ObjectiveToinvestigatetheeffectsofcaveolin1(Cav1)onexpressionsofproinflammatory cytokine interleukin (IL)-1βand IL-6 in the ischemic cortex after permanent middle cerebral artery occlusion in mice. Methods The Cav-1 knockout mice (n=40) and wild-type mice (n=40) were randomly divided into ischemia groups and sham operation groups (n=20 in each group). They w ere also redivided into ischemia or sham operation at 3, 7, 10 and 14 d time points ( n=5 in each time point). A permanent middle cerebral artery occlusion model w as induced by the suture method. Immunohistochemical method w as used to detect the expressions of IL-1βand IL-6 in the ischemic cortex. Results The expression levels of IL-1βand IL-6 in the ischemic cortex at each time point in the ischemia group in Cav1 knockout mice w ere significantly higher than those in the ischemia group in the w ild-type mice ( al P< 0.05 ). Conclusions The upregulations of proinflammatory cytokines IL-1βand IL-6 in the ischemic cortex in Cav1 knockout mice suggests that Cav1 plays an important role in aleviating inflammation after cerebral ischemia.

Objective To investigate the identification and optimal breeding method of caveolin-1 knockout mice, and provide an ideal animal model for further study of the role of caveolin-1 in cerebral ischemic injury and repair. Meth?ods The introduced caveolin-1 gene knockout mice were reared in the SPF laboratory and genomic DNA was extracted from mouse tail tissue by the method of boiling lysis. According to the primer sequences provided by the Jackson Laboratory of America for polymerase chain reaction ( PCR) to detect the genotypes, with the four different ways of mating:caveolin-1 +/ -heterozygote intercrossing, heterozygous and homozygous caveolin-1 -/ -hybrid ( orthogonal and pay) as well as homo-zygous intercrossing. The pregnancy rate, shape characteristics of the filial generation mice and homozygous rate of the pa-rental mice were observed. Results Agarose gel electrophoresis results indicated that the size of molecular weight of the PCR products was about 200 bp and 661 bp, which were consistent with the expected target gene fragment, and identified caveolin-1 gene knockout mice of different genotypes successfully. The results of different mating patterns are basically in a-greement with Mendel rule, and the female and male aveolin-1 -/ -homozygous mice had a certain ability to reproduce, three different genotypes of mice had no significant differences between the shape features. Conclusions PCR can fast and reliably identify the genotypes of caveolin-1 knockout mice using genomic DNA through the method of boiling lysis. Combi- ning the breeding methods of intercrossing of caveolin-1 heterozygous mice and intercrossing of caveolin-1 homozygous mice may be a good way to obtain enough homozygous mice and homologous wild type mice in a short period.

Objective To investigate the CT,MRI features and differential diagnosis of central nervous system primitive neuroec-todermal tumors (cPNET).Methods The CT and MRI findings of 1 5 cases with cPNET proved by pathology were analyzed retro-spectively,and we summarized the imaging features and differential diagnosis.Results For this group,the average age was (8.82± 2.53)and the male to female ratio was 9 ︰6.All lesions located in supratentorial region,which had relatively large volume (average diameter of 6.3 cm),cystic necrosis (12/15),and no or mild peritumoral edema.cPNET showed isodensity,slight hyperdensity or slight hypodensity on CT plain scan,and demonstrated uniform or inhomogeneous enhancement.On MRI plain scan,solid part showed isointensity or slight hypointensity signal on T1 WI,isointensity or slight hyperintensity signal on T2 WI,hyperintensity sig-nal on DWI(12/15),isointensity(9/15)or slightly hyperintensity signal on FLAIR,and showed obvious uniform,honeycombed or irregular enhancement after enhanced scan,no enhancement in cystic necrosis.Conclusion cPNET have certain characteristics,inclu-ding the lower onset age,relatively large volume with well-defined edge and no or mild peritumoral edema,hyperintensity signal on DWI,isointensity signal on FLAIR.

Ultrasonic thrombus ablation is a newly-developed technology for percutaneous arterial recanalization. An ultrasound angioplasty device is described here in detail. The device has an adjustable power output range and distal tip longitudinal displacement range. Experimental data suggest that this ultrasound device is significantly effective in ablating fresh thrombi.
Catheter Ablation ; instrumentation ; Equipment Design ; Expert Systems ; Thrombolytic Therapy ; Transducers ; Ultrasonography, Interventional ; Vibration

Objective To evaluate the efficacy and safety of ESHAP regimen,as a salvage regimen, in treating patients with relapsed or refractory aggressive NHL.Methods 38 patients with relapsed or refrac- tory aggressive NHL were selected to be treated by ESHAP regimen.Results The 38 patients received ES- HAP regimen with a range of 2～6 cycles. The total RR was 55.3 % with complete response(CR)rate of 26.3 %.The major toxicity was myelosuppression with infection,which was tolerable.Conclusion ESHAP regimen is one of safe and effective salvage regimens for the patients with relapsed or refractory aggressive NHL.

Objective To evaluate the protective effect of aminophylline on myocardium in the patients undergoing prothetic valve replacement operation of heart.Mothods Thirty patients undergoing prothetic valve replacement operation of heart were randomized to be treated either with aminophylline(n=15)or without aminophylline treatment(n=15). Aminophylline(5mg/kg)was injected intravenously at 15 minutes after induction of anesthesia.Cardiac troponin I(cTnI), cyclic adenosine monophosphate(cAMP),myeloperoxidase(MPO),ratio of aortic blood neutrophil count to coronary vein sinus blood neutrophil count,hemodynamics,time of aortic cress-clamping and other clinical data were recorded during the operation.Results There were no differences between the two groups in the major perioperative variables.Plasm cTnI concentration in both groups increased after off-clamping than that before CPB,however,it was lower in aminophylline group than that in control group.Concentration of cAMP in both groups after off-clamping was lower than that before CPB, however cAMP concentration in aminophylline group after off-clamping was higher than that in control group.Myocardial MPO activity and neutrophil count ratio after aortic off-clamping in aminophylline group was significantly lower than that in control group.Conclusion These results suggest that aminophylline is helpful to unprotection of myocardial and decreases the sequestration of neutrophil in myocardium.The mechanism of the protection may be related to the cAMP increased in myocardium.

Objective To investigate the effect and mechanism of high concentration glucose on cholesterol absorption of human colon cancer epithelial Caco-2 cells.Methods The experimental study was used.(1) CCK-8 detected cell proliferation:the proliferation rate changes of Caco-2 cells were detected by CCK-8 when different concentrations (12.5,100.0,300.0,700.0,1 000.0,1 388.0 mmol/L) of glucose solution effects on Caco-2 cells in order to ensure the half hindering concentration of glucose concentration on Caco-2 cells.(2)Cholesterol absorption of Caco-2 cells was detected:Caco-2 cells were divided into the cholesterol group,cholesterol plus ezetimibe (cholesterol inhibitor) group and blank control group.Cholesterol group:100 μmol/L cholesterol solution and different concentrations (5.0,25.0,50.0 mmol/L) of glucose solution were added.Cholesterol plus ezetimibe group:100 μmol/L ezetimibe,100 μmol/L cholesterol solution and different concentrations (5.0,25.0,50.0 mmol/L) of glucose solution were added.Blank control group:DMEM culture medium and corresponding concentrations of DMSO were added.The cholesterol absorption amounts of Caco-2 cells were measured.(3) The relative expressions of ATP binding cassette G8 (ABCG8),ATP binding cassette G5 (ABCG5),Nickman-Pick CI Like 1 (NPC1L1) and scavenger receptor class B type Ⅰ (SR-B Ⅰ) were examined by Western blot in the different groups.Caco cells were divided into the glucose group,glucose plus ezetimibe group and control group.The different concentrations (5.0,25.0,50.0 mmol/L) of glucose solution were added into the glucose group,different concentrations (5.0,25.0,50.0 mmol/L) of glucose solution and 100 μmol/L ezetimibe were added into the glucose plus ezetimibe group,and 100 μmol/L ezetimibe were added into the control group.The relative expressions of ABCG8,ABCG5,NPC1L1 and SR-B Ⅰ were detected by Western blot.Measurement data were presented as (x) ±s,repeated measure variance analysis was used to perform variation trend test,and t test was utilized to conduct comparisons among groups.Results (1) CCK-8 results showed:proliferation rates of Caco-2 cells with the glucose solution concentration of 12.5,100.0,300.0,700.0,1 000.0 and 1 388.0 mmol/L were 1.380 ±0.043,1.238 ±0.072,0.736 ±0.035,0.336 ±0.021,0.316 ±0.020 and 0.288 ±0.010,respectively,with a statistically significant difference in the proliferation rates (F =11.019,P ＜ 0.05).The half hindering concentration of glucose solution on Caco-2 cells was 283.54 mmol/L.(2)Cholesterol absorption of Caco-2 cells:① the cholesterol absorption amounts of Caco-2 cells with the glucose solution concentration of 5.0,25.0 and 50.0 mmol/L were 0.282 ± 0.042,0.380 ± 0.063,0.390 ± 0.060 in the cholesterol group and 0.042 ± 0.012,0.197 ± 0.015,0.277 ± 0.029 in the cholesterol plus ezetimibe group,respectively,showing a statistically significant difference between the 2 groups (F =55.566,P ＜ 0.05).②There was a statistically significant difference in cholesterol absorption amounts of Caco-2 cells with different glucose solution concentration in the cholesterol group (F =79.117,P ＜ 0.05).The cholesterol absorption amounts of Caco-2 cells with the glucose solution concentrations of 5.0 mmol/L was lower than that with the glucose solution concentrations of 25.0 mmol/L and 50.0 mmol/L,respectively (t =11.207,11.532,P ＜0.05).There was no statistically significant difference in the cholesterol absorption amounts of Caco-2 cells between the glucose solution concentrations of 25.0 mmol/L and 50.0 mmol/L (t =12.389,P ＞ 0.05).③There were statistically significant differences in cholesterol absorption amounts of Caco-2 cells with the glucose concentration of 5.0 mmol/L and 25.0 mmol/L between cholesterol group and cholesterol plus ezetimibe group (t =10.908,10.644,P ＜ 0.05).(3) The results of Western blot showed:① the relative expression of NPC1L1 protein in Caco-2 cells with the glucose solution concentrations of 5.0,25.0 and 50.0 mmol/L were respectively 0.277 ±0.019,0.558 ±0.015,0.576 ±0.003 in the glucose group and 0.057 ±0.002,0.054 ±0.005,0.077 ±0.005 in the glucose plus ezetimibe group,showing a statistically significant difference (F =482.207,P ＜0.05).② The relative expression of NPC1L1 protein of Caco-2 cells with the different concentration of glucose solution in the glucose group were compared,with a statistically significant difference (F =8.112,P ＜ 0.05).There was a statistically significant difference in the relative expression of NPC1L1 protein in Caco-2 cells with the different concentration of glucose solution in the glucose plus ezetimibe group (F =11.708,P ＜ 0.05).③ The relative expression of NPC1L1 protein in Caco-2 cells with the glucose solution concentrations of 5.0,25.0 and 50.0 mmol/L in the glucose group was statistically different from that in the glucose plus ezetimibe group (t =8.112,11.708,13.920,P ＜ 0.05).Conclusion High concentration glucose solution could promote the reabsorption of cholesterol through increasing NPC1L1 protein expression in Caco-2 cells,and increase the risk of suffering from cholelithiasis in diabetes patients.

AIM:To investigate the proliferation property of stable chemerin gene knockdown vascular smooth muscle cells ( VSMCs) and to explore its mechanism .METHODS:The normal VSMCs , chemerin gene interfering control VSMCs and stable chemerin gene knockdown VSMCs were divided into normal group , PDGF group, control group and knockdown group .The VSMCs in PDGF group were given platelet-derived growth factor-BB ( PDGF-BB) to initiate proli-feration.The cell counting and BrdU assay were employed to investigate the proliferation property of VSMCs .The mitogen-activated protein kinase ( MAPK) signal pathway was determined by Western blot .RESULTS:The cell number and BrdU A value in PDGF-BB-treated VSMCs significantly increased as compared with the normal VSMCs ( P<0.05 ) .Compared with the normal VSMCs , the chemerin knockdown VSMCs showed obviously decreased cell number and BrdU A value ( P<0.05).Simultaneously, no significant difference in the proliferation of VSMCs between the normal VSMCs and the control VSMCs was observed.No significant difference of the protein levels of p-ERK1/2, ERK1/2, p-p38 MAPK and p38 MAPK among 4 kinds of VSMCs was found .The protein level of p-JNK in PDGF-BB-treated VSMCs was up-regulated, while it was down-regulated in chemerin knockdown VSMCs compared with the normal VSMCs .CONCLUSION: Chemerin pro-motes the proliferation of mouse vascular smooth muscle cells by up-regulating p-JNK production .

Hypermethylation of the promoter region is one of the major mechanism of tumor suppressor gene inactivation. In order to provide a research tool for the study on the function of MBD1 gene in DNA methylation and tumorigenesis, antisense MBD1 gene eukaryotic expression plasmid was constructed and transfected into human biliary tract carcinoma cell line QBC-939 to observe its effect on the expression of MBD1 mRNA and protein by using RT-PCR and FCM respectively. Following the transfection, the mRNA level of MBD1 gene decreased from 0. 912 +/- 0.022 to 0.215 +/- 0. 017, and the protein level of MBD1 gene also decreased from (80.19 +/- 5.05) % to (35.11 +/- 4.05) %. There were very significant differences in the expression both at the transcription and post-transcription levels of MBD1 gene between non-tranfection group and the antisense MBD1 gene eukaryotic expression plasmid transfection group (P < 0.01). It was suggested that transfection with the antisense MBD1 gene eukaryotic expression plasmid can significantly reduce the expression level of MBD1 gene in QBC-939, and this study may provide a valid tool for the investigation of the function of MBD1 gene and its role in biliary tract carcinoma.
Biliary Tract Neoplasms/*metabolism ; Biliary Tract Neoplasms/pathology ; Cell Line, Tumor ; DNA Methylation ; DNA-Binding Proteins/*biosynthesis ; DNA-Binding Proteins/genetics ; Eukaryotic Cells/metabolism ; Gene Expression Regulation, Neoplastic ; Genetic Vectors ; Oligonucleotides, Antisense/*genetics ; Plasmids/genetics ; Transcription Factors/*biosynthesis ; Transcription Factors/genetics ; Transfection

Hypermethylation in the promoter region is an important epigenetic mechanism for the transcriptional repression of a number of cancer-associated genes, and over-expression and/or increased activity of DNA methyltransferases are considered to be the main cause of promoter hypermethylation. In order to explore the roles of two methyltransferase members (DNMT1 and DNMT3b) in the cholangiocarcinoma tumorigenesis, antisense eukaryotic expression plasmid of DNMT1 and DNMT3b gene was constructed respectively, and were co-transfected into the human cholangiocarcinoma cell line QBC-939 to observe their biological effects on the cell growth and proliferation ability, apoptosis, cell cycle alteration, and the tumorigenesis ability in the subcutaneous tissue of nude mouse. The results demonstrated that co-transfection with antisense eukaryotic expression plasmid of DNMT1 and DNMT3b gene and single transfection with antisense eukaryotic expression plasmid of DNMT1 gene can suppress the growth and proliferation of QBC-939, block the cell cycle at G1 phase, increase the apoptosis rate, minimize the tumor size in the subcutaneous tissue of nude mouse. The suppressing biological effect of co-transfection is stronger than single transfection with antisense DNMT1. Meanwhile, single transfection with antisense eukaryotic expression plasmid of DNMT3b gene has no effects on the biological characteristics of QBC-939. This study suggests that DNMT1 gene plays a key role in DNA methylation and DNMT3b gene may act as an accessory to support its function in inactivation of tumor suppressor genes. Combination DNMT1 and DNMT3b will increase their biological effects and have the synergistic effect on suppressing the growth of human cholangiocarcinoma cell line QBC-939.
Apoptosis ; Biliary Tract Neoplasms/*metabolism ; Cell Line, Tumor ; Cell Proliferation ; Cholangiocarcinoma/*metabolism ; DNA (Cytosine-5-)-Methyltransferase/*biosynthesis ; DNA (Cytosine-5-)-Methyltransferase/genetics ; Down-Regulation ; Gene Expression Regulation, Neoplastic ; Genetic Vectors ; Mice, Nude ; Neoplasm Transplantation