BACKGROUND: Lentivirus-mediated RNA interference technology has been widely used in the study of gene function because of its specificity, high inhibition efficiency and persistent effects. Virus packaging, transfection, as well as small hairpin RNA (shRNA)-coding sequence could affect the inhibition efficiency. Therefore, impact factors of RNA interference for human Bax inhibitor 1 (BI-1) was studied in this experiment.OBJECTIVE: To find the valid small interference RNA (siRNA) targeting human BI-1 gene by Lentivirus.DESIGN, TIME AND SETTING: Single sample observation was completed in Department of Medical Genetics, Basic Medical College, Peking University Health Science Center from September 2007 to December 2008.MATERIALS: 293T cell and SH-SY5Y cells were preserved in this laboratory; 4 shRNAs were designed through a use of online design tool by Ambion (www.ambion.com). METHODS: Several recombinant plasmids which expressed shRNAs targeting different regions of the BI-1 gene and labeled enhanced green fluorescent protein as a fusion gene were constructed. Four types of shRNA-expressing virions were obtained by cotransfection of shRNA-expressing plasmid and package protein-expressing plasmids into 293T cells. Green fluorescent protein expression was determined using flow cytometry to search the optimal package conditions. Real-time PCR and beta actin mRNA was used as an internal control, four types of recombinant viral supernatant and control viral supernatant were added on the SH-SY5Y cells, knock-down efficacies of the endogenous BI-1 gene were determined, and the RNA interference effective sequences were optimized. MAIN OUTCOME MEASURES: Intracellular green fluorescent protein expression rate of the cells infected with different packaging virus. RESULTS: The optimal infective packaging virus could be obtained when the ratio of plasmids pLentiLox3.7, REV, VSVG and RRE was 2:1:1:1 and the supernatants were collected 48 hours after transfection. Replacement of culture media 24 hours after transfection could increase the infection efficacy of the virions. shRNA targeting-2-17nt (start coding region) of BI-1 gene was most valid interference efficiency and could decrease the gene expression to 40%. CONCLUSION: Influencing factors of Lentivirus-mediated RNA interference technology virus packages were investigated in this study, which could be a foundation for constructing stable BI-1 knocked down neuronal cell model and for studying the abnormal expression of BI-1 related to the neuron apoptosis disease.

Objective To discuss the nursing effect and application value of refinement process of nursing management mode in hospital infection control in the operating room.Methods 620 patients who will carry out surgi-cal treatment were selected as the research subjects.310 cases without detailed process of nursing management were included in the control group,and 310 patients with refinement process of nursing management were selected as obser-vation group.The clinical nursing effects of two groups were recorded.Results Observation group surface 88.94%, medical staff hand 92.54%,disinfectant 95.65%,100.00% sterile items.The control surface 81.38%,80.00% of medical personnel hand,disinfectant 75.93%,90.00% sterile items,the monitoring qualified rate between two groups had statistically significant differences (χ2 =5.021,4.596,7.581,6.725,all P <0.05).The sores of nursing safety, operating room management,form filling,ward nursing in the observation group were (95.37 ±4.66 )points, (94.27 ±5.21 )points,(96.13 ±3.29)points,(95.03 ±4.87)points.Those in control group were (90.12 ± 2.03)points,(89.13 ±2.26)points,(90.03 ±1.88)points,(88.17 ±2.25)points,the differences between two groups were statistically significant (t =18.185,15.935,28.343,22.514,all P <0.05).The scores of nurses and patients communication,nursing procedures,ward environment,care attitude in the observation group were (24.19 ± 0.87)points,(24.03 ±0.64)points,(24.51 ±0.45)points,(24.28 ±0.36)points.Those in the control group were (21.54 ±1.65 )points,(20.88 ±1.72)points,(20.95 ±1.66)points,(21.09 ±0.89)points,the differences between two groups were statistically significant (t =25.013,30.220,36.443,58.502,all P <0.05).Conclusion Refinement process of nursing management mode application helps to control nosocomial infection in operating room of hospital infection control,improve the quality of clinical nursing services,improve the clinical nursing satisfaction, which is worthy of popularization and application in clinic.

Objective:To compare the differences in incidences of insomnia,anxiety(irritability)and manic ep- isode induced by fluoxetine and amitriptyline in treating depression.Method:CBM discs were selected for the data sources. The rates of insomnia,anxiety(irritability)and manic episode from published clinical control trials on depression treated by fluoxetine and amitriptyline were analyzed by applying fixed effect model(FEM)of evidence-based medicine(EBM). Result:Of 1205 cases in 15 studies,the rates of insomnia induced by fluoxetine or amitriptyline were 21.71% and 1.80% ,OR 9.39(95 %CI 5.37-16.44),P0.05.Conclusion:Fluoxetine induces in- somnia and anxiety(irritability)more easily than amitriptyline when treating depression.

BACKGROUND: Inhibiting the degradation of extracellular matrix in the intervertebral disc can delay the degenerative process of intervertebral disc. Matrix metalloproteinases-3 (MMP3) is considered as a key enzyme for degradation of extracelular matrix components such as type II collagen and aggrecan.
OBJECTIVE:To construct the short hairpin RNA lentiviral vector targeting human MMP3 gene and to detect its efficiency of gene silence by infecting human degenerative nucleus pulposus cells.
METHODS:According to the human MMP3 mRNA (NM_002422.4) sequence, four groups of the short hairpin RNA gene sequences targeting MMP3 were designed, synthesized and annealed to form double stranded DNA fragments, which were connected with the LV3 vectors digested by BamHI andEcoRI enzymes, and then transfected into the competent cels. The positive clones were identified by PCR, and analyzed by sequencing. The packaging and titer of lentivirus were determined after transfecting 293T cells. Human degenerative nucleus pulposus cels were infected with lentivirus vector, and the transfection efficiency of each group was observed under inverted fluorescence microscope. The interfering efficiency was detected by real time-PCR and western blot at 72 and 96 hours.
RESULTS AND CONCLUSION:The ds-oligo DNA was successfully inserted into the lentiviral vector as confirmed by electrophoresis and sequence analysis. The recombinant lentivirus was harvested from 293T cels with a viral titer of 1-5 ×108 TU/mL. RNA interference targeting the GCC AGG CTT TCC CAA GCA AAT sequences with the highest interfering efficiency in MMP3 gene at 72 and 96 hours resulted in suppression of MMP3 mRNA expression by 98% and 72%, respectively; and at 96 hours, the interfering efficiency of protein expression was 57.2%. The recombinant lentivirus vector containing RNA interference targeting MMP3 gene is successfuly constructed, which lays a foundation for further studies on the MMP3 function and gene therapy.

Objective To evaluate the efficacy and safety of domestic olmesartan in treatment of mild to moderate essential hypertension in comparison with losartan. Methods Two hundred and thirty-seven patients with mild to moderate essential hypertension were enrolled in a randomized, double-blind, multi-center, paralleded and active-controlled trial, and were divided into olmesartan group (olmesartan 20 mg + losartan 50 mg placebo) and losartan group (losartan 50 mg + olmesartan 20 mg placebo) for a 8-week therapy. Four weeks after treatment, dosages of drugs were doubled in patients with seated diastolic blood pressure ≥90 mmHg (1 mmHg =0.133 kPa). All patients were followed up every two weeks, and the efficacy and adverse effects were observed. Another 32 patients with mild to moderate essential hypertension were enrolled and given olmesartan only, and ambulatory blood pressure monitoring was performed before and 8 weeks after treatment. Results Compared with those before treatment, both systolic blood pressure and diastolic blood pressure significantly decreased in olmesartan group and losartan group 8 weeks after treatment [(15.2 ±13.3) mmHg and (19.5 ±11.8) mmHg, respectively for systolic blood pressure (P <0.001); (15.9 ±7.48) mmHg and (16.2 ± 5.95) mmHg, respectively for diastolic blood pressure (P < 0.01) ], while there was no significant difference between these two groups (P > 0.05). There was no significant difference in total effective rate and incidence of adverse effect between these two groups (86.9% vs 93.7% and 7.63% vs 5.88% , P > 0.05) . Ambulatory blood pressure monitoring demonstrated that trough to peak ratios of systolic blood pressure and diastolic blood pressure were 86% and 71%, respectively. Conclusion Domestic olmesaratan provides an effective, safe and long action in the treatment of mild to moderate essential hypertension.