Objective To investigate the inhibitive effects of biosynthesized short hairpin RNA(shRNA)on the duplication of Influenza A virus(IAV)in human bronchial epithelium(HBE)cells.Methods NP-shRNA and PA-shRNA targeting to the highly conservative sequences of IAV nucleoprotein(NP)and acidic RNA polymerase(PA)were designed and biosynthesized by in vitro transcription,and then were inserted into the plasmid pGenSil-1.After sequencing analysis,the recombinant plasmids NP-shRNA and/or PA-shRNA were transduced into HBE cells followed by infected with IAV A/PR/8/34 H1N1(A/PR8)virus.The cytopathogenic effect(CPE)of the HBE cells,hemagglutination assay(HA)and 50% tissue culture infective dose(TCID50)titer of A/PR8 in the culture supernatants were determined respectively.The mRNA and protein expressions of NP and PA were detected by RT-PCR and Western blotting respectively,and the results were compared in order to evaluate the inhibitive efficiency of NP-shRNA and/or PA-shRNA to A/PR8 duplication in HBE cells.Results The content of NP-shRNA and PA-shRNA biosynthesized by in vitro transcription were respectively 59.4 nmol and 50.6 nmol in each 100 ?l.The purity was more than 2.0 and the sequences were verified to be correct after sequencing analysis.The CPE of HBE cells and the virus titer in the culture supernatants of cells transfected with NP-shRNA,PA-shRNA or NP-shRNA+PA-shRNA were markedly lower than those of the control groups.In 3 h after 1 TCID50 A/PR8 infection,the mRNA level of NP in NP-shRNA group,the mRNA level of PA in PA-shRNA group and those in NP-shRNA+PA-shRNA group were 41.7%,43.4%,68.5% and 73.7% respectively,lower than those of the control group.At 48 h after 1 TCID50 A/PR8 infection,NP synthesis were 92.3%,84.0% and 91.7% respectively of those of the control group.Conclusion Anti-IAV NP-shRNA and PA-shRNA are successfully prepared by in vitro transcription.Both of those markedly inhibit the reproduction of IAV A/PR8 and produce a favorable protective effect on HBE cells.

The studies on the extract processes of Radix Aucklandiae and Rhizoma Coptidis, and the preparation method for Xianglian Drop Pills have been carried out. The preparation method included the adsorption of volatile oil of Radix Aucklandiae with the extract mixture of Radix Aucklandiae and Rhizoma Coptidis. Water in adequate amount was added into the above mixture, The mixture was grinded into homogeneous pasty, then added with melted PEG 4000. After being mixed well on water bath the mixed melt was poured into the home-made drop pill apparatus(at 90℃) preheated, dropped into dimethyl silicone at rate of 24 drops/min through the drop head with internal apeture for 45mm and external apeture for 6. 1mm、and cooled well. The drop pills were obtai ned by removal of dimethylsilicone. Every examination of prepared drop pills coincided with the requests in general notice of drop pills in Pharmacopoeia of the Republic of China(1990). The in vitro antibacterial test showed that Xianglian Drop Pills had the same antibiotic effect as Xianglian Pills did.

Aim To investigate the effect taurine on ischemia-induced neuron apoptosis and its mechanisms. Methods Focal cerebral ischemia was induced by middle cerebral artery occlusion (MCAO). After 6 h permanent MCAO, mRNA expressions of Caspase-3/8 of rat brain were detected with real-time quantitative reverse transcriptase-polymerase chain reaction (real-time QRT-PCR). Oxygen-glucose deprivation (OGD) was used to induce an in vitro ischemia model. Primarily cultured cortical neurons were exposed to 4 h OGD. Intracellular calcium concentrations([Ca~(2+)]_i) was measured with fluorescent Ca~(2+) sensitive probe fura 2 acetoxylmethyl ester (Fura 2/AM). Neuronal apoptosis was assayed with flow cytometry of FITC-annexinV/propidium iodide binding 24 h after OGD. Results mRNA expressions of Caspase-3/8 were upregulated in MCAO model. [Ca~(2+)]_i and neuronal apoptosis were markedly increased in OGD model. Taurine pretreatment reduced the upregulation of mRNA expression of Caspase-3/8 in vivo and ameliorated calcium overload and neuronal apoptosis in vitro. Conclusion Taurine has neuroprotective effect against ischema-induced neuronal apoptosis. This is partly due to its effects on Caspase-3/8 and [Ca~(2+)]_i

Objective To investigate the role of P21 protein and survivin on the cell apoptosis of non-small cell lung cancer (NSCLC) induced by mevastatin. Methods The inhibitory effect of mevastatin on A549 and NCI-H520 cell line was evaluated by MTT assay. The cell cycle distribution and apoptosis induction were determined with flow cytometer and transmission electron microscope. The expression of p21 protein was assessed with flow cytometer. The mRNA expression of p21 and survivin was assessed with RT-PCR. Results Flow cytometry showed that mevastatin induced G_0/G_1 cell arrest in NSCLC cell lines. The results indicated that mevastatin caused apoptosis in concentration-dependent manner. Mevastatin produced no change in expression of P21 mRNA and total P21 protein. Concomitantly, P21 protein localized on cellular membrane was decreased. It was also found that mevastatin suppressed the expression of survivin mRNA in NSCLC cell lines. All these effects were reversed by mevalonate. Conclusions Mevastatin inhibited the proliferation of NSCLC cell lines. Mevastatin exerts growth inhibitory effect and induces apoptosis effect by inhibiting mevalonate synthesis. Its mechanisms might involve blockade of the isoprenylation of p21 protein and down-regulation of the expression of survivin mRNA.

Objective To investigate the effects of p53 on expression and activity of protein kinase R (PKR) as well as biological characters of HeLa cells from cervical carcinoma patients. Methods Recombinant plasmid vector pEGFP-C1/p53 was constructed to over-express p53 then it was transfected into HeLa cells. Transcription levels of p53 and PKR mRNA were detected by reverse transcriptase polymerase chain reaction (RT-PCR) among pEGFP-C1/p53 transfection group, pEGFP-C1 transfection group and blank control group(only transfection reagent was added);Protein expression lev?els of p53, PKR, phosphated PKR(p-PKR)and phosphatedαsubunit of eukaryotic initiation factor 2(p-eIF2α)which is the downstream substrate of PKR were detected by Western Blot among three groups;Proliferation of HeLa cell were deter?mined by methyl thiazolyl tetrazolium(MTT)assay;Invasion of HeLa cell were determined by Transwell cell assay. Re?sults Recombinant plasmid vector pEGFP-C1/p53 was successfully constructed to overexpress p53;Transcription level of p53 and PKR mRNA in pEGFP-C1/p53 transfection group were higher than those in pEGFP-C1 transfection group and in blank control group (P<0.05),and there were no significant difference between their levels in pEGFP-C1 transfection group and in blank control group;Protein expression levels of p53, PKR, p-PKR andp-eIF2α in pEGFP-C1/p53 transfection group were higher than those in pEGFP-C1 transfection group and in blank control group (P<0.05),and there were no sig?nificant difference between those expression levels in pEGFP-C1 transfection group and in blank control group;MTT and Transwell cell results showed that proliferation and invasion of HeLa cells in pEGFP-C1/p53 transfection group were weaker than those in pEGFP-C1 transfection group and in blank control group (P<0.05),and there were no significant difference between proliferation and invasion of HeLa cells in pEGFP-C1 transfection group and in blank control group. Conclu?sion p53 can up-regulate the expression and activity of PKR, promote activation of PKR/eIF2αsignal transduction pas?sage and restrain cell proliferation and invasion of HeLa cells.

Objective To investigate the role of P21 protein and XIAP on apoptosis-inducing effect by mevastatin against A549 cell line and its mechanisms. Methods The cells proliferation was detected by MTT assay. The cell cycle distribution and apoptosis induction were evaluated with flow cytometer and electron microscopy. The mRNA expression of P21 and XIAP was measured with reverse transcription-polymerase chain reaction.The protein expression of P21 and XIAP was tested with flow cytometer and Western Bolt respectively. Results Mevastatin inhibited A549 cell survival in a time-dependent and concentration-depended manner. Flow cytometry showed that mevastatin induced G0/G1 phase arrest and caused apoptosis. Mevastatin did not affect P21 expression at both mRNA and total protein level.Concomitantly,P21 protein localized on cellular membrane decreased.Both mRNA and protein expression in XIAP were down regulated by mevastatin. All these effects were reversed by mevalonate.Conclusion Mevastatin can inhibite proliferation,induce apoptosis and interfere with cell progression of A549 through mevalonate pathway. Its mechanisms are explained by decreasing the expression of XIAP mRNA and protern.Targeting HMG-CoA reductase,Mevastatin blocks the isoprenylation of P21 protein which affects its anchorage on the cellular membrane.

AIM:To explore the effect on stability of hyperforin with different medicinal herbs and the changes of hyperforin content in herb combinations with Hypericum perforatum L.(HP).METHODS:Several common antidepressant medicinal herbs were selected separately to match with HP.The herbal samples consisted of HP group,HP with Curcuma wenyujin Y.H.Chen et C.Ling group,HP with Curcuma longa.L.group,HP with Epimedium brevicornum Maxim group and HP with Wuling powder(WY)group.The contents of hyperforin with each group were determined by reverse phase HPLC.RESULTS:The rates of extraction and retention of hyperforin in HP with Curcuma wenyujin Y.H.Chen et C.Ling group and HP with Curcuma longa.L.group were significantly lower than that of HP group.There were no obvious differences in the extraction rates of the HP group,HP with Epimedium brevicornum Maxim group and HP with WY group.The retention rate of hyperforin in HP group with WY powder was higher than that of HP group.The retention rate of hyperforin in HP with Curcuma wenyujin Y.H.Chen et C.Ling group,was lower than that of HP group.CONCLUSION:The results show that compatibility of HP with traditional Chinese medicine have effects on the stability of active herbal component.

Objective To investigate the expression of VCAM-1 and E-selectin in vessel walls of the patients with thromboangiitis obliterans(TAO).Methods The vessel tissues were taken from 18 cases of amputation operation due to TAO and 16 cases of amputation operation due to non-vascular diseases.The expression of VCAM-1 and E-selectin were assessed by the immunohistochemical method.Results The positive expression rate of VCAM-1 in vessel wall of the patients with TAO was 77.8%(14/18),which of E-selectin was 72.2%(13/18);both of them were obviously higher than 6.3%(1/16) and 6.3%(1/16) in the control group.The positive rate of VCAM-1 and E-selectin had no correlation with the age,smoking amount,smoking time and drug use (P>0.05).Conclusion The expression of VCAM-1 and E-selectin have close correlation with the occurrence and development of TAO.Detecting the expression of VCAM-1 and E-selectin can serve as a diagnostic indicator of TAO.

Objective To investigate the relationship between evidence-based nursing tactics and quality of life of breast cancer patients so as to provide the reference for the best nursing strategy .Methods 87 patients who were diagnosed with breast cancer received surgical treatment were randomly divided into experimental group (44 cases) and control group (43 cases) .Patients in control group were given general nursing ,while those in experimental group were given evidence-based nursing tactics on the ba-sis of general nursing .Both groups were asked to fill in the quality of life questionnaire after 4 months breast cancer treatment and before treatment .Results The quality of life in experimental group was significantly improved after treatment compared with that of before treatment(P< 0 .05) ,the quality of life in the experimental group received evidence-based nursing tactics is significantly better than that of in control group (P< 0 .05) .Conclusion Evidence-based nursing tactics can improve the quality of life in breast cancer patients .

This paper studied the usage and impact of the theory in hospital development.Based on such,special manifestations behind the marginal diminishing effect are taken into consideration to grasp particular demands and requirements of health,and to look for a positive role and a progressive increase of the marginal utility.Furthermore,the explorations in hospitals,such as technical innovations,structure optimizations and management breakthroughs,as well as the experimental and practical construction of micro-economic administrative mechanism in clinical departments were discussed in the text.It has been demonstrated in practice that the practice has a significant effect on the prevention of marginal diminishing utility and an essential reference on hospital transformational development.