AIM To express recombinant human cytochrome P450 3A4 mutants CYP3A4.3, CYP3A4.4, CYP3A4.5 and CYP3A4.18, and to employ them for in vitro metabolism studies of CYP3A4. METHODS Use Bac-to-Bac baculovirus expression system to recombinant baculovirus carrying cDNA of CYP3A4 mutants CYP3A4.3, CYP3A4.4, CYP3A4.5 and CYP3A4.18. Spodoptera frugiperda 9 (Sf9), cells were co-infected by recombinant viruses of CYP3A4 mutants, human NADPH-P450 oxidoreductase and cytochrome b5 to obtain recombinant proteins CYP3A4.3, CYP3A4.4, CYP3A4.5 and CYP3A4.18 with metabolic activity. RESULTS The mRNA transcription of CYP3A4 mutants in Sf9 cells were validated by RT-PCR. Testosterone and 7-benzyloxy-4-(trifluoromethyl) coumarin were metabolized by the lysates of Sf9 cells infected by the recombinant viruses. CONCLUSION CYP3A4 mutants CYP3A4.3, CYP3A4.4, CYP3A4.5 and CYP3A4.18 with metabolic activity were successfully expressed by baculovirus-insect cell expression system. The results indicated that recombinant CYP3A4. 5 showed lower activity comparing to the wild type protein towards testosterone, while CYP3A4. 18 with higher activity, and for CYP3A4.3 and CYP3A4.4 showing similar activity to the wild type protein.

Objective To investigate the correlation of serum alpha-fetoprotein (AFP) with clinical pathological factors and prognosis of hepatocellular carcinoma ( HCC ).Methods Two hundred and twenty patients with HCC who hospitalized in our hospital from January 2007 to June 2009 were enrolled in this study.The relationship of AFP levels before operation to clinical pathological factors such as pathological grade,tumor size,tumor encapsulation,tumor numbers,portal vein tumor thrombus,TNM stage and postoperative recurrence were analyzed.Results Serum AFP concentration before operation were related with tumor size ( x2 =11.76,P ＜ 0.05 ),tumor encapsulation ( x2 =17.07,P ＜ 0.05 ),portal vein tumor thrombus ( x2 =8.944,P ＜ 0.05 ),higher TNM stage ( x2 =4.929,P ＜ 0.05 ) and early postoperative recurrence rate ( x2 =5.569,P ＜ 0.05 ).However,it had no correlation with pathological edmondson grade tumor numbers.Conclusion HCC patients with high serum AFP concentration often has greater tumor without encapsulation,high occurrence rate of portal vein tumor thrombus and higher TNM stage,they are more likely to early postoperative recurrence.Serum AFP before operation is useful not only for diagnosis,but also as a prognostic biomarker to predict prognosis and recurrence.

Objective To evaluate the activities of 18 pairs of antimicrobials combinations against non - duplicate clinical isolates of multidrug resistant Acinetobacter baumannii (MDRAB) in vitro.Methods Collect isolates of Acinetobacter baumannii from different patients from October 2009 to May 2010,which were isolated in Clinical Laboratory Center of Beijing Friendship Hospital,Capital Medical University.Use broth microdilution method to detect MIC of mono-antimicrobial,and checkerboard broth microdilution method to detect combinatied MIC,and calculate fractional inhibitory concentration (FIC) index to determine drug combinations effects.When the performance of the same drug combinations conflicted,appropriate strains were selected for screening of drug-resistant mechanisms by polymerase chain reaction( PCR),including efflux pump genes.Results In tests in vitro,rifampicin and polymyxin B,imipenem and gentamicin,cefepime and levofloxacin showed synergy at high proportion,68.1％,45.5％,40.9％,respectively.Minocycline and rifampicin,ampicillin/sulbactam and tobramycin.Ceftazidime and ciprofloxacin showed additive effect at high proportion,81.8％,68.2％,68.2％,respectively.There were several combinations which appeared the opposite effects to tested strains.Strains No.19 corresponding reaction was synergy and No.21,No.26 corresponding reactions were antagonism.The three strains above were selected for screening resistant mechanisms.The difference is that genotypes of adeS were negative in No.19 and positive in No.21 and No.26.Conclusion Rifampicin and polymyxin B combination showed synergy against the MDRAB in vitro,which can be considered as the treatment choice for critical infections caused by MDRAB.Imipenem and gentamicin,cefepime and levofloxacin also showed synergy in vitro,but in some isolates showed antagonism.This phenomenon may be due to the gene adeS activated by certain antibiotics,and the activated adeS drived efflux pump express or overexpress,which made the drugs in bacterial cells pumped out,causing antagonistic effect.The individual differences in strains should be considered when clinic strain apply these two combinations above.

Objective To evaluate the role of myosin light chain kinase(MLCK)in the increase in the permeability of human pulmonary microvascular endothelial cells(HPMVECs)induced by mechanical stretch.Methods HPMVECs were cultured in vitro and then randomly divided into 3 groups(n = 4 each): mechanical stretch group(group S),ML9 treatment group(group M)and control group(group C).In group S,the monolayer cells and magnetic beads coated with an RGD peptide were incubated in serum-free MCDB131 medium for 2 h,the unbound beads were washed out,and then the cells were exposed to magnetic twisting stimulation(MTS)for 2 h(frequency 3 Hz,intensity 4.2 mT).In group M,the monolayer cells and magnetic beads coated with an RGD peptide were incubated for 2 h in serum-free MCDB131 medium in which MLCK inhibitor ML9(50μmol/L)was added,and the other procedures were the same as in group S.In group C,the cells after washing as in group S were incubated for 2 h without exposing to MTS.Monolayer permeability was detected with FITC-dextran flux in transwell model.The distribution of integrin αVβ3 and actin was detected using the immunofluorescence assay.Results Compared with group C,the permeability of HPMVECs was significantly increased in group S,while no significant change was found in group M.Compared with group S,the permeability of HPMVECs was significantly decreased in group M.In addition,actin polymerized to form stress fiber,and integrin αVβ3 clustered at the end of stress fiber in group S,while actin mainly distributed in the surrounding area of the cell membrane and integrin αVβ3 evenly distributed on the cell surface in group C and M.Conclusion The mechanism by which mechanical stretch induces the increase in the permeability of HPMVECs is related to MLCK activation-mediated stress fiber formation and integrin αVβ3 clustering.

Objective To investigate the self-management self-efficacy,health-related quality of life of hemodialysis patients and explore the relationship between them. Methods Self-management self-efficacy questionnaire and SF-36 were used,and 55 hemodialysis patients participated in this study. Pearson correlate and regression analysis were used to test the relationship between them. Results 55 questionnaires were available. Positively associated between self-management self-efficacy and HRQOL(r=0.268,P＜0.05);significantly positively associated between physical activity self-efficacy and HRQOL(r=0.409,P＜0.01). Regression analysis showed that physical activity self-efficacy is the factor of HRQOL (t=3.261,P＜0.01).Conclusion Physical activity self-efficacy is the factor of HRQOL in hemodialysis patients,increasing self-management self-efficacy is benefit to their HRQOL.

Child ; Clinical Laboratory Techniques ; Humans ; Tuberculosis ; diagnosis

Gastrodin (GAS) is the major bioactive component of the extracts from the rhizome of Gastrodia elata Blume. The aim of this study is to investigate the transport of GAS in Caco-2 cells and the interaction of P-glycoprotein and GAS. The apparent permeability coefficients (Papp) of GAS were measured as a function of directions and concentrations. It was demonstrated that the efflux ratio was < 2.0 over the range of 50-500 micromol x L(-1) of GAS from bi-directional transport studies. The transport rate of GAS was dependent on the concentrations. Papp of GAS was not affected by transport directions, GAS concentration or the classical inhibitors of P-glycoprotein (verapamil and GF 120918). The cellular accumulation of GAS in Bcap37/MDR1 cells transected with hMDR1 gene, was similar to that in Bcap37 cells. The accumulation in both cell lines was concentration dependent. GAS did not affect the accumulation of Rhodamine 123 in Bcap37/MDR1 cells over the range of 50-500 micromol x L(-1). It indicated that the transport of GAS in Caco-2 cell monolayers mainly is by passive paracellular transport pathway. P-glycoprotein did not participate in the absorption of GAS in the intestine or the transport across the blood-brain barrier.

Objective To investigate the ability of theory of mind use in elders,and determine the role of inhibitory control in the use of theory of mind. Methods The participants included 30 young-old people ,30 oldold people and 30 young people. Their ability of theory of mind use and inhibitory control were examined with The Referential Communication Task and Hayling Sentence Completion Test,respectively. Results The correct rate of referential communication task was significantly lower in old-old people (0.43 ± 0.32) than those of young-old people (0.69 ±0.24, P=0. 001 ) and young people (0. 77 ±0. 25, P＜0. 001 ). Partial correlation analysis showed that the total errors in Hayling Test-B was negatively correlated with the correct rate of Referential Communication Task ( r= -0.442, P＜0.001 ). Hiberarchy regression analysis showed that the total errors in Hayling Test-B entered the equation ( β = -0.451, P＜0. 001 ) and could independently explained 16.8% of the variance (△R2 = 0. 168, △F = 19.359, P ＜ 0.001 ) after controlling for age. Conclusion Old-old people show worse performance in the use of theory of mind. The impairment of inhibitory control can partly explain the decline of theory of mind use in old-old people.

The regulatory effect of endothelial cells of the rabbit pulmonary arteriole(PAECs) on the tone of pulmonary artery was studied. The conditioned medium (CM) ofPAECs, cultivated under normal and hypoxic condition for 24 hours, did not induce con-striction of pulmonary artery ring. The aqueous phase was able to induce the constriction.The constriction of pulmonary artery rings was reduced by the heated aqueous phase, but it was disappeared when the heated aqueous phase was treated with trypsin. The constric-tion caused by heated aqueous phase was stronger in hypoxic endothelial cells than innormal cells. Treatment of PAECs with indomethacin and captoril, and administration ofFPL 55712, BN 52021, chlorpheniramine and prazozin to contraction pool did not affectthe constriction of pulmonary artery rings. It was concluded that PAECs might producelong half-life lipid substances which could induce the dilatation of pulmonary artery.Hypoxia inhibted the release of long half-life aqueous heat-unstable vasoconstrictive sub-stances. but enhanced that of the heat-stable fraction which was probably a peptide.

Objective: To understand the cognition status for risk factors and early symptoms of stroke via face to face questionnaire survey in 336 in-hospital patients with atrial ifbrillation (AF).
Methods: A face to face questionnaire survey was conducted by uniformly trained investigators in 336 AF patients who were admitted in our hospital more than 3 days from 2014-04-01 to 2014-10-01.
Results: There were 300/336 AF patients ifnished the questionnaire and among them 131 (43.7%) patients know that AF may increase the risk of stroke and 169 (56.3%) patients unknown or uncertain. There were 41.3%-79.3% patients having cognition for early symptoms of stroke and only 25.3% (76/300) patients may identify all 5 early signs of stroke.
Conclusion: In our research, AF patients had poor cognition for the risk and early signs of stroke. Medical professional should intensify the propaganda and education for stroke prevention.