5-HT通过5-HT2BR促进巨核细胞生长和抗巨核细胞凋亡  

5-HT Promotes Proliferation and Inhibits Apoptosis of Megakarycytes through 5-HT2BR

作　　者：孔惠敏 岑玉蓉 杨默 彭强 黄金棋 KONG Hui-Min;CEN Yu-Rong;YANG Mo;PENG Qiang;HUANG Jin-Qi(Department of Blood Transfusion,The Seventh Affiliated Hospital,Sun Yat-Sen University,Shenzhen 518107,Guangdong Province,China;Department of Hematology,The Affiliated Hospital of Guangdong Medical University,Zhanjiang 524000,Guangdong Province,China;Research Center,The Seventh Affiliated Hospital,Sun Yat-Sen University,Shenzhen 518107,Guangdong Province,China;Emergency Department,Shenzhen Luohu Hospital Group Luohu People's Hospital,Shenzhen 518000,Guangdong Province,China;Department of Hematology,Guangzhou First People s Hospital,Guangzhou 510180,Guangdong Province,China)

机构地区：[1]中山大学附属第七医院输血科,广东深圳518107 [2]广东医科大学附属医院血液内科,广东湛江524000 [3]中山大学附属第七医院科研中心,广东深圳518107 [4]深圳市罗湖医院集团罗湖区人民医院急诊科,广东深圳518000 [5]广州市第一人民医院血液内科,广东广州510180

出　　处：《中国实验血液学杂志》2025年第1期75-81,共7页Journal of Experimental Hematology

基　　金：深圳市基础研究资助项目(JCYJ20190809180213349)。

摘　　要：目的:探讨5-羟色胺(5-HT)对巨核细胞Meg-01增殖、凋亡以及巨核细胞集落形成单位(CFU-MK)的影响及其可能的机制。方法:反相高效液相色谱-电化学检测法检测Meg-01细胞对5-HT的摄取和5-HT在Meg-01细胞中的代谢。免疫荧光染色检测骨髓巨核细胞5-羟色胺2B型受体(5-HT2BR)的表达。将5-HT和/或5-HT2BR抑制剂Ketanserin加入到Meg-01细胞中培养48 h,应用MTT检测法和台盼蓝染色法检测细胞的增殖及细胞活率。Meg-01细胞中加入5-HT和/或Ketanserin孵育72 h,应用流式细胞术检测细胞的早期凋亡情况和凋亡蛋白酶caspase-3的活性。采用CFU-MK培养探究5-HT对巨核细胞分化的影响。结果:Meg-01细胞可以摄取5-HT,并将摄取的5-HT分解成5-羟吲哚乙酸(5-hydroxyindoleacetic acid,5-HIAA)。免疫荧光染色表明骨髓巨核细胞表面有5-HT2BR的表达。5-HT对Meg-01细胞的增殖有促进作用,呈剂量依赖性(r=0.82),5-HT浓度为200 nmol/L时,对Meg-01细胞的促增殖作用最明显(P<0.001)。台盼蓝染色结果也提示200 nmol/L的5-HT对Meg-01细胞的存活率影响最明显(P<0.05)。在5-HT单独存在时,可明显促进Meg-01细胞的增殖(P<0.001),加入Ketanserin后,此作用减弱。5-HT可以明显降低Meg-01细胞的早期凋亡率(P<0.001)和caspase-3的活性(P<0.05),加入Ketanserin之后,Meg-01细胞的早期凋亡率明显增加(P<0.001),caspase-3的活性也有所增加。CFU-MK形成结果表明,5-HT可促进骨髓细胞CFU-MK的形成,且呈剂量依赖性(r=0.89);而在5-HT的基础上加入Ketanserin,5-HT对CFU-MK形成的促进作用减弱(P<0.01)。结论:巨核细胞内有可能存在代谢分解5-HT的单胺氧化酶,将5-HT分解为5-HIAA。5-HT有可能通过5-HT2BR促进巨核细胞的的增殖和分化。此外,在5-HT作用下还可以降低巨核细胞的凋亡,其抗凋亡作用可能是通过5-HT2BR和caspase-3通路介导的。Objective:To investigate the effect of 5-hydroxytryptamine(5-HT)on the proliferation,apoptosis and colony-forming unit-megakaryocyte(CFU-MK)of Meg-01 cells and its possible mechanisms.Methods:The uptake and metabolism of 5-HT in Meg-01 cells were analysed by reverse-phase high-performance liquid chromatography(RP-HPLC)with electrochemical detection.The expression of 5-HT2B receptor(5-HT2BR)in megakaryocytes was detected by immunofluorescence staining.The cell proliferation and viability were measured by MTT and Trypan blue staining after Meg-01 cells were single-cultured or co-cultured with different concentrations of 5-HT/5-HT2BR inhibitor Ketanserin for 48 h.Meg-01 cells were incubated with 5-HT/Ketanserin for 72 h,then the flow cytometry was used to detect early apoptosis of the cells and the activity of caspase-3.Using CFU-MK assay to investigate the effect of 5-HT on the differentiation of megakaryocytes.Results:5-HT could be uptaken by Meg-01 cells,and metabolized into 5-hydroxyindoleacetic acid(5-HIAA).The expression of 5-HT2BR on megakaryocytes could be detected after immunofluorescence staining.5-HT could promote the proliferation of Meg-01 cells at a dose-dependent manner(r=0.82),with the most significant effect observed at a concentration of 200 nmol/L(P<0.001).Trypan blue staining also indicated that 200 nmol/L 5-HT had the most significant effect on the viability of Meg-01 cells(P<0.05).The proliferation of Meg-01 cells treated with 5-HT was increased compared with the untreated control(P<0.001),while the combination of 5-HT with ketanserin downregulated this effect.5-HT significantly reduced the early apoptosis rate(P<0.001)and caspase-3 activity(P<0.05)of Meg-01 cells,while addition of ketanserin significantly increased the early apoptosis rate of Meg-01 cells(P<0.001)and caspase-3 activity also increased to some extent.5-HT promoted the formation of CFU-MK in bone marrow cells in a dose-dependent manner(r=0.89).The addition of ketanserin reduced the promoting effect of 5-HT on CFU-MK formation(P<0

关 键 词：5-羟色胺 Meg-01细胞 5-羟色胺2B型受体 增殖 凋亡 

分 类 号：R331[医药卫生—人体生理学]