2005年 · 第118卷第02期

Background Arsenic trioxide (As2O3) has been identified as a very potent anti-acute leukemic agent. However its role in apoptosis needs to be elucidated. As2O3 interferes with the proliferation and survival of tumor cells via a variety of mechanisms. Drug-target interactions at the level of nuclear matrix (NM) may be critical events in the induction of cell death by As2O3. This study dealt with As2O3 - target interactions at the level of NM in chronic myelogenous leukemia cell line K562 by proteomics. Methods K562 cells were cultured in MEM and treated with different concentrations of As2O3. The nuclear matrix proteins were analyzed by high-resolution two-dimensional gel electrophoresis and computer-assisted image analysis. Results As2O3 significantly inhibited the growth of chronic myelogenous leukemia cell line K562 at low concentrations. While more than 200 protein spots were shared among the nuclear matrices, about 18 distinct spots in the nuclear matrices were found characteristic for As2O3 treated cells. Conclusions: As2O3 induces apoptosis in K562 cells in a dose and time-dependent manner. Our results demonstrated that for the detection of the onset of apoptosis, the alteration in the composition of nuclear matrix proteins was a more sensitive indicator than nucleosomal DNA fragmentation test. These results indicated that As2O3 might be clinically useful in the treatment of chronic myelogenous leukemia. The changes of nuclear matrix proteins in the treated cells can be used as a useful indicator for this treatment. Chin Med J 2004; 118 (2): 100-104

Background An effective purging technique plays an important role in autologous hematopoietic stem cells transplantation. Photodynamic therapy (PDT) provides a novel approach for this purpose. This study dealt with the purging effects of di-sulfo-di-phthalimidomethyl phthalolcyanine zinc (ZnPcS2P2) -based photodynamic therapy (ZnPc-PDT). Methods Fluorescence intensity of cell extracts was measured using a fluoresence spectrophotometry. The proliferative potency of K562 cells and HL60 cells was detected using MTT colorimetric assay, Typan blue dye exclusion method, colony formation test. The proliferative potency of normal hematopoietic cells was evaluated using mixture colony -forming unit (CFU-Mix), granulocyte-macrophage colony-forming unit (CFU-GM), and erythrocyte colony-forming unit (CFU-E) assays. K562 cells were mixed with normal mononuclear cells(MNCs) at ratios of 1: 100 and 1: 1000 for creating the model of simulated remission bone marrow. Colony formation test and nested-RT-PCR were carried out to detect the residual K562 cells in cell mixture. Results After a 5-hour incubation with ZnPcS2P2, the content of ZnPcS2P2 in normal MNCs was the lowest value. At the same time, the content in K562 cells and HL60 cells was very high. Therefore, the time point was selected as the optimal one for irradiating the cell suspensions. ZnPc-PDT could significantly kill proliferative K562 cells and HL60 cells in a dose-dependent manner. At the concentration of 1. 0 (μg/ml, the inhibitory rate of ZnPc-PDT on the colony formation was 100% for K562 cells, 89. 7% for HL60 cells. 0. 25 μg/ml ZnPc-PDT could completely photoinactivate residual K562 cells in the simulated remission bone marrow. Under an identical condition, the inhibitory rates of CFU-Mix, CFU-GM, CFU-E were 18. 0%, 18. 6%, and 17. 8% respectively. Conclusion ZnPc-PDT appears to be a promising approach for bone marrow purging. Chin Med J 2005; 118(2): 105-110

Background This study aimed at investigating the change and significance of nuclear factor-κB (NF-κB) in cardiomyopathy induced by adriamycin (ADR) in rats. Methods Sixty male Wistar rats were randomly divided into three groups: control, ADR and ADR + pyrrolidine dithiocarbamate (PDTC) groups. After 30-day experiment, myocardial histopathological observation was performed. Location and distribution of NF-κB p50 was examined by immunohistochemical assay. Expression of NF-κB p50 protein was examined by immunobolt assay. Electrophoretic Mobility Shift Assay examined activity of NF-κB; Myocardium p53 gene expression was examined by RT-PCR analysis. Results The myocardial lesions of rats were less pronounced in ADR + PDTC group than in ADR group. Compared with control group, there were many myocardium nucleuses, which expressed NF-κB p50 and distribute under epicardium. Expression of NF-κB p50 protein in nucleus increased significantly in ADR group. The NF-κB binding activity increased significantly in ADR group. Myocardium expressions of p53 mRNA increased in ADR group. Conclusions The NF-κB binding activity increased significantly in cardiomyopathy induced by ADR in rats. Moreover, NF-κB plays an important role in causing degeneration of myocardial tissue and regulating expression of related-apoptosis genes. Chin Med J 2005; 118(2): 111-115

Background Many patients with obstructive sleep apnea syndrome (OSAS) have complicated with hypertension and may be prescribed with antihypertension medications to control their blood pressure. But whether antihypertension medications can also decrease arterial stiffness or control the blood pressure increasing following obstructive events is not well described. This study aimed to investigate whether antihypertensive medications can ameliorate the changes in arterial stiffness and blood pressure associated with OSA. Methods Sixty-one OSAS patients [13 women, 48 men, mean age (53. 4 ±12. 3) years], 26 normotensive patients (N), 7 hypertensive patients on no antihypertension medications (H), and 28 hypertensive patients on various combination antihypertension therapy (HM), were prospectively diagnosed with standard nocturnal polysomnography. Beat-to-beat blood pressure was continuously recorded from the radial artery by applanation tonometry during baseline sleep. As a measure of arterial stiffness, arterial augmentation index (AAI) was calculated as the ratio of augmented systolic blood pressure (SBP) to pulse pressure and expressed as a percentage for the following conditions: awakening, the first 10 ("early apnea")and last 10 ("late apnea")cardiac cycles of obstructive events (apnea or hypopnea), and the first 15 cardiac cycles following event termination ("post apnea")for all events with nadir 02 saturation ≤89%. Results Systolic blood pressure (SBP) post-apnea [(142. 74 ±13.06) mmHg (N), (137.06 ±26.56)mmHg (H), (136. 94 ± 14. 1) mmHg (HM)] was significantly increased from awakening [(135. 76 ± 14. 76)mmHg (N), (135.58 ±23. 17) mmHg (H), (129. 77 ±14.00) mmHg (HM)], early apnea [(130.53 ±12. 65) mmHg (N), (124. 47 ±24. 97) mmHg (H), (126. 04 ± 13. 12) mmHg (HM)], and late apnea[(129. 8 ± 12. 68) mmHg (N), (124. 78 ± 25. 15) mmHg (H), (124. 48 ± 13. 82) mmHg (HM)]respectively (P <0. 001, repeated measures ANOVA) . AAI was significantly increased for the N group (P < 0. 001) from awakening to late apnea [(10. 45 ±2. 62) % vs (14. 43 ± 3. 21) %] and from early apnea to late apnea [(10. 61 ± 2. 34) % vs (14. 43 ± 3. 21) %], and also for H group (P <0. 05) from awakening to late apnea [(11. 23 ± 3. 87) % vs (16. 32 ± 8. 02) %] and from early apnea to late apnea [(11. 75 ± 3. 79) % vs(16. 32 ±8. 02) %]. Meanwhile, no significant differences in AAI among awakening, early apnea, late apnea, and post-apnea conditions were found in HM group. Conclusions The current data demonstrate that systemic blood pressure increases significantly during the post-apneic phase of OSAS, compared with that during awakening and intra-apnea phases even with the use of combined antihypertensive therapy which could normalize BP during awakening in the hypertensive patients. However, increases in arterial stiffness during obstructive events could be ameliorated by combined antihypertension medications. Chin Med J 2005; 118(2): 123-129

Background The hemodynamics and oxygenation severely fluctuated during the off-pump coronary artery bypass grafting (OPCABG). This study aimed at investigating whether or not nicardipine combined with esmolol(1: 10) can maintain systemic and tissue oxygenation during OPCABG. Methods Twenty patients scheduled for OPCABG were divided ramdomly into Group nicardipine (N) and Group nitroglycerine (X) respectively combined with esmolol (E) (Dosage ratio: 1 to 10) (Group N + E and Group X + E) with 10 patients in each group. The mixed solution of N + E or X + E were titrated to maintain mean arterial blood pressure between 70 and 80 mmHg following anesthesia induction. The variables of hemodynamics, arterial blood lactate content (Lac) and gastric intramucosal partial pressure of carbon dioxide were measured at the following time points: after induction of anesthesia (T2), pre -revascularization (T2), grafting of left anterior descending (T3), right coronary descending (T4) and left coronary circumflexus branches (T5), post-revascularization (T6), the end of operation (T7) . The delivery of oxygen (DO2),consumption of oxygen (VO2) and gastric intramucosal pH (pHi) were calculated. Results The cardiac index (CI) in Group N + E was significantly increased (P <0. 05) as compared with T1 during OPCABG, while it was mildly decreased in Group X + E. The stroke volumes at T4, T5 in Group N + E and at T3 -T6 in Group X + E were significantly decreased (P <0. 05). The systemic vascular resistance indices in Group N + E were significantly decreased as compared with T1 (P <0. 05) . The heart rates in these two Groups were significantly elevated intraoperatively (P <0. 05) . The DO2 after the infusion of N + E was significantly increased (P <0. 05) or leveled to T1, and the Lac were within the normal range. But the DO2 in Group X + E was decreased throughout the procedure, reaching significant level at T5 (P <0. 05), and the Lac was significantly increased beyond normal range (P <0. 05) . The pHi in Group N + E was maintained above 7. 35 during OPCABG, while it was less than 7. 35 from T4 to T7 in Group X + E. Conclusion Nicardipine combined with esmolol (1: 10) regimen may maintain systemic and tissue oxygenation during OPCABG. Chin Med J 2005; 118(2):130-135

Background Acinetobacter baumannii is one of the main gram-negative bacilli in clinical practice. Nosocomial infections caused by multi-drug resistance Acinetobacter baumannii is very difficult to treat. This study was designed to investigate the antimicrobial resistance characteristics and four resistant gene expressions of aminoglycoside-modifying enzymes including N-acetyltransferases and O-phosphotransferases in Acinetobacter baumannii. Methods Bacterial identification and antimicrobial susceptibility test were performed by PhoenixTM system in 247 strains of Acinetobacter baumannii. Minimal inhibitory concentrations (MICs) of seven aminoglycosides including gentamicin, amikacin, kanamycin, tobramycin, netilmicin, neomycin and streptomycin in 15 strains of multi-drug resistant Acinetobacter baumannii were detected by agar dilution. Four aminoglycoside-modifying enzyme genes were amplified by polymerase chain reaction (PCR) and verified by DNA sequencer. Results The resistance rates of 247 strains of Acinetobacter baumannii against cefotaxime, levofloxacin, piperacillin, aztreonam, tetracycline, ciprofloxacin and chloramphenicol were more than 50%. Imipenem and meropenem showed high antibacterial activities with resistance rates of 3. 2% and 4. 1%. MIC50 and MIC90 of gentamicin, amikacin, streptomycin and kanamycin in 15 strains of multi-drug resistant Acinetobacter baumanii were all more than 1024 mg/L, and the resistance rates were 100%, 100%, 100%and 93. 3%, respectively. But their resistance rates to tobramycin, netilmicin and neomycin were 86. 7%, 93. 3% and 46. 7 %, respectively. Three modifying enzyme genes, including aacCl, aacC2 and aacA4 genes, were found in 15 strains, but aphA6 had not been detected. Their positive rates were 93. 3%, 20. 0% and 20. 0%, respectively. These three genes existed simultaneously in No. 19 strain. Nucleotide sequences of aacCl, aacC2 and aacA4 genes shared 100%, 97. 9% and 99.1% identities with GenBank genes (AY307113, S68058 and AY307114). Conclusion Multi-drug resistant Acinetobacter baumannii strains are rapidly spreading in our hospital, and their resistance to aminoglycosides may be associated with aminoglycoside-modifying enzyme gene expressions. Chin Med J 2005; 118 (2): 141-145

Background Congestive heart failure (CHF) is a major cause of morbidity and mortality worldwide and angiotensin converting-enzyme inhibitor (ACEI) is the cornerstone in its treatment. However, CHF continues to progress despite this therapy, perhaps because of production of angiotensin Ⅱ (Ang Ⅱ) by alternative pathways. The present study was conducted to examine the combined effects of a chronic ACEI, ramipril, and a chronic Ang Ⅱ type 1 receptor blocker, TCV116, on rat CHF after myocardial infarction (MI). Methods Congestive heart failure was caused by MI in rats, which was induced by ligating the left anterior descending coronary artery. The experiment protocol included sham-operated rats (Sham), MI-control rats (MI-control), MI rats treated with ramipril 3 mg/kg (MI-ramipril) or TCV116 2 mg/kg (MI-TCV116) per day, half dosage (MI-1/2R&T) or full dosage (MI-R&T) combination of the two. At 22 weeks, cardiac hemodynamic parameters such as mean arterial pressure (MAP), left ventricular systolic pressure (LVSP), maximal rate of left ventricule pressure development and decline (LV dP/dtmax) and left ventricular end diastolic pressure(LVEDP), and cardiac morphometric parameters such as heart weight (HW), left ventricular weight (LVW)and left ventricular cavity area (LVCA) were measured, mRNA expressions of cardiac molecule genes such as β myosin heavy chain (βMHC), B-type natriuretic peptide (BNP), transforming growth factor-β1, (TGF-β1),collagen I and Ⅲ were quantified with reverse transcription polymerase chain reaction (RT-PCR) in the surviving septum myocardium, and survival rates were calculated. Results There were no significant differences in MI sizes (%) among each MI related experimental groups(33 ±13, 34 ±14, 33 ±13, 35 ±13 and 33 ±14 for MI-control, MI-ramipril, MI-TCV116, MI-1/2R&T and MI-R&T, respectively, no statistical significance for all). Compared with sham-operated rats, MI rats without therapy showed significant increases in morphometric parameters as well as in mRNA expressions of cardiac molecule genes (P <0. 01); while their hemodynamic parameters were significantly impaired (P <0. 01), and in terms of spontaneous deaths survival rate shortened (P <0. 05). Compared with MI rats without therapy, MI rats treated with each single drug showed significant attenuation of mRNA expressions of cardiac molecule genes(P <0. 01); while their hemodynamic parameters were significantly improved (P <0. 05 or P <0. 01), and in terms of spontaneous deaths survival rate prolonged (P <0. 05). Both half and full dosage combined treatments exerted more powerful effects on improvement of cardiac phenotypic changes and on attenuation of βMHC, BNP mRNA expressions (P <0. 05 vs monotherapy);while LVEDP was further lowered (P <0.05 vs monotherapy). However, the total death in MI rats with full dosage combined treatment was more though there were no significant differences when compared with other treatments. Conclusions The results suggest that treatment with appropriate dosage combination of a chronic ACEI and a chronic ARB may further improve cardiac remodeling and cardiac function after MI. Chin Med J 2005; 118(2): 146-154

Background Cecropin-XJ belongs to cecropin-B, which is the most potent antibacterial peptide found naturally. The aim of this study was to investigate the effects of cecropin-XJ on growth and adherence of oral cariogenic bacteria. Methods Four oral cariogenic bacteria (Streptococcus mutans, Lactobacillus acidophilus, Actinomyces viscosus and Actinomyces naeslundii) were chosen for this experiment. The minimal inhibitory concentrations (MICs) and reductive percent of bacterial growth were used to assay the antibacterial activity of cecropin-XJ. Mammalian cytotoxicity of cecropin-XJ was tested with human periodontal membrane fibroblasts by tetrazolium (MTT)colorimetric assay. The bacterial morphological changes induced by cecropin-XJ were examined on scanning electron microscope (SEM). The influence of cecropin-XJ on bacterial adhesion to saliva-coated hydroxyapatite(S-HA) was measured by scintillation counting. Results The MICs of cecropin-XJ for inhibition of the growth of four bacteria ranged from 4. 0 to 42. 8 μmol/L with the highest susceptible to A. naeslundii and the lowest susceptible to L. acidophilus. At pH 6. 8, 5. 5 and 8. 2, 1/2 MIC of cecropin-XJ reduced the number of viable bacteria by 40. 9%, 67. 8% and 32. 8% for S. mutans and by 28. 1%, 57. 2% and 37. 9% for L. acidophilus. The activities against S. mutans and L. acidophilus increased at pH 5. 5 compared with pH 6. 8 (P <0. 01, respectively) . In present of 50% saliva, 1/2 MIC of the peptide decreased the direct count of viable cells by 29. 2% and 14. 4% for S. mutans and L. acidophilus, respectively (P <0. 01 and P> 0. 05, respectively), whereas almost no reduction counts were detected in the presence of 20% serum for both bacteria (P> 0. 05, respectively) . Mammalian cytotoxicity of cecropin-XJ from 1. 0 to 100 μmol/L exhibited no cytotoxicity against human periodontal membrane fibroblasts(P> 0. 05). Bacterial morphological changes induced by MIC of cecropin-XJ examined on SEM showed cell surface disruption. Furthermore, the ability of A. naeslundii adhesion to S-HA decreased significantly with MIC of cecropin-XJ for 10 and 20 minutes (P = 0. 001 and 0. 000, respectively), and S. mutans, A. viscosus to S-HA decreased significantly with MIC of cecropin-XJ for 20 minutes(P=0. 000, respectively). Conclusions Cecropin-XJ exhibited bactericidal action against cariogenic pathogens, and the antibacterial activity enhanced in the acid environment. The results also demonstrate that cecropin-XJ prevents S. mutans and actinomyces adsorption to S-HA. These findings suggest that Cecropin-XJ may have potential to prevent caries. Chin Med J 2005: 118(2): 155-160

Background In previous work, we suggested that some 2-aminosteroids inhibited proliferation and induced differentiation of both human and murine leukemia cells. Here, we reported the actions of another new 2-aminosteroid designated as H89712 on human leukemia cells. Methods Cell colony counting and MTT assay were used to determine proliferation. Cell morphology, histochemical staining, UV detection and cytometry were used to determine differentiation. RT-PCR was used to detect gene expression. Standard statistical method was used to analyze data. Results H89712 inhibited proliferation of HL-60 leukemia cells and the inhibition percentage in MTT assay was 18% at the dose of 10 8 mol/L and 65% at the dose of 10-5 mol/L, respectively. The inhibition for HL-60 in colony assay was 23% at the dose of 10-8 mol/L and 96% at the dose of 10-5 mol/L, respectively. H89712 also induced HL-60 cells toward macrophage-like differentiation. It was verified by flow cytometry that the percentage of positive CD14 expression in differentiated HL-60 cells was about 9 times higher than that of the control at the dose of 10-8 mol/L and 20 times higher than that of the control at the dose of 10-5 mol/L respectively, and this action involved upregulation of MafB gene in HL-60 leukemia cells. On the other hand, H89712 inhibited proliferation of K562 leukemia cells and the inhibition of K562 leukemia cells in MTT assay was shown by 34%at the dose of 10-8 mol/L and 88% at the dose of 10-5 mol/L respectively. The inhibition of K562 leukemia cells in colony assay was 53% at the dose of 10-8 mol/L and 100% at the dose of 10-5 mol/L respectively. H89712 also induced K562 cells toward erythroid-like differentiation and it was verified by flow cytometry that the percentage of positive CD71 expression in differentiated K562 cells was about 9 times higher than that of the control at the dose of 10-8 mol/L and 16 times higher than that of the control at the dose of 10-5 mol/L respectively. This action was related to upregulation of Egr-1 gene in K562 leukemia cells. Conclusions Our results showed the important roles played by MafB in macrophage differentiation and Egr-1 in erythroid differentiation of human myeloid leukemia cells. Chin Med J 2005; 118 (2):91-99