中国组织工程研究

中国组织工程研究 ›› 2019, Vol. 23 ›› Issue (26): 4142-4147.doi: 10.3969/j.issn.2095-4344.1353

• 组织工程血管材料 tissue-engineered vascular materials • 上一篇    下一篇

葡聚糖/聚乳酸-乙醇酸微球联合3种因子促进缺血下肢的血管再生

朱仙花，陈  锋，刘崇栋，周  卫，唐新华
  

  1. 南昌大学第二附属医院血管外科，江西省南昌市  330006
  • 收稿日期:2019-04-04
  • 通讯作者: 陈锋，博士，副主任医师，硕士生导师，南昌大学第二附属医院血管外科，江西省南昌市 330006
  • 作者简介:朱仙花，女，1984年生，江西省上饶市人，汉族，2012年南昌大学医学院毕业，硕士，主治医师，主要从事血管外科相关疾病的研究。
  • 基金资助:

    国家自然科学基金项目(81460083)：复合促血管再生因子/葡聚糖/PLGA纳米微球纤维蛋白胶的血管再生鸡尾酒疗法研究，项目负责人：陈锋；江西省科技厅青年科学基金项目(20142BAB215034)：复合血管再生细胞因子-PLGA纳米微球纤维蛋白胶治疗下肢慢性缺血的实验研究，项目负责人：陈锋；江西省科技厅科技计划一般项目(20141BBG70032)：葡聚糖-PLGA纳米微球携载复合血管生成细胞因子的血管再生治疗研究，项目负责人：陈锋；江西省科技厅青年科学基金重大项目(20151522070093)：动静脉短路法抑制动脉支架内再狭窄的机制研究，项目负责人：陈锋

Dextran/poly(lactic-co-glycolic acid) microspheres combined with three growth factors promote neovascularization in ischemic lower limbs of rats

Zhu Xianhua, Chen Feng, Liu Chongdong, Zhou Wei, Tang Xinhua 
  

  1. Department of Vascular Surgery, Second Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, China
  • Received:2019-04-04
  • Contact: Chen Feng, MD, Associate chief physician, Master’s supervisor, Department of Vascular Surgery, Second Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, China
  • About author:Zhu Xianhua, Master, Attending physician, Department of Vascular Surgery, Second Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, China
  • Supported by:

    the National Natural Science Foundation of China, No. 81460083 (to CF); Jiangxi Science and Technology Department Youth Science Fund, No. 20142BAB215034 (to CF); Science and Technology Planning Project of Jiangxi Province of China (General Program), No. 20141BBG70032 (to CF); Jiangxi Science and Technology Department Youth Science Fund (Major Program), No. 20151522070093 (to CF)

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摘要：

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文题释义：
血管再生治疗：是指向阻塞血管内或缺血的组织器官内，转移具有血管再生作用的细胞因子、基因或干细胞，利用它们促缺血组织血管再生的能力，使缺血组织建立起广泛的新生血管网络，建立丰富的侧支循环，改善缺血组织的血液供应，从而达到临床治疗目的。目前血管再生的细胞因子治疗研究主要问题是：血管再生需要多种细胞因子的协同参与完成，单一的细胞因子诱导生成的血管是不完整不成熟的；细胞因子在体内的生物半衰期较短，为维持其生物学活性必须重复给药。
聚乳酸-羟基乙酸共聚物：是由羟基乙酸和乳酸聚合而成，是一种生物可降解的高分子有机化合物，可在体内缓慢降解为二氧化碳和水，其具有良好的生物相容性、生物可降解性、机械强度和降解速度可调节性、良好的成囊和成膜等可塑性，因此被广泛应用于制药和医用材料领域。
 
 
背景：葡聚糖/聚乳酸-乙醇酸微球药物缓释系统可使药物在作用部位维持有效药物浓度。
目的：探讨携载粒细胞集落刺激因子、促红细胞生成素或血管内皮生长因子葡聚糖/聚乳酸-乙醇酸微球对缺血下肢血管再生的影响。
方法：制备分别携载粒细胞集落刺激因子、促红细胞生成素与血管内皮生长因子的葡聚糖/聚乳酸-乙醇酸微球，通过扫描电镜观察其表面形态，利用ELISA法检测其包封率和累积释放率。将3种携载生长因子的微球与纤维蛋白胶混合，制备携载生长因子葡聚糖/聚乳酸纤维蛋白胶复合物。取24只雄性SD大鼠(江西省中医学院实验动物中心提供)，制作右下肢缺血模型，随机分成2组干预，每组12只：观察组大腿肌肉内侧分5点注射携载3种生长因子葡聚糖/聚乳酸-乙醇酸微球的纤维蛋白胶复合物，对照组大腿肌肉内侧注射葡聚糖/聚乳酸-乙醇酸微球纤维蛋白凝胶复合物。术后第1周取注射部位肌肉组织，免疫组织化学染色观察增殖细胞核抗原、Bcl-2、基质细胞衍生因子1和趋化因子基质细胞衍生因子4的表达；术后第4周取注射部位肌肉组织，碱性磷酸酶与免疫组织化学染色观察毛细血管密度。实验方案经南昌大学医学院动物实验伦理委员会批准。
结果与结论：①葡聚糖/聚乳酸-乙醇酸微球呈球形，表面光滑，直径40-120 μm；搭载粒细胞集落刺激因子、促红细胞生成素与血管内皮生长因子葡聚糖/聚乳酸-乙醇酸微球的包封率分别为84%，85%，82%，4周的累积释放率分别为89.5%，90.3%，91.2%；②观察组增殖细胞核抗原、Bcl-2、基质细胞衍生因子1和趋化因子基质细胞衍生因子4阳性表达高于对照组；③观察组毛细血管密度与α-平滑肌肌动蛋白阳性血管密度高于对照组(P < 0.05)；④结果表明利用葡聚糖/聚乳酸-乙醇酸微球联合多种血管再生因子治疗下肢缺血，是一种有前途的血管再生方法。

关键词: 聚乳酸-乙醇酸共聚物, 粒细胞集落刺激因子, 促红细胞生成素, 血管内皮生长因子, 微球, 缺血, 葡聚糖, 血管再生

Abstract:

BACKGROUND: The dextran/poly(lactic-co-glycolic acid) (PLGA) microspheres as a sustained release drug delivery system enables the drug to maintain an effective concentration at the site of action.
OBJECTIVE: To investigate the effects of dextran/PLGA microspheres carrying granulocyte colony-stimulating factor, erythropoietin and vascular endothelial growth factor on the neovascularization of ischemic lower limb of rats.
METHODS: Dextran/PLGA microspheres carrying granulocyte colony-stimulating factor, erythropoietin, and vascular endothelial growth factor were prepared and their surface morphology was observed by scanning electron microscopy. The encapsulation efficiency and cumulative release rate were detected by ELISA method. Dextran/PLGA microspheres carrying three growth factors were mixed with fibrin glue to prepare a growth factor dextran/PLGA/fibrin glue complex. Right lower limb ischemia models were produced in 24 male Sprague-Dawley rats (Laboratory Animal Center, Jiangsu University of Traditional Chinese Medicine, China). These animal models were randomly divided into two groups (n = 12/group). In the experimental group, dextran/PLGA/fibrin glue complex carrying three growth factors was injected into the inner thigh muscle at 5 points. In the control group, dextran/PLGA/fibrin glue complex without growth factors was identically injected. At 1 week after surgery, the muscle tissue around the injection site was harvested for immunohistochemistry to detect the expression of proliferating cell nuclear antigen, Bcl-2, stromal cell-derived factor 1, and C-X-C chemokine receptor type 4. At 4 weeks after surgery, the muscle tissue around the injection site was harvested for histological and immunohistochemical examination to measure capillary density. This study was approved by Animal Ethics Committee of Nanchang University School of Medicine, China.
RESULTS AND CONCLUSION: The dextran/PLGA microspheres were spherical and had a smooth surface with a diameter of 40-120 μm. The encapsulation efficiency of dextran/PLGA microspheres carrying granulocyte colony-stimulating factor, erythropoietin, and vascular endothelial growth factor was 84%, 85%, and 82%, respectively. The 4-week cumulative release rate was 89.5%, 90.3%, 91.2%, respectively. The expression of proliferating cell nuclear antigen, Bcl-2, stromal cell-derived factor 1, and C-X-C chemokine receptor type 4 in the experimental group was significantly higher than that in the control group. Capillary density and α-smooth muscle actin-positive vascular density in the experimental group were significantly higher than those in the control group (P < 0.05). These results suggest that dextran/PLGA microspheres carrying multiple growth factors for treatment of lower limb ischemia is a promising therapeutic method.

Key words: poly(lactic-co-glycolic acid), granulocyte colony-stimulating factor, erythropoietin, vascular endothelial growth factor, microsphere, ischemia, dextran, angiogenesis

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