β-榄香烯对小鼠黑色素瘤细胞的分化诱导作用

目的 探讨β-榄香烯对黑色素瘤B16细胞的分化诱导作用。方法 以小鼠黑色素瘤B16细胞株为模型,研究β-榄香烯对B16细胞的生长抑制作用和对B16细胞株为模型,研究β-榄香烯对B16细胞的生长抑制作用和对B16细胞黑色素生成能力、酪氨酸酶活力的影响,观察B16细胞的形态改变。结果(1)β-榄香稀显著抑制B16细胞的生长,且呈浓度依赖性;(2)使B16细胞体积增大,变长,多数细胞具有树突样结构;(3)β-榄香烯(10μg/ml,20μg/ml和40μg/ml)组黑色素的生成能力分别为(0.20±0.03),(0.22±0.01)和(0.59±0.09),显著高于空白对照组(0.08±0.01),而维甲酸(10~(-6)mol/ml)组的黑色素的生成能力(0.46±0.08),与β-榄香烯40μg/ml组的生成能力相当;(4)β-榄香烯(20μg/ml、40μg/ml)组的酪氨酶活力分别为(20493±1949)和(21154±568),显著高于空白对照组(11432±3026),与维甲酸(10~(-6)mol/ml)组的酪氨酶活力(23512±2687)相近。结论 β-榄香烯显著抑制小鼠黑色素瘤B16细胞的生长,并从形态上和功能上诱导B16细胞分化。     

盐酸小檗碱对B16细胞的分化诱导作用

目的 测定盐酸小檗碱对小鼠黑色素瘤B16细胞的诱导分化作用。方法 以MTT法测定盐酸小檗碱对B16细胞增殖的抑制作用,通过克隆形成实验、细胞黑色素含量的测定以及B16细胞体内成瘤能力的测定观察盐酸小檗碱对B16细胞的诱导分化作用。结果 盐酸小檗碱对B16细胞具有明显的增殖抑制作用;并使B16细胞生长缓慢,平铺不重叠,呈正常上皮样细胞分化,细胞的克隆形成能力降低,细胞内的黑色素生成能力增强,C57/BL小鼠体内成瘤能力降低。结论 盐酸小檗碱对B16细胞具有诱导分化作用。     

盐酸小檗碱对B16细胞的分化诱导作用

目的测定盐酸小檗碱对小鼠黑色素瘤B16细胞的诱导分化作用。方法以MTT法测定盐酸小檗碱对B16细胞增殖的抑制作用,通过克隆形成实验、细胞黑色素含量的测定以及B16细胞体内成瘤能力的测定观察盐酸小檗碱对B16细胞的诱导分化作用。结果盐酸小檗碱对B16细胞具有明显的增殖抑制作用;并使B16细胞生长缓慢,平铺不重叠,呈正常上皮样细胞分化,细胞的克隆形成能力降低,细胞内的黑色素生成能力增强,C57/BL小鼠体内成瘤能力降低。结论盐酸小檗碱对B16细胞具有诱导分化作用。     

rhIL-24对黑色素瘤B16细胞的体内外抑瘤作用

目的 研究重组人白细胞介素24（rhIL-24）及真核表达基因重组质粒pcDNA3．0-hIL-24对黑色素瘤B16细胞（B16细胞）及其荷瘤小鼠体内外抑肿瘤作用。方法 用rhIL-24蛋白作用于小鼠B16细胞,检测其对B16细胞的生长抑制作用及诱导凋亡作用,并用rhIL-24蛋白及pcDNA3．0-hIL-24治疗荷瘤小鼠。结果 rhIL-24蛋白对体外培养的B16细胞具有显著的抑制生长和诱导凋亡作用,rhIL-24蛋白及pcDNA3．0-hIL-24对C57／BL6小鼠体内形成的恶性黑色素瘤具有明显的抑瘤作用。结论 rhIL-24蛋白具有生物活性,他及真核表达重组质粒对B16细胞及其小鼠种植瘤生长有抑制作用,并能诱导肿瘤细胞凋亡及促进其分化。     

大豆甙元对小鼠B16黑色素瘤细胞的分化诱导作用

大豆甙元在10～40μg·ml~(-1)浓度范围内,能够明显抑制B16细胞的增殖,受药物作用4d的B16细胞,克隆形成能力及体内成瘤能力明显降低,大豆甙元在抑制B16细胞增殖的同时,促进黑色素的生成,且对B16细胞形态具有明显的影响,低浓度时促使细胞平行排列,当浓度增加时,形成网状结构,黑色素颗粒明显增多。     

甘草酸二铵对马兜铃酸致肾小管上皮细胞损害保护作用的实验研究

目的　探讨甘草酸对马兜铃酸 (aristolochic acid,AA)肾损害的保护作用 ,观察甘草酸是否可以减轻 AA对培养的肾小管上皮细胞的损害 ,并初步探讨其可能机制。方法　 5 % FCS RPMI- 16 4 0培养肾小管上皮细胞株 (L L C- PK1) ,采用结晶紫染色法观察细胞增殖 ;荧光染色观察活细胞数目 ;酶动力学检查 L DH活性 ,比色法检测 NAG水平 ;透射电镜观察细胞超微细胞结构。结果　1AA4 0、80、16 0μg/ ml可明显抑制细胞增殖 ,结晶紫 OD值显著减少 ,与无 AA对照组比较 P<0 .0 1。 2甘草酸二铵 (diammoniumglycyrrhizinate,DG)在 5、10、2 5 μg/ ml时结晶紫 OD值与无甘草酸对照组比较 P>0 .0 5 ,甘草酸在 5 0 μg/ ml时 P<0 .0 5 ,10 0 μg/ ml、2 0 0μg/ ml时 P<0 .0 1,提示大剂量甘草酸可改善 AA抑制细胞增殖的作用。 3在 AA4 0μg/ ml作用下随着时间延长 ,无甘草酸组的细胞存活数目逐渐减少 ,而甘草酸 (10 0μg/ ml、2 0 0μg/ ml)作用组 ,活细胞数目明显增多 ,提示高浓度甘草酸对细胞损伤有一定的保护作用。4甘草酸在 5、10、2 5 μg/ ml时 ,L DH释放率、NAG酶水平与无甘草酸对照组比较 P>0 .0 5 ,甘草酸在 5 0 μg/ ml时 P<0 .0 5 ,甘草酸在 10 0μg/ ml、2 0 0μg/ ml时 L DH释放率、NAG酶明显减少 ,与无甘草     

楤白皮总皂苷体内外的抗肿瘤作用

目的:研究楤白皮总皂苷的体内外抗肿瘤活性。方法:采用MTT体外测定楤白皮总皂苷对小鼠B16黑色素瘤、小鼠H22肝癌细胞和小鼠FBL3红白血病细胞的增殖抑制作用;采用小鼠移植性肿瘤S180和H22观察楤白皮总皂苷的体内抑瘤活性。结果:楤白皮总皂苷体外对B16细胞、FBL3细胞的IC50分别为0.077 4 g.L-1和0.199 g.L-1;在0.02～3.2 g.L-1范围内,对H22细胞体外抑制率均为80%以上。在100、200、300 mg.kg-1剂量下,楤白皮总皂苷对S180小鼠肉瘤的抑瘤分别为16.9%、37.1%和27.0%,对H22小鼠肝癌的抑瘤率分别为23.7%、10%和13.4%。结论:楤白皮总皂苷在体内、外均具有一定的抗肿瘤活性。     

Lys-2-rhTNFα单用或联合其它细胞因子抗肿瘤作用的研究

研究了Lys-2-rhTNFα（1）单用或联合^125Ser-rhIL-2(2)和／或rhINFα-2a(3)的体内外抗肿瘤作用。实验结果表明1对人结肠癌细胞株（M7609）、小鼠黑色素瘤细胞株（B16）、小鼠白现细胞株（Yac-1）均有抑制细胞增殖的作用。1合用2及3较它们各自单用及任意两者合用的细胞毒作用明显增强（P＜0．01）。1与2、3联合用药组较它们各自单独用药对小鼠结肠癌Colon26细胞移植性肿瘤的抑瘤作用明显提高（P＜0．05）。     

氯化钴低氧对黑色素瘤细胞系迁移能力及FSTL1表达分泌的影响

目的 探讨氯化钴（CoCl2）化学模拟低氧对黑色素瘤细胞系B16F10迁移的影响,以及Follistatin-like 1（FSTL1）蛋白在此过程中的转录、表达和分泌情况。方法 CoCl2模拟低氧作用于小鼠B16F10细胞,实验分为3组：0 μmol/L CoCl2对照组、50 μmol/L和100 μmol/L CoCl2处理组。用MTT法测定细胞活力;用Transwell法测定细胞迁移能力;qRT-PCR检测Fstl1 mRNA表达;Western blot检测细胞内外FSTL1蛋白表达。结果 CoCl2模拟低氧可以导致B16F10细胞存活率显著下降,并呈浓度和时间依赖性;50 μmol/L CoCl2处理组（0.158±0.006）、100 μmol/LCoCl2处理组（0.203±0.002）B16F10细胞迁移能力均明显高于对照组（0.107±0.001,均P<0.05）;50 μmol/L CoCl2处理组（1.573±0.114）、100 μmol /L CoCl2处理组（2.219±0.085）Fstl1 mRNA表达均显著高于对照组（0.962±0.054,均P<0.05）,而胞内FSTL1蛋白表达与Fstl1 mRNA表达趋势一致。同时也发现,CoCl2处理组细胞外FSTL1蛋白表达均低于对照组,且100 μmol/L CoCl2处理组几乎检测不到FSTL1表达。结论 CoCl2模拟低氧促进黑色素瘤细胞迁移,可能与FSTL1的表达和分泌有关,但其功能和作用机制还需进一步探讨。     

二氧化氯作为消毒剂在医院的应用

本实验证明 ,二氧化氯用于医院内消毒使用浓度及作用时间为 :一般容器用 10 0μg/ ml溶液作用 5min或 50 μg/ ml溶液作用 2 0 min;物品表面擦拭用 2 0 0 μg/ ml溶液作用 10 min;普通病房空气喷雾消毒用 2 0 0 μg/ ml溶液 8ml/ m3,作用 30 min;医疗器械浸泡用 10 0μg/ ml溶液作用 10 min以上或 2 0 0μg/ ml作用 5min以上。     

Apoptotic activity of betulinic acid derivatives on murine melanoma B16 cell line

The mitochondrion plays a crucial role in the process of apoptosis and has thus become one of the targets for the search for potential chemotherapeutic agents. Betulinic acid [3beta-hydroxy-lup-20(19)lupaen-28-carbonic acid], a lupane-type triterpene which is abundant in many plant species, has been shown to exert a direct effect on the mitochondria and subsequent apoptosis in melanoma cells. Chemical synthesis and modification of betulinic acid are being explored to develop more potent derivatives. We present here the apoptotic activity of several natural derivatives of betulinic acid which were isolated from the roots of a Chinese medicinal herb, Pulsatilla chinensis (Bge) Regel [Ye, W., Ji, N.N., Zhao, S.X., Liu, J.H., Ye, T., McKervey, M.A., Stevenson, P., 1996. Triterpenoids from Pulsatilla chinensis. Phytochemistry 42, 799-802]. Of the five compounds tested, 3-oxo-23-hydroxybetulinic acid was the most cytotoxic on murine melanoma B16 cells (IC50=22.5 microg/ml), followed by 23-hydroxybetulinic acid and betulinic acid (IC50=32 and 76 microg/ml, respectively), with lupeol and betulin exhibiting the weakest cytotoxicity (IC50> or =100 microg/ml). Exposure of B16 cells to betulinic acid, 23-hydroxybetulinic acid and 3-oxo-23-hydroxybetulinic acid caused a rapid increase in reactive oxidative species production and a concomitant dissipation of mitochondrial membrane potential in a dose- and time-dependent manner, which resulted in cell apoptosis, as demonstrated by fluorescence microscopy, gel electrophoresis and flow-cytometric analysis. Cell cycle analysis further demonstrated that both 3-oxo-23-hydroxybetulinic acid and 23-hydroxybetulinic acid dramatically increased DNA fragmentation at the expense of G1 cells at doses as low as 12.5 and 25 microg/ml, respectively, thereby showing their potent apoptotic properties. Our results showed that hydroxylation at the C3 position of betulinic acid is likely to enhance the apoptotic activity of betulinic acid derivatives (23-hydroxybetulinic acid and 3-oxo-23-hydroxybetulinic acid) on murine melanoma B16 cells.     

桦木酸的抗肿瘤作用及其诱导KB细胞凋亡的研究

目的研究桦木酸(betulinic acid,BetA)体内对S180肉瘤的抑制作用和体外对人口腔上皮癌(KB)细胞系凋亡的诱导活性。方法建立小鼠体内荷S180肉瘤模型,测定BetA的体内抑瘤率;采用MTT分析、细胞形态学观察、原位末端标记和流式细胞仪检测等方法检测BetA对KB细胞生长状态和细胞周期的影响以及诱导细胞凋亡的作用。结果BetA 800和1 200 mg.kg-1给荷瘤小鼠灌胃,对S180肉瘤有显著的抑制作用;体外对KB细胞生长呈剂量依赖性抑制作用(其IC50为11.60μmol.L-1),并出现大量凋亡细胞。结论BetA体内对S180肉瘤、体外对KB细胞增殖均有抑制作用,诱导肿瘤细胞死亡的主要途径可能是凋亡。     

SBF-1, a synthetic steroidal glycoside, inhibits melanoma growth and metastasis through blocking interaction between PDK1 and AKT3

In the present study, we demonstrate that SBF-1, a synthetic steroidal glycoside, has a strong antitumor activity against melanoma cells in vitro and in vivo. SBF-1 induced cell cycle arrest with a reduced expression of various cell cycle related proteins in B16BL6 melanoma cells without causing apoptosis. SBF-1 dramatically inhibited kinase activity of 3-phosphoinositide dependent protein kinase 1 (PDK1) and thus down-regulated phosphorylation of protein kinase B (AKT). Among three known isoforms of AKT, PDK1 only interacted with AKT3 in B16BL6 melanoma cells, and SBF-1 almost completely blocked this interaction. In addition, adhesion to fibronectin and expression of integrin α4 were significantly reduced in a concentration-dependent manner. Knockdown of AKT3 resulted in the decrease in integrin α4 expression and cell adhesion. Moreover, SBF-1 inhibited the growth of melanoma xenografts and down-regulated the phosphorylation of AKT in vivo. In a mouse model of spontaneous metastasis, SBF-1 at very low doses of 1 and 3 μg/kg enormously inhibited melanoma metastasis into draining popliteal lymph nodes. Taken together, this study shows a small molecular compound SBF-1 with a very strong anti-melanoma activity both in vitro and in vivo. Its mechanism underlying such antitumor effect is related to the blockage of the interaction between PDK1 and AKT3.     

三氧化二砷抑制小鼠B16黑色素瘤生长作用及其机制

目的　探讨三氧化二砷 (As2 O3 )对小鼠B16黑色素瘤的生长及其血管生成的抑制作用 ;同时观察其对B16细胞增殖活性、细胞形态、细胞周期及凋亡的影响。方法　选用小鼠黑色素瘤B16细胞接种C5 7BL/ 6J小鼠 ,观察腹腔注射As2 O3 对实体瘤的重量及成瘤率的影响 ;应用HE染色、Ⅷ RAg免疫组化染色观测瘤组织内新生血管密度 ;采用CellTiter 96AqueousOne试剂检测B16细胞增殖活力 ;Giem sa染色、Feulgen染色观察细胞形态学变化 ;流式细胞术分析细胞周期及细胞凋亡。结果　As2 O3 能显著抑制小鼠B16黑色素瘤的生长 ,治疗组成瘤率为 37 5 % ,抑瘤率达81 6 1% ,并能显著抑制瘤组织内血管生成 ;体外实验观察到As2 O3 能抑制B16细胞增殖 ,并存在浓度依赖效应 ,IC50 为32 99μmol·L-1;细胞形态学观察结果显示As2 O3 使B16细收稿日期 :2 0 0 4-0 2 -17,修回日期 :2 0 0 4-0 3 -2 8基金项目 :安徽省教育厅资助项目 ,No 2 0 0 4kJ2 79作者简介 :夏　俊 ( 1965 -) ,女 ,硕士 ,副教授 ,硕士生导师 ,研究方向 :肿瘤分子生物学 ,Tel:0 5 5 2 3 0 664 12 2 0 97,E mail:xia jun1965 @yahoo .com .cn崔秀云 ( 1941-) ,女 ,教授 ,博士生导师 ,研究方向 :癌基因与抑癌基因 ,Tel:0 411 472 0 64 8,E mail:cuixy @dlmedu .edu .     

YoshixolTR inhibits B16 melanoma cell growth in vivo and induces apoptosis-like (quantum thermodynamic) cell death.

In this report, antitumor effects of YoshixolTR in vivo and in vitro were investigated in B16 melanoma cells. For in vivo experiments, the present study shows a dramatic inhibition of tumor growth of B16 melanoma transplanted on the leg or intraperitoneal cavity after treatment with YoshixolTR intraperitoneally. A proliferation of B16 cells in vitro was inhibited by YoshixolTR in a dose-and time-dependent manner. YoshixolTR induced apoptosis-like cell death in histological observations (phase-contrast, scanning and transmission electron microscopy), DNA fragmentation, and a smaller increase in lactate dehydrogenase (LDH) as a marker of cell leakage. Immunohistochemical investigation of cytoskeletal components, such as actin and tubulin, showed a cell wall disruption of B16 melanoma cells and a nuclear extrusion after the treatment with YoshixolTR. Treatment with YoshixolTR in vitro showed an arrest at the G0/G1 stage of the cell cycle, followed by a flow cytometric measurement. As a possible physiological mechanism of YoshixolTR on B16 melanoma cells, intracellular Ca++ was measured with Fura-2 technique. An adequate concentration of YoshixolTR, which induces apoptosis-like cell death, showed a decrease in intracellular free Ca++ concentration. In conclusion, YoshixolTR has an antitumor potency with a new biological mechanism of cell growth, proliferation, and differentiation, including cellular signalling pathways, and is a new candidate for an ideal chemotherapeutic agent against malignant tumors.     

南瓜蛋白对B16细胞的诱导分化作用

目的测定南瓜蛋白对小鼠黑色素瘤B16细胞的诱导分化作用。方法以MTT法测定南瓜蛋白对B16细胞的增殖抑制作用,并以形态学、黑色素含量及细胞周期变化为指标,观察南瓜蛋白诱导B16细胞分化的作用。结果南瓜蛋白对B16细胞具有明显的增殖抑制作用,并阻止B16细胞由G1期向S期过渡从而停滞于G1期;细胞形态向正常上皮样细胞分化,生长缓慢,平铺不重叠,甚至形成网状结构,胞质内细胞器丰富,黑色素小体明显增多;黑色素含量增加。结论南瓜蛋白对B16细胞具有明显的诱导分化作用。     

4味中药乙醇提取物祛黄褐斑作用的比较研究

目的比较白附子、银花、白芨、六月雪4味中药乙醇提取物祛黄褐斑作用的差异,选出作用最强的药材。方法以MTT法测定小鼠B16黑素瘤细胞增殖抑制率、酪氨酸酶活性、黑素含量变化;采用多重比较法进行统计分析。结果银花的浓度为62.5μg.mL-1,白芨、白附子各浓度(500、250、125、62.5μg.mL-1)均对B16鼠黑素瘤细胞增殖抑制作用较强。白附子的浓度为250μg.mL-1或银花的浓度为62.5μg.mL-1对小鼠B16黑素瘤细胞酪氨酸酶活性抑制作用最佳。白附子的浓度为125μg.mL-1对黑素合成抑制作用最好。结论白芨与白附子可作为祛黄褐斑有效成分的候选药材。     

Piper longum inhibits VEGF and proinflammatory cytokines and tumor-induced angiogenesis in C57BL/6 mice

The antiangiogenic activity of Piper longum was studied using in vivo as well as in vitro models. In vivo, antiangiogenic activity was studied using B16F-10 melanoma cell-induced capillary formation in C57BL/6 mice. Intraperitoneal administration of the extract (10 mg/dose/animal) significantly inhibited (50.6%) the number of tumor-directed capillaries induced by injecting B16F-10 melanoma cells on the ventral side of C57BL/6 mice. The cytokine profile in the serum of these animals showed a drastically increased level of proinflammatory cytokines such as IL-1beta, IL-6, TNF-alpha, GM-CSF and the direct endothelial cell proliferating agent, VEGF. Administration of the methanolic extract of P. longum could differentially regulate the level of these cytokines. The level of IL-2 and tissue inhibitor of metalloprotease-1 (TIMP-1) was increased significantly when the angiogenesis-induced animals were treated with the extract. The extract of P. longum at non-toxic concentrations (10 microg/ml, 5 microg/ml, 1 microg/ml) inhibited the VEGF-induced vessel sprouting in rat aortic ring assay. Moreover, P. longum was able to inhibit the VEGF-induced proliferation, cell migration and capillary-like tube formation of primary cultured human endothelial cells. Hence, the observed antiangiogenic activity of the plant P. longum is related to the regulation of these cytokines and growth factors in angiogenesis-induced animals.