldehyde ; analogs & derivatives ; Glucosides ; Metabolome ; Metabolomics ; Monoterpenes ; Paeonia ; chemistry ; Plant Roots ; chemistry

Objective To investigate the effects of ropivavaine on gamma -aminobutyric acid(GABA)-activated membrane cur-rents in isolated dorsal root ganglion(DRG) neurons of the rats with ischiadic nerve chronic constriction injury (CCI) and to discuss the possible analgesia mechanism of ropivacaine .Methods The whole-cell patch clamp technique was used to record and compare the changes of GABA receptor activation currents of acute isolated DRG neurons after 30 s of ropivacaine preperfusion in the oper-ating side and the operative opposite side of the CCI model rats and the sham-operation group .Results (1)Compared with the oper-ative opposite side ,the sham-operation group and the control group ,the thermal withdrawal latency in the operative side group of the CCI model rats was notablely shortened(P<0 .05);(2)the amplitude of GABA-activated currents with different concentration GABA(0 .1-1 000μmol/L) in the operative opposite side group of the CCI operation was significantly greater than that of the op-erative side group and the sham-opeartion group ;(3)DRG neurons after ropivacaine preperfusion (0 .1-1 000μmol /L) showed va-rying degrees of enhancement effect on the 100 μmol/L GABA-activated currents ,the enhancement amplitude in the CCI operative opposite side group was significantly greater than that in the operative side group and the sham-operation group ;(4)The dose-re-sponse curve of DRG neurons GABA (0 .1-1 000μmol/L) activated current in the operative side group of the CCI rats after ropiva-caine pre-perfusion (100 μmol/L) was shifted to the left ,the difference between two EC50 had no statistical significance (P>0 .05) .Conclusion Ropivacaine has the enhancement effect on GABA activated currents in the DRG neurons of the CCI model rats , which could be one of reasons for ropivacaine producing the anesthetic and analgesic effect .

Objective To evaluate the efficacy of fiberoptic bronchoscope (FOB)-guided tracheal intubation with laryngeal mask airway i-gel (LMA i-gel) in patients undergoing cervical spine surgery. Methods Forty ASA Ⅰ or Ⅱ patients, aged 36-62 yr, weighing 57-78 kg, scheduled for cervical spine surgery under general anesthesia, were randomly divided into 2 groups (n = 20 each): FOB-guided tracheal intubation with oropharynx ventilation tube group (group O) and FOB-guided tracheal intubation with LMA i-gel (group I). Anesthesia was induced with midazolam 0.05 mg/kg, propofol 2 mg/kg, fentanyl 2-3 μg/kg and rocuronium 0.9 mg/kg. The intubation time, fiberoptic bronchoscope score, the number of successful intubation, hypertension, tachycardia and hypoxemia were recorded. All the patients were followed up postoperatively for adverse effects like sore throat or hoarseness, etc. Results The rate of successful LMA i-gel placement at first attempt was 100%, placement time was (10 + 3) s, and the rate of successful intubation in the two groups was 100%. The intubation time was significantly shorter, the rate of successful intubation at first attempt and fiberoptic bronchoscope score were significantly higher in group I than in group O (P ＜ 0.05). Hypertension, tachycardia and hypoxemia were not found in the two groups. There was no significant difference in the incidence of adverse effects between the two groups (P ＞0.05). Conclusion FOB-guided tracheal intubation with LMA i-gel can provide adequate ventilation during operation, improve the success rate of FOB-guided intubation and shorten the intubation time in patients undergoing cervical spine surgery.

The main objective of this research is to investigate the effects of astragaloside IV, calycosin separately glucoside, formononetin on oxidative stress in Chang Liver cells induced by H2O2. In the experiments, Chang Liver cells (a kind of normal human hepatocytes) were used as the research object, bifendate which has a clear hepatoprotective effect was used as the positive control drug, then the oxidative damage model of Chang Liver cells were established by H2O2. Cells were divided into six groups: blank control group, oxidative stress group, astragaloside IV group, calycosin separately glucoside group, formononetin group and positive control group. Then endogenous antioxidant system related indexes were detected by micro plate and colorimetric method; intracellular reactive oxygen species (ROS) were detected by DCFH-DA fluorescent probe; and the expressions of CYP2E1 were evaluated by liver microsomes, mRNA, and protein, respectively with spectrophotometry, Real-time PCR method, and Western blot technique. Results showed that H2O2 decreased antioxidant activity, and increased ROS level and expression of CYP2E1. The above oxidative stress status had been changed with protections of the three components of Astragalus membranaceus (compared with oxidative stress group, P < 0.05, P < 0.01), which taken as a whole had equivalent effects as the drug of positive control group( bifendate). Taken together, three Astragalus membranaceus ingredients all had significant or extremely significant inhibiting effects on oxidative damaged Chang Liver cells which were induced by H2O2, and the oxidative damage of Chang Liver cells had been relieved.
Astragalus membranaceus ; chemistry ; Cells, Cultured ; Cytochrome P-450 CYP2E1 ; metabolism ; Humans ; Isoflavones ; pharmacology ; Liver ; drug effects ; Oxidative Stress ; drug effects ; Reactive Oxygen Species ; metabolism ; Saponins ; pharmacology ; Triterpenes ; pharmacology

To investigate the mechanism of macrophages apoptosis induced by stressed Salmonella typhi,macrophages were co-cultivate with inhibitors caspase 3,8,9 and anti-TNF-α antibody and then S.typhi was added to construct the infection model..The rate of macrophage apoptosis was assayed by flow cytometry and the contens of caspase 3.8 ,9 and anti-TNF-α antibody as well as NO were then determined respectively.It was found that the apoptotic rates of macrophages were significantly inhibited by caspase 3 and caspase 8 inhibitors and antibody against TNF-α respectively (P<0.01).A significantly enhanced generation of caspase 3 and caspase 8 activities during macrophage apoptosis induced by S.typhi correlated with the increased generation of TNF-αand NO (P<0.01).These results indicate that the inactive NO and TNF-α mediated inhibitors caspase 3 and caspase 8 participate the exogenous apoptotitic pathway of macrophages induced by S.typhi.

Objective To detect beta-human papilloma virus (HPV) and determine its genotype in actinic keratosis (AK) lesions.Methods Tissue specimens were collected from the lesions of 39 patients with AK and normal skin of 40 healthy controls.A nested PCR was performed to detect alpha-HPV and beta-HPV DNA in these specimens.The genotype of beta-HPV was determined in beta-HPV DNA-positive specimens by a common PCR using specific primers targeting 12 HPV genotypes,including HPV 5,8,15,17,19,20,21,23,36,38,49 and 80.Results The detection rate of beta-HPV DNA was 84.6％ (30/39) in the patients with AK,and 30.0％ (12/40) in the healthy controls (x2 =6.76,P ＜ 0.05),while no significant difference was observed in the detection rate of alpha-HPV DNA between the two groups (12.8％ vs.7.5％,x2 =0.91,P ＞ 0.05).HPV 38 was the predominant genotype of beta-HPV in these patients with a detection rate of 36％ (12/33),followed by HPV36.The prevalence of all the 12 genotypes of HPV was consistently low in the healthy controls.Mixed HPV infections were observed in 10 AK lesions,but in none of the healthy controls.No statistical difference was noted in the positivity rate of beta-HPV among patients at different ages,of different genders,with different occupations or clinical courses (x2 =0.53,0.94,0.81,0.73,respectively,all P ＞ 0.05).Conclusions Compared with healthy controls,the patients with AK showed a higher beta-HPV infection rate,with HPV38 as the predominant genotype.

Objective To study the damage of rat articular cartilage induced by different doses of T-2 toxin, and to explore the relationship between mini-dose T-2 toxin and articular cartilage damage. Methods A total of 120 Wistar rats, weighing 50 - 70 g, were randomly divided into four groups according to their body weights: T-2 toxin group 0(control), 100, 200, 300 μg/kg, 30 rats in each group. Animals in the control group were fed standard rat chow, and animals in the three T-2 toxin groups were fed T-2-toxin-contaminated chow (the dose was 100, 200, 300 μg/kg, respectively). After 6 months, rats were euthanized by ether asphyxiation. The bilateral knee joints were collected and section prepared. The articular cartilage was examined by light and electronic microscope. Results Light microscope showed, the rat articular chondrocytes were clear and arranged orderliness in the control group. The rat articular chondrocytes were disarranged in 100 μg/kg T-2 toxin group.Degeneration and necrosis were found in 200 μg/kg group. Chondrocytes were shrunken with hypereosinophilia cytoplasm and fragmented pyknotic nuclei, extensive areas of chondrocyte loss and chondrocyte clones were visible in 300 μg/kg group. Scanning electronic micrograph(SEM) showed, the rat articular chondrocytes were clear, well formed and arranged tidy in the control group. The surface of articular cartilage was rough in 100 μg/kg group.Collagen fasciculi ruptured and stacked up in 200 μg/kg group. Presented a typical articular dryness phenomenon,the cartilage surface collapsed and many pits appeared in 300 μg/kg group. Transmission electronic microscope (TEM) showed that chondrocytes were abundant with cytoplasm, well-developed rough endoplasmic reticulum in the control group; agglomerate chromatin scattered along the karyotheca, nuclear membrane was thickening, with vacuolar degeneration of the endoplasmic reticulum in the 100 μg/kg group; endoplasmic reticulum expended, with protein retention and organelles breaks in the 200 μg/kg group. A large number of chondrocytes lost organdles, the membrane structures disrupted and the cartilage matrix stromatolyzed in the 300 μg/kg group. Conclusions Within the range of 100 - 300 μg/kg, T-2 toxin induces dose-related articular cartilage injury, the greater the dose, the more serious damage.

Objective This study evaluated the effects of combined procedure of surgical treatment and cell transplantation on dilated cardiomyopathy. Methods Eight patients (5 men and 3 women) with moderate to severe mitral regurgitation from end-stage dilated cardiomyopathy underwent surgery. Four patients were in functional class (FC) Ⅳ, six were in FC Ⅲ. There age ranged from 15 to 56 years. The preoperative ejection fraction (EF) ranged from 0.15 to 0.32 (mean 0.26 ± 0.08). Mitral valve replacement was performed in 5 patients and mitral valve repair in 3. Auto-bone marrow monenuclear cells were harvested, isolated, washed, and resuspended for direct injection after surgical procedure. Results All patients survived and were discharged from the hospital. After a mean follow-up period of 18 months ( 12 - 42 months). Echocardiography showed postoperative ejection fraction and wall movement velocity increased after 6 months. Radionuclide ventriculography showed myocardial perfusion improved significantly. Conclusion Combined procedure of surgical treatment and cell transplantation led to significant improvement in cardiac function in patients with dilated cardiomyopathy.