1.Improvements in the establishment and evaluation of cerebral ischemia models made by thread occlusion in rats
Bin HE ; Jinsong ZHANG ; Binxia SHAO ; Jie CHENG ; Fei HUANG ; Hang XIAO ; Rang GAO ; Xiang LU
Chinese Journal of Emergency Medicine 2009;18(12):1248-1251
Objective To modify the methods of operation and establishment of cerebral ischemia rat models made by thread occlusion. Method We randomly divided 120 male SD rats into a common group (n = 50), an improvement group (n = 60) and a sham-operated group (n - 10). Rats in the common and improvement groups were made into models using the common and improvement methods separately. All models were evaluated on the basis of physical sign indices and 2,3,5-triphenyltetrazolium chloride (TTC) staining. The TTC staining results were taken as gold standards. Then, we compared the achievement ratios of the two groups, and computed the sensitivity and specificity of every physical sign index based on these standards. The χ~2 or correction χ~2 test was used to compare the ratios. Results (1) The achievement ratio in the improvement group was significantly higher than that in the common group (71.67% vs. 52.00%, P = 0.034). (2) The sensitivity of evaluation for both common and improvement methods was 98.55%. However, the specificity of evaluation for the improvement method was significantly higher than that for the common method (100.00% vs. 40.00%, P =0.000). Conclusions The establishment achievement ratio and evaluation correctness of models are obviously elevated by modification of the operation and establishment methods.
2.Repairation of bone and skin defect in leg with vascularized tibial bone-skin flap graft
Fei REN ; Chun-Sheng CHENG ; Hong-Wei JIA ; Song-Feng LU ; Shao-Jun LUO ;
Chinese Journal of Microsurgery 2000;0(03):-
Objective To evaluate the efficacy of tibial bone-skin flap grafts in the management of se- vere traumatic osteomyelitis complicated with bone and skin defect in leg to avoid amputation.Methods From March 1998 to Aug.2004,12 cases of the traumatic osteomyelitis complicated with bone and skin defect in leg were treated with vascularized tibial bone-skin flap graft.The longest flap was 17cm,widethest 10cm, The longest bone flap was 12cm.They were followed up for 0.6 to 5 years.Results All the tibial bone-skin flaps survived completely,2 cases of osteomyelitis recurred.The followed-up,from 0.5 to five years,showed good bone union in all cases,averageing 15 weeks.The infection was under control.The leg function and con- tour were satisfactory.Conclusion The tibial bone-skin flap has the advantages of having distinguished sign of anatomy,highly vascularized,easy to obtain,simply and flexible procedure,improving circulation,short- ens hospitalization and suitable for treatment of traumatic osteomyelitis complicated with bone and skin defect in leg.
3.Expression and its significance of aquaporin 8 in patients with functional constipation or constipated irritable bowel syndrome
Sisi QIU ; Shiyang MA ; Yan CHENG ; Hui SHAO ; Lei DONG ; Fei DAI
Chinese Journal of Digestion 2016;36(8):538-542
Objective To detect the expression level of aquaporin 8 (AQP8) in patients with functional constipation(FC) or constipated irritable bowel syndrome (IBS-C),and the correlation between the expression of AQP8 and clinical features.Methods From March to December 2014,a total of 16 patients with IBS-C and 19 patients with FC met Rome Ⅲ criteria were collected,and nine healthy individuals were assigned to control group at the same period.The ascending and decending colonic tissues mucosa of FC,IBS-C and control group were taken under endoscope.The expression of AQP8 at mRNA and protein level was detected by real-time polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC).The differences in AQP8 mRNA expression and AQP8 relative area were analyzed by Kruskal-Wallis test among groups,and Pearson correlation coefficient was performed for correlation analysis between the expression and clinical features.Results The relative expressions of AQP8 mRNA of ascending colon and descending colon of FC patients (1.38,0.61 to 4.09;2.65,0.82 to 7.52) and IBS-C patients (2.23,0.82 to 4.67;1.35,0.51 to 2.03) were higher than those of control group (0.56,0.19 to 0.97;0.38,0.21 to 1.19),and the differences were statistically significant (ZFc =-2.435,-3.149,ZIBS-C =-2.690,-2.152;all P<0.05).AQP8 mRNA expression of descending colon in patients with FC was higher than that of patients with IBS-C,and the difference was statistically significant (Z =-2.003,P =0.045).The expression of AQP8 in patients with FC and IBS-C was positively correlated with disease course (ascending colon r=0.57 and 0.53;descending colon r=0.49 and 0.54,all P<0.05),and was negatively correlated with frequency of defecation (ascending colon r=-0.82 and-0.61;descending colon r=-0.49 and-0.53,all P<0.05).There was no correlation between the expression of AQP8 and age,gender,onset age,presence of abdominal symptoms of the patients (all P> 0.05).Most of AQP8 of FC group was expressed in cytoplasm of colonic mucosa epithelial cells,while that of IBS-C group and control group was mostly expressed at apical membrane and basal membrane of epithelial cells.The results of semi-quantification demonstrated that AQP8 relative area of descending colon of FC and IBS-C group increased compared with that of control group (3.42% (1.24% to 5.61%),2.45%(1.72% to 4.27%) vs 1.18% (0.35% to 2.81%);Z=-2.534,-2.151,both P<0.05).Meanwhile,AQP8 relative area of ascending colon of FC group increased compared with that of control group (2.46%(1.48% to 4.18%) vs 1.14%(1.29% to 2.15%) Z=-2.041,P<0.05).Conclusion There are differences in AQP8 expression quantity and location in cells of descending colon between patients with FC and patients with IBS-C,which is a way for differentiation these two diseases.
4.A cadaveric study of relationships among rotational alignment reference axes of distal femur and tibial mechanical axis.
Bao-hui ZHAO ; Bai-cheng CHEN ; De-cheng SHAO ; Fei WANG ; Shi-jun GAO ; Bo LU
Chinese Journal of Surgery 2008;46(14):1085-1087
OBJECTIVESTo investigate the relationships among rotational alignment reference axes of distal femur and tibial mechanical axis, and determine the safest rotational alignment reference axis.
METHODSDigital photos were taken of 30 cadaveric lower extremities with knee in extension and flexion at 90 degrees , angles were measured among tibial mechanical axis and a line perpendicular to clinical epicondylar axis, a line perpendicular to surgical epicondylar axis, Whiteside's line and femoral mechanical axis. Statistical analysis of relationships among those axes were performed.
RESULTSThe angles among the tibial mechanical axis and a line perpendicular to the clinical epicondylar axis, a line perpendicular to the surgical epicondylar axis, Whiteside's line and femoral mechanical axis were 0.6 degrees varus, 3.9 degrees varus, 0.2 degrees valgus and 3.0 degrees varus respectively. The angle between the femoral mechanical axis and the tibial mechanical axis was significantly larger than the angles among the tibial mechanical axis and a line perpendicular to the clinical epicondylar axis, the Whiteside's line (P < 0.05). There was no significant difference compared with the angle between a line perpendicular to the surgical epicondylar axis and the tibial mechanical axis. Angles of the clinical epicondylar axis, the surgical epicondylar axis and the Whiteside's line between knee extension and flexion were 2.3 degrees valgus, 0.9 degrees varus and 3.1 degrees valgus respectively.
CONCLUSIONThe surgical epicondylar axis rather than the clinical epicondylar axis or the Whiteside's line is the safest femoral rotational alignment reference axis intraoperatively.
Arthroplasty, Replacement, Knee ; Biomechanical Phenomena ; Femur ; anatomy & histology ; surgery ; Humans ; Knee Prosthesis ; Rotation ; Tibia ; anatomy & histology ; surgery
6.Construction of subtracted cDNA library by suppression subtractive hybridization for differentially expressed genes in eosinophils from asthma patients.
Jin-song XU ; Shao-xi CAI ; Fei ZOU ; Wan-cheng TONG ; Wei-ren WAN ; Hai-jin ZHAO
Journal of Southern Medical University 2006;26(1):82-85
OBJECTIVETo construct a subtracted cDNA library of differentially expressed genes in eosinophils from asthma patients.
METHODSSuppression subtractive hybridization (SSH) was used to isolate the cDNA fragments of differentially expressed genes in the eosinophils of asthma patients before and after treatment. The cDNA fragments were directly inserted into T/A cloning vector to establish the subtractive library, followed by amplification of the library through E. coli transformation with calcium chloride and screening of blue and white clones of the transformants. One hundred positive bacterial clones were randomly picked and identified by colony PCR.
RESULTSThe amplified library contained more than 3,000 positive bacterial clones. Analysis of the randomly selected 100 white clones by PCR showed that 90% of the clones contained 100-500 bp inserts, which might be the cDNA fragments of differentially expressed genes in eosinophils of asthma patients before treatment.
CONCLUSIONA subtracted cDNA library of differentially expressed genes in the eosinophils of asthma patients before and after treatment is constructed successfully by SSH and T/A cloning techniques, which lays a solid foundation for screening and cloning new specific differentially.expressed genes in the eosinophils of asthma patients.
Asthma ; blood ; genetics ; DNA, Complementary ; genetics ; Eosinophils ; metabolism ; Gene Expression Profiling ; Gene Expression Regulation ; genetics ; Gene Library ; Humans ; Male ; Middle Aged ; Nucleic Acid Hybridization
7.Reverse 201Tl myocardial redistribution induced by coronary artery spasm
Ding-cheng, XIANG ; Ji-lin, YIN ; Zhi-hua, GONG ; Zhen-hong, XIE ; Jin-he, ZHANG ; Yan-fei, WEN ; Shao-dong, YI
Chinese Journal of Nuclear Medicine 2010;30(5):300-303
Objective To investigate the mechanism of reverse redistribution (RR) on dipyridamole 201Tl myocardial perfusion studies in the patients with coronary artery spasm. Methods Twenty-six patients with coronary artery spasm and presented as RR on dipyridamole 201Tl myocardial perfusion studies were enlisted as RR group, while other 16 patients with no coronary artery stenosis nor RR were enlisted as control group. Dipyridamole test was repeated during coronary angiography. Corrected thrombolysis in myocardial infarction (TIMI) frame count (CTFC) and TIMI myocardial perfusion grade (TMPG) were measured at RR related and non-RR related coronary arteries before and after dipyridamole infusion respectively.All of the data were analyzed by Student's t-test orχ2-test and correlation analysis. Results Coronary artery angiography showed slower blood flow and lower myocardial perfusion in RR related vessels when compared with non-RR related vessels in RR group, but there was no significant difference among the main coronary arteries in control group. The perfusion defects of RR area at rest were positively related to slowerblood velocity at corresponding coronary arteries ( r = 0.79, t = 10.18, P < 0.001 ). In RR related vessels,CTFC were (36 ±6) frames and (26 ±7) frames (t =4.15, P <0.01 ), while TMPG were (2.02 ±0.39)grades and (2.92 ± 0.12) grades ( t = 2.25, P < 0.05 ) before and after dipyridamole infusion, respectively.In non-RR related vessels, CTFC were (29 ±7) frames and (25 ±5) frames (t =2.31, P <0.05), while TMPG were (2.56 ± 0.31 ) grades and (2.96 ± 0.06) grades ( t = 2.17, P < 0.05 ) before and after dipyridamole infusion, respectively. However, there were no significant changes of CTFC and TMPG before and after dipyridamole infusion in control group ( t = 0.932, 0.867, respectively, both P > 0.05 ). Conclusion RR is related to the decreased blood flow and myocardial perfusion induced by coronary artery spasm at rest,which may be improved by stress test such as intravenous dipyridamole infusion.
8.Metabolic heterogeneity of gastric cancer cell lines
Xia-Xia FAN ; Ying-Li ZHU ; Bei-Bei SHAO ; Lu-Lu LI ; Ya-Fei LI ; Ao JIA ; Ai-Feng WANG ; Yong-Cheng MA
Chinese Journal of Pharmacology and Toxicology 2018;32(4):278-279
OBJECTIVE Gastric cancer is one of the most common malignant tumors,and the inci-dence rate is the highest in all kinds of tumors in China. However,it remains unclear that how signifi-cantly gastric cells are dependent on glycolysis,and which type of gastric cells are sensitive to glycolysis inhibition. In this study, several kind of gastric cancer cell lines were used as the research object, and the metabolic characteristics of different cell lines were systematically analyzed to provide theoretical support for the accurate treatment of gastric cancer. METHODS We examined the energy metabolism of four gastric cancer cell lines(MGC-803,SGC-7901,HGC-27 and BGC-823)by using glycolysis inhibitor, 2-deoxy-D-glucose(2-DG)and inhibitor of oxidative phosphorylation,oligomycin.Oxygen consumption rates(OCR)and L-lactate were also measured with an XF96 Analyzer(Seahorse Biosciences)to deter-mine the significance of metabolism of oxidative phosphorylation and aerobic glycolysisin gastric cells. In addition, western blot was used to detect the contribution of AMP-activated protein kinase (AMPK), and anti-apoptotic proteins(Bcl-2 and survivin)to clarify the mechanism of death or survival of gastric cancer cells treated by 2-DG or oligomycin. RESULTS In this study, it was shown that the growth of gastric cell lines were suppressed by 2-DG.However,the sensitivity to 2-DG was quite different among cell lines:IC 50 of 2-DG was from 3.28 mmol·L-1(MGC-803)to 15.57 mmol·L-1(BGC-823).MGC-803 was relatively sensitive to 2-DG (IC 50:3.28 mmol·L-1), consumed more glucose and produced more lactate (waste product of glycolysis) than the three other cell lines. Consequently, MGC-803 could be more dependent on glycolysis than other cell lines, which was further confirmed by the fact that glucose (+)FCS(-)medium showed more growth and survival than glucose(-)FCS(+)medium.Alternatively, BGC-823, most resistant to 2-DG (IC50: 15.57 mmol·L- 1), was most sensitive to oligomycin, and showed more growth and survival in glucose(-)FCS(+)medium than in glucose(+)FCS(-)medium. Thus,we had reasons to think BGC-823 cells depended on oxidative phosphorylation for energy production. In BGC-823,AMPK,which is activated when ATP becomes limiting,was rapidly phosphorylated by 2-DG, and expression of Bcl-2 was augmented,which might result in resistance to 2-DG.Interestingly,AMPK phosphorylation and augmentation of Bcl-2 expression by 2-DG were not observed in MGC-803,which is 2-DG sensitive. CONCLUSION There is a large metabolic difference between gastric cancer cell lines,which will facilitate the future gastric cancer therapy by targeting metabolic pathways.
9.Anti-esophagus cancer activity and mechanism of DN3,a novel natural diterpenoid derivative,as a dual inhibitor of glycolysis and oxidative phosphorylation
Yong-Cheng MA ; Ying-Li ZHU ; Xia-Xia FAN ; Ao JIA ; Ya-Fei LI ; Bei-Bei SHAO ; Ai-Feng WANG
Chinese Journal of Pharmacology and Toxicology 2018;32(4):297-297
OBJECTIVE To probe into the anti-esophagus cancer activity and mechanisms of DN3, a novel natural diterpenoid derivative. METHODS The anti-tumor activity in vitro of DN3 was evaluated by MTT, and by using human esophageal carcinoma cells xenografted into athymic mice model in vivo. The specific mechanisms of DN3, as a dual inhibitor of glycolysis and oxidative phos-phorylation(OXPHOS)were explored through cell and molecular biology techniques.For instance,the manner of cancer cell death induced by DN3 was characterized by hoechst33342, FITC-Annexin V/PI staining and flow cytometric analysis,then these changes of glucose consumption,glucose uptake and lactate production in glycolysis, as well as oxygen consumption rate (OCR) and ATP content in OXPHOS caused by DN3 were performed separately through related kits and SeahorseBioscience XF24 Extra-cellular Flux Analyzer.Furthermore,in order to obtain a clear understanding of the inhibition of DN3 to glycolysis and OXPHOS, these regulatory factors were investigated by Western blot, such as PI3K/AKT, c-Myc and p53 of glycolysis, Bax and HK2 of mitochondrial function. RESULTS DN3 inhibited the growth of esophagus cancer cell EC9706, EC109 and EC1 cells in a dose and time dependent manner,but showed no significant effects on human esophageal epithelial cells(HEECs).DN3 caused significant G2/M arrest of esophagus cancer cell lines and induced apoptosis of these cell lines, which indicated DN3 inhibited the growth of esophagus cancer cell through blocking cell cycle and inducing apoptosis in a dose and time-dependent manner. Importantly, 8 μM DN3 decreased the extracellular acidification rate (ECAR) by 45% in EC109, which indicated glycolysis was inhibited by DN3. Mean-while, DN3 decreased the oxygen consumption rate (OCR) and the OCR linked to intracellular ATP production in EC109 cells,but that was not obvious in HEECs,so which indicated that DN3 could selec-tively block OXPHOS of cancer cells. In addition, the accumulation of reactive oxygen species (ROS) and the drop of mitochondrial membrane potential (MMP) were also observed in EC109 incubated by DN3,which suggested mitochondrial biological function was disturbed.Furthermore,the expression of PI3K/AKT, c-Myc and HK2 related to glycolysis were down-regulated by DN3, but the p53 and Bax were up-regulated in esophageal carcinoma cells. The changes of these enzymes accounted for the decreased glycolysisand OXPHOS in esophageal carcinoma cells treated by DN3. CONCLUSION The new compound DN3 has a strong anti-esophageal carcinoma activity,and it is tolerable that DN3 is seen as a dual inhibitor of glycolysis and oxidative phosphorylation.
10.Antitumor activityand mechanism of JD20,a newly synthetic analog of natural ent-kaurane diterpenoid
Ying-Li ZHU ; Xia-Xia FAN ; Ya-Fei LI ; Ao JIA ; Bei-Bei SHAO ; Ai-Feng WANG ; Yong-Cheng MA
Chinese Journal of Pharmacology and Toxicology 2018;32(4):346-346
OBJECTIVE To study the anti-tumor activity and molecular mechanism of natural diter-pene derivative JD20 in vitro. METHODS Screening the sensitive of gastric carcinoma cell lines to JD20 by cytotoxicity test for 24 h.Cell morphology was evaluated by using DAPI.After staining of can-cer cells with PI or annexin V-FITC/PI respectively,the cell cycle and apoptosis induced by JD20 were detectded by flow cytometry. The change in cell membrane potential was detected by JC-1 test kit. Western blot method was used to detect the apoptosis-related protein. RESULTS The novel natural kaurane diterpene derivative JD20 had a significant inhibitory effect on tumor cells and was particularly active on gastric cancer cell lines HGC-27 (IC50=4.72 ± 1.37 μmol·L- 1) and MGC-803 (IC50=7.36 ± 0.86 μmol·L-1).Further studies found that JD20 resulted in thecell cycle arrest in the G2/M phase,and induced a significant apoptosis in HGC-27. In addition, JD20 also caused the drop of mitochondrial membrane potential of HGC-27 within a short time (3 h). Furthermore, the Western blotting analysis showed that JD20 could induce the up-regulation of p53,Bax and Bim protein expression in gastric can-cer cells,and the releasing of cytochrome c from the mitochondria into the cytoplasm,as well as the ac-tivation of casepase-9/3.CONCLUSION The natural kaurane diterpene derivative JD20 can inhibit the proliferation of various human cancer cells by blocking the cell cycle and inducing apoptosis, and its mechanism of inducing apoptosis may be related to the mitochondria-mediated apoptosis pathway.