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中华细胞与干细胞杂志(电子版)

中华细胞与干细胞杂志(电子版) ›› 2017, Vol. 07 ›› Issue (04) : 202 -207. doi: 10.3877/cma.j.issn.2095-1221.2017.04.003

所属专题： 文献；

论著

心肌样细胞移植对大鼠心肌梗死的疗效观察
王秀力1, 姜世强2, 刘颖1, 王洁1, 吕茜1, 王雷1, 李春梅1, 宫海滨1,†()   
  1. 1. 221009　徐州市心血管病研究所　徐州市中心医院中心实验室　徐州市医学重点实验室
    2. 221009　徐州仁慈医院麻醉科
  • 收稿日期:2016-12-19 出版日期:2017-08-01
  • 通信作者: 宫海滨
  • 基金资助:
    江苏省生命健康科技专项资金资助项目(BL2012019); 徐州市科技发展项目(XM13B052)

Curative effect of transplantion of cardiomyocyte-like cells on myocardial infarction rats

Xiuli Wang1, Shiqiang Jiang2, Ying Liu1, Jie Wang1, Qian Lyu1, Lie Wang1, Chunmei Li1, Haibin Gong1,†()   

  1. 1. Department of Xuzhou Cardiovascular Disease Institute, Xuzhou Central Hospital Central Laboratory, Xuzhou Key Lab of Medica, Xuzhou 221009, China
    2. Department of Anesthesiology, Xuzhou Mercy Hospital, Xuzhou 221009, China
  • Received:2016-12-19 Published:2017-08-01
  • Corresponding author: Haibin Gong
  • About author:
    Corresponding author: Gong Haibin, Email:
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引用本文：

王秀力, 姜世强, 刘颖, 王洁, 吕茜, 王雷, 李春梅, 宫海滨. 心肌样细胞移植对大鼠心肌梗死的疗效观察[J]. 中华细胞与干细胞杂志(电子版), 2017, 07(04): 202-207.

Xiuli Wang, Shiqiang Jiang, Ying Liu, Jie Wang, Qian Lyu, Lie Wang, Chunmei Li, Haibin Gong. Curative effect of transplantion of cardiomyocyte-like cells on myocardial infarction rats[J]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2017, 07(04): 202-207.

目的

观察心肌样细胞（CLC）移植对心肌梗死大鼠的疗效。

方法

制备大鼠心肌梗死动物模型，4周后超声检测大鼠血流动力学变化；体外分离、纯化大鼠骨髓间充质干细胞（BMSC），15 ng/ml的胰岛素样生长因子1（IGF-1）诱导BMSC分化为CLC 3，7，14和21 d，采用Western blot技术检测心肌标志性蛋白α-SMA，Ttroponin-T和Troponin-I的表达。CLC移植治疗心肌梗死大鼠，实验共分4组：心肌梗死组，生理盐水对照组，BMSC组及CLC组。移植治疗4周，超声检测血流动力学变化后取材大鼠心肌，冰冻切片，HE染色后检测大鼠心肌梗死的面积。统计分析采用单因素方差分析，实验组之间比较采用t检验。

结果

心肌梗死模型制备4周，模型组大鼠的LVSP数值为（118.74±11.56）mmHg，低于假手术组（130.42±12.58）mmHg，模型组大鼠的LVEDP数值为（14.29?±?7.63）mmHg，高于假手术组（2.44±1.11）mmHg，+dp/dt数值为（2645.17±468.16）mmHg/s低于假手术组（4087.29?±525.17）mmHg/s，-dp/dt数值为（2487.80±348.21）mmHg/s低于假手术组（3865.81±614.37）mmHg/s，差异均具有统计学意义（P < 0.01）。IGF-1诱导BMSC 3，7，14和21d后均检测到α-SMA、Troponin-T和Troponin-I的表达，与第3天比较P < 0.01。移植治疗4周后发现生理盐水对照组大鼠血流动力学没有任何改变，BMSC组及CLC组大鼠的LVEDP数值分别为（9.15±4.54）mmHg和（4.67±3.46）mmHg，低于心肌梗死组（14.29±7.63）mmHg，+dp/dt数值分别为（3245.45±368.76）mmHg/s和（3612.78±468.54）mmHg/s，高于心肌梗死组（2645.17±468.16）mmHg/s，-dp/dt数值分别为（3014.11±412.12）mmHg/s和（3347.21±398.56）mmHg/s，高于假手术组（2487.80±348.21）mmHg/s,同时CLC组大鼠的LVEDP（4.67±3.46）mmHg低于BMSC组（9.15±4.54）mmHg，±dp/dt数值分别是（3612.78±468.54）mmHg/s和（3347.21±398.56）mmHg/s高于BMSC组（3245.45±368.76）mmHg/s和（3014.11±412.12）mmHg/s，差异均具有统计学意义（P < 0.01）。BMSC组和CLC组的心梗面积与心肌梗死组比较明显减小，CLC组的梗死面积与BMSC组比较也明显减小，差异均具有统计学意义（P < 0.01）。

结论

IGF-1诱导BMSC分化为CLC后移植治疗心肌梗死大鼠比单独移植BMSC疗效更好。

关键词: 骨髓, 间质干细胞, 胰岛素样生长因子Ⅰ, 肌细胞，心脏, 心肌梗死, 细胞移植
Objective

To observe the effect of transplanting cardiomyocyte-like cells (CLC) to treat myocardial infarction in rats.

Methods

Animal models of myocardial infarction were prepared, and hemodynamics changes of rats models were tested by ultrasonic method 4 weeks later. Bone mesenchymal stem cells (BMSC) were isolated and purified in vitro, which were induced to CLCs by 15 ng/ml insulin-like growth factor 1 (IGF-1) , and then the expression of myocardial iconic protein α-SMA, Troponin-T and Troponin-Ⅰwere detected with Western blotting method in 3 d, 7 d, 14 d and 21 d. The experimental rats were randomly assigned into 4 groups (a myocardial infarction group, a normal saline group, a BMSC group and a CLC group) . The area of myocardial infarction was detected on rats 4 weeks after transplantation therapy by HE coloring. Data among multiple groups were compared by one-way ANOVA and data between two independent groups were compared by t-test.

Results

After myocardial infarction models were prepared for 4 weeks, the LVSP (118.74 ± 11.56) mmHg of the model group were significantly lower than those of the sham operation group (130.42 ± 12.58) mmHg, the LVEDP (14.29 ± 7.63) mmHg of the model group were significantly higher than those of the sham operation group (2.44 ±1.11) mmHg, +dp/dt (2645.17 ± 468.16) mmHg/s of the model group were significantly lower than those of the sham group (4087.29 ± 525.17) mmHg/s, and -dp/dt (2487.80 ± 348.21) mmHg/s of the model group were also significantly lower than those of the sham group (3865.81 ± 614.37) mmHg/s (P < 0.01) . The expressions of α-SMA, Troponin-T and Troponin-Ⅰwere detected when BMSCs were induced by IGF-1 on 3 d, 7 d, 14 d and 21 d (P < 0.01) . There were not any changes in haemodynamics of the normal saline group 4 weeks after transplantation therapy, the LVEDP of the BMSC group (9.15?±?4.54) mmHg and CLC group (4.67 ± 3.46) mmHg were significantly lower than those of the myocardial infarction group (14.29 ± 7.63) mmHg, the +dp/dt of BMSC group (3245.45?±?368.76) mmHg/s and CLC group (3612.78?±?468.54) mmHg/s were significantly higher than those of the myocardial infarction group (2645.17?±?468.16) mmHg/s, the -dp/dt of BMSC group (3014.11?±?412.12) mmHg/s and CLC group (3347.21?±?398.56) mmHg/s were also significantly higher than the sham group (2487.80?±?348.21) mmHg/s. In the meantime, the LVEDP of the CLC group (4.67?±?3.46) mmHg was significantly lower than that of the BMSC group (9.15?±?4.54) mmHg P < 0.01, the +dp/dt of the CLC group (3612.78?±?468.54) mmHg/s were significantly higher than those of the BMSC group (3245.368?±?468.16) , the -dp/dt of the CLC group (3347.21?±?398.56) mmHg/s were also significantly higher than those of the BMSC group (3014.11?±?412.12) mmHg/s. The myocardial infarction area of the BMSC group and CLC group was reduced obviously compared with the myocardial infarction group, and the myocardial infarction area of CLC group was less than that of the BMSC group (P < 0.01) .

Conclusion

It may be of a better effect that transplanting BMSC induced by IGF-1 into CLC than only transplanting BMSC on myocardial infarction rats.

Key words: Bone marrow, Mesenchymal stem cells, Insulin-like growth factor I, Myocytes, cardiac, Myocardial infarction, Cell transplantation
表1 模型组和假手术组大鼠心功能血流动力学指标变化比较（±s）
分组 动物数 LVSP（mmHg） LVEDP（mmHg） +dp/dt（mmHg/s） -dp/dt（mmHg/s）
假手术组 30 130.42±12.58 2.44±1.11 4087.29±525.17 3865.81±614.37
模型组 40 118.74±11.56 14.29±7.63 2645.17±468.16 2487.80±348.21
t值 ? 3.979 -9.605 11.905 11.028
P值 ? < 0.01 < 0.01 < 0.01 < 0.01
图1 倒置显微镜下观察分离培养的BMSCs（×400）
图2 倒置显微镜下观察BMSC的鉴定（免疫细胞化学染色方法，×400）
表2 IGF-1诱导BMSC不同时间分化为心肌样细胞后α-SMA、Troponin-T和Troponin-I的表达结果（±s）
分组 第3天 第7天 第14天 第21天 F值 P值
α-SMA 1.00±0.00 1.14±0.08 1.58±0.09 1.60±0.05 250.69 < 0.01
Troponin-T 1.00±0.00 1.35±0.06 1.92±0.10 1.85±0.07 445.55 < 0.01
Troponin-I 1.00±0.00 1.17±0.26 1.89±0.08 1.89±0.10 106.36 < 0.01
表3 移植BMSC组与CLC组大鼠和心肌梗死组大鼠心功能血流动力学指标变化比较（±s）
分组 动物数 LVSP（mmHg） LVEDP（mmHg） +dp/dt（mmHg/s） -dp/dt（mmHg/s）
心肌梗死组 10 118.74±11.56 14.29±7.63 2645.17±468.16 2487.80±348.21
生理盐水对照组 10 119.67±10.41 14.12±3.89 2754.24±478.25 2498.14±354.34
BMSC组 10 121.25± 9.45 9.15±4.54a 3245.45±368.76a 3014.11±412.12a
CLC组 10 125.14± 6.78 4.67±3.46ab 3612.78±468.54ab 3347.21±398.56ab
F值 ? 1.023 42.441 14.367 13.758
P值 ? 0.393 < 0.01 < 0.01 < 0.01
表4 心肌梗死大鼠注射BMSC与CLC后梗死面积的比较（±s）
分组 动物数 LVSP（mmHg） LVEDP（mmHg） +dp/dt（mmHg/s） -dp/dt（mmHg/s）
心肌梗死组 10 118.74±11.56 14.29±7.63 2645.17±468.16 2487.80±348.21
生理盐水对照组 10 119.67±10.41 14.12±3.89 2754.24±478.25 2498.14±354.34
BMSC组 10 121.25± 9.45 9.15±4.54a 3245.45±368.76a 3014.11±412.12a
CLC组 10 125.14± 6.78 4.67±3.46ab 3612.78±468.54ab 3347.21±398.56ab
F值 ? 1.023 42.441 14.367 13.758
P值 ? 0.393 < 0.01 < 0.01 < 0.01
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