Microbial lipases have been widely used in traditional industries,as well as in emerging biocatalysed areas owing to their ability to catalyze a variety of reactions in aqueous and non-aqueous media.Therefore,it is very important to enhance amount of lipase production by recombinant overexpression for meeting market demand.A critical review of main and novel strategies which have been employed for recombinant expression of microbial lipases are presented,including codon optimization,fusion and co-expression,dual expression system based on hybrid promoters,homologous overexpression,cell surface displaying and high-throughput screening based on gene library of expression.These new technologies are gradually coming to the forefront in the recombinant expression of lipase,especially for cell surface displaying and high-throughput screening based on gene library of expression.Meanwhile,several recombinant expressions for representative microbial lipases were also introduced and discussed,which are available for consultation when attempting to overexpress any lipases by scientists and industrialists.

Purpose:To study the effect of inhibition of telomerase activity and cell cycle by transcriptase telomerase inhibitors (3′ azido 3′ deoxythymidine, AZT) on squamous cell carcinoma of tongue in vitro.Methods:Human squmous cell carcinoma of tongue cell line Tca8113 was used as target cell. Telomerase activity was determined by TRAP PCR ELISA in untreated and treated Tca8113 by AZT, cell cycle phases were analyzed by flow cytometry. Results:Telomerase activity of Tca8113 was significantly inhibited when treated with AZT, and the effect of inhibition was dose dependant (rate of telomerase activity treated with AZT in 0.3, 0.6, 1.0, 1.5mol 10 -1 was 0.69 0.03, 0.61 0.08, 0.53 0.11, 0.50 0.02 respectively, rate of telomerase activity treated without AZT was 0.76 0.06). Cell cycle of treated Tca8113 was changed with marked increase in G 2 /M phase compared with untreated Tca8113 (62.8% vs 19.7%, P

Objective:To develop an HPLC method for the determination of capsaicine, dihydrocapsaicin, imperatorin and isoim-peratorin in Guanjie Jietong plasters. Methods:A CAPCELL PAK C18 column was used as the chromatographic column, and the flow rate was 0. 8 ml·min-1. The mobile phase A consisted of methanol-acetonitrile(1∶1), the mobile phase B was of 1% phosphoric acid. The detection wavelength was 280 nm and 254 nm, respectively. Results:There was a good linear relationship between the con-centrations and the peak areas within the range of 0. 298-5. 956μg(r=0. 9998) for capsaicine, 0. 152-3. 044μg(r=0. 999 6) for di-hydrocapsaicin, 0. 018-0. 352 μg(r=0. 999 5) for imperatorin and 0. 010-0. 204 μg(r=0. 999 3) for isoimperatorin. The average re-covery was 97. 8%(RSD=1. 02%), 97. 0%(RSD=0. 76%), 96. 6%(RSD=0. 65%) and 98. 1%(RSD=1. 35%), respectively. Conclusion:The method is convenient, accurate, sensitive and repeatable, which can be used in the determination of capsaicine, di-hydrocapsaicin, imperatorin and isoimperatorin in Guanjie Jietong plasters.

Objective To investigate the transactivating effect of hepatitis C virus(HCV)core protein on inducible nitric oxide synthase(iNOS)gene promoter and the molecular biological mecha- nisms of HCV pathogenesis.Methods Polymerase chain reaction(PCR)technique was employed to amplify the sequence of iNOS promoter by using HepG2 genomic DNA as template,and the product was cloned into pGEM-T vector.The iNOSp gene was cut from T-iNOSp by KpnⅠand XhoⅠ,and then was cloned into pCAT3-Basic,the constructed vector was named as pCAT3-iNOSp,pCAT3-iN- OSp was transfected into the LO_2 cell line.LO_2 cell was also cotransfected with pcDNA3.1(-)-core and pCAT3-iNOSp by FuGENE 6 transfection reagents.The LO_2 cells transfected with pCAT3-Basic was used as negative control.The activity of CAT in LO_2 cells was detected by an ELISA kit after 48 hours,which reflected the transactivating function of HCV core protein to iNOS gene promoter.Re- sults The expressive vector pcDNA3.1(-)-core and report vector pCAT3-iNOSp had been construc- ted and confirmed by restriction enzyme digestion and sequencing.The expression of CAT in LO_2 cells transfected with pCAT3-iNOSp and peDNA3,1(-)-core was 11 times as higher as that of pCAT3-bas- ic,and 6 times as higher as that of pCAT3-iNOSp.Conclusion It is suggested that HCV core protein can transactivate iNOS gene promoter.

The fermentation conditions of lipase production by Geotrichum candidum Y162 were optimized. Initially, the most suitable carbon olive oil, nitrogen source soybean flour and NH4Cl, salt BaCl2 and MgCl2 were selected according to single factorial experiments respectively. Based on the result, screening methodology Plackett-Burman design was used to evaluate the effects of twelve factors related to lipase production and three statistically significant factors olive oil, BaCl2 and NH4Cl were selected. The path of steepest ascent was used to approach the optimal region of lipase production subsequently. Then, the optimal combined concentration for maximum enzyme activity were further optimized by response surface methodology and determined as follows: olive oil 2.35%, BaCl2 0.36%,and NH4Cl 4.69%.The optimization of culture conditions of G.candidum Y162 led to a 2.25-fold increase in lipase production relative to initial result 14.16 U/ml, which indicate that single factor in combination with response surface methodology is an effective method for optimization of lipase production conditions by G.candidum Y162.

Adult ; Aged ; Humans ; Middle Aged ; Pneumoconiosis ; diagnostic imaging ; Radiography, Thoracic ; Retrospective Studies

Human bocavirus (HBoV) 1-4 have been detected both in respiratory and stool samples since the first HBoV was discovered in 2005. HBoV-1 is mostly associated with respiratory infection, while HBoV 2-4 are usually associated with intestinal tract infection. A variety of signs and symptoms have been described in patients with HBoV infection, including cough, wheezing, pneumonia, and diarrhea, but the research on pathogenic mechanism of HBoV is limited because HBoV cannot be cultured in vitro due to the lack of appropriate host cells. Three-dimensional epithelial cell culture, reverse genetics, and viral metagenomics are identified as novel tools that may promote the research on pathogenic mechanism of HBoV and the discovery of new viruses. This review summaries currently available diagnostic approaches such as electron microscopy, cell culture, PCR, and immunoassay in order to provide a method reference for indepth research on HBoV.
Animals ; Human bocavirus ; genetics ; growth & development ; isolation & purification ; pathogenicity ; Humans ; Parvoviridae Infections ; diagnosis ; virology ; Viral Proteins ; genetics ; metabolism ; Virology ; methods ; Virulence ; Virus Cultivation

OBJECTIVE To have an retrospective analysis of the prevalent fungi infected in patients in ICU,and their pathologic sites and drug resistance and to find out the hazard factors.METHODS The research used API 20C AUX and Rosco disk diffusion method to test the drug resistance of fungi isolated from the infected samples collected from ICU patients in our hospital between Jan 2006 and Mar 2007.RESULTS Totally 123 fungi strains were spotted,the majority of them being Candida albicans,accounting for 34.1%;Candida glabrata 26.8% and Candida tropicalis 18.7%.All the fungi were sensitive to amphotericin B and nystatin(Mycostatin) and partially resistant to fluconazole,itraconazole and ketoconazole.Patients with long term use of wide-spectrum antibiotics and catheters and elder patients would have higher fungi infection rate.CONCLUSIONS Candida are the major pathogens of fungal infections in ICU.The major infected site is lower respiratory tract.Amphotericin B and nystatin have good sensitivity but higher toxicity.Conazole medicines also have good sensitivity to C.albicans and C.tropicalis.But they are resisted largely by C.glabrata and C.krusei.Reasonable use of bacteriophage and reduction of unnecessary diagnosis and treatment procedures,and early discovery,diagnosis and treatment are keys to prevent and cure the invasive fungal infections.

Objective To assess the diagnostic value of anti-mutated citrullinated vimentin (MCV)antibody for rheumatoid arthritis (RA).Methods Meta analysis.The databases,including PubMed,The Excerpta Medica Database (Embase) and China National Knowledge Infrastructure (CNKI),from January 1990 to January 2013,were employed to search for the studies related to diagnostic value of anti-MCV for RA.The software Meta-Disc1.4 was used to perform Meta-analysis,investigate the sources of heterogeneity and draw the summary receiver operating characteristic (SROC) curve.The Begg's test was applied to evaluate the publication bias by use of Stata1 1 software.Results A total of 27 studies were included.Pooled sensitivity was 0.74[95％ confidence interval (CI) 0.73 to 0.76] and pooled specificity was 0.83 [95％ CI (0.82 to 0.85)].The area under the summary receiver operating characteristic curves was 0.896.Twenty-three literatures which detected both anti-MCV antibody and anti-cyclic citrullinated peptide(antikCCP) antibody show that:the area under curre of anti-MCV antibody and anti-CCP antibody were 0.887 and 0.924 respectively,but there was no significant difference (Z =1.525,P =0.064).Conclusions Results of the meta-analysis demonstrated that the MCV antibody may be a useful parameter in diagnosing RA as it shows high sensitivity and specificity.But there was no statistical difference between the results of anti-MCV antibody and anti-CCP antibody for the RA diagnosis.

Objective To investigate the value of procalcitonin (PCT) on predicting the severity and prognosis in patients with early acute respiratory distress syndrome (ARDS).Methods A prospective observation study was conducted. A total of 113 patients with ARDS undergoing mechanical ventilation admitted to intensive care unit (ICU) of Affiliated People's Hospital ofJiangsu University from October 2012 to April 2016 were enrolled. Based on oxygenation index (PaO2/FiO2), the patients were classified into mild, moderate, and severe groups according to Berlin Definition. Twenty-five healthy volunteers were served as controls. Demographics, acute physiology and chronic health evaluation Ⅱ (APACHE Ⅱ) score, and Murray lung injury score were recorded. Within 24 hours after diagnosis of ARDS, the serum levels of PCT and C-reactive protein (CRP) were determined by enzyme-linked fluorescence analysis (ELFA) and immune turbidimetric method, respectively. The patients were also divided into survival and non-survival groups according to clinical outcome within 28-day follow-up, and the clinical data were compared between the two groups. Spearman rank correlationwas applied to determine the correlation between variables. The predictive value of the parameters on 28-day mortality was evaluated with receiver operating characteristic curve (ROC). Kaplan-Meier survival curve analysis was used to compare different PCT levels of patients with 28-day cumulative survival rate. Results After excluding patients who did not meet the inclusion criteria and loss to follow-up, the final 89 patients were enrolled in the analysis. Among 89 ARDS patients analyzed, 27 of them were mild, 34 moderate, and 28 severe ARDS. No significant differences were found in age and gender between ARDS and healthy control groups. Infection and trauma were the most common etiology of ARDS (55.1% and 34.8%, respectively). Compared with healthy control group, both CRP and PCT in serum of ARDS group were higher [CRP (mg/L): 146.32 (111.31, 168.49) vs. 6.08 (4.47, 7.89), PCT (μg/L): 3.46 (1.98, 5.56) vs. 0.02 (0.01, 0.04), bothP < 0.01], and the two showed sustained upward trends with the ARDS course of disease. Compared with mild group, severe group had significantly higher APACHE Ⅱ and Murray scores. Spearman rank correlation analysis showed that both serum PCT and CRP in patients with ARDS was correlated well with APACHE Ⅱ score (r values were 0.669 and 0.601, respectively, bothP < 0.001), while PCT was weakly but positively correlated with Murray score (r = 0.294,P = 0.005), but not the case of CRP (r = 0.203,P = 0.052). APACHE Ⅱ score and serum PCT in non-survival group (n = 38) were significantly higher than those of the survival group [n = 51; APACHE Ⅱ score: 26.00 (23.00, 28.50) vs. 21.00 (17.00, 25.00), PCT (μg/L): 6.38 (2.82, 9.49) vs. 3.09 (1.08, 3.57), both P < 0.01], but Murray score and CRP level were similar between survivors and non-survivors. The areas under ROC curve (AUC) of APACHE Ⅱ score and PCT for predicting 28-day mortality were 0.781 and 0.793, respectively, which were better than those of AUC of Murray score and CRP (0.606 and 0.561, respectively, allP < 0.05). The AUC of APACHE Ⅱ score combined with PCT was significantly higher than that of PCT (0.859 vs. 0.793,P = 0.048) or APACHE Ⅱ score alone (0.859 vs. 0.781,P = 0.038). Using a PCT cut-off value of > 4.35μg/L for predicting 28-day mortality, the sensitivity and specificity was 92.2% and 63.2%, respectively, and the positive and negative likelihood ratios were 2.50 and 0.12 respectively. Kaplan-Meier survival curve analysis indicated that the patients whose PCT more than 4.35μg/L, had lower 28-day cummulative survival rate as compared with those with PCT ≤ 4.35μg/L (log-rank test: χ2 = 5.013,P = 0.025).Conclusion The elevated serum PCT level in patients with ARDS seems to be correlated well with the severity of lung injury, and appears to be a useful prognostic marker of outcome in the early phases of ARDS.