摘要
目的:以糖尿病小鼠(db/db)为研究对象,比较单一使用AMD3100与AMD3100联合粒细胞集落刺激因子(granulocyte colony-stimulating,G-CSF)及两者不同使用顺序对骨髓内皮祖细胞(endothelial progenitor cells,EPCs)的动员效果,为临床获取糖尿病人EPCs进行细胞治疗提供思路。方法:采用AMD3100、重组人粒细胞集落刺激因子注射液(recombinant human granulocyte colony-stimulating,rh G-CSF)参考国内外文献所述剂量和方法进行骨髓EPCs的动员。选用5~6周雄性小鼠,随机将80只db/db小鼠分为A组(AMD3100+NS动员组),B组(AMD3100+G-CSF动员组),C组(G-CSF+AMD3100动员组)和D组(NS空白对照组);80只db/+小鼠为相应对照组,分组对应为A’组,B’组,C’组和D’组。每组在动员后第3、7、10、14天4个时间点于心脏部位取外周血,流式细胞仪检测CD34+/CD133+/Flk-1+细胞的比例。结果取平均值用均值±标准差(单位%)表示,统计学方法处理后进行比较和分析。结果:(1)D组和D’组在动员后第3、7、10、14天4个时间点,CD34+/CD133+/Flk-1+细胞比例无差异(P>0.05),但D’组中CD34+/CD133+/Flk-1+细胞比例高于D组(P=0.003)。(2)A组、B组和C组db/db小鼠在动员后第3、7、10、14天4个时间点,CD34+/CD133+/Flk-1+细胞比例逐渐上升,A组至第7天达到峰值,随后逐渐下降;B组和C组至第10天达到峰值,随后逐渐下降,且在达到峰值时CD34+/CD133+/Flk-1+细胞比例与其余3个时间点相比差异有意义(P=0.000)。在第10天,B组和C组db/db小鼠外周血中CD34+/CD133+/Flk-1+细胞比例较A组和D组高(P=0.000),但B组和C组db/db小鼠外周血中CD34+/CD133+/Flk-1+细胞比例之间的差异无统计学意义(P>0.05)。(3)A’组、B’组和C’组db/+小鼠在动员后第3、7、10、14天4个时间点,CD34+/CD133+/Flk-1+细胞比例逐渐上升,至第7天达到高峰,随后逐渐下降,且在第7天CD34+/CD133+/Flk-1+细胞比例与其余3个时间点相比差异有意义(P=0.000)。在达到峰值的第7天,C’组CD34+/CD133+/Flk-1+细胞比例较A’组、B’组和D’组高(P=0.000)。结论:AMD3100联合G-CSF动员骨髓EPCs的效果优于单一使用AMD3100的动员效果,在非糖尿病状态下,先使用G-CSF再联合AMD3100的动员作用更优,但在糖尿病状态下先使用AMD3100再联合G-CSF的动员作用更优。
Objective:To investigate the role of AMD3100 and granulocyte colony-stimulating(G-CSF)used alone or in different sequences in the mobilization of bone marrow endothelial progenitor cells(EPCs)in diabetic mice(db/db mice),and to provide thoughts for collecting EPCs from diabetic patients for clinical treatment. Methods:With reference to the dose and methods described in related articles in China and foreign countries,AMD3100 and recombinant human granulocyte colony-stimulating(rh G-CSF)injection were used for the mobilization of bone barrow EPCs. A total of 80 male db/db mice aged 5-6 weeks were randomly divided into group A(AMD3100+NS mobilization group),group B(AMD3100+G-CSF mobilization group),group C(G-CSF +AMD3100 mobilization group),and group D(NS blank control group). A total of 80 db/+male mice were selected as corresponding controls and were divided into groups A’,B’,C’,and D’. Peripheral blood samples were collected from the heart on days 3,7,10,and 14 after mobilization,and flow cytometry was used to measure the proportion of CD34+/CD133+/Flk^^-1+cells. The results were expressed as mean±standard deviation(%),and comparison and analysis were performed after statistical analysis. Results:On days 3,7,10,and 14 after mobilization,there was no significant difference in the propor tion of CD34+/CD133+/Flk^-1+cells between groups D and D’(P>0.05),but group D’had a higher proportion than group D(P=0.003).In group A,the proportion of CD34+/CD133+/Flk^-1+ cells gradually increased and reached the highest value on day 7 and then gradually decreased;in groups B and C,the proportion of CD34+/CD133+/Flk^-1+ cells gradually increased and reached the highest value on day10 and then gradually decreased;the proportion of CD34 +/CD133 +/Flk^-1+ cells at the peak was significantly higher than that at the other three time points(P=0.000). On day 10,groups B and C had a significantly higher proportion of CD34+/CD133+/Flk^-1+ cells in peripheral blood of db/db mice than groups A and D(P=0.000),but there was no significant difference between groups B and C(P>0.05). In groups A’,B’,and C’after mobilization,the proportion of CD34+/CD133+/Flk^-1+ cells gradually increased and reached the peak on day 7 and then gradually decreased;the proportion of CD34+/CD133+/Flk^-1+ cells on day 7 was significantly higher than that at the other three time points(P=0.000). On day 7,the C’group had a significantly higher proportion of CD34+/CD133+/Flk^-1+ cells than groups A’,B’,and D’(P=0.000). Conclusion:AMD3100 combined with G-CSF has a better effect in the mobilization of bone marrow EPCs than AMD3100 alone. In non-diabetic conditions,G-CSF followed by the addition of AMD3100 has a better mobilization effect,but in diabetic conditions,AMD3100 followed by the addition of G-CSF has a better mobilization effect.
作者
林晓莹
汪虹
Lin Xiaoying;Wang Hong(Department of Burns ,the Second Affiliated Hospital of Kunming Medical University)
出处
《重庆医科大学学报》
CAS
CSCD
北大核心
2019年第3期247-254,共8页
Journal of Chongqing Medical University
基金
国家自然科学基金地区科学基金资助项目(编号:81660321)
云南省科技厅-昆明医科大学应用基础研究联合专项资助项目[编号:2017FE468(-177)]
