Objective To supply scientific basis for the reasonable development and utilization of Rubus multibracteatus Levl.et Vant.resources.Method The total flavonoids was extracted from Rubus multibracteatus Levl.et Vant.with 60% methanol and its content was determined by spectrophotometry.The reliability and repeatability of the method were tested through stability,repeatability,accuracy and recovery experiments.Result Both standard and tested solutions had the maximum absorbance at 510 nm.The RSD values of repeatability,accuracy and recovery experiments were 2.11%,0.97% and 2.4% respectively.The results of content determination showed that the total flavonoids in Rubus multibracteatus Levl.et Vant.was 3.8%.Conclusion Extracted with 60% methanol,colored with aluminiumnitrate and determined by spectrophotometry at 510 nm were suitable for the determination of total flavonoids in Rubus multibracteatus Levl.et Vant..

Objective To investigate the effects of remifentanil on sinoatrial (SA) node autorhythmicity in rabbits.Methods Twenty-four healthy rabbits of both sexes weighing 1.8-2.2 kg were sacrificed.Their hearts were removed and sinoatrial nodes were dissected and placed in Tyrode solution saturated with 95 ％ O2-5 ％ CO2 at 36 ℃.The action potentials of the sinus node pacemaker cells were recorded by intracellular glass microelectrode technique.The experiment was performed in 3 parts (n =8 each).Part Ⅰ:the sinoatrial node was exposed to remifentanil 2,4,8,16 and 32 ng/ml respectively.The action potentials were recorded after the sinoatrial nodes were exposed to each concentration of remifentanil for 15 min.Part Ⅱ and Ⅲ:the sinoatrial nodes were first exposed to Ca2+ channel agonist Bay K8644 0.5 μmol/L or K+ channel blocker TEA 20 nmol/L for 15 min.Then remifentanil was added until the concentration reached 16 ng/ml (final concentration) and 15 min later the action potentials were recorded.The action potential parameters included,amplitude of action potential (APA),rate of pacemaker firing (RPF),action potential duration at 90％ repolarization (APD90) and velocity of diastolic depolarization (VDD).Results Remifentanil significantly decreased,APA,RPF,VDD and prolonged APD90 in a concentration dependent manner as compared with the baseline values.Pretreatment with Bay K8644 could block the effects of remifentanil on SA node pacemaker cells,while TEA did not affect the electrophysiologic effects of remifentanil on SA node pacemaker cells.Conclusion Remifentanil exerts a negative chronotropic action on SA node pacemaker cells.These effects are likely produced by decrease in Ca2+ current,while opening of K + channels is not involved in these effects.

Objective To investigate the effects of remifentanil on lung ischemia-reperfusion (I/R) injury and expression of angiotensin-converting enzyme (ACE) mRNA in rats.Methods Forty pathogen-free adult male Sprague-Dawley rats,weighing 300-350 g,were randomly divided into 5 groups (n =8 each):sham operation group (group S),group I/R and different doses of remifentanil groups (groups R1-R3).Lung I/R was produced by occlusion of the left hilum of lung for 45 min followed by 120 min reperfusion.Remifentanil was infused intravenously at a rate of 0.2,0.6 and 1.2 μg·kg-1 ·min-1 until 120 min of reperfusion after a loading dose of 1 μg/kg at 15 min before occlusion of the left hilum of lung in groups R1,R2 and R3,respectively.The equal volume of normal saline was given in groups S and I/R.The rats were scarified at 120 min of reperfusion and lungs were removed for microscopic examination and determination of wet/dry lung weight ratio (W/D ratio) and the expression of ACE mRNA (by RT-PCR) in the lung tissue.The pathological changes of the lung were scored.Results Compared with group S,the pathological score and W/D ratio were significantly increased and the expression of ACE mRNA was up-regulated in groups I/R,R1 and R2,and the pathological score was significantly increased (P ＜ 0.05),and no significant change was found in W/D ratio and the expression of ACE mRNA in group R3 (P ＞ 0.05).Compared with group I/R,the pathological score and W/D ratio were significantly decreased and the expression of ACE mRNA was down-regulated in groups R2 and R3,and the pathological score and W/D ratio were significantly decreased (P ＜ 0.05) and no significant change was found in the expression of ACE mRNA in group R1 (P ＞0.05).Compared with group R1,the pathological score and W/D ratio were significantly decreased and the expression of ACE mRNA was down-regulated in group R3,and the pathological score and W/D ratio were significantly decreased (P ＜ 0.05) and no significant change was found in the expression of ACE mRNA in group R2 (P ＞ 0.05).The pathological score,W/D ratio and expression of ACE mRNA were significantly lower in groups R3 than in group R2 (P ＜ 0.05).The pathological changes of the lung were significantly reduced in groups R1-R3 as compared with group I/R.Conclusion Remifentanil can attenuate lung I/R injury in rats in a dose-dependent manner and down-regulation of ACE mRNA expression may be involved in the mechanism.

Objective Mongolian gerbil can make themself urine concentration for saving water and adapt to the harsh desert environment, due to their very unique moisture control system in the body.Methods Mongolian gerbil is resistant to drought on account of their special kidney. Histology of the kidney, ureter and bladder in Meriones Unguiculataus were observed by light microscopy using HE staining.Results The results showed that compared with rats and mice, the Mongolian gerbils have more developed distal tubules, and well developed inner renal medulla.Conclusions We hope that the findings of this study enrich our understanding of the histology of urinary system in Mongolian gerbils and provide support for the laboratory animalization of this animal.

AIM　To investigate the effects of ginkgolide B on arachidonic acid (AA) metabolizing enzymes and the level of intracellular calcium in rat polymorphonuclear leukocytes. METHODS　Intracellular free calcium was quantitated by Fura-2 fluoresence technique. Phospholipase A2 (PLA2) activity was determined by incorporating 3H-arachidonic acid in leukocytes. 5-Lipoxygenase (5-LO) activity was evaluated by RP-HPLC. RESULTS　In comparison with control, ginkgolide B at final concentration of 0.1-10 μmol*L-1 inhibited A23187 induced AA release by 10.9%-22.2%; at final concentration of 0.1-50 μmol*L-1, ginkgolide B inhibited LTB4 and 5-HETE synthesis stimulated by PAF by 29.4%-88.8% and 26.2%-89.3% respectively. At the final concentration of 0.1-100 μmol*L-1, ginkgolide B decreased the rise of intracelluar calcium level induced by pletelet activating factor (PAF) and N-formyl-methionine-leucine-phenglalanine (fMLP) by 13.9%-51.4% and 2.2%-36.6%, respectively. CONCLUSION　Ginkgolide B was found to significantly inhibit PLA2 and 5-LO activities, as well as the increase of the intracellular calcium induced by PAF.

BACKGROUND:Mesenchymal stem cells (MSCs) exist in human tissues.Presently,cell source is single;culture method has great differences;obtained results are not consistent.Thus,it cannot verfy that isolated and cultured cells are identical calls,which is difficult to compare.OBJECTIVE:To compare the biological features of MSCs derived form bone marrow (BM),perpheral blood (PB) and cord blood (CB) under in vitro culture conditions.DESIGN,TIME AND SETTING:The cytological in vitro controlled study was performed at the Department of Hematology,Navy General Hospital of Chinese PLA from June 2007 to December 2008.MATERIALS:A total of 10 donors of hemopoietic stem cell transplantation at the Department of Hematology,Navy General Hospital of Chinese PLA were selected.MB and PB cells were obtained from the same donor,and cell volumes were respectively 20 mL and 2 mL.CB cells (30 mL) were obtained from healthy primipara at the Department of Obstetrics,Navy General Hospital of Chinese PLA.METHODS:MSCs were obtained from BM,PB and CB by Percoll density gradient + adherence method,and then incubated in DMEM/F12 medium containing 10% fetal bovine serum.When 80%-90% confluency,cells were digested in trypsin-EDTA and made into 5×10~8/L cell suspension as P_0.Above-described operation was performed as P_1,and the rest may be deduced by analogy as P_2-P_5.MAIN OUTCOME MEASURES:The following parameters were measured:cell growth morphology;results of Wright-Giemsa staining;results of cytochemistry;cell proliferation amount;cell surface markers using flow cytometry.RESULTS:Time of adherence,time to 50% confluency and time to 80% confluency of BMSCs were earlier comarped with the PBMSCs and UCMSCs.Adherent cells from BM grew in whirpool-like type,while CB and PB did not at 5-7 days.Majority of aderent cells from BM were fibroblast-like cells,and small parts were endothelioid cells.Aderent cells from PB and CB at the fifth generation contained more endothelioid cells and mononuclear and macrophage-like cells besides fibroblast-like cells.PAS stain,Sudan black B stein,alkaline phosphatase (AKP) staining of adherent cells from BM,PB and CB were negative from P_1 to P_5.Compared with P0 cells,number of BMMSCs till P5 was significantly more in PBMSCs and UCMSCs (P ＜ 0.05).Positive rates of CD29,CD44,CD90,CD71,CD105,CD166 and HLA-ABC were 55.9% 92.8% at P0 to P5,but ≤6% following BMMSCs were incubated;19.7%-33.4% at P0 to P5,but ≤10% following PBMSCs were incubated;35.4%-93.2% at P_0 to P_5,but ≤20% following CBMSCs were incubated.Positive rates of CD34,CD45 and HLA-DR were low in BM-,PB-and CB-MSCs.Positive rates of CD14 and CD31 were low in BMMSCs;12.1%-28.3% in PBMSCs,and 8.1%-21.3% in CBMSCs.CONCLUSION:MSCs can be attained from BM,PB and CB.Quantities of MSCs form BM are the highest,with single component,followed by CBMSCs and PBMSCs,with multiple components.

Objective To compare the sensitivity, specificity and consistency of five kits for quantitative detection of hepatitis B virus (HBV) DNA. Methods Four domestic fluorescence kits, A, B, C, and D, and Roche Cobas Ampliprep/Cobas TaqMan HBV test for quantitation of HBV DNA in serum, were applied to detect the National Standards, an additional plasma for sensitivity (7 times dilution), 15 plasma samples from healthy blood donor and 45 plasma samples from chronic hepatitis B patients. Results All five kits showed the correct results for the 9 positive specimens and 8 negative specimens from the National Standards. The quantitative results of specimens for sensitivity met requirements of the National Standards. The lowest concentration of HBV DNA by these three kits was 15.6 IU/mL. The lowest detection level of HBV DNA for domestic kit B was ≤500 IU/mL. There was linear correlation between the results by Roche kit and domestic kit C (r> 0.98). Kit D showed 2 false-negatives results in the 15 healthy blood donor samples. The coincidence rates between 4 domestic kits and the Roche kit ranged from 50% to 96% (A: 61%, B: 50%, C: 96%, D: 83 %). The consistency rate of kit C with the Roche kit was significantly higher than those of kit D, A and B (X2=5.62, P<0.05, X2=28. 93, P<0.01, X2=44.31,P<0.01, respectively). The consistency rates among these 5 kits were highest when testing samples with HBV DNA levels between 1×104-1×107 IU/mL. Conclusions The qualities of domestic kits vary remarkably and kit C has the greatest consistency rate with the Roche kit. Therefore, the qualities of domestic kits need to be further improved.

ObjectiveTo observe the effects of different angles between tibial tunnel and tibial platform on “killer turn” in posterior cruciate ligament (PCL) reconstruction,and primarily discuss a safe and reasonable tunnel technology. Methods Eighteen fresh tendon grafts were used to reconstruct the PCL on the tibial side of fresh cadavers.The tibial tunnels of all specimens were built via anteromedial approach.Based on the different angles between tibial tunnel and tibial platform,all specimens were divided into Group A (30°),Group B (40°) and Group C (50°),with six specimens in each group.Area of tibial tunnel exit,pressure of tibia tunnel exit and circulation characteristics of tendons under the cyclic load before and after biomechanical test were recorded.ResultsThe area of tibial tunnel exit had statistical difference among three groups after the test ( F =8.80,P ＜ 0.05 ).The pressure of tibial tunnel exit had statistical difference among three groups (F =3.91,P ＜ 0.05 ).The cyclic frequency and fatigue strength of the transplanted tendons had statistical difference among three groups under the same cyclic load ( 256 N ) and same frequency ( 126 Hz ) ( F =4.25,P ＜ 0.05 ).Conclusions The angle between tibial tunnel and tibial platform has negative correlation with the area and pressure of tibial tunnel exit,and has positive correlation with the cyclic frequency and fatigue strength of the transplanted tendons under cyclic load.The ideal anatomy position of the tibial tunnel is the anteromedial tunnel with the angle of 40° between the tibial tunnel and the tibial platform.

ObjectiveTo assess the diagnostic value of ultrasonography in the diagnosis and pathological classifi cation of appendicitis with pathological results.MethodsThe study included 111 cases who confi rmed by surgery and pathology in the Six Affi liated Hospital of Sun Yat-sen University from June2012 to December 2013. The image characteristics of ultrasonic images performed before surgery were analyzed retrospectively. The cases were divided into five groups based on the pathology: 6 acute simple appendicitis, 45 acute suppurative appendicitis, 15 acute gangrenous appendicitis, 33 chronic appendicitis, and 12 acute attack of chronic appendicitis. The length, width and wall thickness of the appendicitis measured by preoperative ultrasound between various pathological type were compared with Kruskal-Wallis statistical methord, and also used the same method to compared the one which had statistically signifi cance between the fi ve groups. As in the above case, Fisher Exact Test was used to compare the characters′ of ultrasonography included gradations of the appendix, the tube cavity expansion, stercorolith, the appendiceal abscess, mergering lymph node enlargement and the appendix around effusion, if there was statistically signifi cance, the same methord was used to the further comparison between thefi ve groups.ResultsTotally 90 of 111 cases of were reminded appendicitis by preoperative ultrasound, the diagnostic accuracy rate was 81.08%, including acute suppurative appendicitis 91.11% (41/45), acute gangrenous appendicitis 93.33% (14/15), acute simple appendicitis 83.33% (5/6), chronic appendicitis 60.61% (20/33), chronic appendicitis onset acute 83.33% (10/12). There were no statistical differences of preoperative ultrasound measured between various pathological type of appendicitis in length, while the appendix width measured by preoperative ultrasound of acute gangrenous appendicitis was wider than the acute simple group and chronic group, and there were statistically signifi cance between them (P<0.05), in the aspect of wall thickness, the acute suppurative appendicitis and the acute gangrenous appendicitis groups were thicker than the chronic group, and these differences had statistically signifi cance (P<0.05). The stercorolith, lymph node enlargement and the appendiceal abscess in different types of appendicitis had no statistical difference. There were statistically differences (P<0.05) between the acute suppurative goup, acute gangrenous group and the chronic group respectively, both in gradations of the appendix and the tube cavity expansion. Also there were statistically difference (P<0.05) between the acute gangrenous appendicitis group and the chronic group in the aspect of the appendix around effusion.ConclusionsUltrasound is valuable in the diagnosis of appendicitis, especially for acute gangrenous appendicitis and acute purulent appendicitis. The measurement of appendix with wide diameter, wall thickness by ultrasond preoperatively, and the characteristics of the wall layers, lumen expansion degree and the appendix around effusion are valuable in identifying chronic appendicitis, acute suppurative appendicitis, and acute gangrene appendicitis; The ultrasonic measurement of appendix wide diameter could identify acute gangrenous appendicitis and acute simple appendicitis. Ultrasound had limited value in identifying acute suppurative, acute gangrenous appendicitis, acute simple appendicitis, chronic appendicitis, and chronic appendicitis onset acute.

[Objective] To investigate the effects and mechanisms of Nrf2-ARE (nuclear factor erythroid-2 related factor-anti-oxidant response element) pathway on uremic serum-mediated endothelial dysfunction in human aortic endothelial ceils.[Methods] Human aortic endothelial cells were incubated in endothelial cell medium containing 10％ normal serum,10％ non-diabetic nuremic serum or 10％ diabetic uremic serum respectively,and 20 μmol/L tertiary butyl hydroquinone (tBHQ) were pretreated with cells to active Nrf2-ARE pathway.The cells apoptosis rate were measured by flow cytometry,and the synthesis of NO was detected by flow cytometry and immune fluorescent confocal,while the expression of P-eNOSer1177/eNOS,and quinone oxidoreductase-1 (NQO1) were measured by western blotting.The levels of malondialdehyde,superoxide dismutase,eatalase,and glutathione in these cells were also measured with kits.[Results] Aortic endothelial cells incubated with uremic serum had a higher level of apoptosis rate and MDA (P ＜ 0.05),and a lower level of NO systhesis,P-eNOSSer1177/eNOS expression,CAT,SOD,GSH (P ＜ 0.05).Pretreated with tBHQ can reduce the apoptosis rate and MDA level (P ＜ 0.05),improve the amount of NO systhesis,the expression of P-eNOSSer1177/eNOS,the levels of CAT,SOD,and GSH in these cells (P ＜ 0.05).[Conclusion] Activation of Nrf2-ARE pathway can improve endothelial dysfunction in aortic endothelial cells induced by uremic serum,and its mechanism might be related with enhancement of the antioxidant stress.