Abstract Objective:To evaluate the value of measurement of serum intercellular adhesion molecule-l(sICAM-l) and sE-selectin in di-agnosis and postoperative monitoring for hepatocellular carcinoma. Methods: sICAM-1 and sE-selectin were measured in patients with chronic hepatitis,liver cirrhosis,HCC and healthy controls by ELISA. Results:The mean levels of sICAM-1 and sE-selectin in 79 patients with HCC was significantly higher than that of other three groups ( P

Objective To investigate the effects and possible mechanism of action of inhibiting hepatitis B virus X protein (HBx) expression on liver cancer metastasis. Methods The suppression of HBx expression in MHCC97H cells was performed by siRNA interference technique, and the effects of HBx suppression on the metastasis of MHCC97H cells were detected by Matrigel invasion assays and in a lung-metastasis mouse model. The expression levels of related epithelial-mesenchymal transition (EMT) and apoptosis proteins were examined by Western blotting. Results Introduction of HBx-siRNA into MHCC97H cells inhibited the expression of HBx and the ability to metastasize,downregulated the expression of Twist and N-cadherin, and upregulated E-cadherin expression. These changes resulted in inhibiting EMT of MHCC97H cells. Meanwhile, apoptosis involved in the Twist-P53 pathway was also found. Conclusions Inhibiting expression of HBx can decrease the metastatic a-bility of MHCC97H cells by changing EMT and inducing apoptosis.

Objective To investigate the relationship of androgen receptor (AR) CAG repeat polymorphism and coronary artery disease (CAD) in elderly men and its potential mechanism.Methods Totally 296 elderly men undergoing coronary angiography were enrolled in this study.Serum total testosterone (TT) and free testosterone (FT) levels were measured.Androgen receptors (ARs) in peripheral lymphocytes were determined by flow cytometry.Genome DNA was extracted from peripheral leucocytes using standard techniques.Gene fragments containing AR CAG repeats were amplified by PCR with specific fluorescent labeled primers.PCR products were separated with agarose gels.CAG repeat number of each sample was obtained by genotyping.Results AR CAG repeats varied from 11 to 28 (P25-P75:18-22; median:20) in elderly male patients.They were divided into the long AR group (CAG repeats≥22,n=82) and the short AR group (CAG repeats＜22,n=214).Compared with the long AR group,serum FT level was much lower in the short AR group [(24.1±23.1) ×10-6mmol/L vs.(31.2±27.8)×10-6mmol/L,P＜0.05].The prevalence of coronary artery disease was higher in the short AR group than in the long AR group [84.1％ (180 cases) vs.69.5％(57 cases),P＜0.05].The FT level was lower in the short AR group combined with CAD than in the control group [(22.4±20.5) ×10-6mmol/L vs.(33.6±32.4)×10 6mmol/L,P＜0.01].There were no significant differences in serum TT and AR levels between the long and short AR groups.No significant correlations were found in the AR CAG repeats polymorphism with FT,TT or AR levels.Age was the main risk factor for FT and AR levels.Logistic regression analysis showed that FT level was negatively correlated with CAD (OR=0.98,95 ％ CI:0.973-0.998,P=0.01),and short AR increased the risk of CAD in elderly male patients (OR=3.44,95％CI:1.887-6.264,P＜0.01).Conclusions Serum FT level is correlated with age and is significantly decreased in elderly male patients with short AR repeats,which may increase the risk of CAD in elderly men.

Objective To optimize the formulation in preparation of dropping pills with total glucosides in Radix Paeoniae Alba and evaluate its accumulative release percentage in vitro.Methods With the hardness,roundness,and adhesion as the evaluation,orthogonal design was conducted.With dissolution rate of 12 h as the indices,according to the uniform design,the optimum coating formulation of Eudragit RL and RS was established.Results The dropping pills met the criterion of formulation,and the preparation release met the characteristics of the sustained release of the first order kinetics.Conclusion The optimal formulation is simple,which provides the basis for further development of new preparations of total glucosides in Radix Paeoniae Alba.

Objective:The immunogenicity of DNA vaccine immunogenicity can be improved by fusing antigenic genes to IgG Fc fragment.It is critical for enhancing immunogenicity of DNA vaccine to construct the secreting eukaryotic expressing vector.The purpose of this study is to construct a secreting eukaryotic expressing vector ligated with IgG Fc gene by amplifying human IgG Fc fragment and fusing the fragment with human CD5 signal peptide sequence for highly efficient expression.Methods:Human lymphocytes were isolated from the tonsil obtained by removal surgery.The total RNA of lymphocytes was extracted using Trizol reagent.Human IgG Fc cDNA was amplified by RT-PCR from lymphocytes and then inserted into pMD-18T vector.The Fc encoding sequence fused with CD5 signal peptide(sp) sequence was constructed by cross PCR using Fc fragment and CD5sp sequence and cloned in pcDNA-CD5 plasmid as the template.CD5sp-Fc fragment was inserted into pcDNA3.1 expressing vector to construct the pcDNA-CD5sp-Fc plasmid.The plasmids were transfected into HEK293T cells mediated by calcium phosphate.Western blot was used to detect expressed Fc protein in cultural supermatant of the cells.Results:The amplified sequence of human IgG Fc was consistent with that previously published.The secretory expression of Fc in HEK293T cells was achieved and the expressing level reached 50 ?g/106 cells at 48 h culture after transfection.Conclusion:The human IgG Fc gene is amplified by RT-PCR methods and the secretory expression of Fc gene mediated by CD5 signal peptide in 293T cells is achieved.The results provide a experimental basis for further construction of antigen genes fused to IgG Fc fragment and investigation on DNA vaccines and Fc as a biological adjuvant.

Objective To improve patient postoperative comfort of vitrectomy with tamponnade in the prone position, design a new prone position headrest to reduce complications caused by improper body position and observe its clinical effect. Methods According to the postoperative position of the patients, 360 cases were collected. The patients were divided into the control group and the observation group with 180 cases of each group. Observation group was treated with the new prone position headrest nursing, control group were treated with routine prone position. The comfort of patients, postoperative adverse reactions, success rate of retina reattachment were observed. Results According to simplified comfortable situation scale, physiological, psychological, social culture and environment of each individual score respectively was (2.74±0.21), (3.54±0.29) , (3.25±0.23), (3.36±0.27) points in observation group and (2.30± 0.19), (2.92±0.31), (2.93±0.26), (2.79±0.30) points in control group, and there were significant differences (t=12.368-20.845, all P<0.05). The daily posture duration in postoperative first time and 5 days was respectively (220.00±25.08), (1008.00 ± 20.32) min in observation group and (85.00±28.07), (650.00± 30.12) min in control group, and there were significant differences(t=48.117, 133.194, all P<0.01). The incidence of corneal edema, conjunctival congestion, water turbidity in observation group were lower than those in control group at 4 weeks after surgery, and there were statistically significant difference (U=6.308,8.130, 6.875, P < 0.01). The incidence of high intraocular pressure, recurrent retinal detachment rate and reduction rate in observation group were lower than those in control group at 4 weeks after surgery, and there were statistically significant difference (χ2=9.000, 10.540, 11.770, P < 0.01). Conclusions The new prone headrest can effectively improve the resection of vitreous body with tamponade patients in comfort, ensure the operation effect.

Objective:To research on the immune recognition mechanism of synthetic pyrethroids and generic specific antibody.Methods:We studied on quantitative structure-activity relationship ( QSAR ) of synthetic pyrethroids and their analogs as well as antibody activity ( IC50:fifty percent inhibition concentration ) using stepwise multiple linear regression method.Based on calculating structure descriptors of synthetic pyrethroids and their analogs , two-demensional QSAR ( 2D-QSAR ) model was established.The main factors affecting antibody activity were screened using 2D-QSAR,and predictive ability of QSAR models were evaluated by the method of leave-one-out( LOO) cross-validation.Meanwhile, the structure parameters of synthetic pyrethroid fragments were calculated and then analyzed using partial least squares ( PLS) assay.And then hologram QSAR ( H-QSAR) model was constructed on molecular substructure and antibody activity.The fragments contribution to antibody activity were illustrated by encoding different colors.Results:Decision coefficent (R2) of 2D-QSAR model and HQSAR model were 0.920 and 0.917 individually,cross-validation coefficient ( Q2 ) of two QSAR models were 0.875 and 0.660 respectively ,which showed two models had good predictive abil-ity.The result from 2D-QSAR model was also obtained that smaller was hydrophobicity of pyrethroids , easier was recognized by antibody.In addition,the optimum HQSAR model was constructed after we tried many combinations of these parameters .The fragment size in optimum HQSAR model was between 4 to 10,a hologram length was 61,optimum principle component was 4,and the fragment type of B/C/Ch was selected.However ,the fingerprint encoded results of synthetic pyrethroids weren′t consistent completely with exper-imental IC50 values.Conclusion:Hydrophobicity of synthetic pyrethroids is the largest correlation factors in antibody recognization .

BACKGROUND: Various etiological mechanisms are involved in cerebral infarction. Both free radicals and lipid peroxidation participate in the atherosclerosis and damage of neural cells after cerebral ischemia. Compound danshen(Radix Salviae Miltiorrhizae) is a common prescribed Chinese herb, acting on activating blood circulation and removing stasis for cerebral infarction and coronary heart disease, but its mechanism has been unknown in many aspects.OBJECTIVE: To observe the effect of compound danshen on neural function defect and free radicals in patients with cerebral infarction so as to probe into its possible mechanisms.DESIGN: A randomized controlled trial.SETTING: Neurological Internal Department of a hospital affiliated to one university.PARTICIPANTS: Totally 538 inpatients were collected in Neurological Internal Department of First Hospital affiliated to Jinzhou Medical College from February to December 2002, their diagnosis compiled with "Diagnostic Keys on Every Type Cerebral Vascular Disorders" adopted on the 4th National Acadenic Meeting on Cerebral Vascular Disorders, and determined by cerebral CT scan. All of those were the first attack of atherosclerosis cerebral infarction in 72 hours. The patients with cardiac infarction, heart failure, auricular fibrillation, insufficiency of liver and kidney function,hemorrhage of digestive tract, vascular dementia and bulbar paralysis and the patients who could not be well cooperated were not included. A total of 68 patients compiled with the standards, of which, 38 patients were male and 30 patients female, aged varied from 52 to 78 years, at the average of (64. 62 ±5.80) years. The patients selected were randomized into study group and the control by lot-drawing nethod according to the hospitalized sequence and volunteer principle of the control.METHODS: The basic treatment was same in two groups. In study group,compound danshen injection was added together with physiological saline 250 mL for intra-venous drip, once daily, continuous 14 days made one course. In the control, thrombosis removing injection 15 mL was added together with physiological saline 250 mL for intra-venous drip, once daily,continuous 14 days made one course.Level of serum lipoperoxide(LPO) and activity of superoxide dismutase (SOD).RESULTS: Statistical differences presented in declined scores of severity of neural function defect after treatment in two groups compared with their own controls(in study group: 28.62 ±6.76 vs 13.84 ± 8.16; in the control:28.58±7.05 vs 21.52±8.24, t=8.134, t=3.796 respectively, P＜ 0. 001 ). The score in study group was declined more obviously compared with the control after treatment, indicating very significant difference ( t = 3. 861, P ＜ 0. 001 ). The effective rate of compound danshen injection was 88.24% in treatment of cerebral infarction, which significantly superior to that in the control(67.65% ) (x2 =4.19, P ＜ 0.05). Compound danshen remarkably reduced serum LPO level[ (8.69 ± 1.28) nmol/L vs (5.86 ± 1.42) nmol/L, t =8. 628, P ＜ 0. 001 ] and statistical differences presented compared with the result in the control after treatment[(5.86±1.42) nmol/L vs(8.56±0.95) nmol/L, t=9.125, P ＜0.001] . Simultaneously, SOD activity in serum was significantly increased, [ (26. 25±4. 64) mkat/g vs(30. 01 ± 3.87) mkat/g, t = 3. 629, P ＜ 0. 001] indicating statistical differences compared with the result in the control after treatment[ (30. 01 ±3.87) mkat/g vs(26.33 ±4. 14) mkat/g, t =3. 778,P ＜ 0.001].CONCLUSION: Compound danshen improves significantly neural function defect in patients with cerebral infarction, with definite therapeutic effects on the treatment. It can reduce serum LPO content and increase serum SOD activity in patients with cerebral infarction. It is predicted that removing free radicals and anti-lipid peroxidation damage is probably one of the important mechanisms of it, which provides a further theoretic evidence for the treatment of cerebral infarction clinically.

OBJECTIVEThis study was conducted to investigate the function of the first intron of porcine growth hormone (pGH) gene in the gene expression.

METHODSPCR method was used to amplify the first intron from pig genomic DNA. The intron was then inserted into pGH cDNA to construct pGH cDNA-intron (pGH cDNA-in). The recombinant adenoviruses containing pGH cDNA and pGH cDNA-in genes under control of CMV promoter were generated by homologous recombination method in HEK 293 cells respectively. The effect of the first intron on gene expression was evaluated by comparing the expression levels of pGH cDNA-in and pGH cDNA mediated by adenovirus vectors in CHO cells.

RESULTSThe expression level of pGH cDNA containing the first intron increased by 117%, which was significantly higher than that of pGH cDNA without the intron (P < 0.001).

CONCLUSIONThe first intron of pGH gene has the function to improve pGH gene expression.

Adenoviridae ; genetics ; Animals ; CHO Cells ; Cricetinae ; DNA Transposable Elements ; DNA, Complementary ; DNA, Recombinant ; Genetic Vectors ; Growth Hormone ; genetics ; Introns ; Swine

To prepare recombinant fox growth hormone (fGH), we amplified its cDNA from silver fox pituitary tissue by RT-PCR and cloned into yeast shuttle vector pPIC9K down stream of a-factor signal peptide sequence by SnaB I and Not I restriction sites. The recombinant secretion vector pPIC9K/fGH, linearized by Sal I, was transformed into histidine-deficient Pichia pastoris strain GS115 by electroporation. We selected His+ -transformed methylotropic (His+, Mut+) yeast using histidine-absent medium containing dextrose (MD) or methanol (MM) as the only carbon source, and then screened the recombinant GS115 with multi-copy fGH genes by G418. The secretive expression of fGH was performed under the induction of methanol in shaking flask culture. The results showed that the fGH cDNA sequence amplified in this paper was basically in consistence with the published in GenBank. We achieved the secretive expression of recombinant fGH identified by SDS-PAGE and Western blotting. The fGH expression level was 119 mg/L, accounted for 34% of total proteins in fermentation medium.
Animals ; Base Sequence ; Cloning, Molecular ; DNA, Complementary ; genetics ; Electroporation ; Foxes ; genetics ; Genetic Vectors ; genetics ; Growth Hormone ; biosynthesis ; genetics ; Molecular Sequence Data ; Pichia ; genetics ; metabolism ; Recombinant Proteins ; biosynthesis ; genetics