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• 2013 Volume 23 Issue 6
  Published: 30 November 2013

    

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  2013, 23(6): 1.

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  Effects of FoxM1 downregulation by small interfering RNA on chemosensitivity of human colon cancer cell

  HE Ya-Long-1, Wang-Xiao-Yan-1, Zhang-You-Li-1 , Fan-Yu-2

  2013, 23(6): 461.

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  Objective: To study the effects of Forkhead box protein M1（FoxM1）small interfering RNA（siRNA）on chemosensitivity and mechanism of human colon cancer cell. Methods: Three FoxM1 siRNAs were designed and constructed. The best effect of siRNA downregulation was evaluated by Real time PCR.All cancer cells were divided into different dose of siRNA groups,and RPMI 1640 instead of FoxM1 siRNA as the blank control group (ConA); after transfected for 48 h,all groups cells were treated with  irinotecan(Irinotecan, CPT11, 7.5 μmol/L),respectively.The proliferation and chemosensitivity of cancer cells were evaluated by MTT assay,and the phosphorylation of Akt protein was exmined by Western blot.Results: FoxM1 siRNA could downregulate the FoxM1 expression in a dose and time dependent manner. The MTT results showed that the inhibit rates were  18.36%,39.78%,58.18%,78.28% in different groups\[ConA,siRNA 3.125 nmol/L, siRNA 6.25 nmol/L and siRNA 12.5 nmol/L, respectively. Statistical analysis showed that inhibition rates were in a dose dependent(r=0.775，P<0.05）. The phosphorylation of Akt protein was inhibited compared with ConA group. Conclusion: FoxM1 siRNA could sensitize human colon cancer cells to chemotherapy through inducing phosphorylation of Akt protein.  ［Key words］colon carcinoma; forkhead box protein M1; chemosensitivity; Akt
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  Isolation, culture and identification of rat bone marrow derived endothelial progenitor cells

  LU Yao-Liang-1, Kang-Tao-1, Li-Xiao-Qiang-2, Han-Song-1

  2013, 23(6): 465.

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  Objective: To explore the culture, differentiation and identification method and biological characteristics of the rat bone marrow derived endothelial progenitor cells (EPCs)．Methods: We separated the rat bone marrow mononuclear cells using density gradient centrifugation combined with differential sidewall separation, cultured endothelial cell with EGM-2, obtained adherent cells within 48 h (early EPCs) and adherent cells after 48 h(late EPCs), respectively. The cell morphological images were observed under inverted microscope; EPCs surface antigen markers, CD34, CD133 and VEGFR-2 were identified by the flow cytometry, the phagocytosis of EPCs were detected by laser scanning confocal microscope, the internal ultrastructure of EPCs were observed by transmission electron microscope. Results: Mononuclear cells were small round after the first incubation, the late EPCs was given priority to spindleshaped, and there was obvious stem cell colony after culture in 7 days; till the 6th generations of EPCs, the cellular morphology was gradually close to the endothelial cells, appeared the tube cavity structure. CD34, CD133, VEGFR2 expressed positive, could absorb Dil-acLDL combined with FITC-UEA-1. There was characteristic organelles Weible-Palade (W-P) small body in the EPCs under inverted microscope. Conclusion: The EPC was successfully isolated from bone marrow, and purified, cultured by density gradient centrifugation combined with differential adherent method; it had strong proliferation ability, abundant cell count and stable biological characteristics.  ［Key words］endothelial progenitor cells; bone marrow; stem cells
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  Value of staining with TTC to instantly judge the area of  coagulation by microwave

  ZHANG Jun-1, Zhang-Chi-Min-2

  2013, 23(6): 470.

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  Objective: To investigate the significance of staining with 2,3,5Triphenyltetrazolium chloride (TTC) to judge the area of coagulation by microwave quickly. Methods: Rabbits undergo hepatic coagulation at 45W for 3 minutes under US-guided percutaneous microwave coagulation, and to observe TTC as well as other results under the light, electron microscope and ultrasound. Results: The areas of coagulation necrosis can not be stained with TTC, but the noncoagulated area stained red with TTC. The cell morphology of liver stained with TTC can be observed under the light microscope.Conclusion: TTC can judge the area coagulated by microwave immediately, simply and truly. The area without TTC is coagulated area, and red area is non-coagulated area.  ［Key words］triphenyltetrazolium chloride staining； microwave/therapeutic use； ultrasonography; rabbit
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  Effects of hydrogenrich saline solution on renal ischemia/reperfusion injury in aging rats

  LIU Ji-Pu, Xu-Xian-Lin, He-Xiao-Zhou, Fan-Min

  2013, 23(6): 475.

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  Objective: To study the effects and mechanism of hydrogenrich saline solution on renal ischemia/reperfusion injury(RIRI) in aging rats. Methods: Thirty Sprague-Dawley rats 20 months aged were randomly divided into three groups: sham-operated group, ischemia/reperfusion group(IR), hydrogen-rich saline solution treatment group(H2  group)； each group had 10 rats.Once established the ischemia/reperfusion models,the venous blood samples of each group were taken for determination of serum BUN and Cr.The left kidney were removed for examination such as microscopic pathology by HE stain,MDA,superoxide dismutase(SOD),8-OHDG content,mRNA level of hemeoxygen-1(HO-1) and Caspase-3 and protein expression by RT-PCR and immunohistochemistry. Results: The BUN,Cr,MDA and 8-OHDG of IR group were higher than the sham-operated group and the H2  group(P＜0.05),but the level of SOD in IR group was lower than the sham-operated group and the H2  group(P＜0.01).The pathology change in the H2 group were significantly decreased compared with the IR group,the protein and mRNA expression of Caspase-3 in IR group was higher than that of sham group(P＜0.05) and  H2 group (P＜0.01).The expression of HO-1 in H2 group was higher than that of other groups(P＜0.01). Conclusion: The hydrogen-rich saline solution had protective affection to the RIRI and such effect may be related to the anti-oxidant and inhibiting cell apoptosis of the hydrogen.Ischemia reperfusion can cause significant oxidative stress of kidney tissues,causing damage.  ［Key words］hydrogen; ischemia/reperfusion injury; aged rats; renal; antioxidant agent  
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  High performance liquid chromatography determination method for three monomers of ginkgolic acid in mouse plasma

  YANG Xiao-Ming-1, Shen-Jie-Hong-2, Li-Yue-Ying-3, Lin-Jian-Wei-1, Huang-Bing-Zhong-4

  2013, 23(6): 480.

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  Objective: To develop a high performance liquid chromatography (HPLC) method for determining three monomers of ginkgolic acids, GA 13∶0, GA 15∶1 and GA 17∶1,  in mouse plasma. Methods: Plasma samples were treated with acetonitrile for protein precipitation. The analytical column was SinoChrom ODSAP C18(250 mm×4.6 mm, 5 μm).The mobile phase consisted of a mixture of methanol3% acetic acid (92∶8) at a flow rate of 1.0 mL· min^-1. The detection wavelength was at 310 nm. Results: Standard curves of GA 13∶0, GA 15∶1 and GA 17∶1 were linear in the range  of 0.135-20 μg·mL^-1, 0.129-20 μg·mL^-1, and 0.211-22 μg·mL^-1, respectively (both r＞0.9990). The lower limit of quantification were 0.135, 0.129 and 0.211 μg·mL^-1, respectively. The extraction recoveries＞70.2% (n=5) and the accuracies were 95.2%-101.5%. The intra-day and inter-day precisions were lower than 6.2% and 9.5 % (n=5), respectively. Conclusion: The HPLC method was successfully built for determining GA 13∶0, GA 15∶1 and GA 17∶1, which was simple and  sensitive, accurate with  reproducible, and suitable for clinical pharmacokinetic study.  ［Key words］ginkgolic acid monomer; high performance liquid chromatography; mouse
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  Bioinformatics analysis of rpoS gene in Serratia plymuthica

  GUO Song, Liu-Xiao-Guang

  2013, 23(6): 485.

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  Objective: To analyze the rpoS gene in Serratia plymuthica G3 using bioinformatics approaches to unravel its nucleotide and amino acid sequences,physics, chemistry and structure features with hope in providing help in further studies of the biological function of RpoS in G3.Methods:  A series of bioinformatics software were used to predict information on sequence, physical, biochemistry and structure features of RpoS in G3. Results:  RpoS was a hydrophilic protein without trans-membrane domain or N-signal peptide. Further analysis revealed that RpoS contained two N-glycosylation sites and 22 phosphrylation sites with a primary secondary structure of α-helix and random coil. Conclusion:  The sigma subunit RpoS in strain G3 characterized by a high level of phosphrylation and the secondary structure composed of α-helix and random coil was consistent in its biological function as a signal integrator and network master regulator to be involved in the general stress response in bacteria.  ［Key words］Serratia plymuthica; RpoS; general stress response; master regulator; bioinformatics
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  Effects of postconditioning with sevoflurane on iNOS/NO pathway  in rats with acute lung injury induced by lipopolysaccharide

  YANG Fen, Zhao-Jian-Hua

  2013, 23(6): 489.

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  Objective: To investigate the effects of postconditioning with sevoflurane on inducible nitric oxide synthase(iNOS)/nitric oxide(NO) pathway in rats with acute lung injury induced by lipopolysaccharide. Methods:  Thirty rats were randomly divided into 5 groups(n=6)，NS group and LPS group, rats received intratracheal instillation of normal saline(1 mL/kg) and lipopolysaccharide(5 mg/kg), respectively; sevoflurane groups(S 0.5, 1, 2 h), 4 h after lipopolysaccharide intratracheal instillation(i.e. after the occurrence of acute lung injury), rats received sevoflurane inhalation(2.4%) for 0.5, 1, 2 h, respectively, simulating a postconditioning scheme. Lung tissue samples were harvested 6 h after normal saline or lipopolysaccharide intratracheal instillation. Histological examination by light microscopy was performed; iNOS expression and NO concentration in lung parenchyma were measured by immunohistochemistry and with NO Assay Kit. Results: Compared with the NS group, iNOS expression and NO concentration in the LPS group were significantly increased(P<0.01), with more severe lung injury. Compared with the LPS group, iNOS expression and NO concentration in S groups were greatly reduced(P<0.01 or P<0.05), with attenuated lung injury, especially in S1h and S2h groups. Conclusion: Postconditioning with sevoflurane could attenuate lipopolysaccharideinduced acute lung injury in rats through inhibiting iNOS/NO pathway activation and reducing the inflammatory response.  ［Key words］sevoflurane; lipopolysaccharide; acute lung injury; postconditioning; inducible nitric oxide synthase; nitric oxide  
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  Relationship between HBV basic core promoter coexistent mutation of  1762/1764 sites and viral genotypes or HBeAg seroconversion

  XU Jun-1, Zhang-Yuan-Hai-2, Zheng-Zhong-Wei-1, Wu-Jian-Ye-1, Ge-Guo-Hong-2, Shao-Jiang-Bo-2

  2013, 23(6): 493.

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  Objective: Based on the results of the incidence rates of the basic core promoter (BCP) coexistent mutation of 1762/1764 sites and the expression of hepatitis B virus (HBV) e antigen (HBeAg) and its antibody in chronic hepatitis B(CHB) patients with common viral genotypes in China, to investigate the relationship between the coexistent mutation and both of viral genotypes and HBeAg seroconversion in the natural course of chronic HBV infection.  Methods: The sera of 168 CHB patients randomly selected with no previous history of antiviral therapy were detected. Their viral genotypes and coexistent mutation were determined by probe-specific real-time polymerasechain reaction (PSRT-PCR) that applied TaqMan MGB probe which had been designed and made exactly in past time; and HBeAg and its antibody(HBeAb) titer were measured by chemiluminescence microparticle immunoassay(CMIA). Results: The coexistent mutation in BCP 1762/1764 sites was significantly more frequent in genotype C HBV patients than that in genotype B patients (70.65% vs 29.17%, P=0.000). The wild genotype B group had a significantly higher proportion of HBeAg-negative HBV patients than the coexistent mutation genotype B group (73.53% vs 28.57%, P=0.004) and the wild genotype C group (73.53% vs 22.22%, P=0.000); the coexistent mutation genotype C group had a significantly higher proportion of HBeAg-negative HBV patients than the wild genotype C group(63.08% vs 22.22%, P=0.000) and the coexistent mutation genotype B group(63.08% vs 28.57%, P=0.018). In all HBeAg-negative patients, the wild genotype B group had a more higher HBeAbpositive rate than the coexistent mutation genotype B group(84.00% vs 0.00%, P=0.003) and the wild genotype C group (84.00% vs 16.67%, P=0.004), while there was no significant difference between the coexistent mutation genotype C group and the wild genotype C group(19.51% vs16.67%, P=1.000) and the coexistent mutation genotype B group(19.51% vs 0.00%, P=1.000). Conclusions: During the natural course of chronic HBV infection in China, the BCP coexistent mutation of 1764/1762 sites was more common in patients with genotype C, compared with genotype B; and the thorough HBeAg seroconversion occurred more frequently in wild genotype B HBeAg-negative patients, with HBeAb-positive; while the halfway HBeAg seroconversion occured in coexistent mutation genotype C HBeAg-negative patients, without HBeAb-positive.  ［Key words］hepatitis B virus; basic core promoter; mutation; genotype; hepatitis B e antigen seroconversion
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  Changes of cyclophilin A levels in patients with coronary heart disease

  YUAN Wei, Yan-Jin-Chuan, Wang-Dan-Qing, Liang-Yi, Wu-Jun

  2013, 23(6): 498.

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  Objective: To investigate the peripheral blood of cyclophilin A levels in the patient with coronary heart disease (CHD) and its correlation with BNP、TnI. Method:A total of 127 patients who underwent coronary angiography were selected as research subjects. They were divided into three groups by coronary angiography: 41 cases of patients with stable angina as SAP; 43 cases of patients with acute coronary syndrome as ACS, including two groups, 20 cases of unstable angina as UAP, 23 cases of myocardial infarction as AMI; 43 cases of patients with non-coronary heart disease as control group. The blood of all patients were collected after in hospital, the AMI group was collected blood again after 1 week as recovery period. The cyclophilin A  levels of all patients were measured with Elisa.Result:The cyclophilin A level in the ACS group was（24.2±2.7）ng/mL, and was higher than that in the SAP group (16.2±2.4)ng/mL and the control group (7.8±1.1)ng/mL（P<0.05）, but there was no significant difference between the AMI group (24.1±2.4)ng/mL and UAP group(24.5±3.1)ng/mL. The cyclophilin A level in the AMI group (after 1 week) was lower than that in the acute stage \[(17.9±3.0 )ng/mL vs（24.1±2.4）ng/mL）]（P<0.05）. In the ACS group, the BNP, TnI levels were higher than the control group; the LVEF was lower than the control group. The cyclophilin A levels had positive correlation with BNP, TnI（r=0.893，P<0.05；r=0.765，P<0.05）, and had negative correlation with LVEF（r=－0.528，P<0.05）. Conclusion:The cyclophilin A levels had close relation to the stability of coronary heart disease, and maybe could predict the occurrence of thrombotic events in patients with CHD.  ［Key words］coronary heart disease； cyclophilin A； Btype natriuretic peptide；  troponinI
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  Analysis of the MT2A-838G/C and the MCP-1 -2518G/A gene  polymorphism in carotid arteriosclerosis

  PAN Li-Li, Zhang-Jun, Wei-Jiang, Shi-Yuan-Ping, Feng-Yue-Hua, Yu-Yang, Luo-Guang-Hua

  2013, 23(6): 501.

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  Objective: To assess the potential association of the MT2A-838G/C and the MCP-1 -2518G/A gene in patients with carotid arteriosclerosis (CA) in Chinese han population of Jiangsu. Method:  A total of 103 consecutive patients with a diagnosis of CA and compared with 46 healthy controls. The polymorphism were detected by basequenched probe technique, the genotypes and allele frequencies of MT2A-838G/C and MCP-1-2518G/A were calculated. Results: The genotype GG, GC, CC distribution of MT2A were 50.49%, 36.89%, 12.62% in patients with CA, and were 52.17%, 36.96%, 10.87% in controls, respectively (P>0.05).Allele G and C frequency of MT2A in patients with CA were 70.65%, 29.35%, and were 68.93%, 31.07% in controls, respectively (P>0.05). The genotype AA, AG, GC distribution of MCP-1 were 20.39%, 44.66%, 34.95% in patients with CA, and were 15.22%,52.17%, 32.61% in controls, respectively (P>0.05). Allele A and G frequency of MCP-1 in patients with CA were 41.30%, 58.70%,and were 42.72%,57.28% in controls, respectively (P>0.05).  Conclusion: The 838G/C of MT2A gene and the 2518G/A of MCP-1 gene may be not the independent risk factor for CA in Chinese Han population of Jiangsu.  ［Key words］carotid arteriosclerosis; metallothioneins; single nucleotide polymorphism
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  Effects of tirofiban treatment on emergency percutaneous coronary  intervention in patient with acute coronary syndrome

  LIU Yi-Na, Zhu-Jian-Hua, Wu-Xiang, Wei-Mei-Fang

  2013, 23(6): 505.

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  Objective: To observe the effects of tirofiban treatment in emergency percutaneous coronary intervention(PCI) of acute coronary syndrome(ACS) patients.  Methods: A total of 210 hospitalized patients with ACS were selected, randomly divided into tirofiban group(108) and control group(102). Clinical data, interventional treatment, major adverse cardiac events during hospitalization, incidence of arrhythmia, bleeding complications and related events of two groups were compared. Results:The differences of clinical information, interventional treatment and bleeding complications with the two groups of patients were not statistically significant(P>0.05).Compared with the control group, the incidence of major adverse cardiac events of tirofiban group was significantly reduced during hospitalization. The incidence of bradycardia of tirofiban group was significantly lower than that of control group. Earlier treatment with  betablockade in our study was one of the outcomes  facilitated by tirofiban treatment. The patients put on tirofiban also experienced less ventricular arrhythmia mandating electric cardioversion/defibrillation. Conclusion: Tirofiban improved the effects of emergency interventional therapy and reduced arrhythmia during operation period in acute coronary syndrome patients.  ［Key words］tirofiban; acute coronary syndrome; percutaneous coronary intervention； arrhythmia
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  Mutation of Kras gene in lung cancer tissues obtained by fiberoptic  bronchoscopy and its clinical value

  CHEN Ping, Li-Jian, Zhu-Wen-Jing, Hang-Jian-Ming

  2013, 23(6): 509.

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  Objective: To explore the mutation of K-ras in biopsy samples of fiberoptic bronchoscopy(FB) in patients with lung cancer  and evaluate its clinical value of lung cancer.Method: PCR-single strand conformational polymorphism(PCR-SSCP) methods and silver staining technique was used to examine the mutation at K-ras for biopsy samples of FB in 52 patients with lung cancer and 9 patients with benign pulmonary disease. Results: The mutation rate of K-ras gene was 23.1%(12/52)and 0%(0/9) in lung biopsy samples in patient with lung cancer and benign pulmonary disease, respectively. The mutation of K-ras was obvious in adenocarcinoma cell carcinomas, but not significantly associated with patients age, sex, smoking history, cell differentiation or clinical stage of lung cancer.Conclusions: K-ras gene mutation may play important role in genesis and progression of lung cancer and may be an early event of lung cancer.Mutation of K-ras gene in biopsy samples of FB may be used to diagnose lung cancer.  ［Key words］lung cancer; fiberoptic bronchoscopy; K-ras gene; mutation
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  Expressions of estrogen receptor，progesterone receptor and human  epithelial receptor2 in elderly breast cancer patients and
   their clinical significance

  LU Li-1, Cui-Fei-Lun-2, Xiao-Xiu-Di-1, Xu-Xue-Song-1

  2013, 23(6): 513.

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  Objective: To analyze the expressions of estrogen receptor (ER), progesterone receptor (PR) and human epithelial receptor2(CerbB2) in elderly patients with breast cancer, and to investigate its clinical significance. Methods:  One hundred and twenty-five female breast cancer patients aged  65 years and older were enrolled in the study. Clinical data of all the patients were retrospectively reviewed. Expressions of ER, PR and CerbB2 were detected by immunohistochemistry. Correlation between expressions of ER, PR and CerbB2 and tumor size, histological grade, tumor location, lymph node metastasis were statistically analyzed. Results: The positive expressions of ER, PR and CerbB2 were 84(67.2%), 83(66.4%) and 25(20.0%), respectively. The expressions of ER and PR were correlated with histological grade, while CerbB2 expression was correlated with histological grade, tumor size and lymph node metastasis, respectively（both P<0.01）. Three and five years overall survivals of elder patients were 74.9% and 62.0%; the mean overall survival was 6.9 years. Conclusions:  The increased expressions of ER, PR and decreased  expression of CerbB2 in female breast cancer patients reflected the pathobiological features of the cancer, which would be helpful for the treatment and surgical operation of breast neoplasms.  ［Key words］estrogen receptor; progesterone receptor; human epithelial receptor2; elderly patients; breast cancer
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  Expression of HSP27 and TGF-β1 in papillary thyroid carcinoma and its significance

  CHEN Xia, Yuan-Guo-Yue, Wang-Dong, Wang-Ying, Li-Mei

  2013, 23(6): 517.

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  Objective: To investigate the expression and clinic significance of heat shock protein 27(HSP27) and TGF-β1 in papillary thyroid carcinoma.  Methods: The expression of HSP27 and TGF-β1 was detected by immunohistochemistry in 50 cases with papillary thyroid carcinoma， 50 cases with thyroid adenoma and 30 cases with normal thyroid tissues，which had undergone operation in department of surgery，and confirmed by pathology. Results: The cells which express HSP27 and TGF-β1 were stained brown or dark brown in their cytoplasm.We choose the cells stained biopsy to detect. The positive expression rate of HSP27 was 70%(35/50) with papillary thyroid carcinoma，48%(24/50) with thyroid adenoma，46.7%(14/30) with normal thyroid tissues and of TGF-β1 was 80%(40/50) with papillary thyroid carcinoma， 62%(31/50) with thyroid adenoma，56.7%(17/30) with normal thyroid tissues.The positive rates of HSP27, TGF-β1 were significantly higher in the papillary thyroid carcinoma than in thyroid adenoma(P<0.05). The positive rates of HSP27, TGF-β1 were significantly higher in the papillary thyroid carcinoma than in normal thyroid tissues(P<0.05). The positive rates in the thyroid adenoma was close to the positive rates in normal thyroid tissues(P>0.05). Conclusion： The expression of HSP27 and TGFβ1 may be important biological markers reflecting the carcinogenesis, progression and prognosis of the papillary thyroid carcinoma.  ［Key words］heat shock protein 27; transforming growth factorβ1; papillary thyroid carcinoma; thyroid adenoma
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  Duration of proneposition ventilation influence the therapeutic  effect on patients with early acute respiratory distress syndrome
   caused by pulmonary diseases

  YAN Jun, Ji-Mu-Sen, Jin-Zhao-Chen

  2013, 23(6): 520.

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  Objective: To determine the efficacy of mechanical ventilation on prone position (PPV) on oxygenation and respiratory mechanics in patients with early acute respiratory distress syndrome (ARDS) resulting from  pulmonary cause (ARDSp).  Methods: Twenty-six ARDSp patients were placed in prone position for 6 h. The effect of different time (pre-PPV, PPV 0.5, 2, 4,6 h) on oxygenation and respiratory mechanics were observed. Results: Compared with pre-PPV, in ARDSp patients, partial pressure of oxygen in artery(PaO₂)/ inhaled oxygen concentration (FiO₂) was increased after 2 h PPV (224.86±30.84 vs 192.45±46.81, P=0.03). And 4 h after PPV, the PaO₂/FiO₂ still further raised compared with PPV 2 h (286.38±36.67 vs 224.86±30.84, P=0.00). No significant difference in the PaO₂/FiO₂ was found between PPV 4 h and PPV 6 h (286.38±36.67 vs 287.46±38.84, P=0.98). There was no significant difference in the respiratory system compliance between before and after the prone position ventilation(F=1.00, P=0.40). Conclusions: The PaO₂/FiO₂ was improved when patients with early ARDSp were ventilated on prone position, especially for 4 h PPV.  ［Key words］ acute respiratory distress syndrome; prone position; oxygenation; respiratory mechanics