Objective To construct an expression vector harboring CYP2B1 suicide gene, and detect its expressions in tumor cell lines. Methods PCR amplification was performed using primers based on murine CYP2B1 gene sequence from gene bank and pc3/2B1 as template. PCR product was directly inserted an eukaryotic expression plasmid pcDNA3.0. The recombinants were analyzed and identified by restriction enzyme analysis, PCR and sequencing. Then the recombinant vector pcDNA3.0/CYP2B1 was transfected into three tumor cell lines by liposome-mediated method. The expressions of CYP2B1 gene in all the cell lines were detected by RT-PCR method. Results pCDNA3.0/CYP2B1 vector was successfully constructed, and could express CYP2B1 mRNA in the three tumor cell lines. Conclusion Eukaryotic expression vector pcDNA3.0/CYP2B1 containing CYP2B1 gene under the control of a CMV promoter is an novel effective expression vector for tumor gene therapy.

AIM: To study the protective effects of n on -mitogenic human fibroblast growth factor (nm-haFGF) on cardiomyocytes injured b y reactive oxygen free radicals. METHODS: The cardiomyocytes were isolated from neonatal SD mouse by trypsin digestion. The cardiomyocytes injury model was established by expos ing the cells to hydrogen peroxide (H 2O 2), and the injury status in cardiomy ocytes were evaluated by examining the cellular viability, measuring cell apopto sis and observing the change of cellular morphology. nm-haFGF was added to the c ulture medium, and the changes of cellular viability, superoxide dismutase (SOD) , malondialdehyde (MDA) and celluar apoptosis were observed. RESULTS: A dose-dependence relation between the concentration of H 2O 2 and the cardiomyocytes injury was observed. 10-80 ?g/L nm-haFGF dose -dependently increased cardiomyocyte viability and the general SOD activity, as well as decreased the content of MDA and the quantity of cardiomyocyte apoptosis . CONCLUSION: The higher the concentration of H 2O 2, the mor e serious the cardiomyocyte injury. nm-haFGF may have a good protective effects on cardiomyocytes treated with H 2O 2.

AIM: To investigate the effect of basic fibroblast growth factor (bFGF) on radiation-induced splenocytic apoptosis of mice. METHODS: At 14 h after whole body irradiation with 0.5 Gy and 1.0 Gy,splenocytes were cultured with and without bFGF,and splenocytic apoptosis was quantitatively analysed by flow cytometry. Cell proliferation was determined by the method of [3H]-TdR incorporation. RESULTS: bFGF(1 ?g/mL and 2 ?g/mL) could reduce the rate of cell apoptosis ,and promote the proliferation of splenocytes. CONCLUSION: bFGF could inhibit radiation-induced splenocytic apoptosis and promote the proliferation of splenocytes and then enhance body immunity.

To investigate the mechanism of inhibitory effect of a novel bFGF antagonist peptide isolated from the phage display random heptapeptide library on cell proliferation induced by basic fibroblast growth factor. The effect of P7 on cell morphology was observed under an inverted microscope. Flow cytometry was applied to analyze the effect of P7 on cell cycle progress of bFGF-stimulated cells. The effect of P7 on bFGF-induced activation of MEK and Erk1/2 in MAPK pathway was detected by Western blotting. The results showed that no significant cell morphology change was observed in the range of detected concentrations of P7. Cell cycle analysis showed that P7 decreased S-phase cell population and arrested cell cycle at the G0/G1 phase of bFGF-stimulated cells. The results of MAP kinase activation assay indicated that P7 decreased bFGF-induced MEK and Erk1/2 phosphorylation in a dose-dependent manner. P7 inhibited proliferation of bFGF-stimulated Balb/c 3T3 cells possibly via cell cycle arrest at the G0/G1 phase and down-regulation of signal molecular activation in MAPK pathway.

Aim To observe the effect of Sinomenine on the activation of NF-?B and the degradation of its inhibitor I-?B in peritoneal macrophages of BALB/c mice in vitro and provide experimental evidence for further evaluation of the antiinflammatory and antirheumatic effects of Sinomenine. Methods Effect of Sinomenine on the activation of NF-?B p65 in the cells was investigated by using fluorescence-labelling and laser confocal scanning microscopy; Effect of Sinomenine on the degradation of I?B-? was investigated by Western blot. Results SIN (0.25,1.25 mmol?L -1) attenuated the activation of NF-?B p65 in peritoneal macrophages of BALB/c mice induced by LPS in vitro. SIN(0.25,1.25 mmol?L -1) inhibited the degradation of I-?B-? in peritoneal macrophages of BALB/c mice induced by LPS in vitro. Conclusion SIN can partially inhibit the activation of NF-?B p65 and the degradation of I?B-? in peritoneal macrophages of BALB/c mice induced by LPS in vitro.

Objective To explore the correlation of serum homocysteine(Hcy)levels and hypertrophy of left ventricle in very elderly hypertensive patients.Methods According to plasma Hcy levels,patients with essential hypertension (n=378) were divided into non H-type hypertension group (n=142) and H-type hypertension group (n=236). Height, weight, reg?ular medication, blood pressure, renal function, blood lipid profile and the concentration of plasma Hcy were recorded. Color Doppler ultrasonic equipment was used to determine the morphology and structure of left ventricle. The correlation between plasma Hcy and left ventricle remodeling was analyzed. Results The ratio of left ventricular hypertrophy was higher in H-type hypertension group than that in non H-type hypertension group(45.8%vs 24.6%,χ2=16.81,P<0.001). Patients in H-type hypertension group had higher systolic blood pressure, higher plasma level of Hcy and larger left ventricular posterior wall thickness(LVPWT), larger interventricular septal thickness (IVST) and increased left ventricular mass index (LVMI) compared to those in non H-type hypertension group(162.20 ± 14.97)vs(149.70 ± 5.06)mmHg,(19.76 ± 5.83)μmol/L vs (9.53±0.72)μmol/L,(9.77±2.35)vs(9.21±2.68)mm,(9.74±3.15)vs(8.51±2.42)mm,(118.64±39.38)vs(101.85±41.71)g/m2 respectively, all P<0.05). There was a positive correlation between LVMI and Hcy(r=0.381,P<0.001). Multivariable Lo?gistic regression analysis showed that hyperhomocysteinemia was an independent risk factor of LVMI. Conclusion High plasma Hcy level is an independent risk factor of LVMI, which works together with hypertension to promote left ventricular re?molding.

To investigate the killing effect of PNP/MeP-dR suicide gene system on hepatoma cells,pcDNA3. 0/PNP, an eukaryotic expression vector harboring E. coli PNP gene, was transfected into human hepatoma HepG2 cells by liposome-mediated method. A HepG2 cell line with stable PNP gene expression, HepG2/PNP, was established with presence of G418 selection. The cell growth curves were determined with trypan blue staining. The sensitivity of HepG2/PNP to MePdR and bystander effects were assayed by MTT and FCM methods. The enzymatic activity of the product of PNP gene was determined by HPLC method. The cytotoxic effects of MeP-dR on HepG2/PNP cells were obvious (IC50 =4.5μmol/L) and all HepG2/PNP cells were killed 4 days after the treatment with 100μmol/L MeP～dR. In mixed cultures containing increasing percentages of HepG2/PNP cells, total population killing was demonstrated when HepG2/PNP cells accounted for as few as 5% of all HepG2 cells 8 days after the treatment with 100μmol MeP-dR. Highpressure liquid chromatography (HPLC) demonstrated that the PNP enzyme could convert MePdR into 6-MP. PNP/MeP-dR suicide gene system had an advantage over traditional suicide gene systems for hepatoma gene therapy. Our e results suggest that high-level bystander effects of this system result in significant anti-tumor responses to hepatoma gene therapy, especially in vivo.

Objective To survey a prevalence of osteoporosis and prevalence of osteoporosis combined with hyponatremia in elderly hospitalized patients,and their risk factors.Methods We enrolled 2496 elderly hospitalized patients with detected plasma levels of sodium,calcium,25 (OH) D3,PTH,plasma PINP,and β-CTX.At the same time,sex,age,height,weight,smoking history,drinking history and BMI(kg/m2) in form of a questionnaire were recorded and calculated.The risk factors for osteoporosis were analyzed using multivariate Logistic regression method.Results The osteoporosis prevalence was 12.2％ (305/2496 inpatients)with 31.5 ％ (96/305)in male,68.5％ (209/305)in female(x2 =4.651,P=0.031).The prevalence of osteoporosis with hyponatremia was 27.5 ％ (84/305),with 24.8 ％ (21/84) in male and 75.2 ％ (63/84) in female(x2 =9.251,P=0.025).As compared with three groups of non-osteoporosis,normal serum Na+ with and without osteoporosis,the osteoporosis patients with hyponatremia were more aged,in a higher proportion of women and smokers,in lower BMI,and in low levels of serum sodium,BMD 25(OH)D3 (F=13.783,0.861,7.146,24.520,0.548,x2 =15.113、4.472;P =0.001,0.000,0.021,0.015,0.003,0.021,0.005).Multivariate Logistic regression analysis showed that aging,female,low BMI,smoking history,drinking history,low plasma 25(OH)D3 level,low plasma PINP level,and high plasma β-CTX level were the risk factors for osteoporosis(OR 4.215,2.271,3.176,2.013,1.237,3.987,1.843,1.972;all P＜0.05).Conclusions The osteoporosis prevalence is high in elderly patients,especially in old women.The risk factors for osteoporosis are diverse,and clinical conditions of osteoporosis patients with hyponatremia are much more severe than the others.More efforts should be given to them and need to be focused on the complications of osteoporosis.

Objective:To investigate the effects of endoplasmic reticulum(ER)stress on the stability of atherosclerotic plaques in mice by examining the action of lipopolysaccharide(LPS)-induced Toll-like receptor 4(TLR4)on the protein expression levels in the ER stress pathway in atherosclerotic plaques of polipoprotein E gene knockout (ApoE -/-) mice. Methods:From October 2015 to February 2016, 24 ApoE -/-mice were randomly divided into the control group, the LPS group and the TAK group after 10 weeks of high-fat feeding(n=8, each group). After 10 weeks of intervention, peripheral blood was extracted by removing the eyeballs for the measurement of total cholesterol(TC), triglycerides(TG)and oxidized low density lipoprotein(ox-LDL). Then mice were sacrificed to obtain carotid and aortic specimens.Immunohistochemistry was used to detect the expression of carotid plaque macrophages(MOMA-2), smooth muscle actin(α-actin), TLR4, interleukin-1β(IL-1β), interleukin-6(IL-6), tumor necrosis factor-α(TNFα)and nuclear factor-κ-gene binding(NFκB). Western blotting was used to determine the expression of PKR-like eukaryotic initiation factor 2αkinase(PERK), C/EBP-homologous protein(CHOP)and glucose-regulated protein 78(GRP78). Results:The levels of TC, TG and ox-LDL were elevated in the LPS group, compared with the control and TAK groups[(25.0±2.3) mmol/L vs. (20.2±1.6) mmol/L and (20.8±2.6) mmol/L, (1.3±0.1) mmol/L vs.(1.3±0.1) mmol/L and (1.0±0.1) mmol/L, (17.4±1.3) mmol/L vs.(15.8±1.6) mmol/L and (12.1±1.1) mmol/L, P<0.05]. The comparison of plaque morphology and pathology showed that the LPS group had a wider range of atherosclerotic plaques, more macrophages and fewer vascular smooth muscle cells than the control and TAK groups( P<0.05). The expression of TLR4, IL-1β, IL-6, TNFα, NFκB, PERK, CHOP and GRP78 was higher in the LPS group than in the control and TAK groups( P<0.05). Compared with the control group, the expression of PERK, CHOP and GRP78 was lower in the TAK group( P<0.05). The expression of TLR4, PERK, CHOP and GRP78 was higher in the LPS group. Conclusions:LPS-induced TLR4 can up-regulate the expression of proteins in the ER stress pathway, increase the secretion of inflammatory cytokines downstream of the ER stress pathway, aggravate lipid metabolism disorders and increase the instability of atherosclerotic plaques.

Stroke has become the leading cause of death in China, and carotid atherosclerosis is an important risk factor for ischemic stroke. In current carotid atherosclerosis diagnosis and treatment guidelines, patients are stratified based on the degree of vascular stenosis. However, the presence of carotid atherosclerotic vulnerable plaques can cause ischemic stroke at any time, regardless of the degree of carotid stenosis. The development of MRI technology, especially the advent of high-resolution MRI, enables non-invasive assessment of carotid plaque structure and properties, and provides optimized treatment strategies for high-risk stroke population in the early stage to achieve the goal of prevention and treatment of stroke.