【作者】 刘敏；

【导师】 宋艳秋；

【作者基本信息】 吉林大学 ， 内科血液学， 2006， 硕士

【摘要】 目的:1.建立K562/NOD-SCID白血病小鼠模型;2.通过体内实验证明人参皂甙Rg3能抑制白血病细胞体内增殖。方法:1.首先用k562细胞接种全身照射后的裸鼠,待皮下成瘤后取出局部瘤块,选取无坏死的瘤组织制成瘤细胞悬液,腹腔接种全身照射后的NOD/SCID小鼠。每周剪尾法采血行外周血白细胞计数,同时外周血涂片,染片分类。取小鼠主要脏器及腹腔内局部瘤块行病理检查。2.在建立NOD/SCID小鼠人慢粒急变白血病模型基础上,给予Rg3灌胃9周,分成预防组和治疗组,低量组(10mg/kg)和高量组(20mg/kg),预防组为灌胃2周后再接种白血病细胞,每周采血行外周血白细胞计数及涂片,并计算各组小鼠生存期。结果:1.成功建立全身播散的白血病模型,4周时外周血涂片可见白血病细胞,晚期浸润肝、脾、骨髓等造血器官,白细胞上升到接种前的8-10倍,血涂片分类白血病细胞达20-30%。腹腔局部可有瘤块,多位于腹腔内或大网膜,较少累及其他器官。2.从第4周开始对照组的外周血白细胞数高于预防组和治疗组,第8周时除治疗组低量组外其他各用药组与对照组外周血白细胞数相比有显著差异(P<0.05),但各用药组间无明显差异,从第4周开始对照组的白血病细胞比例基本上高于预防组和治疗组,第8周时对照组外周血白血病细胞比例高于各用药组,但无显著差异(P>0.05),各用药组间无明显差异。第9周时对照组全部死亡,而用药组均有小鼠存活。结论:1.腹腔接种k562瘤细胞于全身照射后的NOD/SCID小鼠能建更多还原

【Abstract】 Leukemia mouse model is an important tool for research on theetiology , pathology and therapy strategies for leukemia. Non-ObeseDiabetic/Severe Combined Immunodeficiency Mice is immunodeficientin multiple aspects, which has a weakening activity of natural killercell and complement,decreasing mature macrophagus besidesdeficiency of both cell-mediated and humoral-mediatedimmunity.Therefore,NOD/SCID is the best animal species to establishin vivo model of human leukemia.There have been many domestic reportson the establishment of disseminated human leukemia in NOD/SCIDafter human leukemia cells cultured in vitro,most of which are acutemyelogenous leukemia cell lines such as HL-60 and K562 are inoculatedinto the recipient mice.Research on ginsenosides Rg3 have manifested that its anti-tumoreffects are brought into full play aiming many targets and throughmultiple mechanisms.It effects on every links of the development oftumor such as inhibiting the proliferation of tumor cells,inducingthe apoptosis of malignant cells,preventing tumors frominfiltration and lymphangitic spread,immuno-modulation, antimut-ation as an chromosome stabilizing agent and changing the expressionof genes.It can also show angio-neogenesis effects to inhibit themetastasis along blood vessels.However,most of research are focusedon solid tumors whereas research on leukemia are rare. There arereports that Rg3 can induce the apoptosis of certain human leukemiacell lines such as K562 in vitro while we are not informed of reportsin vivo so far.The purpose of our experiment is :1.to establish a K562/NOD-SCIDleukemia mouse model;2.to validate the effect of Rg3 to inhibit theproliferation of leukemia cells in vivo.In this study we establish a K562/NOD-SCID leukemia mouse modelsuccessfully .First inoculate irradiated nude mice with K562 cellssubcutaneouly,then take out of the local K562 tumor and grind thetumor tissue to single-cell suspension , which were inoculated intoirradiated NOD/SCID mice by intraperitoneal injection. On the fourthweek after inoculation we can find leukemia cells on peripheral bloodsmear ,and leukemia cell infiltration in liver,spleen and bonemarrow .On the brink of death,the number of peripheral WBC was 8—10 times as much as that before inoculation.The percentage ofleukemia cell on peripheral blood smear is 20—30% of average .There were local tumors in the abdominal cavity or on the greateromentum. The K562/NOD-SCID leukemia mouse model imitates the processof leukemia ,so it is a good tool for research of new drug effecttest and target or gene therapy.On the base of successful establishment of K562/NOD-SCIDleukemia mouse model with ip route,we give K562/NOD-SCID Rg3 byintragastric administration for a period of 9 weeks and divide theminto prophylaxis and treatment group,low dose(10mg/kg)and high dosegroup(20mg/kg).The prophylaxis group have been given Rg3 for 2 weeksbefore inoculation. We carry out the numbering of peripheral WBC andsurvey of peripheral blood smear once a week.Moreover,we calculatethe life span of K562/NOD-SCID of every group.Since the fourth weekafter inoculation, the number of peripheral WBC of control group ishigher than that of prophylaxis and treatment group.On the eighthweek there is a significant difference between all the other groupswith the application of Rg3 except the treatment group with low doseand control group(P<0.05),But there is no significant differencebetween prophylaxis and treatment group, low dose and high dosegroup.On the eighth week,the proportion of leukemic cell ofperipheral blood of the control group is higher than that of everygroup with the application of Rg3,but there is no significantdifference(P>0.05), there is also no significant differencebetween every group with the application of Rg3. On the ninth weekafter inoculation, K562/NOD-SCID of control group died out whilethere was survival of K562/NOD-SCID of all the groups with theexertion of Rg3. Ginsenosides Rg3 can inhibit the proliferation ofleukemia cells in vivo ,manifesting some therapeutic effect.K562/NOD-SCID leukemia mouse model provides us with a favourablemodel system for the research on the mechanisms of proliferation,differentiation and regulation of leukemia cells,the exploitationof new drugs and evaluation of therapeutic effects of new treatmentplan.There has been rare reports on Rg3,s effects on leukemia in vivoat home and abroad.Rg3 is worth further exploitation with itsadvantage of nature and security.At the same time,we should payattention to the followings:1).There is difference between murineand human cytokines ,adhesion molecules, extracellular matrix;2).Nomatter how good effects that new drug has shown in the mouse,detailedpharmacodynamic research should be carried out for sometimes drugconcentration in the mouse may be ineffective for human.Animalexperiment in great numbers should be undertaken on the effects ofginsenosides Rg3 in the inhibition of proliferation of leukemiacells in vivo and the possible mechanisms should be investigated.更多还原