1.Changes in expression of transforming growth factor beta-1 during myogenic differentiation of pulmonary microvascular endothelial cells in rats with hepato-pulmonary syndrome
Zhi WANG ; Bin YI ; Bin GUO ; Kaizhi LU
Chinese Journal of Anesthesiology 2011;31(7):862-864
ObjectiveTo investigate the changes in the expression of transforming growth factor beta-1 (TGF-β1) during differentiation of pulmonary microvascular endothelial ceils (PMVECs) into smooth muscle cells in rats with hepato-pulmonary syndrome (HPS).MethodsPrimary PMVECs were harvested from healthy adult SD rats of both sexes aged 3-4 months and inoculated in low-glucose DMEM culture medium (1(6/cm2 ) and randomly divided into 2 groups ( n =24 dishes each):control group ( group C) and HPS group.HPS was produced by chronic ligation of common bile duct.In group C serum obtained from normal rats was added to PMVECs,while in HPS group serum obtained from rats with HPS was added.The final concentration of serum was 10%.After being incubated for 24,48 and 72 h,the expression of SM-MHC,SM-α-actin and calponin protein and TGF-β1 mRNA and protein in PMVECs was determined by RT-PCR and Western blot analysis.ResultsThe expression of SMMHC,SM-α-actin and calponin protein.was positive in HPS group whereas the expression of SM-α-actin and calponin protein was negative and the expression of SM-MHC protein was barely detectable in group C.The expression of SM-MHC,TGF-β1 mRNA and protein was significantly higher in HPS group than in group C.The expression of SM-MHC,SM-α-actin and calponin protein and TGF-β1 mRNA and protein was increasing with duration of incubation from T1 to T3 in group HPS.ConclusionTGF-β1 plays an important role in the myogenic differentiation of PMVECs in rats with HPS.
2.Role of 15-F2t-isoprostane in intestinal ischemia/reperfusion injury in rats
Shihong WEN ; Yi LI ; Cai LI ; Yunsheng LI ; Ying LIU ; Wenqi HUANG ; Kexuan LIU
Chinese Journal of Anesthesiology 2011;31(7):850-853
ObjectiveTo investigate the role of 15-F2t-isoprostane in intestinal injury induced by intestinal ischemia/reperfusion (I/R) in rats.MethodsThirty-two pathogen free adult male SD rats weighing 230-255 g were randomly divided into 4 groups ( n =8 each):group sham operation (group S) ; group intestinal I/R; group SQ-29548 (TXA2 receptor antagonist) (group SQ) and group DMSO (the solvent).Intestinal I/R was induced by 60 min occlusion of superior mesenteric artery (SMA) followed by 120 main reperfusion in groups I/R,SQ and DMSO SQ-29548 2 μmol/kg and DMSO were injected subcutaneusly at abdominal wall at 30 min before SMS in groups SQ and DMSO respectively.Arterial blood samples were taken at 120 min of reperfusion for determination of serum diamine oxidase (DAO) activity and 15-F2t-isoprostane,endothelin-1 (ET-1) and thromboxane B2 (TXB2) concentrations.Intestinal tissues were removed for microscopic examination and determination of myeloperoxidase (MPO) and SOD activities,MDA and lactate contents.Intestinal damage was assessed and scored according to Chiu (0 =normal,5 =disruption of tunica propria,bleeding and ulceration).ResultsIntestinal I/R significantly increased Chiu's score,MDA and lactate contents and MPO activity and decreased SOD activity in intestine in group I/R as compared with group S.SQ-29548 pretreatment significantly decreased Chiu's score,lactate content and MPO activity in intestine and increased intestinal SOD activity and decreased serum DAO activity and ET-1 concentration in group SQ as compared with group I/R.Conclusion15-F2t-isoprostane is involved in the development of intestinal injury induced by intestinal I/R by activating TXA2 receptor,increasing ET-1 production and promoting neutrophil infiltration.
3.Effects of tetramethylpyrazine on neuropafhdc pain in rats
Chinese Journal of Anesthesiology 2011;31(7):840-843
ObjectiveTo evaluate the effects of tetramethylpyrazine on neuropathic pain (NP) in rats.MethodsFifty-four male SD rats weighing 180-220 g were randomly divided into 3 groups ( n =18 each):sham operation group (group S); group NP and tetramethylpyrazine group (group T).NP was induced by chronic constrictive injury (CCI) in groups NP and T.Four ligature were placed on right sciatic nerve at 1 mm intervals with 4-0 chromic catgut.In group T tetramethylpyrazine 100 mg/kg was administered intraperitoneally once a day for 14 consecutive days after operation.Thermal and mechanical pain threshold was measured at 1 day before and 3,7,and 14 d after operation.Six animals in each group were sacrificed after pain threshold measurement on the 3rd,7th and 14th days after operation and the lumbar segment ( L(4).5 ) of the spinal cord was removed for determination of apoptosis (by TUNEL) and Bcl-2 and caspase-3 expression in spinal dorsal horn (by immuno-histochemistry).Apoptosis rate (AR =the number of apoptotic cells/the total number of cells examined) was calculated.Results CCI significantly decreased the thermal and mechanical pain threshold,up-regulated Bcl-2 and caspaas-3 expression in spinal dorsal horn and increased AR in groups NP and T.Tetramethylpyrazine significantly increased the thermal and mechanical pain threshold,up-regulated Bcl-2 expression and down-regulated caspase-3 expression and decreased AR in group T as compared with group NP.ConclusionTetramethylpyrazine can attenuate NP through inhibition of apoptosis in the spinal cord.
4.Effects of etomldate postconditlonlng on apoptotds In a rnt model of focal cerebral lschemia-reperhrslon In- jury
Chinese Journal of Anesthesiology 2011;31(7):881-883
ObjectiveTo investigate the effects of etomidate postconditioning on apoptosis in a rat model of focal cerebral ischemia-reperfusion(I/R) injury.MethodsThirty-two pathogen-free male SD rats weighing 250-300 g were randomly divided into 4 groups ( n =8 each) using random number table:group sham operation ( group S); group focal cerebral I/R; group lipid emulsion (vehicle for etomidate) (group L) and group etomidate postconditioning (group Ep).Focal cerebral I/R was induced by inserting a nylon thread with rounded tip into right internal carotid artery.The thread was advanced cranially until resistance was met.Middle cerebral artery was occluded for 2 h in groups I/R,L and Ep.Normal saline,lipid emulsion and etomidate emulsion 20 mg/kg were injected peritoneally at the end of ischemia in groups I/R,L and Ep respectively.The animals were sacrificed at 24 h of reperfusion and their brains were removed for microscopic examination,assessment of apoptosis (by TUNEL) and detection of Bcl-2 and Bax expression ( by immuno-histochemistry).Apoptosis index ( AI =the number of apoptofic neurons/the total number of neurons examined × 100% ) and Bcl-2/Bax ratio were calculated.Results I/R induced microscopic changes,significantly increased AI and Bcl-2 Bax ratio and up-regulated Bcl-2 and Bax expression in group I/R as compared with group S.Etomidate postconditioning significantly amefiorated brain damage,decreased AI,increased Bcl-2/Bax ratio,up-regulated Bcl-2 expression and down-regulated Bax expression in ischemic cerebral hemisphere in group Ep as compared with group I/R.There was no significant difference in brain damage,AI and Bcl-2 and Bax expression and Bcl-2/Bax ratio between groups I/R and L.ConclusionEtomidate postconditioning can attenuate focal cerebral ischemia-reperfusion injury in rats by inhibiting apoptosis and modulating Bcl-2 and Bax expression.
5.Effect of aminoguanidine on expression of inducible nitric oxide synthase in neurons in the small intestinal nerve plexus of starved rats
Xinoping ZHANG ; Zhihui LU ; Aiguo CHENG
Chinese Journal of Anesthesiology 2011;31(7):878-880
ObjectiveTo evaluate the effect of aminoguanidine on expression of inducible nitric oxide synthase (iNOS) in neurons in the small intestinal nerve plexus of starved rats.MethodsNinety male SD rats weighing 230-270 g were randomly divided into 3 groups:group normal control (group C,n =10) ; group starvation (group S,n=40) and group starvation + aminoguanidine (group A,n =40).The animals were allowed free access to water but no food during starvation in S and A groups.In group A the animals were given aminoguanidine 150 mg·kg-1 ·d-1 intraperitoneally during starvation.Ten animals were sacrificed at 3,5,7 and 9 d of starvation respectively and intestine specimens were taken for determination of ratio of intestinal transit using dextran blue-2000 as indicator.Then the specimens of intestinal myenteric nerve plexus of ileum were collected and stained by histochemistry with nicotinamide-adenine dinucleotide phosphate-d for determination of iNOS expression.ResultsStarvation significantly reduced the small intestinal transit and increased iNOS expression in neurons in the myenteric nerve plexus of small intestine in proportion to days of starvation in group S compared with group C.Intraperitoneal aminoguanidine significantly attenuated the starvation-induced changes in intestinal transit and iNOS expression.ConclusionAminoguanidine can attenuate the up-regulation of the expression of iNOS in neurons in the myenteric nerve plexus of small intestine induced by starvation and is helpful in promoting the intestinal transit in starved rats.
6.Effects of diazoxide pretreatment on expression of phosphatidylinositol 3-kinase mRNA and protein serine/ threonine kinase mRNA in rat myocardial microvascular endothelial cells exposed to hypoxia-reoxygenation
Su CAO ; Qiuping CHEN ; Huanju KANG ; Shiren SHEN
Chinese Journal of Anesthesiology 2011;31(7):871-873
ObjectiveTo investigate the effects of diazoxide pretreatment on expression of phosphatidylinositol 3-kinase(PI3K) mRNA and protein serine/threonine kinase(Akt) mRNA in rat myocardial microvascular endothelial cells exposed to hypoxia-reoxygenation (H/R).MethodsThe SD rat myocardial microvascular endothelial cells were cultured.The cells were seeded in 96-well plates ( 100μd/hole) or in 6 cm diameter dishes (2 ml/dish) with the density of 1 × 106/ml and randomly divided into 4 groups ( n =25 each):normal control group (group C),H/R group,diazoxide pretreatment group (group DZ) and diazoxide pretreatment + 5-hydroxydecanoate (5-HD,a mitochondrial ATP-sensitive potassium channel blocker) group (group DZ + 5-HD).The cells were exposed to 2 h hypoxia followed by 2 h reoxygenation.Diazoxide 100 μmol/L and diazoxide 100 μmol/L + 5-HD 100 μmol/L were added to the culture medium 2 h before hypoxia in groups DZ and DZ + 5-HD respectively.The cell viability,apoptotic rate and expression of PI3K mRNA and Akt mRNA were detected at the end of reoxygenation.ResultsCompared with group C,the cell viability was significantly decreased,while the apoptotic rate increased and expression of PI3K mRNA and Akt mRNA up-regulated in group H/R (P <0.05 or 0.01).Compared with group H/R,the cell viability was significantly increased,while the apoptotic rate decreased and expression of PI3K mRNA and Akt mRNA up-regulated in group DZ (P < 0.05 or 0.01).5-HD could inhibit diazoxide pretreatment-induced changes mentioned above (P < 0.05 or 0.01 ).ConclusionDiazoxide pretreatment can reduce H/R injury by promoting PI3K gene and Akt gene transcription through activation of mitochondrial ATP-sensitive potassium channels in rat myocardial microvascular endothelial cells.
7.Individualized lung protective ventilation regimen based on pressure-volume curve for one lung ventilation in patients undergoing thoracic surgery
Jianluo SI ; Yue SU ; Shaotuan SONG
Chinese Journal of Anesthesiology 2011;31(7):847-849
ObjectiveTo evaluate the effectiveness of setting PEEP and tidal volume (VT ) according to pressure-volume (P-V) curve during one lung ventilation (OLV) in patients undergoing thoracic surgery.Methods Twenty-five ASA Ⅰ or Ⅱ patients of both sexes aged 44-64 yr weighing 57-75 kg undergoing lobectomy under general anesthesia were enrolled in this study.Double-lumen tube was inserted.Correct positioning was verified by flberoptic bronchoscopy.The patients were mechanically ventilated.P-V curve was measured by SSS system during OLV.Lower inflection point (LIP)and upper inflection point (UIP) were determined.The pressure at LIP (PLIP) and volume at UIP (VUIP) were measured.Bilateral lungs were ventilated for 30 min (T0) at first before OLV was started.PEEP was set at PLIP + 0.196 kPa and VT was set at VUIP,and the patients were ventilated for 30 min (T1).VT was then reduced to 80% of VUIP.OLV was performed for another 30 min (T2).VT was then further reduced to 60% of VUIP and the patients were ventilated for 30 min (T3).PEr CO2 was maintained at 4.67-6.00 kPa.Arterial blood and central venous samples were taken at T0-3.Blood gas analysis was performed.Qs/Qt was calculated.MAP,HR,CVP,peak airway pressure (Peak),airway resistance (Rsw) and lung compliance (CL) were measured and recorded at T0-3.ResultsHR,Ppeakk,Rsw and Qs/Qt were significantly increased while CL and PaO2 decreased at T1-3,CVP was significantly increased at T1.2 and MAP and PaCO2 were increased at T3 as compared with the baseline values at T0.Ppeak and Rsw were significantly decreased at T2.3 and PaO2 was significantly increased while Qs/Qt decreased at T2,CVP was decreased,MAP and PaO2 were increased at T3 as compared with the values at T1.ConclusionsMechanical ventilation with VT set at 80% of VUIPandPEEPatPUIP+0.196kPa provides best ventilatory efficacy for OLV in terms of PaO2 and hemodynamics.
8.The role of PI3K-Akt signal pathway in protection of myocardium agalnst ischemia-reperfusion injury by diazoxide postconditioning in rats
Qihong ZHAO ; Ying ZHANG ; Hengfei LUAN ; Yinming ZENG ; Ying YE ; Guoli GONG
Chinese Journal of Anesthesiology 2011;31(7):874-877
ObjectiveTo investigate the role of phosphatidylinositol 3-kinase/protein-serine-threonine kinases (PI3K-Akt) signal pathway in the protection of myocardium against ischemia-reperfusion (I/R) injury by diasoxide postconditioning in rats.MethodsThirty-six male SD rats weighing 250-300 g were randomly divided into 4 groups using random number table (n =9 each):group I/R; group diazoxide (group D); group wortmannin (PI3K inhibitor) (group W) and group wortmannin + diazoxide (group WD).The animals were anesthetized with intraperitoneal 10% chloral hydrate 4 ml/kg,intubated and mechanically ventilated.Myocardial I/R was produced by 30 min occlusion of left anterior descending coronary artery followed by 120 min reperfusion.0.1% dimethyl sulfoxide (the vehicle for diazoxide and wortmannin)/diazoxide at 0.47 mg· kg-1 · min-1/wortmannin at 1 μg·kg-1 ·min-1 was infused for 15 min starting from 25 min of ischemia in groups I/R,D and W respectively.In group DW wortmannin was infused at 5 min before diazoxide infusion.Blood samples were collected from left ventricle at the end of 120 min reperfusion for measurement of lactate dehydrogenase (LDH) activity.Myocardial infarct size was measured.Myocardial specimens were obtained for determination of apoptosis index (the number of apoptotic cells/the total number of cells examined) and expression of Bcl-2 and Bax.Bcl-2/Bax ratio was calculated.ResultsDiazoxide postconditioning significantly decreased plasma LDH activity,myocardial infarct size,apoptosis index and Bax expression in myocardium and increased myocardial Bcl-2 expression and Bcl-2/Bax ratio in group D compared with group I/R.Wortmannin partly counteracted the protective effects of diazoxide postconditioning against myocardial infarct.There was no significant difference in the above variables between groups I/R and W.ConclusionPI3K-Akt signal pathway is involved in the protective effects of diazoxide postconditioning on myocardium against I/R injury.
9.Effect of remifentanil on hepatic ischemia-reperfusion injury in rats with liver cirrhosis
Rui JING ; Wan MA ; Gang MA ; Jianzhen WANG ; Yuhua LIU
Chinese Journal of Anesthesiology 2011;31(7):865-867
ObjectiveTo investigate the effect of remifentanil on hepatic ischemia-reperfusion (I/R) injury in rats with liver cirrhosis.MethodsThirty male SD rats weighing 260-300 g were randomly divided into 3 groups (n =10 each):group liver cirrhosis (group C); group liver cirrhosis + hepatic I/R (group I/R) and group remifentanil (group R).Liver cirrhosis was produced in all animals in the 3 goups.I/R injury was induced by 20 min occlusion of the hepatic artery and portal vein entering the middle and left lobes of the liver followed by 4 h reperfusion at 1 week after establishment of hepatic cirrhosis in I/R and R groups.In group R remifentanil was infused iv at 1 μg·kg-1 ·min-1 starting from 10 min before ischemia until the end of 4 h reperfusion.Venous blood samples were taken from inferior vena cava at the end of 4 h reperfusion for measurement of serum ALT and AST activities.The animals were then sacrificed and liver specimens were taken from middle lobe for determination of Bcl-2 and Bax expression (by immuno-histochemistry) and hepatocyte apoptosis (by TUNEL) and microscopic examination.Apoptosis index (percentage of apoptotic cells) was calculated.ResultsI/R significantly increased serum ALT and AST activities,Bax expression and apoptosis index and decreased Bcl-2 expression in group I/R as compared with group C.Remifentanil significantly attenuated the I/R-induced changes in serum ALT and AST activities,Bax and Bcl-2 expression and apoptosis in group R as compared with group I/R.Remifentanil also ameliorated I/R-induced liver damage.ConclusionRemifentanil can auenuate hepatic I/R injury in rats with liver cirrhosis by up-regulating Bcl-2 expression and down-regulating Bax expression and inhibiting apoptosis.
10.Effects of sevoflurane and isoflurane on blockade of adult type nicotinic acetylcholine receptors of skeletal muscle of rats by rocuronium
Liu LI ; Min SU ; Li WEI ; Wei KE ; Cao JUN ; Dong JUN
Chinese Journal of Anesthesiology 2011;31(7):805-808
ObjectiveTo investigate the effects of sevotlurane and isoflurane on blockade of adult nicotinic acetylcholine receptor (ε-nAChR) of skeletal muscle of rats by rocuronium.MethodsHEK293 cell line was provided by Molecular Biology Laboratory,Chonqing Medical University and cultured in DMEM liquid culture medium containing 10% bovine calf serum at 37 ℃in incubator filled with 5% CO2.ε-nAChR was expressed heterologously in HEK293 cell using liposome transfection technology.The whole cell patch clamp technology was used to establish the concentration-effect curves for inhibition of acetylcholine (ACh)-induced current.The 5% inhibitory concentration values ( IC5 ),25 % inhibitory concentration values ( IC25 ) and 50% inhibitory concentration values (IC50) for sevoflurane,isoflurane and rocuronium were calculated.Firstly the inhibitory effect of rocuronium at a concentration of IC25 was studied in the presence of sevoflurane and isoflurane at concentrations of IC5,IC25 and IC50.Subsequently the inhibitory effect of rocuronium at concentrations of IC5,IC25 and IC50 was studied in the presence of sevoflurane and isoflurane at a concentration of IC25.Inhibition of ACh-induced current amplitude was recorded.ResultsAt the concentrations of IC5,IC25 and IC50 sevoflurane or isoflurane had synergistical inhibitory effect on the current amplitude of ε-nAChR with rocuronium at the concentrations of IC5 and IC25,while had addition inhibitory effect on the current amplitude of ε-nAChR with rocuronium at the concentration of IC50.The inhibitory effect of sevoflurane at the concentration of IC5 on the current amplitude of ε-nAChR with rocuronium at the concentration of IC25 was weaker than that of isoflurane,while the inhibitory effect of sevoflurane at the concentration of IC50 on the current amplitude of ε-nAChR with rocuronium at the concentration of IC25 was stronger than that of isoflurane.The inhibitory effect of sevoflurane at the concentration of IC25 on the current amplitude of ε-nAChR with rocuronium at the concentration of IC5 was weaker than that of isoflurane.ConclusionEither sevoflurane or isoflurane has synergistical blocking effect on ε-nAChR with low concentration of rocuronium,while has addition blocking effect with high concentration of rocuronium.Potentiation of blocking effect of rocuronium on ε-nAChR by low concentration of sevoflurane is weaker than that by isoflurane,while the potentiation by sevoflurane at high concentration is stronger than that by isoflurane.