BACKGROUND: Hematogenesis of a body mainly depends on the proliferation of hematopoietic progenitor cell (HPC). Hematopoietic functional impairment will occur when hematopoietic cells are injured by radioactive ray or chemical drug. The proliferation of HPC is the key link of promoting hematogenesis.OBJECTIVE: To study the effects of nine monomers extracted from Spatholobus suberectus Dunn (SSD) on proliferation of HPC in marrow-depressed mice.rnDESIGN: Randomized controlled trial.SETTING: Department of Pharmacy, General Hospital of Chinese PLA.MATERIALS: This experiment was conducted in the Department of Clinical Pharmacology, General Hospital of Chinese PLA from November 2002 to February 2003. Totally 348 healthy Kunming mice, weighing 22-25g, clean grade, of irrespective gender, were selected in this study (certification: SCXK-2001-001). The animal experiment was approved by the local ethics committee. SSD was provided by Dispensary of Traditional Chinese Medicine, General Hospital of Chinese PLA; monomers (gallocatechin, formononetin, catechin, pyromucic acid, syringic acid, Demethylvestitol, 1,3,5-benzenetriol, ononin, and epicatechin) were extracted from SSD acetoacetate; TGL-16 centrifuger was made in Shanghai 6th Medical Equipment Factory; CO2 incubator was made in SANYO Company, Japan; MK inverted microscope was provided by OLYMPUS Company, Japan.METHODS: Experimental grouping: Mice were randomly divided into 29 groups, including normal group; control group; gallocatechin high-, medium-, low-dose groups; formononetin high-, medium-, low-dose groups; catechin high-, medium-, low-dose groups; pyromucic acid high-, medium-, low-dose groups; syringic acid high-, medium-, low-dose groups; Demethylvestitol high-, medium-, low-dose groups; 1,3,5-benzenetriol high-, medium-, low-dose groups; ononin high-, medium-, low-dose groups; epicatechin high-, medium-, low-dose groups with 12 mice in each group. Experimental intervention: All the mice except the mice in normal group had been given total body sublethal dose of irradiation by 60Co γ-ray (215.3 rontgen/min, 4 Gy dose rate, irradiation time of 107.5 seconds). Normal saline was injected intraperitoneally into 8 mice in normal group and control group at the third day after inadiation. Stored solution 2,0.4,0.08g/L of each monomer was intraperitoneally injected into the mice in each monomer high-, medium-, low-dose groups, respectively, at the third day after irradiation. Experimental evaluation: Thirty minutes after administration, blood of 8 mice in normal, control group and 12 mice in other groups was collected and normal, control and each dose monomer-containing serums were obtained after centrifugation for 15 minutes, filtering through 0.45μm filter membrane. Then 4 mice in normal and control group were killed to study the effects of nine monomers on proliferation of HPC in marrow-depressed mice by counting erythrocyte colony-forming unit (CFU-E), burst-forming uniterythroid (BFU-E), granulocyte-macrophage colony-forming unit (CFU-GM), and megakaryocyte colony-forming unit (CFU-Meg).MAIN OUTCOME MEASURES: CFU-GM, CFU-E, BFU-E, and CFU-Meg in each group.rnRESULTS: Totally 348 mice were included in the final analysis. CFU-E: The quantity of CFU-E in high-dose of catechin, gallocatechin, syningic acid, and epicatechin groups was significantly higher than that in the control group (P<0.05-0.01) while the quantity of CFU-E in medium-and low-dose of catechin and medium-dose of gallocatechin was also significantly higher than that in the control group (P<0.05). CFU-GM: Except pyromucic acid and ononin groups, the amount of CFU-GM in other groups was significantly higher than that in the control group (P<0.01). BFU-E: Compared with control group, the amount of BFU-E remarkably increased under the effect of each dose of catechin, gallocatechin, syringic acid and high-, medium-dose of epicatechin (P<0.05). CFU-Meg: The amount of CFU-Meg in high-, low-dose syringic acid groups, low-dose gallocatechin groups and each dose group of catechin and epicatechin was significantly higher than that in the control group (P<0.05). Amount of all colonies in the control group was significantly lower than that in the normal group (P<0.01).rnCONCLUSION: Nine monomers extracted from SSD can promote the proliferation of HPC in bone marrow depressed mice. In particular, the activity of catechin to stimulate proliferation is the strongest.%背景:机体造血主要依靠造血祖细胞来扩增,当受到放射线、化学药物等损伤时,即会发生造血功能障碍.目的:观察从传统中药鸡血藤中提取的九个单体化合物对骨髓抑制小鼠造血祖细胞增殖的影响.设计: 随机对照实验.单位:中国人民解放军总医院临床药理研究室.材料:实验于2002-11/2003-02在中国人民解放军总医院药理研究室完成,选用348只健康昆明种小鼠,体质量22.25g,清洁级,雌雄各半,由解放军军事医学科学院实验动物中心提供,许可证号码为SCXK-2001-001.实验过程中对动物的处置符合动物伦理学标准.鸡血藤由中国人民解放军总医院中药房提供,单体化合物没食子儿茶素、芒柄花素、儿茶素、焦性粘液酸、丁香酸、Demethylvestitol、间苯三酚、芒柄花甙、表儿茶素由中国医学科学院药物研究所从鸡血藤乙酸乙酯部位中分离.TGL-16型离心机为上海医疗器械六厂生产,CO2培养箱为日本SANYO公司产品,MK型倒置显微镜为日本OLYMPUS公司产品.方法:[1]实验分组:随机抽签法将小鼠分为29组:鸡血藤九种单体化合物没食子儿茶素、芒柄花素、儿茶素、焦性粘液酸、丁香酸、Demethylvestitol、间苯三酚、芒柄花甙、表儿茶素高剂量组、中剂量组及低剂量组共27组,同时设立正常组及对照组,每组12只.[2]实验干预:除正常组外,小鼠均给予全身60Co γ射线照射,剂量为4Gy.照射后第3天,9种单体化合物高、中、低剂量组小鼠分别腹腔注射2,0.4,0.08g/L相应单体化合物药液0.5mL,正常组和对照组小鼠腹腔注射等体积的生理盐水.[3]实验评估:给药30min后分別取正常组及对照组各8只及其他组全部小鼠静脉血制备正常、对照及各单体含药血清,取对照组剩余4只小鼠的骨髓细胞进行造血祖细胞培养,培养分为对照组和九个单体化合物高、中、低剂量组,分别在各组培养体系中加入对照及各单体含药血清,另取正常组小鼠4只进行骨髓造血祖细胞培养作为细胞培养的正常组,通过计数各组CFU-E、BFU-E、CFU-GM、CFU-Meg集落数来观察九个单体化合物对骨髓抑制小鼠造血祖细胞增殖的影响.主要观察指标:各组CFU-E、BFU-E、CFU-GM、CFU-Meg集落数.结果:纳入小鼠348只,均进入结果分析,无脱失值.[1]CFU-E集落数:儿茶素、没食子儿茶素、丁香酸、表儿茶素高剂量组CFU-E显著高于对照组(P<0.05-0.01),儿茶素中、低剂量组及没食子儿茶素中剂量组CFU-E也显著高于对照组(P<0.05).[2]CFU-GM集落数:除焦性粘液酸、芒柄花甙各剂量组外,鸡血藤其余单体化合物各剂量组CFU-GM均较对照组明显增加(P<0.01).[3]BFU-E集落数:儿茶素、没食子儿茶素、丁香酸各剂量组及表儿茶素高、中剂量组的BFU-E较对照组明显增加(P<0.05).[4]CFU-Meg集落数:儿茶素和表儿茶素各剂量组、丁香酸高、低剂量组及没食子儿茶素低剂量组的CFU-Meg显著高于对照组(P<0.05).对照组所有集落数均明显少于正常组,差异有统计学意义(P<0.01).结论从中药鸡血藤中提取的单体化合物可刺激骨髓抑制小鼠造血祖细胞的增殖,其中儿茶素的刺激增殖活性相对最强.
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