黑枸杞花青素联合人脂肪源性血管外膜细胞支持脐血造血干/祖细胞的增殖  

Anthocyanins from Lycium ruthenicum Murr combined with human adipose-derived pericytes/perivascular cells support proliferation of umbilical cord blood hematopoietic stem/progenitor cells

作　　者：申娅媚 牛云霞 杨婷婷[3] 马洁[4] 胡代宏 郑波 Shen Yamei;Niu Yunxia;Yang Tingting;Ma Jie;Hu Daihong;Zheng Bo(Clinical Medicine College,Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Ningxia Key Laboratory of Stem Cell and Regenerative Medicine,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Diagnosis and Treatment Engineering Technology Research Center of Nervous Disease of Ningxia,General Hospital of Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Department of Hematology,General Hospital of Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China)

机构地区：[1]宁夏医科大学临床医学院,宁夏回族自治区银川市750004 [2]宁夏回族自治区干细胞与再生医学重点实验室,宁夏回族自治区银川市750004 [3]宁夏医科大学总医院,宁夏神经系统疾病诊疗工程技术研究中心,宁夏回族自治区银川市750004 [4]宁夏医科大学总医院血液内科,宁夏回族自治区银川市750004

出　　处：《中国组织工程研究》2025年第1期58-64,共7页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(81860028),项目负责人:郑波;宁夏自然科学基金项目(2023AAC03527),项目负责人:郑波。

摘　　要：背景:黑枸杞花青素(Anthocyanins in Lycium ruthenicum Murr,ALRM)是黑枸杞中重要的活性成分之一,具有抗氧化、免疫调节等功效。人脂肪源性血管外膜细胞(CD146^(+)hAD-PCs)是骨髓间充质干细胞的前体细胞,在体外具有促进造血干/祖细胞增殖与分化的功能。ALRM联合CD146^(+)hAD-PCs对脐血造血干/组细胞的体外支持作用有待于研究。目的:探讨ALRM联合CD146^(+)hAD-PCs对脐血CD34^(+)造血干/祖细胞体外扩增的支持作用。方法:CCK-8法检测不同质量浓度ALRM(0,200,400,600,800,1000 mg/L)对CD146^(+)hAD-PCs增殖的影响;流式细胞术检测ALRM对CD146^(+)hAD-PCs细胞周期的影响。共培养实验分为空白组、ALRM组、CD146^(+)hAD-PCs组、ALRM+CD146^(+)hAD-PCs组,分析ALRM联合CD146^(+)hAD-PCs对脐血CD34^(+)造血干/祖细胞的体外支持作用。共培养1,2,4周,比较扩增后细胞数量、集落形成单位数量,流式细胞仪检测细胞免疫表型,ELISA检测细胞因子水平。结果与结论:(1)ALRM质量浓度为200 mg/L时,CD146^(+)hAD-PCs活力最高,CD146^(+)hAD-PCs的G_(0)/G_(1)期细胞比例下降,S期、G_(2)/M期细胞比例上升(P<0.01)。(2)脐血CD34^(+)造血干/祖细胞数量变化:在共培养1,2,4周时ALRM+CD146^(+)hAD-PCs组高于ALRM组(P均<0.05),在共培养2,4周时ALRM+CD146^(+)hAD-PCs组高于CD146^(+)hAD-PCs组(P均<0.05),ALRM组与空白组随着共培养时间延长细胞数量逐渐减少。(3)集落形成能力及免疫表型分析:在共培养1,2周时ALRM+CD146^(+)hAD-PCs组的集落形成单位数量高于CD146^(+)hAD-PCs组和ALRM组(P均<0.05);在共培养1,2,4周时ALRM+CD146^(+)hAD-PCs组CD45^(+)、CD34^(+)CD33^(-)细胞比例高于CD146^(+)hAD-PCs组(P均<0.01)。(4)细胞因子变化:在共培养4周时ALRM+CD146^(+)hAD-PCs组的白细胞介素2水平高于ALRM组、CD146^(+)hAD-PCs组(P<0.05);在共培养2,4周时ALRM+CD146^(+)hAD-PCs组白细胞介素3水平高于CD146^(+)hAD-PCs组(P<0.05);在共培养1周时ALRM+CD146^(+)h AD-PCs组的粒细胞�BACKGROUND:Anthocyanin is one of the most important active components in Lycium ruthenicum Murr,which has antioxidant and immunomodulatory effects.CD146~+human adipose-derived pericytes/perivascular cells(CD146~+hAD-PCs)are the progenitors of bone marrow mesenchymal stem cells,which can promote the proliferation and differentiation of hematopoietic stem/progenitor cells in vitro.The support effect of anthocyanin in combination with CD146~+hAD-PCs on umbilical cord blood hematopoietic stem/progenitor cells remains to be studied.OBJECTIVE:To investigate the supporting effect of anthocyanins in Lycium ruthenicum Murr(ALRM)combined with CD146~+hAD-PCs on umbilical cord blood CD34~+hematopoietic stem/progenitor cells(UCB CD34~+HSPCs)in vitro.METHODS:The CCK-8 assay was used to detect the effect of different concentrations(0,200,400,600,800,1000 mg/L)of ALRM on the proliferation of CD146~+hAD-PCs.Flow cytometry was used to detect the effect of ALRM on the cell cycle of CD146~+hAD-PCs.The co-culture experiments were divided into blank group,ALRM group,CD146~+hAD-PCs group,and ALRM+CD146~+hAD-PCs group to analyze the in vitro supporting effect of ALRM combined withCD146~+h AD-PCs on UCB CD34~+HSPCs.The number of expanded cells and the number of colony-forming units were compared at 1,2,and 4 weeks of co-culture.The immunophenotype of cells was detected by flow cytometry.The level of cytokines was detected by enzyme-linked immunosorbent assay.RESULTS AND CONCLUSION:(1)The cell viability of CD146~+hAD-PCs was highest at an ALRM concentration of 200 mg/L,the proportion of G_0/G_(1)phase cells decreased and the proportion of S and G_2/M phase cells increased in CD146~+hAD-PCs(P<0.01).(2)The change in number of UCB CD34~+HSPCs cells in the ALRM+CD146~+h AD-PCs group was higher than that in the ALRM group at 1,2,and 4 weeks of co-culture(all P<0.05),and higher than that in CD146~+hAD-PCs group at 2 and 4 weeks of co-culture(all P<0.05).The number of cells in the ALRM group and blank group decreased gradually with the extension

关 键 词：黑枸杞花青素 人脂肪源性血管外膜细胞 脐血 造血干/祖细胞 共培养 体外扩增 

分 类 号：R459.9[医药卫生—治疗学] R394.2[医药卫生—临床医学] R542.2