三氧化二砷诱导K562细胞凋亡过程中血管内皮因子的变化

目的 通过观察三氧化二砷(As2O3)对人类慢性髓系白血病(CML)细胞株K562细胞的作用.探讨其治疗CML的作用机制.方法 将K562细胞与不同浓度As2O3共同孵育,于24、48、72 h用MTT方法检测K562细胞存活率,用AnnexinV-FITC检测凋亡细胞,同时用酶联免疫吸附法(ELISA)检测K562细胞上清液中血管内皮因子(VEGF)的浓度.结果 As2O3＜2 μmol·L-1时,对K562细胞增殖抑制和诱导凋亡的作用与空白对照组比较,无统计学意义(P＞0.05);As2O3＞2 μmol·L-1时则具统计学意义(P＜0.05);在相同作用时间下,AS2O3 浓度升高对K562细胞的增殖抑制率及细胞凋亡率亦升高;当As2O3＞8 μmol·L-1时,对K562细胞的抑制及细胞凋亡率不再上升. As2O3＜2 μmol·L-1时上清液中VEGF浓度与空白对照组比较无显著性(P＞0.05);As2O3＞2 μmol·L-1时VEGF的浓度差异有显著性(P＜0.05),且随As2O3 浓度的增加VEGF浓度上升;当As2O3＞8 μmol·L-1时则变化不显著(P＞0.05).结论 As2O3可抑制K562细胞增殖,并具诱导凋亡作用, As2O3在2.0～10.0 μmol·L-1梯度浓度,作用在24～72 h时段,表现为时间和剂量依赖性,还可下调VEGF表达水平.     

三氧化二砷通过JNK信号通路诱导K562细胞凋亡

目的:探讨c-Jun氨基末端激酶(JNK)信号通路在三氧化二砷(As2O3)诱导K562细胞凋亡中的作用及机制。方法:体外培养K562细胞,用As2O3及特异性JNK抑制剂SP600125对K562细胞进行处理;倒置相差显微镜下观察细胞形态学变化;MTT法检测不同时间点细胞增殖抑制率;AnnexinV/PI染色结合流式细胞术检测细胞凋亡率;ELISA检测p-JNK蛋白表达的变化;流式细胞术检测突变型P53表达。结果:ELISA显示4μmol/LAs2O3作用48h后p-JNK蛋白表达增强,经SP600125预处理后,As2O3诱导的K562细胞p-JNK蛋白表达明显减弱(P<0.01),As2O3诱导的细胞增殖抑制率和细胞凋亡率均下降,与As2O3单作用组相比突变型P53表达增加(P<0.05)。结论:JNK信号转导通路在As2O3诱导K562细胞凋亡过程中发挥重要作用,是As2O3诱导K562细胞凋亡的主要途径之一。     

三氧化二砷诱导K562/ADM耐药细胞凋亡中活性氧的作用

目的：探讨三氧化二砷（As2O3）诱导K562／ADM细胞凋亡中活性氧（ROS）的作用和其与耐药基因radr1及其编码的P-糖蛋白（P—gP）表达的关系。方法：应用噻唑蓝（MTT）比色法检测K562／ADM细胞增殖活性；Annexin V／PI染色法检测细胞凋亡；RT-P（鼠检测mdr1基因mRNA的表达；流式细胞术测定P-gP表达、ROS活性和细胞内阿霉素（ADM）含量。结果：As2O3显著抑制K562／ADM细胞的增殖，Annexin V／PI双染检测显示凋亡细胞明显增加：ROS活性明显下降；mdr1 mRNA和P-gP表达明显降低，P-gP功能受抑，细胞内ADM含量显著增高。结论：As2O3抑制K562／ADM耐药细胞的增殖活性和促进其凋亡，其机制可能为通过降低细胞ROS水平、抑制radr1/P-gp的表达和功能，进而逆转mdr1/P-gP介导的多药耐药和凋亡抑制。     

As2O3诱导K562细胞凋亡过程中细胞表面电荷变化的研究

目的 :研究三氧化二砷 (As2 O3 )诱导K562 细胞凋亡时细胞表面电荷的变化 ,阐明As2 O3 诱导K562 细胞凋亡的可能机制 ,为As2 O3 在临床上的应用提供理论依据。方法 :应用细胞电泳仪检测As2 O3 诱导K562 细胞凋亡中细胞电泳率的变化。通过细胞增殖、活力检测 ,形态学观察 ,亚G1期细胞含量和DNA凝胶电泳等鉴定细胞凋亡。结果 :1.0～ 2 0 μmol·L-1As2 O3 作用于K562 细胞 ,在细胞形态、DNA凝胶电泳、FCM显示出典型的细胞凋亡特征之前 ,试验组细胞表面电荷已在 1.6h左右开始下降 ,6h内降低最明显 ,6h后虽有下降 ,但幅度明显减弱。与对照组比较 ,低于 1.0 μmol·L-1的As2 O3 对细胞表面电荷影响不大。结论 :细胞表面电荷的下降是K562 细胞凋亡的早期事件。细胞表面电荷的下降有一定的时间范围 ,超过此范围 ,细胞表面电荷下降趋向无限大。     

4-[4″-(2″,2″,6″,6″-四甲基-1″-哌啶氮氧自由基)氨基]-4′-去甲表鬼臼毒素对K562/ADM细胞的抑制作用

目的比较4-[4″-(2″,2″,6″,6″-四甲基-1″-哌啶氮氧自由基)氨基]-4′-去甲表鬼臼毒素(GP-7)对多药耐药人慢性粒细胞白血病K562的多柔比星耐药株细胞(K562/ADM细胞)的抑制作用是否优于依托泊苷。方法以依托泊苷和K562细胞为对照,用不同浓度GP-7处理K562/ADM细胞不同时间,MTT比色法测定细胞增殖,流式细胞仪测定细胞周期和细胞凋亡率,普通光学显微镜观察细胞凋亡形态,琼脂糖凝胶电泳观察细胞DNA凋亡性降解。结果8~128mol.L-1GP-7处理48h或64μmol.L-1GP-7处理24~72h,GP-7对K562/ADM细胞的增殖抑制呈剂量依赖性(r=0.947,P<0.05)和时间依赖性(r=0.999,P<0.01)。GP-7及依托泊苷对K562/ADM的IC50分别为(45.9±1.8)及(68.7±4.6)μmol.L-1;64μmol.L-1GP-7作用48h可使G2/M期细胞明显增多,相同情况下依托泊苷则使S期细胞明显增多;GP-7可引起K562/ADM和K562细胞凋亡,但其引起的K562/ADM和K562细胞凋亡率与依托泊苷无明显差异;GP-7可引起K562/ADM和K562细胞典型的凋亡形态学变化和DNA凋亡性降解,但GP-7引起的K562/ADM细胞DNA凋亡性降解弱于K562细胞;128及256μmol.L-1GP-7或依托泊苷处理K562/ADM和K562细胞48h,GP-7诱导DNA凋亡性降解的作用强于依托泊苷,但32和64μmol.L-1时作用则相反。结论GP-7可抑制多药耐药白血病细胞株K562/ADM的增殖,诱导细胞凋亡。GP-7抑制多药耐药白血病细胞株K562/ADM的作用优于依托泊苷。     

三氧化二砷对胃癌细胞SGC7901/ADR GST-π和TopoⅡ表达的影响

目的探讨三氧化二砷(As2O3)对胃癌细胞SGC7901/ADR阿霉素(ADM)耐药性的逆转作用和对GSTπ和TopoⅡ表达的影响。方法用MTT法检测As2O3的非细胞毒性浓度和SGC7901/ADR细胞对ADM的敏感性,用流式细胞仪检测细胞内药物浓度及用免疫组织化学法检测细胞GSTπ和TopoⅡ的表达。结果与结论0.4~0.8μmol.L-1As2O3对耐药细胞SGC7901/ADR无明显毒性(P<0.01),As2O3可下调GSTπ表达,提高SGC7901/ADR细胞内ADM浓度,部分逆转SGC7901/ADR细胞对ADM的耐药性。     

孤啡肽逆转人白血病K562/ADM细胞耐药性的研究

目的 探讨孤啡肽对人白血病K562/ADM细胞对阿霉素耐药性的逆转作用.方法 测定孤啡肽的细胞毒性及其对K562/ADM细胞药物敏感性的影响,计算耐药逆转倍数.瑞士染色观察药物作用后K562/ADM细胞形态的改变;流式细胞仪检测阿霉素单用或与孤啡肽联用K562/ADM细胞凋亡百分率;DNA琼脂糖凝胶电泳观察孤啡肽与阿霉素联用后K562/ADM细胞内DNA的损伤程度.结果 非细胞毒性剂量的孤啡肽(10-7mol/L)作用于K562/ADM细胞时,对阿霉素耐药起到了部分逆转作用,使K562/ADM细胞的IC50由原来的(46.99±0.25)μg/ml降低至(23.11±0.29)μg/ml,其逆转倍数为2.03倍;孤啡肽(10-7mol/L)和阿霉素(20μg/ml)联合处理K562/ADM细胞48h,K562/ADM细胞呈典型的凋亡形态改变;细胞的凋亡率达到(18.73±3.90)%,明显高于两种药物在相同条件下单独作用的效果,P＜0.01;结论 孤啡肽能诱导K562/ADM细胞凋亡,从而能逆转K562/ADM细胞的耐药性,提高阿霉素的敏感性.     

MEK抑制剂联合三氧化二砷对髓系白血病细胞凋亡的研究

目的:研究MEK抑制剂PD98059联合三氧化二砷(As2O3)对髓系白血病细胞凋亡的影响及其作用机制。方法:将PD98059、As2O3单独或联合作用于髓系白血病细胞系HL-60、K562细胞,用AnnexinV-FITC法检测细胞凋亡,用流式细胞术检测Bcl-2、Caspapse-3表达。结果:联合组与单用组相比,细胞凋亡率明显增高。Bcl-2在HL-60、K562细胞均高水平表达。As2O3明显抑制HL-60细胞Bcl-2表达,对K562细胞Bcl-2无明显抑制作用。单用PD98059、As2O3及两药合用在诱导HL-60、K562细胞凋亡过程中,活化caspapse-3均明显上升,两药合用较单用PD98059或As2O3活化caspapse-3明显升高。结论:PD98059联合As2O3同时抑制ERK/MAPK和Bcl-2,激活Caspase酶,对HL-60细胞有协同促凋亡效应。两药联合同时靶向作用ERK/MAPK和BCR/ABL,活化Caspase酶,协同诱导K562细胞凋亡。PD98059可增强As2O3对髓系白血病细胞的凋亡诱导作用。     

钙调素拮抗剂O-(4-乙氧基-丁基)-小檗胺对人白血病细胞K562和K562/A02生长的抑制作用

目的研究钙调素拮抗剂O-(4-乙氧基-丁基)-小檗胺(EBB)体外抗白血病作用及机制。方法四唑盐(MTT)比色法评价EBB的细胞毒作用;流式细胞术检测细胞凋亡;荧光分光光度计检测细胞ROS水平;激光共聚焦显微镜检测[Ca2+]i含量。结果EBB明显抑制敏感K562和耐药K562/A02细胞增殖,IC50分别为(8.22±1.67)mol.L-1和(8.97±1.48)mol.L-1。EBB作用早期诱导凋亡,晚期引起坏死。K562/A02细胞的ROS静息值是K562细胞的2倍,EBB诱导两种细胞ROS生成呈浓度和时间依赖性,加入SOD明显降低坏死率。K562/A02细胞的[Ca2+]i低于K562细胞。EBB能明显增加两种细胞的[Ca2+]i。结论EBB明显抑制K562和K562/A02细胞增殖。信号分子Ca2+、ROS参与了EBB的细胞毒作用。     

联合应用化疗药物对三氧化二砷耐药白血病细胞的毒性实验研究

目的:探讨化疗药物联合应用对三氧化二砷(As2O3)耐药白血病细胞(K562/AS2)的毒性作用。方法:细胞毒实验采用MTT法,二药合用时细胞毒性作用采用ChouTalalay联合指数法分析,细胞表面P糖蛋白(Pgp)和细胞内柔红霉素(DNR)浓度测定采用流式细胞术测定。结果:K562/AS2细胞对三氧化二砷、柔红霉素、鬼臼乙叉苷(VP16)、三尖杉酯碱(H)、米托蒽醌(NVT)和阿糖胞苷(AraC)的耐药倍数分别为7.4、2.9、3.8、3.6、2.8和1.1。K562细胞和K562/AS2细胞的细胞表面Pgp或细胞内任意荧光强度无显著的统计学意义(P>0.05)。As2O3与DNR、VP16、H或NVT联合应用时,对K562、K562/AS2和Pgp表达的白血病细胞(K562/A02)细胞的联合指数均大于1。异搏定与DNR联合应用时,对K562和K562/AS2细胞的联合指数均大于1,但是对K562/A02细胞的联合指数均小于1。结论:K562/AS2细胞对As2O3、DNR、VP16和NVT耐药,其机制与Pgp表达无关。异搏定联合应用DNR可以逆转K562/A02对DNR的耐药性,不能逆转DNR对As2O3耐药细胞的耐药性。As2O3与DN、VP16、H和NVT联合应用时,对K562、K562/AS2和K562/A02细胞的毒性均为拮抗作用。     

Synthesis,Cytotoxicity and Antileishmanial Activity of Some N-(2-(indol-3-yl)ethyl)-7-chloroquinolin-4-amines

We report herein the condensation of 4,7-dichloroquinoline (1) with tryptamine (2) and D-tryptophan methyl ester (3) . Hydrolysis of the methyl ester adduct (5) yielded the free acid (6) . The compounds were evaluated in vitro for activity against four different species of Leishmania promastigote forms and for cytotoxic activity against Kb and Vero cells. Compound (5) showed good activity against the Leishmania species tested, while all three compounds displayed moderate activity in both Kb and Vero cells.     

Nachweis einiger Heilmittel in der Kniegelenksynovialflüssigkeit

Zusammenfassung Mittels Gaschromatographie und Dünschichtchromatographie wiesen die Autoren 11 Substanzen nach, welche durch Injektion oder nach Verabreichung per os in die Kniegelenksynovialflüssigkeit eindrangen. In ihrer Aufstellung konnten sie eine direkte Beziehung zwischen Struktur sowie chemischphysikalischen Eigenschaften der Substanz und ihrer Fähigkeit, aus dem Blut in die Kniegelenksynovialflüssigkeit einzudringen, nicht nachweisen, außer der Tatsache, daß Substanzen mit starker Affinität zu Eiweißstoffen erst in höheren Dosen nachweisbar waren.     

Lung disease and PKCs

The lung offers a rich opportunity for development of therapeutic strategies focused on isozymes of protein kinase C (PKCs). PKCs are important in many cellular responses in the lung, and existing therapies for pulmonary disorders are inadequate. The lung poses unique challenges as it interfaces with air and blood, contains a pulmonary and systemic circulation, and consists of many cell types. Key structures are bronchial and pulmonary vessels, branching airways, and distal air sacs defined by alveolar walls containing capillaries and interstitial space. The cellular composition of each vessel, airway, and alveolar wall is heterogeneous. Injurious environmental stimuli signal through PKCs and cause a variety of disorders. Edema formation and pulmonary hypertension (PHTN) result from derangements in endothelial, smooth muscle (SM), and/or adventitial fibroblast cell phenotype. Asthma, chronic obstructive pulmonary disease (COPD), and lung cancer are characterized by distinctive pathological changes in airway epithelial, SM, and mucous-generating cells. Acute and chronic pneumonitis and fibrosis occur in the alveolar space and interstitium with type 2 pneumocytes and interstitial fibroblasts/myofibroblasts playing a prominent role. At each site, inflammatory, immune, and vascular progenitor cells contribute to the injury and repair process. Many strategies have been used to investigate PKCs in lung injury. Isolated organ preparations and whole animal studies are powerful approaches especially when genetically engineered mice are used. More analysis of PKC isozymes in normal and diseased human lung tissue and cells is needed to complement this work. Since opposing or counter-regulatory effects of selected PKCs in the same cell or tissue have been found, it may be desirable to target more than one PKC isozyme and potentially in different directions. Because multiple signaling pathways contribute to the key cellular responses important in lung biology, therapeutic strategies targeting PKCs may be more effective if combined with inhibitors of other pathways for additive or synergistic effect. Mechanisms that regulate PKC activity, including phosphorylation and interaction with isozyme-specific binding proteins, are also potential therapeutic targets. Key isotypes of PKC involved in lung pathophysiology are summarized and current and evolving therapeutic approaches to target them are identified.     

Gender-related symptoms and correlates of alcohol dependence among men and women with a lifetime diagnosis of alcohol use disorders

This study explored gender-related symptoms and correlates of alcohol dependence in a crosssectional study of 150 men and 150 women with a lifetime diagnosis of alcohol use disorders (AUD). Participants were recruited in equal numbers from treatment settings, correctional centres and the general community. Standardized measures were used to determine participants' use of substances, history of psychiatric disorders and psychosocial stress, their sensation seeking and family history of substance use and mental health disorders. Multivariate analyses were used to detect patterns of variables associated with gender and the lifetime severity of AUD. Men had a longer history of severe AUD than women. Women had similar levels of alcohol dependence and medical and psychological sequelae as men, despite 6 fewer years of AUD. More women than men had a history of severe psychosocial stress, severe dependence on other substances and antecedent mental health problems, especially mood and anxiety disorders. There were differences in family history of alcohol-related problems approximating same-gender aggregation. The severity of a lifetime AUD was predicted by its earlier age at onset and the occurrence of other disorders, especially anxiety, among both men and women. The limitations in the generalizability of these findings due to sample idiosyncrasies are discussed.     

Biochemical and molecular characterisation of cubozoan protein toxins

Class Cubozoa includes several species of box jellyfish that are harmful to humans. The venoms of box jellyfish are stored and discharged by nematocysts and contain a variety of bioactive proteins that are cytolytic, cytotoxic, inflammatory or lethal. Although cubozoan venoms generally share similar biological activities, the diverse range and severity of effects caused by different species indicate that their venoms vary in protein composition, activity and potency. To date, few individual venom proteins have been thoroughly characterised, however, accumulating evidence suggests that cubozoan jellyfish produce at least one group of homologous bioactive proteins that are labile, basic, haemolytic and similar in molecular mass (42-46 kDa). The novel box jellyfish toxins are also potentially lethal and the cause of cutaneous pain, inflammation and necrosis, similar to that observed in envenomed humans. Secondary structure analysis and remote protein homology predictions suggest that the box jellyfish toxins may act as α-pore-forming toxins. However, more research is required to elucidate their structures and investigate their mechanism(s) of action. The biological, biochemical and molecular characteristics of cubozoan venoms and their bioactive protein components are reviewed, with particular focus on cubozoan cytolysins and the newly emerging family of box jellyfish toxins.     

Dose tolerability of chronically inhaled voriconazole solution in rodents

Invasive pulmonary aspergillosis (IPA) is a fungal disease of the lung associated with high mortality rates in immunosuppressed patients despite treatment. Targeted drug delivery of aqueous voriconazole solutions has been shown in previous studies to produce high tissue and plasma drug concentrations as well as improved survival in a murine model of IPA. In the present study, rats were exposed to 20 min nebulizations of normal saline (control group) or aerosolized aqueous solutions of voriconazole at 15.625 mg (low dose group) or 31.25 mg (high dose group). Peak voriconazole concentrations in rat lung tissue and plasma after 3 days of twice daily dosing in the high dose group were 0.85 ± 0.63 μg/g wet lung weight and 0.58 ± 0.30 μg/mL, with low dose group lung and plasma concentrations of 0.38 ± 0.01 μg/g wet lung weight and 0.09 ± 0.06 μg/mL, respectively. Trough plasma concentrations were low but demonstrated some drug accumulation over 21 days of inhaled voriconazole administered twice daily. Following multiple inhaled doses, statistically significant but clinically irrelevant abnormalities in laboratory values were observed. Histopathology also revealed an increase in the number of alveolar macrophages but without inflammation or ulceration of the airway, interstitial changes, or edema. Inhaled voriconazole was well tolerated in a rat model of drug inhalation.