摘要
目的探讨升麻整甲汤治疗急性髓系白血病(AML)的可能作用机制。方法48只NSG免疫缺陷小鼠随机分为空白组、模型组、升麻鳖甲汤全方组、升麻鳖甲汤拆方组,每组12只。除空白组外,其余各组小鼠采用小动物辐照仪予200cGy^(60)Co全身照射后,尾静脉注射对数生长期的KG-la细胞1×10^(6)个(0.2ml),建立AML小鼠模型。1周后模型组和空白组小鼠予生理盐水每次0.1ml灌胃,升麻整甲汤全方组小鼠予浓度为1.73g/ml的升麻整甲汤药液每次0.1ml灌胃,升麻鳖甲汤拆方组小鼠予浓度为1.43g/ml的升麻鳖甲汤拆方药液每次0.1ml灌胃,各组均每日灌胃2次,连续28天。每日观察小鼠一般情况,每隔两日记录各组小鼠生存情况。干预28天后检测外周血象(包括白细胞绝对值、血红蛋白、血小板计数);采用HE染色法观察小鼠肝、脾、骨髓组织病理变化;分别采用Western Blot法、免疫荧光染色法检测骨髓中线粒体分裂相关蛋白动力相关蛋白1(Drp1)、线粒体分裂蛋白1(Fis1)及线粒体自噬相关蛋白PTEN诱导激酶1(Pink1)、E3泛素连接酶(Parkin)表达水平。结果空白组小鼠生存率为83.33%(10/12),模型组为25.00%(3/12),升麻整甲汤全方组为66.67%(8/12),升麻整甲汤拆方组为41.67%(5/12),差异具有统计学意义(P<0.05)。模型组小鼠骨髓、脾脏、肝脏组织中可见白血病细胞浸润,脾脏组织结构破坏,肝脏组织可见散在局灶性变性坏死。各药物干预组小鼠骨髓、脾脏及肝脏组织中白血病细胞浸润均有所减轻,脾脏组织结构轻度破坏,其中升麻鳖甲汤全方组改善更为显著。与空白组比较,其余各组小鼠外周血白细胞绝对值升高,血红蛋白、血小板计数以及骨髓Drpl、Fis1、Pink1、Parkin蛋白表达降低(P<0.05)。与模型组比较,升麻整甲汤全方组白细胞绝对值降低,血红蛋白、血小板计数以及骨髓Drpl、Fis1、Pink1、Parkin蛋白表达升高;升麻鳖甲汤拆方组白细胞绝对值降低,血红蛋白、血小板计数以及骨髓Fis1、Parkin蛋白表达升高(P<0.05)。升麻整甲汤全方组白细胞绝对值低于升麻整甲汤拆方组,骨髓Drpl、Pink1蛋白表达高于升麻鳖甲汤拆方组(P<0.05)。结论升麻鳖甲汤能够改善AML模型小鼠生存状态和生存时间,改善肝脏、脾脏、骨髓组织病理损伤,且升麻整甲汤全方相较于拆方的治疗作用更加显著,其机制可能与调高线粒体动力学分裂、诱导细胞自噬有关。
Objective To investigate the possiblemechanism of Shengma Biejia Decoction(升麻鳖甲汤,SBD)in the treatment of acute myeloid leukemia(AML).Methods Forty-eight NSG immunodeficient mice were randomly divided into blank group,model group,whole and disassembled SBD groups,with 12 mice in each group.Except for the blank group,the whole body of the mice in the other groups were irradiated by a small animal irradiator(200 cGy Co),and then injected with 1x10°(0.2 ml)of KG-la cells in the logarithmic growth phase into the tail vein to establish the AML mouse model.One week later,the successfully modeled mice in the the model group and the blank control group were given O.Imlof normal saline per time by gavage,whilethose in the whole and disassembled SBD groups were given 0.1 ml of SBD with a concentration of 1.73 g/ml and 1.43 g/ml per time by gavage,respectively,twice daily for 28 days.The general condition of the mice was observed every day,and the survival condition was recorded every two days.After 28 days,the peripheral blood indicators including the absolute value of white blood cells,hemoglobin value,and platelet count were detected.HE staining was used to observe the pathological changes of liver,spleen and bone marrow in mice.Western blot and immunofluorescence(IF)were used to detect the expression levels of mitochondrial mitosis related proteins including dynamin-related protein 1(Drp1)and mitochondrial fission protein 1(Fis1),as well as mitophagy-related proteins PTEN-induced kinase(Pink1)and E3 ubiquitin ligase(Parkin).Results The survival rate of the blank group was 83.33%(10/12),while that of the model group,the whole and dissembled SBD groups was 25.00%(3/12),66.67%(8/12),and 41.67%(5/12),respectively,showing statistically significant difference(P<0.05).In the model group,infiltrated leukemia cells were found in the bone marrow,spleen and liver,and spleen tissue structure was damaged,with scattered focal degeneration and necrosis in the liver.In the SBD groups,the infiltration of leukemic cells in bone marrow,spleen and liver of mice in each group was alleviated,and the tissue structure was slightly damaged;more improvements were seen in the whole SBD group.Compared to those in the blank group,the absolute value of peripheral while blood cells in the other groups increased,while the hemoglobin level,platelet count,and the expression of Drpl,Fisl,Pinkl and Parkin decreased significantly(P<0.05).Compared to those in the model group,the absolute value of while blood cells decreased,while hemoglobin,platelet count and expression of proteins Drpl,Fisl,Pinkl,and Parkin increased in the whole SBD group(P<O.05);the absolute value of while blood cells decreased,while hemoglobin,platelet count,and the expression of protein Fisl and Pinkin increased in the dissembled SBD group(P<0.05).The absolute value of white blood cells was lower,and the bone marrow Drpl and Pinkl protein expressions were higher in the whole SBD group than in the dissembled SBD groupthan(P<0.05).Conclusion SBD can improve the survival condition and survival time of AML model mice,and improve the histopathological damage of liver,spleen and bone marrow,and the whole SBD group shows better effect than the disassembled prescription.Its mechanism may be related to the up-regulation of mitochondrion dynamic division and the promotion of autophagy.
作者
司雨平
崔思远
魏文健
殷学伟
李宗宏
马国栋
王琰
郑伟
王振振
刘月莉
王敬毅
徐瑞荣
SI Yuping;CUI Siyuan;WEI Wenjian;YIN Xuewei;LI Zonghong;MA Guodong;WANG Yan;ZHENG Wei;WANG Zhenzhen;LIU Yueli;WANG Jingyi;XU Ruirong(The First School of Clinical Medicine,Shandong University of Traditional Chinese Medicine,Jinan,250014;The Affiliated Hospital of Shandong University of Traditional Chinese Medicine;School of Clinical Medicine,Shandong University)
出处
《中医杂志》
CSCD
北大核心
2023年第14期1475-1482,共8页
Journal of Traditional Chinese Medicine
基金
国家自然科学基金(81903984)
山东省自然科学基金(ZR2019PH018)
山东省医药卫生科技发展计划(202003040045,2018WS184)。
关键词
急性髓细胞白血病
升麻鳖甲汤
自噬
线粒体分裂
阴阳毒
acute myeloid leukemia
Shengma Biejia Decoction(升麻鳖甲汤)
autophagy
mitochondrial fission
yin-yangtoxin
