Objective: To construct a recombinant lentivirus harboring RNAi sequence targeting rat nogo receptor gene and to observe its infection efficiency of 293T cells. Methods: Lentivirus shuttle plasmid containing siNgR199 cDNA was constructed by gene engineering and was used to transfect 293T cells in the presence of packaging plasmids. Forty-eight hours later the supernatant was collected and the titer of virus was determined. The recombinant lentivirus and the standard lentivirus were used to transfect 293T cells at 1 MOI,3 MOI,5 MOI,10 MOI and 20 MOI. Polymerase chain reaction (PCR) was used to detect the recombinant vector; enhanced green fluorescent protein (EGFP) expression was used to determine the titer and the infection rate of the recombinant lentivirus under fluorescent microscope. Results: Restriction endonuclease and PCR analysis confirmed that the siNgR199 cDNA was successfully inserted into the lentivirus vector. The titer of the recombinant lentivirus harboring siNgR199 was 1×108 ifu/ ml and the best MOI was 3. Conclusion: The recombinant lentivirus containing siNgR199 gene has been successfully constructed, which lays a foundation for future axon regeneration in treatment of spinal cord injury.

The effects of ketamine on the hemodynamics in the postshock stage in 10 severely burn patients were observed.Their average burn area was 53.7?14% TBSA.Anesthesia was inducedwith 2 mg/kg of ketamine and then maintained with 50?g/kg/minute of ketamine.All the patients kept on spontaneous breathing.Measurements were performed before anesthesia,5 and 30 minutes after the induction of anesthesia.and 20 minutes after the discontinuation of ketamine.Our findings indicate that the severely burn patients in the postshock stage were in a state of active hemodynamics with increased cardiac output and decreased peripheral resistance.The plasma concentrations of epinephrine and norepinephrine were apparently increased.A small dose of ketamine could further increase the release of epinephrine,which is beneficial to the restoration of normal cardiovascular functions.

Objective To establish a quality standard for Biqing Suppository. Methods Rhizoma Dioscoreae Septemlobae, Rhizoma Acori Tatarinowii, Radix Et Rhizoma Salviae Miltiorrhizae and Cortex Phellodendri Chinensis were qualitatively identified by TLC method. The content of berberine hydrochloride in Biqing Suppository was determined by HPLC method. The chromatographic column was Waters C18 (250 mm×4.6 mm, 5 μm);the mobile phase was acetonitrile∶water (0.2%phosphoric acid-0.02%three triethylamine)=25∶75;the flow rate was 1 mL/min;the detection wavelength was 265 nm. Results The spots of qualitative identification method were clear without interference. Berberine hydrochloride had good linear relationship in the range of 0.235-9.40 μg;the regression equation was Y=40 033 998.176 3X-85 021.2, r=0.999 7;the average recovery rate was 99.16%, RSD=3.45%. Conclusion The method is simple, flexible and reproducible, which can be used as the quality control standard for Biqing Suppository.

AIM: To study the endothelial progenitor cell markers and biological characteristics of human umbilical cord blood CD133+ cells isolated by immunomagnetic bead (IMB) method. METHODS: The experiment was carried out in the Laboratory of Burn Surgery, Xijing Hospital of the Fourth Military Medical University of Chinese PLA from November 2002 to August 2003.①CD133+ cells were enriched from human umbilical cord blood of placenta through spontaneous delivery or cesarean delivery (offered by Department of Gynecology and Obstetrics, Xijing Hospital of the Fourth Military Medical University of Chinese PLA), with the informed consents of parturients.②Human umbilical cord blood was anticoagulated by ACD-B and then diluted, 7 mL of cord blood combined with 3 mL lymphocyte isolation liquid were centrifuged for 30 minutes at 2 100 r/min. Low-density mononuclear cells coloring buffy were selected to wash off platelet and inoculated for 30 minutes by the IMB and FITC-CD133 antigen at room temperature to wash excessive antigens. The FITC-CD 133-coated IMB was added into mononuclear cells which were cultured for 30 minutes at room temperature and placed in magnetic field for 1 minute to remove the cell suspension. IMB-cell isolation liquid was supplemented to release CD133+ endothelial progenitor cells, which were cultured with EGM-2MV media in dishes coated with collagen I. And 24 hours later those unadherent cells were detached, culture liquid was removed half after 3 hours, and total culture duration was 3-4 weeks.③The morphologic change of CD133+ cells isolated with IMB were observed; The percentage of CD133+ cells in cord blood monocytes and sorting efficiency of IMB were analyzed by fluorescence activated cell sorting (FACS). The cells cultured in vitro were identified by fluorescence immunoassay and enzymo-histochemistry while transmission electron microscope (TEM) was used to observe the W-P body of mature vascular endothelial cells. RESULTS: ①Morphologic observation of CD133+ cells culture in vitro: The adherent cells expanded pseudopods on days 4 and 5, and grew rapidly in clonal manner 7 days later. The cells showed a typical spindle-shaped morphology during 2-3 weeks and cluster areas of cobblestone-like cells after 2-week culture and confluence.②Isolation rate of CD133+ cells: The percentage of CD133+ cells in cord blood monocytes was 0.91%. And after IMB sorting, the percentage of CD133+ cells was 85.52%.③On the fifth day, few cells were positive for CD133 while most cells were positive for CD 34 and vWF; On the tenth day, all cells were negative for CD133 and most cells were still positive for CD 34 and vWF.④The cells after 10-day culture had typical W-P body in TEM. CONCLUSION: Immunomagnetic sorting can efficiently select enriched CD133+ endothelial progenitor cells from human umbilical cord blood. Our data of CD133, CD34, vWF antibodies and TEM support the hypothesis that endothelial progenitor cells may contribute to mature vascular endothelial cells.

Objective Aprotinin, a serine proteinase inhibitor, has been reported to reduce blood loss significantly in patients undergoing cardiac surgery with CPB, heart and liver transplantation. The aim of this study was to evaluate the effect of aprotinin on intraoperative blood loss, transfusion requirement and blood coagulation during liver cancer resection.Methods Eighty-two ASA Ⅰ -Ⅲ patients ( 51 male, 31 female ) aged 33-65 yr undergoing liver cancer resection ( 61 partial hepatectomy, 21 extirpation of liver cancer) were studied. The patients were randomly divided into 2 groups : aprotinin group received a bolus of aprotinin 1 112 EPU after induction of anesthesia, followed by continuous aprotinin infusion at 278 EPU?h-1 until 2 h after operation ( n = 40); control group received normal saline instead of aprotinin ( n = 42) . The patients were premedicated with sodium luminal, droperidol-fentanyl and atropine. Anesthesia was induced with midazolam 2 mg, thiopental 5 mg?kg-1 and succinylcholine 1.5 mg? kg-1 . After tracheal intubation the patient was mechanically ventilated (VT = 8-12 ml?kg-1 ) and PaCO2 was maintained at about 35 mm Hg, Anesthesia was maintained with N2O/O2 , fentanyl and vecuroniurn. Venous blood samples were taken before induction of anesthesia (baseline) , 0.5 h, 2 h and 4 h after skin incision and 6 h and 12 h after operation for routine blood tests, thromboelastography ( TEG), and determination of activated partial thromboplastin time (APTT), thromboplastin time (TT) prothrombin time (PT) and plasma fibrinogen concentration (Fig) . Intraoperative blood loss and amount of blood transfused were recorded. Results The preoperative hypercoagulable state was ameliorated and coagulation was maintained within the normal range in aprotinin group; while in control group the hypercoagulable state was aggravated during operation and at the end of operation it changed to hypocoagulable state. The intraoperative blood loss and amount of blood infused were significantly less in aprotinin group than in control group. Conclusion The use of aprotinin during liver cancer resection results in reduction in intraoperative blood loss and less transfusion requirement.

AIM To observe the effect of Yishenjiangzhuo decoction on the NE,5-HT contents of hippocampus in cerebral ischemia reperfusion rats. METHODS High pressure lipuid chromatography-electrochemical process was used to measure the NE and 5-HT contents on the d 1, d 7 and d 15 after cerebral ischemia reperfusion by common carotid artery occlusion. RESULTS The NE contents of hippocampus were respectively 364.25?66.47, 349.76?59.38, (344.59?70.31) ?g?g -1 and the 5-HT contents were respectively 646.72?83.33,629.11?90.64,(596.68?99.47) ?g?g -1 on the d 1, d 7 and d 15 in the model group, which significantly decreased compared with the control group( P

Objective To investigate the clinical value of serum homocysteine (Hcy),Cystatin C (CysC) detection in coronary heart disease prevention,diagnosis and treatment.Methods A total of 200 cases of coronary heart disease patients including 50 cases of acute myocardiac infarction (AMI) (AMI group),85 cases of unstable angina pectoris (UAP)(UAP group.) and 65 cases of stable angina pectoris (SAP) (SAP group) were selected,and 120 cases of healthy controls were selected as control group.The serum levels of Hcy and CysC were detected and compared.Results The serum levels of Hcy and CysC in AMI group,UAP group and SAP group were significandy higher than those in control group[Hcy:(22.35 ± 8.18),(16.54±7.56),(14.52±6.38) μmol/Lvs.(9.35±3.23) μmol/L; CysC:(1.32±0.27),(1.88± 0.66),(1.19 ± 0.46) mg/L vs.(0.80 ± 0.33) mg/L],and there were significant differences between two groups (P＜ 0.01).The serum level of Hcy in AMI group was higher than that in UAP group and SAP group [(22.35 ± 8.18) μ mol/L vs.(16.54 ± 7.56),(14.52 ± 6.38) μ mol/L],and there was significant difference (P ＜ 0.01).Conclusions The relationship between Hcy,CysC level and coronary heart disease is close,and Hcy levels increase with increasing degree of coronary heart disease.Detection of Hcy and CysC has some clinical value on prevention,diagnosis,treatment of coronary heart disease.

Objective To analyze the effect of overweight on appendicitis and laparoscopic surgery.Methods Clinical data of 200 patients with laparoscopic appendectomy were analyzed retrospectively.Overweight and non-overweight patients were compared.The risk factors of appendicitis were analyzed.Results The operation time [(56.19 ± 30.42) min vs.(48.40 ± 21.03) min] and length of stay [(5.35 ± 2.04) d vs.(4.52 ± 1.49) d] between the overweight and non-overweight groups had statistically significant differences (t =2.040,P =0.043,t =3.219,P =0.002).Overweight was correlated with appendix inflammation (OR =0.149,P =0.000).Conclusion Over weight patients are prone to develope into complicated appendicitis.It should take surgeon's attention for more difficult operation and more complication.

OBJECTIVE:To study the protective effect of Compound pueraria tablets on the aging model rats. METHODS:The rats were divided into normal group,model control group,positive control group (vitamin E) and Compond pueraria tablets high-dose,medium-dose and low-dose (800,400,200 mg/kg) groups. The aging model rats were made by injection of D-galac-tose solution for 30 d in latter 5 groups,and they were given relevant medicine at same time every 7 days for consecutive 56 days since 31st day. SOD activity and MDA content in the serum and myocardium and LPF contents in the myocardium were determined after the last dosing,and apoptotic cells were counted and the pathology of cerebral tissues were observed. RESULTS:Compared with normal group,the activity of SOD in the serum and myocardium were decreased significantly in model control group,MDA and LPF contents in the myocardium and the number of apoptotic cells were increased significantly(P<0.01);significant myocar-dial cell injury was found. Compared with model control group,the activity of SOD in the serum and myocardium were increased significantly,while MDA and LPF content in the myocardium and the number of apoptotic cells in Compound pueraria tablets high-dose,medium-dose and low-dose groups were decreased significantly(P<0.05 or P<0.01);myocardial cell morphology was improved significantly. CONCLUSIONS:Compound pueraria tablets has obvious protective effect on myocardial cells in aging rats induced by D-galactose,the mechanism maybe related to the increase of SOD activity and the decrease of the content of MDA and LPF.

OBJECTIVE:To optimize the purification process of the extract from Yiqi guben granules. METHODS:The purifi-cation effect of the process was investigated with transfer rate of polysaccharide,calycosin glucoside and dry paste as evaluation in-dexes,using ZTC1+1 type Ⅱ,and shell poly sugar and 95% ethanol as clarifying agents. The purification process of the extract from Yiqi guben granules was optimized by orthogonal test using the ratio of material to liquid,the amount of clarifying agent and standing time as factor. The validation test was conducted. RESULTS:Selecting ZTC1+1 type Ⅱ as a clarifying agent,the best trans-fer rate of effective component had been obtained;optimal purification process was as follows as the ratio of material to liquid 1:2, the ratio of ZTC1+1 type Ⅱ A liquid 5%,the ratio of B liquid 10%,standing time of 5 h. The results of verification test showed transfer rates of dry paste in 3 tests were 71.54%,70.98%,69.21%,respectively;those of polysaccharide were 82.55%, 81.78%,82.15%,respectively;those of calycosin glucoside were 91.92%,92.34%,91.58%,respectively (all RSD≤1.72%, n=3). CONCLUSIONS:The optimized purification process is effective,stable and practical,and can be used for the purification of the extract from Yiqi guben granules.