Objective To investigate antimicrobial susceptibility of Klebsiella pneumoniae (K .pneumoniae )to aminoglycosides and detection of 16S rRNA methylase genes in K .pneumoniae .Methods Ninety-six non-repetitive clinical K .pneumoniae isolates were collected from Xiangya hospital of Central South University from January to Ju-ly 2009,minimal inhibitory concentrations (MICs)of gentamycin,amikacin and tobramycin were determined by agar dilution method ;genotype of 16S rRNA methylase genes (armA,rmtA,rmtB ,rmtC,rmtD ,npmA)were detec-ted by polymerase chain reaction(PCR).Results MIC50 of amikacin,gentamycin and tobramycin was 256μg/mL, 512μg/mL and 512μg/mL respectively;and MIC90 were all>512μg/mL;antimicrobial resistance rate was 21 .88%, 63.54%,and 41 ,67% respectively.68 isolates (70.83%)were resistant to at least one kind of antimicrobial agent, 21 isolates(21 .88%)were resistant to three kinds of antimicrobial agents.22 isolates(22.92%)carried armA,but rmtA,rmtB ,rmtC,rmtD and npmA were not detected;of 22 isolates harboring armA 16S rRNA methylase genes, 17(77.27%)were highly resistant to gentamicin,amikacin and tobramycin,the homology of armA positive isolate and armA (FJ410928.1 )was 100%.Conclusion armA 16S rRNA methylase gene harbored in K .pneumoniae plays an important role in aminoglycoside resistance.

miRNA was 20～25 nt endogenous non-coding RNA. miRNAs are encoded small RNAs that hybridize with messenger RNAs, resulting in degradation or translational inhibition of targeted transcripts. In order to investigate the function of miR-138 on mouse mammary epithelial cells, technique for gene silencing-miRNA inhibitor (anti-miRNA) was applied to make miR-138 silence, qRT-PCR was showed valid for inhibitor miR-138. And Western blot, CASY(r)-technology was put in use to study some change of mouse mammary epithelial cells after miR138 inhibitor. It was shown that miR-138 suppresses the exepress of PRL-R(P

Electromagnetic Fields ; Electromagnetic Radiation ; Humans ; Mitochondria ; radiation effects

AIM: To investigate the effect of Dendranthema morifolium (Ramat) Tzvel (DM) on isolated rat heart and ventricular myocytes during ischemia/anoxia and reperfusion/reoxygenation.METHODS: The Langendorff perfused rat hearts were used to measure intraventricular pressure and coronary flow. The cell contraction and intracellular calcium transient in enzymatically isolated ventricular myocytes were determined. RESULTS: (1) DM (0.5 g/L) significantly attenuated the inhibitory effects induced by ischemia/reperfusion on left ventricular developed pressure (LVDP), ?dp/dt max, coronary flow and LVDP?HR, meanwhile increased the content of SOD and decreased the content of MDA in the myocardium; (2) DM (0.5 g/L) attenuated the inhibitory effects of anoxia and reoxygenation on [Ca 2+]i transient and cell contraction in isolated ventricular myocytes. CONCLUSION: DM attenuated the effects on contractility and intracellular calcium induced by ischemia/anoxia and reperfusion/reoxygenation in the isolated rat heart and the ventricular myocytes. The mechanism might be related to increase in SOD activity and maintaining [Ca 2+]i homeostasis.

AIM: To investigate the effect of interleukin-2 (IL-2) on the intracellular calcium in electrically stimulated adult rat ventricular myocytes during anoxia and reoxygenation. METHODS: The isolated cardiac ventricular myocytes were exposed to 5 min anoxia followed by 10 min reoxygenation. Chemical anoxia was introduced by Krebs-Henseleit (K-H) solution containing 10 -3 mol/L sodium dithionite. The spectrofluorometric method was used to verify intracellular calcium transient with fura-2/AM as calcium fluorescence probe. RESULTS: It was shown that during anoxia, the amplitude of Ca 2+ transient was decreased, diastolic [Ca 2+ ] i, time to peak and time to relaxation of Ca 2+ transient were increased. All the parameters were got back but did not returned to the pre-anoxia level during reoxygenation. IL-2 at 2?10 5 U/L administrated during anoxia aggravated the effect of rexoxygenation on [Ca 2+ ] i transient. Pretreatment with a specific ? opioid antagonist, nor-BNI (10 -8 mol/L), abolished the effect induced by IL-2 during anoxia on the [Ca 2+ ] i transients, whereas specific ? opioid antagonist, naltrindole (10 -6 mol/L), did not cancel the effect. CONCLUSION: It is concluded that administration of IL-2 during anoxia aggravated the effect of reoxygenation on the [Ca 2+ ] i transients of isolated ventricular myocytes, which was mediated by cardiac ? opioid receptor pathway.

Objective To investigate the appropriate setting up of normal reference ranges of lymphocyte subsets in some flow cytometry laboratories and to study the effects of different flow cytometers and various reagents by different manufacturers on the analysis of peripheral blood lymphocyte subsets. Methods Three FCM labs (named A, B and C) in Beijing region were selected representing 3 commonly used flow cytometers (Beckman Coulter Epics XL, Beckman Coulter Cytomics FC500, BD FACS Calibur). 50 samples from healthy donors were distributed to 3 labs and tested according to individual lab's standard operating procedure to verify whether the normal reference ranges of peripheral blood lymphocyte subsets established were appropriate. The application of internal quality control was also investigated. Commercial blood quality control reagents were given to the 3 FCM labs and tested within 20 working days paralleled with routine samples. In addition, 20 patients' samples were prepared using 4 different combinations of reagents ( a , b , c and d). The results from combination a, which used the Beckman Coulter reagents and instrument, were compared to the results from combination b, c and d, which used reagents from different manufacturers. Then the prepared samples were tested on Beckman Coulter Epics XL to evaluate the effects of different combinations of reagents on the results of peripheral blood lymphocyte subsets analyzed by the same instrument. Furthermore, 24 patients' samples prepared by same reagents from Beckman Coulter company were tested on both Beckman Coulter Epics XL and BD FACS Calibur respectively to assess the effects of different instruments on peripheral blood lymphocyte subsets. 20 patients' samples prepared by same reagents and instruments were analyzed by Beckman Coulter Epics XL analytic system and BD FACS Calibur analytic system respectively to assess the effects of the two analytic systems on the lymphocyte subsets. Results Over 10% of the results for NK and T4/T8 in lab A as well as T4 in labs B and C fell outside of their normal reference ranges. The probabilities exceeding corresponding normal reference ranges were 16% ( 9/50 ), 24% ( 12/50 ), 22% (11/50) and 12% ( 6/50 ), respectively. The results using internal blood quality control in 3 FCM labs within 20 working days were all within the reference ranges of the quality control provided by the kit. The biases from b and c reagent combinations were substantial compared with that of reagent a combination. Among the biases from b and c reagent combinations, the lowest probability of bias exceeding 10% was T8 of combination b, which had probability of 70% (14/20). The highest probabilities of hias exceeding 10% were T3 and T4 of b and c reagent combinations, which reached 100% (20/20) . Furthermore, the biases of T3, T8 and B of d reagent combination compared with that of reagent a combination were also substantial. The probabilities of bias exceeding 10% were 35% (7/20) ,85% (17/20) and 75% (15/20), respectively. Comparing the results of samples prepared and analyzed by reagents and instruments from different manufacturers to that of samples prepared and analyzed by the same company's reagents and instruments showed that there were great discrepancies in T3, T4 , T8 , B and NK. The probabilities of bias exceeding 10% were 71% ( 17/24), 80% (19/24) ,38% (9/24), 33% (8/24) and 92% (22/24), respectively. The biases of T8, NK and B were substantial when compared the results from Beckman Coulter Epics XL analytic systems and BD FACS Calibur analytic systems. The probabilities of bias exceeding 10% were 55% (11/20 ), 70% ( 14/20 ) and 55% (11/20), respectively. Conclusions FCM labs should set up their own normal reference range for peripheral blood lymphocyte subsets. The normal reference range should be verified periodically. It is important to apply internal blood quality control regularly and accumulate the quality control results. The reagents and instrument for preparing peripheral blood samples should be from the same manufacturers.

Objective To investigate the role of outer membrane protein in clinical isolated car-bapenem resistance Acinetobacter baumannii. Methods Carbapenem resistance and sensitive strains were collected from the same patient. After MIST and REP-PCR analysis, carbapenemases were detected by isoe-lectric focusing. Different expressed membrane proteins were identified by two-dimension electrophoresis and mass spectrometry analysis. We also used efflux pump inhibitor PAβN(Phe-Arg-β-naphthylamide) to con-firm the phenotype. Results Carbapenem resistance and sensitive strains were attributed to the same pat-tern. At positions of P17.6 and P19.0, two β-lactamases were expressed in two investigated strains, no cabapenemases were detected. Six differential expressed membrane proteins were identified, a 34 × 10~3 membrane protein that was confirmed by efflux pump inhibitor PAβN experiment (imiponem MIC decreased from far above 32 μg/ml to 8μ/ml) and OprD and CarO. Conclusion Up-regulation of exported protein accompanied with down-regulation of OprD and CarO other than carbaponemases are responsible for carbap-enem resistance in A. baumannii.

Objective To investigate the correlation between RDW and disease activity in patients with IBD and evaluate clinical significance of RDW as a potential indicator to monitor IBD activity. Methods 256 patients with IBD were divided into two groups. One was UC group including 136 patients with 80 active period cases and 56 emission period cases. Another was CD group including 120 patients with 75 active period cases and 45 emission period cases. 60 bacillary dysentery diseases and 80 healthy controls were selected as bacillary dysentery group and healthy control group. RDW, Hb, WBC, PLT, CRP, ESR, MCV were tested and monitored with development of disease at different stages. We compared RDW with CRP,ESR, PLT, Hb, MCV parameters. By ROC curve analysis, the sensitivity and specificity of RDW was estimated in identifying the IBD's activity. Results The mean values of RDW in active UC group, remission UC group, bacillary dysentery group and control group were ( 16. 1 ± 2. 7), ( 13.5 ± 2. 1 ), ( 13.0 ± 2. 0)and ( 12. 8 ± 1.8), respectively. There was significant difference among four groups ( F = 51.9, P < 0. 01 ).RDW in active UC group was significant higher than that in remission UC group, bacillary dysentery group and healthy control group ( t = 8. 12, 9. 67, 11.85, P < 0.05) and RDW in remission UC group was significant higher than that in bacillary dysentery group and healthy control group as well ( t = 2. 45, 2. 67,P <0. 05). The mean values of RDW in active CD group, remissive CD group,bacillary dysentery group and control group were ( 16. 9 ± 2. 2 ), ( 13. 8 ± 1.1 ), ( 13.0 ± 2. 0), ( 12. 8 ± 1.8 ). There was significant difference among four groups ( F = 113.9, P < 0. 01 ). RDW in the active CD group was significant higher than that in remission CD group, bacillary dysentery group and healthy control group (t = 11.47,18.63,18. 72, P < 0. 05 ) and RDW in remission CD group was also significant higher than that in bacillary dysentery group and healthy control group ( t = 3.60, 3. 72, P < 0. 05 ). RDW in UC and CD groups demonstrated a positive correlation with CRP and ESR (r=0. 484, 0. 525, 0. 286, 0. 358 and P<0. 01, <0. 01, < 0. 05, < 0. 01, respectively) but an inverse correlation with Hb and MCV (r = -0. 378, -0. 271,- 0. 329, - 0. 298 and P < 0. 01, < 0. 01, < 0. 05, < 0. 01, respectively). In UC groups RDW represented a larger area under ROC curve (0. 8.54) compared with CRP, ESR, PLT, Fib and MCV. When the cut-off value of RDW was 14. 0, the sensitivity and specificity for identifying active UC were 82% (65/80) and 72% (40/56) respectively. In CD groups, the area of RDW under ROC curve was the largest (0. 925 )among all indicators. When the cut-off of RDW was 14.5, the sensitivity and specificity for identifying active CD was 88% (66/75) and 82% (37/45) respectively. Conclusion RDW in patients with IBD is a useful indicator to estimate the IBD activity and predict disease progression.

Objective To evaluate the biological variations of 8 lymphocyte subsets using flow cytometric double-platform method.Methods Twenty healthy adults were recruited from Peking Union Medical College Hospital in September 2013.At 8:00 AM,12:00 PM,and 4:00 PM on days 1,3,and 5,venous blood was collected from the volunteers.The percent and absolute lymphocyte subset counts were measured using duel-platform method.The sample collection and handling techniques were standardized.Before each batch analysis,the instrument quality controls were performed using the same lots of reagents.The intra-individual coefficient of variation (CVI) and inter-individual coefficient of variation (CVG) were calculated by nested ANOVA with SPSS 13.0 software.The analytical coefficient of variation (CVA),index of individuality (Ⅱ) and reference change value (RCV) were calculated by Excel2003.A mean pairwise comparison was determined by one-way ANOVA and variance analysis.The values between groups were analyzed by independent sample t test.Results For T cells (CD3 +),helper T cells (CD3 + CD4 + CD8-),suppressor T cells (CD3 + CD4-CD8 +) and B cells (CD3-CD19 +),the intra-individual coefficient of variation (CVI) and inter-individual coefficient of variation (CVG) were 0.03,0.06,0.05,0.14 and 0.12,0.16,0.23,0.31 respectively,which were all similar to those in previous studies.However,the Ⅱ and RCV of the four lymphocyte subsets were very different from those in previous studies,which were 0.26,0.40,0.22,0.44 and 8.77,16.86,14.93,39.69,respectively.Moreover,variations in absolute count,CVI,CVG,and analysis coefficient of variation (CVA) of all 8 lymphocyte subsets were greater than those of relative count.Variations in the percent and absolute counts for the CD3 + CD4-CD8-,CD3 + CD4 + CD8 +,and CD3+ CD16+ CD56+ cell subsets were relatively high.The CVI,CVG and CVA for the cells of CD3 + CD4-CD8-were 0.12,0.49 and 0.16.The CVI,CVG and CVA for the cells of CD3+ CD4+ CD8+ were 0.40,0.93 and 0.55.The CVI,CVG and CVA for the cells of CD3 + CD16 + CD56 + were 0.28,1.11 and 0.16.Conclusions Investigation on the CVI,CVG and CVA may allow us to obtain Ⅱ and RCV,by which we can determine the utility of traditional population based reference ranges.Documentation of the RCV indices may be used as objective delta-check values in quality management and decide whether clinical significance existed in the continuously detected results.

AIM: To study the effect of salvia miltiorrhiza (SM) on cell contraction and intracellular calcium of enzymatically isolated rat ventricular myocytes during normoxia and anoxia/reoxygenation. METHODS: Contraction and intracellular calcium were determined with video tracking system and spectrofluorometric method, and the chemical anoxic method was employed. RESULTS: The ?d L /d t max , dL of cell contraction and the amplitude of [Ca 2+ ] i in the cardiomyocytes following SM treatment were decreased in a dose-dependent manner. During anoxia, the ?d L /d t max , dL and amplitude of [Ca 2+ ] i were decreased, while the diastolic Ca 2+ level was elevated compared with control group. All the contractile parameters and the diastolic Ca 2+ level were back toward pretreatment values during reoxygenation, but could not return to control level. After the treatment with SM (3 g/L), ?d L /d t max and dL of cell contraction and the amplitude of [Ca 2+ ] i were higher and the diastolic Ca 2+ level was lower than those in anoxia/reoxygenation group. CONCLUSION: SM antagonized effects of anoxia and reoxygenation on cell contraction and intracellular calcium in isolated ventricular myocytes.