OBJECTIVE To determine the distribution of bacterial flora in hospital infection and to provide laboratory(evidence) for controlling hospital infection and selecting rationally antibiotics in clinic practice.METHODS All(isolates) were identified by routine procedure.MRSA and ESBLs-producing rate of Escherichia coli and Klebsiella pneumoniae were(examined.) RESULTS Among all these clinical infectious specimens,there were 202 strains of Gram negative bacilli,(accounting) for 40.9%(202/495);166 strains of fungi,accounting for 33.5%;621 strains of Gram positive cocci,for 20.6%(102/495).Candida albicans,E.coli,Pseudomonas aerugionosa,C.tropicalis and C.glabrata took the first five bacteria in infection.Analysis of drug resistant bacteria suggested that the isolated rate of ESBLs-producing strains in Staphylococcus aureus be 47.6%,be CNS in MRCNS 78.1% and MRSA in SA be 42.3%.CONCLUSIONS Multidrug resistance and fungus infection are the main risk factors in our hospital.We must improve means of treatment on clinical work and use antibiotic rationally to reduce the infection rate.

For the problems that 3 first-class ternary hospitals which are not directly affiliated to medical universities are facing in cultivating high-educated staff's scientific abilities,analyze the importance to carry out scientific work in clinic and discuss how to improve their scientific abilities from hospitals,departments and high-educated staff themselves.

The aim of this study was to explore the effect of small hairpin loop RNA (shRNA) silencing hypoxia-induced factor 1alpha (HIF-1alpha) gene on the expression of vascular endothelial growth factor (VEGF) and pigment epithelium derived factor (PEDF) in human retinal pigment epithelium (RPE) cells under hypoxic condition. Two target sites of HIF-1alpha mRNA were chosen and two kinds of shRNA were designed and synthesized against the target sites. Then the two kinds of shRNA were transfected into human RPE cells in vitro, respectively. These cells were cultured under hypoxic condition that was simulated by using 150 mumol/L CoCl(2). The mRNA expressions of HIF-1alpha, VEGF and PEDF were tested by semi-quantitative reverse transcription PCR (RT-PCR). The protein levels of HIF-1alpha, VEGF and PEDF were analyzed by Western blotting. After the two kinds of HIF-1alpha-specific shRNA were transfected into RPE cells respectively, the expression of HIF-1alpha mRNA and the levels of HIF-1alpha protein were decreased significantly in RPE cells under hypoxic condition. The expression of VEGF mRNA and the levels of protein significantly were also decreased. However, the levels of PEDF protein was significantly increased, but the expression of PEDF mRNA showed no significant changes. In conclusion, HIF-1alpha-specific shRNA can effectively silence the HIF-1alpha gene, and consequently down-regulate VEGF and up-regulate PEDF expression against hypoxia. These results reveal that HIF-1 is associated with posttranslational mechanism for down-regulating PEDF under hypoxia and provide an explanation for hypoxia-provoked increases in VEGF/PEDF ratios. These results also suggest that HIF-1 is one of the key cytokines to retinal neovascularization.

In order to explore the effect of high glucose concentration and high glucose concentration with hypoxia on the production of hypoxia-inducible factor-1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF), human RPE cells were cultured in 5.56 mmol/L glucose (control group), 5.56 mmol/L glucose with 150 micro mol/L CoCl2 (hypoxic group), 25 mmol/L glucose (high glucose group) and 25 mmol/L glucose with 150 micro mol/L CoCl2 (combination group). RT-PCR was used to detect the expression of HIF-1alpha and VEGF mRNAs. Western blot analysis was used to measure the levels of HIF-1alpha and VEGF proteins. Although the small amount of HIF-1alpha protein was able to be detected in high glucose group but not in control group, there was no significant difference between the expression of HIF-1alpha mRNA of RPE cells in high glucose group and that of RPE cells in control group. As compared with RPE cells in control group, the mRNA expression and the protein synthesis of VEGF in high glucose group were up-regulated. As compared with RPE cells in hypoxic group, the expression of HIF-1alpha mRNA of RPE cells in combination group was not different, but the protein synthesis of HIF-1alpha, the mRNA expression and the protein synthesis of VEGF were more obviously up-regulated. In conclusion, high concentration glucose mainly influence the protein synthesis of HIF-1alpha of RPE cell, and HIF-1alpha protein is able to be accumulated in high concentration glucose. Under hypoxia, the HIF-1alpha protein induced by high concentration glucose is more stable, and the expression of VEGF is obviously increased. It is suggested that high concentration glucose may play a role in retinal neovascularization, especially at ischemia stage of diabetic retinopathy.

BACKGROUND:Immortalized cervical epithelial cels H8 can become cancerous under the induction of carcinogenic agent, and may cause cervical cancer when there is a cofactor interaction. However, there is stil a lack of effective intervention for female patients with precancerous lesions, and this treatment is blank in the clinic. OBJECTIVE: To explore the effects and mechanism of astragalus injection on apoptosis of human immortalizedcervical epithelial cels H8. METHODS: This study contained two groups: astragalus drug group and the blank control group. (1) Enzyme linked immunosorbent assay (ELISA) was used to detect DNA fragments of apoptotic H8 after astragalus injection. (2) Enzyme-labeling instrument was used to analyze the changes in caspase-3 and caspase-9 activities a fter astragalus injection. (3) Western blot assay was used to detect the protein expression changes of caspase-3, caspase-9 and PARP in H8 cels after astragalus injection. RESULTS AND CONCLUSION: (1) ELISA results showed that at 0, 6, 12 and 24 hours after 20 g/L astragalus injection, DNA fragments were gradualy increased with time prolonged in a time-dependent effect (P < 0.05). (2) Enzyme-labeling instrument demonstrated that at 0, 6, 12 and 24 hours after 20 g/L astragalus injection, caspase-3 and caspase-9 activities increased in a time-dependent manner (P < 0.05). (3) At 0, 6, 12 and 24 hours after 20 g/L astragalus injection, the expression of cleaved caspase-3 and cleaved caspase-9 were gradualy increased in H8 cels (P < 0.05). Cleaved PARP protein expression was gradualy decreased (P < 0.05). These findings indicate that astragalus injection could obviously induce H8 apoptosis, which may be associated with the upregulated protein expression of caspase-3 and caspase-9.

AIM To investigate the effect of fenofibrate and simvastatin on the serum free fatty acids of alcoholic fatty liver in rats. METHODS The rat model of alcoholic fatty liver was reproduced by chronic ethanol ingestion plus olive oil diet. The model rats were divided into three groups as follows: finofibrate treatment group(finofibrate 80 mg?kg -1 po, once a day),simvastatin treatment group (simvastatin 4 mg?kg -1 po, once a day)and control group without either above-mentioned treatment. Experimental rats were treated for four weeks and then sacrificed for blood sampling. Serum free fatty acids were analyzed by gas chromatography. RESULTS Fenofibrate significantly ameliorated the decrease in polyunsaturated fatty acids induced by ethanol [oleic acid:(38.212?7.788) ?g?L -1 vs (31.620?6.142) ?g?L -1,linoleic acid:(37.269?8.065) ?g?L -1 vs (30.254?9.063) ?g?L -1,arachidonic acid:(11.646?2.601) ?g?L -1 vs (9.012?1.236) ?g?L -1] accompanied by the improvement of the fat infiltration of the liver, but demonstrated no effect on the increase in serum saturated fatty acids by ethanol. In the contrast, simvastatin can aggravate the decrease in polyunsatrurated fatty acids and significantly increase the levels of satrurated fatty acids in serum induced by ethanol along with the pathological aggravation of alcoholic fatty liver. CONCLUSION The results of present study revealed that fenofibrate and simvastatin exerted different effect on the serum free fatty acids of alcoholic fatty liver. Polyunsatrurated fatty acids in the serum play an important role in the pathogenesis and treatment response of alcoholic fatty liver.

BACKGROUND:Large-scale expansion of undifferentiated and multipotential adipose-derived stem cells using serum-free culture system is a difficult issue to be resolved. OBJECTIVE:To establish an in vitro culture system combined with the extracellular matrix in order to investigate the efficiency, effectiveness and security of extracellular matrix on expanding adipose-derived stem cells. METHODS:In vitro isolated adipose-derived stem cells were seeded in traditional two-dimensional plastic plates and extracellular matrix-coated plates supplemented with serum-free medium respectively. After in vitro expansion, total cellnumber, expression of cellsurface markers, cellsenescence degree and multipotent differentiation ability (adipogenic, osteoblastic and chondrogenic differentiation) of adipose-derived stem cells cultured under both conditions were detected and compared. Moreover, the clinical safety of adipose-derived stem cells expanded in extracellular matrix-coated plates was investigated. RESULTS AND CONCLUSION:Total cellnumber of passage 5 adipose-derived stem cells cultured in extracellular matrix-coated plates was 10 times more than that in traditional two-dimensional plastic plates. Flow-cytometric analysis showed that adipose-derived stem cells cultured with extracellular matrix expressed stem cellsurface markers. cellular senescence examination showed that almost al of passage 15 adipose-derived stem cells cultured with extracellular matrix showed no aging, while most passage 5 adipose-derived stem cells cultured by the two-dimensional system aged and lost their proliferation ability. Multidirectional induction of adipose-derived stem cells showed that passage 15 adipose-derived stem cells cultured with extracellular matrix could stil differentiate into adipocytes, osteoblasts and chondrocytes as passage 5 adipose-derived stem cells did, which performed much better than the induced differentiations of passage 5 adipose-derived stem cells cultured by the two-dimensional system. Karyotype analysis and in vivo invasion experiment insured the clinical safety of adipose-derived stem cells expanded with extracellular matrix. Al above results suggest a safe and more efficient expansion system of extracellular matrix for clinical application using the serum-free culture system combined with extracellular matrix.

Objective To investgate prevalence and clinical significance of antiproteinase 3(PR3)an- tibodies in Wegener's granulomatosis(WG)and other vasculitis patients.Methods One hundred and eleven systemic vasculitis patients with WG(9 cases,including 21 serums of tracking WG patients)and other systemic vasculitis(102 cases),403 secondnary vasculitis CTD(SLE 213 cases,RA 135 cases),nephritis 62 cases,30 healthy subjects were examined for anti-PR3 and anti-MPO antibody by enzyme-linked immunosorbent assay (ELISA)and ANCA by indirect immunofluorescence(IIF)was performed.Result Anti-PR3 positive were 23 in 588 serums of patients.The prevalence of anti-PR3 positive was WG(16/21,71.4 %),other systemic vas- culitis were not found anti-PR3,SLE(6/213,2.8%),RA(1/135,0.7%).In particular,the prevalence of anti- PR3 and cANCA with WG tended to be higher in the patients with other systemic and secondnary vasculitis (P＜0.05).The sensitivity and specificity of anti-PR3 for diagnosis of WG were 71.42% and 98.58%.The sensi- tivity and specificity of combination anti-PR3 and cANCA were 61.90% and 99.82%.Anti-PR3 and cANCA are associated with treament of WG.Conclusion Anti-PR3 antibody has high specificity for diagnosis of RA. Detection of anti-PR3 and cANCA at the same time can improve the specificity considerably.As sensitive markers of WG,anti-PR3 antibody may be useful for diagonosis and early treament.Anti-PR3 also may be useful for activity and relapse of WG.

Metastasis is a multistep process involving modification of morphology to suit migration, reduction of tumor cell adhesion to the extracellular matrix, increase of cell mobility, tumor cell resistance to anoikis, and other steps. MicroRNAs are well-suited to regulate tumor metastasis due to their capacity to repress numerous target genes in a coordinated manner, thereby enabling their intervention at multiple steps of the invasion-metastasis cascade. In this study, we identified a microRNA exemplifying these attributes, miR-124, whose expression was reduced in aggressive MDA-MB-231 and SK-3rd breast cancer cells. Down-regulation of miR-124 expression in highly aggressive breast cancer cells contributed in part to DNA hypermethylation around the promoters of the three genes encoding miR-124. Ectopic expression of miR-124 in MDA-MB-231 cells suppressed metastasis-related traits including formation of spindle-like morphology, migratory capacity, adhesion to fibronectin, and anoikis. These findings indicate that miR-124 suppresses multiple steps of metastasis by diverse mechanisms in breast cancer cells and suggest a potential application of miR-124 in breast cancer treatment.
Anoikis ; Breast Neoplasms ; genetics ; metabolism ; pathology ; Cell Adhesion ; Cell Line, Tumor ; Cell Movement ; Connective Tissue Growth Factor ; metabolism ; DNA Methylation ; Down-Regulation ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; MicroRNAs ; genetics ; metabolism ; Neoplasm Metastasis ; rho GTP-Binding Proteins ; metabolism ; rho-Associated Kinases ; metabolism

To develop polyclonal antibodies against predicted B cell epitopes in HIV-1 accessory protein Vpr, the prepared consensus Vpr amino acid sequence was used to predict potential B cell epitopes by online softwares (ABCpred and Bcepred), the synthesized polypeptides of B-cell epitopes were subsequently conjugated with keyhole limpet hemocyanin (KLH) and then used to immunize rabbits. The antibody titers were determined by ELISA, and antibody specifity was analyzed by Western-Blotting and immunoprecipitation, respectively. Amino acid residues 3-19 (N) and 82-95 (C) of Vpr were predicted as the potential B cell epitopes. After inoculation of the conjugation of synthesized peptide to KLH, the antibody titers in rabbit sera against N and C peptides reached more than 1:100000 by ELISA. Western-Blotting analysis showed that the polyclonal antibodies reacted with both wild Vpr and fusion protein of GFP with Vpr, no matter Vpr was derived from HIV-1 B subtype or CRF07_BC recombinant form; Immunoprecipitation analysis showed similar reactions to Western-Blotting results. Two B cell epitopes of Vpr were successfully predicted by Bio-informatics methods and polyclonal antibodies against those peptides could be successfully prepared.
Animals ; Antibodies, Viral ; blood ; immunology ; Cell Line ; Enzyme-Linked Immunosorbent Assay ; HIV Infections ; blood ; immunology ; virology ; HIV-1 ; genetics ; immunology ; Humans ; Peptides ; genetics ; immunology ; Rabbits ; vpr Gene Products, Human Immunodeficiency Virus ; genetics ; immunology