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.

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.

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 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.

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.

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 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.

Objective To determine whether the 12-lead-resting electrocardiogram(ECG) is a predictor of left vertricular(LV) recovery after successful recanalization of chronic total coronary occlusions(CTO).Methods The 12-lead-resting ECG was evaluated for Q-wave areas and parameters of QT dispersion.Impairment of regional wall motion was evaluated by real-time three-dimensional echocardiography(RT-3DE) at baseline and at follow-up.Results The wall motion score index(WMSI) was improved from 1.56?0.31 to 1.12?0.21(P0.05).Conclusion In patients with recanalization of CTO,recovery of regional wall motion is reliably predicted by analysis of the12-lead-resting ECG.

Objective To investigate the clinical value of aprotinin blood anesthesia in the radical excision of esophageal carcinoma. Methods A total of 90 patients with esophageal carcinoma undergoing radical excision were divided into two groups according to using aprotinin or not. Patients in experiment group (group A, 40 patients) were injected with 1 112 EPU aprotinin followed by constant pumped infusion of 278 EPU/h until 2 h after operation. Patients in the control group (group B, 50 patients) were treated with constant pumped infusion of 0.9% saline. The venous blood was collected for blood routine examination, thromboelastography(TEG) and normal coagulable function test at the following time points: before induction, at 2 h and 4 h after the beginning of operation, at the end of operation and at 12 h after operation. The changes of TEG and normal coagulable state were monitored during the whole surgical process. The intraoperative volume of hemorrhage, perioperative transfusion rate and average volume of transfusion in the two groups were compared. Results The preoperative coagulable state in experiment group was kept relatively stable during the operation. Volume of intraoperative hemorrhage, perioperative transfusion rate and average volume of blood transfusion in experiment group were significantly lower than those in the control group. Conclusion Aprotinin blood anesthesia can stabilize the coagulable state, reduce the volumes and rates of hemorrhage and transfusion, and hence can find wide application in the radical excision of esophageal carcinoma.