3.Study of the mechanisms of curcumin on mitochondrial permeability transition of hepatocytes in rats with sepsis
Pei TAO ; Haiyan YIN ; Yonghui MA
Chinese Critical Care Medicine 2014;26(9):666-670
Objective To study the effects of mitochondrial permeability transition (MPT) on hepatocyte apoptosis and mitochondrial damage,and investigate the effects of curcumin on MPT and the related mechanisms in septic rat.Methods Fifteen healthy male Sprague-Dawley (SD) rats were randomly divided into three groups:sham group,sepsis group and curcumin group,with 5 rats in each group.Septic rat model was reproduced by cecal ligation and puncture (CLP).The rats in sham group were flipped the cecum without perforation and ligation.The rats in the curcumin group were treated with curcumin 100 mg·kg-1·d-1 (dissolved in saline to 10 mL/kg) by oral gavage for 7 days,while the other groups were treated with normal saline.Tissue samples were harvested in each group at 12 hours after operation.Pathological changes in hepatic mitochondria were observed under electron microscopy,concentration of free calcium was examined with confocal laser scanning microscope.After Fluo-3/AM staining,protein and mRNA expression of active caspase-3,Bcl-2 and Bax were examined by Western Blot and reverse transcription-polymerase chain reaction (RT-PCR).Results Under the transmission electron microscope,intact cell membrane,adqulis cytoplasm,and normal and clear mitochondrion was found in the sham group.Mitochondria in sepsis group swelled obviously with mitochondrial cristae broken or disappearance,unclear bilateral membrane structure,while the curcumin group showed much less pathological changes,with few mitochondria swell,and smear bilateral membrane structure.The fluorescence intensity index of sham group,sepsis group and curcumin group was raised successively (417.33 ± 15.88,772.95 ± 42.37,1 560.84 ± 160.78,respectively,F=184.149,P=0.000).The protein and mRNA expression of active caspase-3 and Bax had the highest level in sepsis group,followed by the curcumin group,and that in the sham group was the lowest [active caspase-3 protein (gray scale):1.698 ± 0.061,0.694 ± 0.045,0.246 ± 0.027,F=1 289.667,P=0.000; active caspase-3 mRNA (2-Δ△Ct):1.031 ±0.135,0.578 ± 0.144,0.183 ±0.036,F=66.958,P=0.000; Bax protein (gray scale):1.826 ±0.126,1.254 ±0.140,0.623 ±0.901,F=94.536,P=0.000; Bax mRNA (2-ΔΔCt):2.774 ±0.338,1.661 ±0.226,0.656 ±0.114,F=124.710,P=0.000],all of these values had statistical significance among the three groups (all P<0.01).While Bcl-2 protein and mRNA had the highest level in curcumin group and lowest level in the sham group [Bcl-2 protein (gray scale):0.716 ±0.091,1.328 ±0.147,1.656 ±0.104,F=84.918,P=0.000; Bcl-2 mRNA (2-△ΔCt):0.617 ±0.118,1.393 ±0.096,1.650 ±0.167,F=83.846,P=0.000].The protein and mRNA expressions of Bcl-2/Bax ratio were lowest in sepsis group,then sham group,and highest in curcumin group [Bcl-2/Bax protein (gray scale):0.726 ± 0.055,1.150 ± 0.043,1.333 ± 0.163,F=46.265,P=0.000; Bcl-2/Bax mRNA (2-Δ△Ct):0.505 ±0.041,0.944±0.097,1.006 ±0.168,F=12.211,P=0.001].Conclusions MPT can lead to mitochondrial dysfunction and further cause hepatocyte apoptosis.Mechanism of effect of curcumin on MPT may be related to reduction of intracellular calcium concentration,promotion of anti-apoptotic Bcl-2 gene expression,inhibition of caspase-3 activation and Bax gene.
4.Lentiviral vector-mediated transfection of bone morphogenetic protein 2 gene into endothelial progenitor cells from rat bone marrow
Xiuru YIN ; Ling PEI ; Zuodi LIANG
Chinese Journal of Tissue Engineering Research 2014;(32):5197-5202
BACKGROUND:Gene therapy has become a new trend for disease therapy and brought promise for some refractory diseases. The key point is to choose the proper cell, gene and vector. OBJECTIVE:To investigate the effectiveness and feasibility of bone morphogenetic protein 2 (BMP2) gene transfected into endothelial progenitor cells (EPCs) from rat bone marrow for gene therapy. METHODS:The EPCs were isolated, cultured and identified from the bone marrow of Sprague-Dawley rats. Empty vector (LV-eGFP) or BMP2 gene (LV-eGFP-BMP2) was transferred into EPCs by the constructed lentiviral vector (LV). We examined the transfection efficiency by eGFP fluorescence, BMP2 secretion by ELISA, BMP2 expression by Western blot, and compared the capacities of migration, proliferation and anti-apoptosis after transfection in the three groups of normal EPCs, empty vector-EPCs, and BMP2-EPCs. RESULTS AND CONCLUSION:The transfection efficiency of lentiviral vector was 90%. BMP2 gene-transferred EPCs secreted and expressed more BMP2 proteins (P<0.01), and showed enhanced anti-apoptotic ability (P<0.05). The proliferation and migration capacity did not change obviously (P>0.05). After successful transfection with lentivirus-BMP2 gene, EPCs can secrete and express more BMP2 protein and show enhanced anti-apoptotic ability without obvious influence on the proliferation and migration capacity.
5.The study on curative effect of acupuncture therapy with the Shu-points of five Zangs and Ge-shu point on elderly depression
Jie CHEN ; Jie ZHANG ; Yin PEI
International Journal of Traditional Chinese Medicine 2010;32(4):339-340
Objective To observe the clinical effect of acupuncture on elderly depression. Methods A total of 63 patients were designated into a treatment group and a control group by using a random number table. Acupuncture on Ge-shu point and Shu-points of Five-Zangs were administrated for the treatment group, and Fluoxetine was used for the control group at a dosage of 20 mg, once daily. The treatment of both groups lasted for 6 weeks. Results The curative effect of acupuncture was sound on relieving depression and the scores of HAMD in the treatment group exhibited a decreasing trend. There was no statically significant difference between the two groups on the total effective rate and the cure rate. Conclusion It has a good anti-depression result to needle Shu-points of Five-Zangs.
7.Protective effect of ulinastatin on intestinal barrier function of septic rats
Xiaoling YE ; Pei TAO ; Yue'e CHEN ; Haiyan YIN ;
Chinese Journal of Clinical Infectious Diseases 2015;8(6):549-553
Objective To observe the protective effect of ulinastatin (UTI) on the intestinal barrier function of septic rats.Methods Septic rat model was established using Sprague-Dawley rats by cecal ligation and puncture (CLP) method.Thirty Sprague-Dawley rats were randomly divided into 3 groups (n =10 for each grop) : sham group, septic group and UTI group.All rats received intraperitoneal injections of 0.9% saline (10 mL/kg) after and 8 h after surgery.In UTI group, UTI (10 × 104 U/kg in 10 mL/kg saline) was injected after and 8 h after surgery.Collect blood samples after 0, 8, 12 h after surgery to examine levels of procalcitonin (PCT), intestinal fatty acid binding protein (iFABP) and diamine oxidase (DAO) by enzyme-linked immunosorbent assay (ELISA) method.Rats were killed 12 h after surgery to collect intestine tissue samples.Pathological changes of intestine were observed under microscopy, and the expression of tight junction protein-1 (ZO-1) and occludin were analyzed by Western blot.Results In sham group, the mucosa structure was complete and the shape was normal, and villi stood neatly.In septic group, intestinal was expanded, intertinal mucosal was atrophic, villi were scanty.An inflammatory infiltrate with numerous nuetrophils was found in the mucosal.In UTI group, the level of severity was relatively slight.The relative optical density of Western blot images were decreased on ZO-1 and occludin in CLP and UTI groups, and decreased more in CLP group (F =43.15 and 52.23, P < 0.05).At 0h after surgery, the plasma values of PCT, iFABP and DAO were similar in three groups (F =11.17, 22.45 and 13.58, P > 0.05).At 8h and 12h after surgery, values of PCT, iFABP and DAO in septic and UTI groups were much higher than those in the sham group, and those in UTI group were also significantly higher than those in septic group (F8h=85.26, 44.59 and 101.47, F12h =59.44, 49.26 and 69.57, all P<0.05).PCT, iFABP and DAO levels were first increased and then fell down in sham group, those in septic group were keeping increasing, and those in UTI group were first increased and then kept stable.Conclusion UTI shows protective effect to intestinal barrier function in rats with sepsis.
8.Protective effects of propofol preconditioning on myocardium against hypothermia ischemia normothermia reperfusion injury on isolated rat hearts
pei-liang, WANG ; shan-shan, ZHU ; yin-ming, ZENG
Journal of Shanghai Jiaotong University(Medical Science) 2006;0(08):-
Objective To investigate the protective effects of propofol preconditioning on myocardium against hypothermia ischemia normothermia reperfusion injury on isolated rat hearts. Methods The Langendorff apparatus was used.Sixty SD rat hearts were divided randomly into 5 groups after 20-minute equilibrium(n=12): control(Con) group,hearts were continually perfused with K-H buffer for 175 min;ischemia/reperfusion(I/R) group,hearts were perfused with K-H buffer for 40 min,then subjected to global ischemia at 27 ℃ for 75 min,and followed reperfusion at 37 ℃ for 60 min;propofol preconditioning group 1(P1),group 2(P2),and group 3(P3),hearts were perfused with K-H buffer including 50,100,and 150 ?mol/L propofol for 10 min and followed reperfusion like I/R group,respectively.Heart rate(HR),left ventricular end-diastolic pressure(LVEDP), left ventricular developed pressure(LVDP) and ?dp/dtmax at the end of equilibration,pre-ischemia and at the end of reperfusion were recorded.The contents of creatine kinase(CK) and lactate dehydrogenase(LDH) in coronary effluent were measured at the end of equilibration and 1,10,20,30,and 60 min during reperfusion.The activity of superoxide dismutase(SOD) and the contents of maleic dialdehyde(MDA) were measured at the end of reperfusion.The area of infarct region was determined at the end of reperfusion. Results HR,LVDP,?dp/dtmax and SOD activity in P2 and P3 group were higher than those in I/R group(P
9.Protective effect of Ulinastatin on intestinal barrier function of elderly patients with sepsis
Haiyan YIN ; Pei TAO ; Xiaoling YE ; Yuee CHEN
Chinese Journal of Emergency Medicine 2016;25(2):177-181
Objective To observe the effect of Ulinastatin (UTI) on intestinal barrier function of elderly patients with sepsis in order to explore an optimized program of early treatment for such patients instituting a guidelines for clinical practice.Methods All selected patients randomly (ramdom number) divided into conventional treatment group (n =34) and UTI treatment group (n =34).Peripheral venous blood samples of elderly patients with sepsis were routinely taken at days 0,2,3,7 (D0,D2,D3,D7) after admission for detecting the changes of ptasma values of Intestinal fatty acid binding protein (IFABP),Diamine oxidase (DAO),and D-lactate (D-Lac).While patients' APACHE Ⅱscores,length of ICU stay and 28 d mortality also were recorded at the same time.Results At admission,there were no significant differences in patients' APACHE Ⅱscores,plasma values of IFABP,DAO and D-Lac between two groups (P > 0.05).During the course of treatment,the patients' APACHE Ⅱscores,plasma values of IFABP,DAO and D-Lac were declining in both groups,while those declinations in UTI group showed significantly more prominent.Compared with conventional therapy group,IFABP level was significandy lower in UTI group (P < 0.05),and the plasma values of DAO and D-Lac were significantly lower at D2 and D3 in UTI group (P < 0.05),but not at D7 (P > 0.05).The plasma levels of IFABP,DAO and D-Lac were positively associated with APACHE Ⅱscores,but not with length of ICU-stay (P >0.05).After treatment,28d mortality did not differ significantly between the two groups (P >0.05).Conclusions UTI has a protective effect on intestinal barrier function in elderly patients with sepsis.UTI can improve the prognosis of elderly patients with sepsis and reduce ICU stay,while has no significant effect on reducing 28 d mortality.
10.Expression of Stat5 in thyroid carcinoma and its relationship with EMT
Yun XUE ; Daoming LI ; Jing ZHANG ; Pei HUANG ; Yuhui YIN
The Journal of Practical Medicine 2017;33(6):905-908
Objective To study the expression of Stat5,E-cadherin and Vimentin in thyroid carcinoma and the correlation among them. Method The expression of Stat5,E-cadherin and Vimentin in 149 cases of thyroid specimens(including 23 cases of nodular goiter and 126 cases of thyroid carcinoma)was detected by immunohisto-chemistry SP method and the correlation analysis was conducted with the main clinical pathological parameters. Results The positive rate of Stat5 and Vimentin in thyroid carcinoma was 85.71% and 77.78% respectively, which was significantly higher than that in nodular goiter(26.09%and 8.70%)(P<0.05)and the positive rate of E-cadherin in thyroid carcinoma was 25.40%,lower than that in nodular goiter(86.96%)(P<0.05). Three kinds of protein expression were not significantly correlated with gender and age (P > 0.05),but obviously with lymph node metastasis and pathological type(P<0.05). Stat5 was negatively correlated with the expression of E-cadherin (r=-0.335,P=0.000)but positively with the expression of Vimentin(r=0.218,P<0.05). Conclusion The high expression of Stat5 and EMT in thyroid carcinoma tissues indicate that Stat5 may be involved in the EMT process of thyroid carcinoma ,thus it can promote the invasion and metastasis of thyroid carcinoma.