人胚胎脑组织中低分子肿瘤抑制物的实验研究

人胚胎脑组织提取液可以抑制人白血病细胞系的生长。分析表明:肿瘤抑制活性主要分布在小于10kDa组分,经Sephadex-G25凝胶过滤可初步分离为单一组分。这种低分子肿瘤抑制物具有广谱的抗肿瘤效应,在体外可以抑制人白血病、肝癌、胃癌细胞系和小鼠粒、单核和淋巴系白血病细胞,但对人骨髓CFU-GM和小鼠骨髓CFU-GM的抑制作用较弱,说明人胎脑低分子肿瘤抑制物对肿瘤细胞具有一定的选择性抑制作用。     

苏云金芽孢杆菌Bt9875晶体蛋白诱导HL-60细胞凋亡

[目的]研究苏云金芽孢杆菌(Bacillus thuringiensis,Bt)Bt9875菌株晶体蛋白对人急性髓细胞性白血病细胞HL-60的影响.[方法]采用MTT比色、荧光显微观察、DNA凝胶电泳、流式细胞术等方法来检测不同浓度的Bt9875晶体蛋白处理后HL-60细胞的凋亡特征.[结果]Bt9875晶体蛋白对HL-60细胞的生长具有明显的抑制作用,且随着蛋白质浓度的增加对HL-60细胞生长抑制愈加明显,而对正常人外周血单个核细胞(PBMC)无作用;荧光显微镜下观察发现经该蛋白作用后HL-60细胞核的形态呈现凋亡特征;流式细胞术分析表明,HL-60细胞经100 μg/mL晶体蛋白作用后,凋亡率达到52%;琼脂糖凝胶电泳显示细胞DNA呈梯状降解.[结论]初步证明了Bt9875晶体蛋白在体外能够明显抑制HL-60细胞的增长,并诱导其凋亡,这为苏云金芽抱杆菌晶体蛋白的应用开创了新的思路.     

低分子抑瘤物净化白血病细胞的实验研究及临床应用

胎儿肝脏中存在一类小分子量（分子量〈１０ｋＤ）的肿瘤抑制物,其在体外对ＨＬ－６０等多种白血病细胞系具有明显的选择性抑制作用,对急性白血病患者骨髓的白血病祖细胞也具有这种选择性抑制效果;分离纯化获得两种天然低分子抑瘤物（７－ＫＣ和７－β－ＨＣ）和一种外源性低分子抑瘤物（ＤＢＰ）,研究证实其抑制白血病细胞生长的作用机制是诱导白血病细胞凋亡;将其应用于白血病及恶性淋巴瘤自体骨髓移植时的体外净化,完成１２     

氟苯达唑对人急性髓系白血病HL-60细胞的增殖抑制作用研究

目的:研究氟苯达唑对人急性髓系白血病HL-60细胞增殖的抑制作用,明确氟苯达唑对HL-60细胞周期,凋亡发生的作用机制。方法:噻唑蓝法(MTT)检测氟苯达唑对人急性髓系白血病HL-60细胞的生长抑制作用,流式细胞术检测氟苯达唑对HL-60细胞周期,DNA片段化的影响,免疫印迹法检测Caspase, Raf, Bcl-2家族蛋白表达。结果:氟苯达唑抑制人急性髓系白血病HL-60细胞生长,HL-60细胞G2/M期增加,与阴性对照组相比,在一定的剂量和时间内,差别具有显著统计学意义;DNA片段化上升,0.25,0.5,1μM组与对照组相比差别具有显著统计学意义,促使Cleaved PARP,Cleaved-caspase 3,Cleaved-caspase 9蛋白表达量趋势增加;Bag-1和Bcl-2蛋白表达量降低;b-raf,c-raf磷酸化蛋白表达水平逐渐降低。结论:氟苯达唑通过诱导HL-60细胞阻滞于G2/M期,增加DNA片段化水平,激活Caspase, Raf, Bcl-2家族介导的凋亡相关通路抑制人急性髓系白血病HL-60细胞增殖,诱导人急性髓系白血病HL-60细胞发生凋亡而发挥抗肿瘤作用。     

DADS对DJ-1核定位高表达人白血病HL-60细胞的影响

最近研究表明,DJ-1在许多肿瘤中过表达,而且DJ-1的核表达与肿瘤的生物学行为有关. 本文主要研究二烯丙基二硫（DADS）对DJ-1核定位高表达人白血病HL-60细胞生物行为学的影响,阐明DJ-1在细胞核内的功能,为临床诊断及治疗过程提供一个潜在的治疗靶点. 通过基因转染技术建立核内高表达HL-60细胞株（DJ-1/HL-60）,利用软琼脂集落形成实验、 MTT法、间接免疫荧光细胞化学实验、硝基蓝四氮唑( NBT)还原比色实验评估HL-60细胞的增殖与分化,DJ-1核定位过表达促进HL-60细胞的增殖并抑制其分化. Transwell迁移侵袭小室实验表明DJ-1核定位过表达可以促进HL-60的迁移和侵袭能力. Western blot结果表明DADS具有抑制HL-60细胞中核内DJ-1蛋白表达的能力. 说明DJ-1核定位高表达具有促进HL-60细胞增殖和迁移侵袭及抑制HL-60细胞分化的作用,DADS可以诱导DJ-1核定位高表达HL-60细胞分化及抑制迁移侵袭, DJ-1核定位高表达可减弱DADS抑制HL-60细胞增殖的作用.     

DADS对DJ-1核定位高表达人白血病HL-60细胞的影响（英文）

最近研究表明,DJ-1在许多肿瘤中过表达,而且DJ-1的核表达与肿瘤的生物学行为有关.本文主要研究二烯丙基二硫(DADS)对DJ-1核定位高表达人白血病HL-60细胞生物行为学的影响,阐明DJ-1在细胞核内的功能,为临床诊断及治疗过程提供一个潜在的治疗靶点.通过基因转染技术建立核内高表达HL-60细胞株(DJ-1/HL-60),利用软琼脂集落形成实验、 MTT法、间接免疫荧光细胞化学实验、硝基蓝四氮唑(NBT)还原比色实验评估HL-60细胞的增殖与分化,DJ-1核定位过表达促进HL-60细胞的增殖并抑制其分化. Transwell迁移侵袭小室实验表明DJ-1核定位过表达可以促进HL-60的迁移和侵袭能力. Western blot结果表明DADS具有抑制HL-60细胞中核内DJ-1蛋白表达的能力.说明DJ-1核定位高表达具有促进HL-60细胞增殖和迁移侵袭及抑制HL-60细胞分化的作用,DADS可以诱导DJ-1核定位高表达HL-60细胞分化及抑制迁移侵袭, DJ-1核定位高表达可减弱DADS抑制HL-60细胞增殖的作用.     

新生牛肝中低分子抑瘤物影响白血病细胞生长的实验研究

新生牛肝经匀浆、离心和分级超滤后,得到分子量≤1.0kDa的、耐热的成分——新生牛肝抑瘤物(new-bom calfliver suppressor,BLS-1)。对BLS-1的生物学活性进行了检测,结果表明在体外液体和半固体琼脂培养条件下,它对人和小鼠白血病细胞系(如HL-60、L833 和 P388 等)的生长均有明显的抑制作用,并存在着一定的量效关系;与同样条件下BLS-1对人和小鼠正常粒-巨噬系祖细胞的抑制作用相比较,BLS-1对白血病细胞生长的抑制作用具有较强的选择性。进一步观察了BLS-1对5例急性非淋巴细胞白血病病人骨髓的原代白血病细胞集落(leukemic blast progenitor,L-CFU)生成的影响,结果表明,当BLS-1的浓度达到500μg/ml时,L-CFU的生成受到明显抑制。上述研究结果提示,在新生牛肝中可能存在类似于人胎儿肝脏中的一类低分子天然抑瘤物。     

新生牛肝中三种低分子化合物牛磺酸、鸟氨酸、肌肽对HL-60细胞凋亡的诱导作用

目的:研究三种新生牛肝源低分子化合物:牛磺酸、鸟氨酸、肌肽对HL-60白血病细胞增殖的抑制作用并探讨其调控机理.方法:用MTT法分别检测三种活性成分作用后HL-60细胞和正常人淋巴细胞的存活率.分别用琼脂糖凝胶电泳,顺磁共振ESR技术,免疫组化法测定其对HL-60细胞的核DNA、氧自由基活性和细胞周期蛋白水平的影响.结果:三种化合物能够有效地抑制HL-60细胞的增殖,而对正常的人淋巴细胞的生长没有抑制作用;三种化合物使HL-60细胞的核DNA产生30 kb片段,使HL-60细胞内的氧自由基活性降至痕量水平,并下调HL-60的p45/skp2的水平,而上调p27/kip的水平.结论:牛磺酸、鸟氨酸、肌肽能够通过调控细胞周期蛋白的水平而选择性的抑制肿瘤细胞的增殖.     

NADPH氧化酶源性活性氧介导epinodosin诱导HL-60细胞分化及细胞骨架重组

该文分析研究了对映-贝壳杉烷二萜epinodosin通过调节NADPH氧化酶源活性氧诱导急性早幼粒细胞白血病细胞HL-60向成熟粒细胞分化的特征。结果显示,epinodosin在4.0~8.0μmol/L浓度时抑制HL-60细胞生长,导致其S期阻滞,诱导HL-60细胞形成肾形核和多叶核细胞,使细胞吞噬能力和对硝基四氮唑蓝(nitrotetrazolium blue chloride,NBT)的还原力显著增强、细胞表面抗原CD11b表达上调,表明epinodosin可诱导HL-60细胞向成熟粒细胞分化;epinodosin还诱导HL-60细胞骨架重组,显著促进微管的成束组装以及提高波形纤维含量,上述分化特征与临床诱导分化治疗剂全反式维甲酸(all-trans retinoic acid,ATRA)对HL-60细胞的分化诱导特性极为相似。进一步检测显示,抗氧化剂NAC(N-acetyl-L-cysteine)以及NADPH氧化酶抑制剂夹竹桃麻素(apocynin,APO)和联苯基三价碘(diphenyleneiodonium,DPI)均可抑制epinodosin对HL-60细胞的分化诱导作用,表明epinodosin导致HL-60细胞活性氧浓度的升高与NADPH氧化酶活性关联,提示epinodosin通过调节NADPH氧化酶源性活性氧诱导HL-60向粒细胞分化。     

白藜芦醇对肿瘤细胞HL-60代谢热化学研究

目的：用微量热法研究了白藜芦醇对白血病细胞HL-60代谢作用的热化学特征。方法：等温量热仪测定HL-60细胞在白藜芦醇作用下生长代谢的热功率输出曲线。结果：白藜芦醇在20μg/mL-80μg/mL范围内,热曲线一直处于缓慢下降状态,对该肿瘤细胞有明显抑制作用,当计量达到100μg／mL-120μg／mL时,热功率曲线值呈缓慢增加趋势,抑制效果变弱。结论：白藜芦醇对细胞HL-60代谢有明显的抑制作用,为白藜芦醇在防治肿瘤中的应用提供了可靠的信息。     

人胎肝中低分子抑瘤物的分离纯化、结构鉴定及其对肿瘤细胞生长的选择性抑制作用

在胎儿组织中存在一类低分子肿瘤抑制物。胎肝细胞的甲醇－丙酮提取物中保留有大部分抑瘤活性。用体外液体培养条件下对人急性粒系白血病细胞系（ＨＬ－６０）的抑制作用为指标,跟踪指导分离过程。提取物经反相Ｃ１８中压液相色谱、ＳｅｐｈａｄｅｘＬＨ－２０凝胶色谱及氨基键合相高效液相色谱分离得到一纯活性物质,经ＮＭＲ和ＭＳ鉴定为７－酮基胆固醇（７－ｋｅｔｏｃｂｌｅｓｔｅｒｏｌ,７－ＫＣ）。体外琼脂培养条件下７－ＫＣ对小鼠ＷＥＨ１－３和Ｓ－１８０细胞较对正常小鼠骨髓粒一巨噬系祖细胞有更强的抑制增殖及集落生成作用。７－ＫＣ对ＨＬ－６０细胞增殖较对正常人骨髓ＣＦＵ－ＧＭ有更强的抑制作用。     

人胎肝细胞分泌的低分子抑瘤物对白血病细胞的抑制作用

本工作证明了在胎儿组织中存在一类低分子天然肿瘤抑制物,它是胎儿组织细胞生成和分泌的,对瘤株细胞和原代白血病细胞有选择性抑制作用。初发期或复发期急性非淋巴细胞白血病患者的骨髓在体外液体培养条件下与肝细胞上清及其甲醇提取物共同孵育4d,可使所有病例的骨髓AML—CFU降低到不可检出的程度。因而,低分子天然抑瘤物是天然肿瘤免疫中的一个组成部分,它在肿瘤的诊断和治疗中有着深远的意义。     

Recombinant human interferon sensitizes resistant myeloid leukemic cells to induction of terminal differentiation

Recombinant human leukocyte interferon (IFN-alpha A) inhibits growth of the human promyelocytic leukemic cell line HL-60 without inducing these cells to differentiate terminally. When IFN-alpha A is combined with agents capable of inducing differentiation in HL-60 cells, such as 12-O-tetradecanoyl-phorbol-13-acetate (TPA), cis or trans retinoic acid (RA) or dimethylsulfoxide (DMSO), growth suppression and induction of differentiation are dramatically increased. By growing HL-60 cells in increasing concentrations of TPA, RA, or DMSO, a series of sublines have been developed which are resistant to the usual growth inhibition and induction of differentiation seen when wild type HL-60 cells are exposed to these agents. Treatment of these resistant HL-60 cells with the combination of IFN-alpha A and the appropriate inducer results, however, in a synergistic suppression in cell growth and a concomitant induction of terminal differentiation. The ability of interferon to interact synergistically with agents which promote leukemic cell maturation may represents a novel means of reducing resistant leukemic cell populations.     

Antileukemic effect of coral-prostanoids clavulones from the stolonifer Clavularia viridis on human myeloid leukemia (HL-60) cells

To elucidate the biological activities of coral-prostanoids, clavulones, discovered from the Japanese stolonifer Clavularia viridis, we examined the effect of clavulone on the cell growth of human cancer (human promyelocytic leukemia (HL-60) cells and HeLa cells) and normal (Chang liver cells and lung fibroblasts) cells in vitro. Clavulone showed strong antiproliferative and cytotoxic activities in the human cells and it had some selectivity to leukemic (HL-60) cells over other HeLa cells or normal cells on the basis of the IC50 values and cytotoxic effect of the cells. The IC50 value of clavulone in the HL-60 cells was about 0.4 microM (0.2 micrograms/ml). Over 1.0 microM (0.5 micrograms/ml), clavulone showed a significant cytotoxic activity on the HL-60 cells. The data on DNA synthesis and flow cytometric analysis revealed that clavulone arrests the cells in the G1-phase and inhibits the cell growth of HL-60 cells by inhibiting S-phase DNA synthesis. These results suggest that clavulone has a potent antileukemic effect on HL-60 cells.     

Human thymic epithelial cells produce granulocyte and macrophage colony-stimulating factors

The development of culture conditions for growing normal human thymic epithelial (TE) cells free from contamination with other stromal cells has allowed us to identify and characterize TE cell-derived cytokines. In this study, we report that cultured human TE cells produced CSF that supported the growth of clonal hematopoietic progenitor cells in the light density fraction of human bone marrow cells. Thymic epithelial supernatants (TES) induced growth of granulocyte/macrophage colonies (CFU-GM), mixed granulocyte/erythrocyte/monocyte/megakaryocyte colonies (CFU-GEMM), and early burst-forming unit erythroid colonies (BFU-E). In addition, TES induced differentiation of the promyelocyte leukemic cell line HL-60 and stimulated growth of both granulocyte (CFU-G) and monocyte (CFU-M) colonies from murine bone marrow cells. Using anion exchange column chromatography, pluripotent CSF activities in TES were separated and shown to be distinct from an IL-1-like cytokine that has been shown as a TE cell-derived cytokine (TE-IL-1). Colony-stimulating activity supporting the growth of bone marrow CFU-GEMM, BFU-E, and CFU-GM co-eluted at 150 to 180 mM NaCl. A separate peak of CFU-GM-stimulating activity eluted early in the gradient at 20 mM NaCl. In Northern blot analysis of enriched RNA, synthetic oligonucleotide probes complementary to human G-CSF and M-CSF coding sequence each hybridized with a single RNA species of 1.7 and 4.4 kb, respectively. These data suggest that normal human TE cells synthesize G-CSF and M-CSF that promote differentiation of non-lymphoid hematopoietic cell precursors.     

甲胎蛋白在体外对人肝癌细胞生长的影响

本文用MTT比色法观察了甲胎蛋白(AFP)在体外对人肝癌细胞生长的影响。结果表明,AFP能促进SMMC-7721人肝癌细胞的生长。当AFP与AFP抗体合用时,AFP抗体能减弱AFP对SMMC-7721细胞生长的促进作用;AFP抗体单用对此种细胞的生长亦有抑制作用。另一方面,在相同的实验条件下,AFP和AFP抗体对HL-60人白血病细胞的生长无明显影响;提示AFP的促生长作用具有一定的肿瘤细胞特异性,并非一种蛋白质对培养细胞的非特异性营养作用。此外,AFP亦能促进MCF-7人乳腺癌细胞的生长,AFP抗体对此种细胞的生长有抑制作用。由于MCF-7细胞存在功能性AFP受体,也能合成和分泌AFP。这就提示,人肝癌细胞中的AFP很可能与其受体特异性结合,产生促生长效应。确切机制尚待进一步阐明。     

A protein complex expressed during terminal differentiation of monomyelocytic cells is an inhibitor of cell growth

A protein complex (PC) composed of the MRP8 and MRP14 proteins has previously been shown to be a specific inhibitor of casein kinase I and II. This PC is expressed during the late stages of terminal differentiation induced in human promyelocytic HL-60 leukemia cells by 1 alpha,25-dihydroxyvitamin D3 and in human monocytic THP-1 leukemia cells by phorbol 12-myristate 13-acetate. This expression is associated with terminal cell differentiation because incubation of HL-60 cells with an agent or condition that causes suppression of growth but not induction of differentiation does not result in expression of the PC. At concentrations of 5-15 nM, the purified PC inhibited the growth of HL-60 cells and THP-1 cells, as well as other cell types belonging to different cell lineages. This growth inhibition was preceded by a reduction in [32P]phosphate incorporation and, at the higher PC concentrations, was associated with a reduction in [3H]thymidine, [3H]uridine, and [32S]methionine incorporation. The specific expression pattern and growth-inhibitory character of the PC suggests that the complex may have a role in suppressing cell growth during monomyelocytic terminal differentiation induced by specific chemical stimuli and during physiological and pathological events associated with monomyelocytic cell functions.