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富半胱氨酸蛋白61对大鼠自体脂肪移植存活影响及其机制的实验研究

陈兆欢 段然 黄晓璐 李青峰

陈兆欢, 段然, 黄晓璐, 等. 富半胱氨酸蛋白61对大鼠自体脂肪移植存活影响及其机制的实验研究[J]. 器官移植, 2021, 12(4): 403-411. doi: 10.3969/j.issn.1674-7445.2021.04.006
引用本文: 陈兆欢, 段然, 黄晓璐, 等. 富半胱氨酸蛋白61对大鼠自体脂肪移植存活影响及其机制的实验研究[J]. 器官移植, 2021, 12(4): 403-411. doi: 10.3969/j.issn.1674-7445.2021.04.006
Chen Zhaohuan, Duan Ran, Huang Xiaolu, et al. Experimental study of effect and mechanism of cysteine rich protein 61 on survival of adipose tissues in rats after autologous fat grafting[J]. ORGAN TRANSPLANTATION, 2021, 12(4): 403-411. doi: 10.3969/j.issn.1674-7445.2021.04.006
Citation: Chen Zhaohuan, Duan Ran, Huang Xiaolu, et al. Experimental study of effect and mechanism of cysteine rich protein 61 on survival of adipose tissues in rats after autologous fat grafting[J]. ORGAN TRANSPLANTATION, 2021, 12(4): 403-411. doi: 10.3969/j.issn.1674-7445.2021.04.006

富半胱氨酸蛋白61对大鼠自体脂肪移植存活影响及其机制的实验研究

doi: 10.3969/j.issn.1674-7445.2021.04.006
基金项目: 

国家自然科学基金 81620108019

详细信息
    作者简介:

    陈兆欢,女,1995年生,硕士,研究方向为器官修复重建与再生,Email:chzhh2019@163.com

    通讯作者:

    黄晓璐,女,博士,研究方向为器官修复重建与再生,Email:xl_huang@alumni.sjtu.edu.cn

    李青峰,男,博士研究生导师,教授,研究方向为器官修复重建与再生,Email:dr.liqingfeng@yahoo.com

  • 中图分类号: R617, R62

Experimental study of effect and mechanism of cysteine rich protein 61 on survival of adipose tissues in rats after autologous fat grafting

More Information
  • 摘要:   目的  探索富半胱氨酸蛋白61即CCN家族成员1(CCN1)对自体脂肪移植术后脂肪组织存活的影响及其作用机制。  方法  建立大鼠自体脂肪移植模型1周后,随机分为CCN1组与对照组,每组各20只。比较两组大鼠移植脂肪存活情况; 比较两组大鼠移植脂肪组织形态、活性脂肪细胞比例及新生血管数量; 利用高通量测序技术比较两组大鼠移植脂肪组织差异表达信使核糖核酸(mRNA)并进行聚类分析; 比较两组大鼠移植脂肪组织促炎因子的表达水平。  结果  与对照组比较,CCN1组脂肪组织质量保留率较高,差异有统计学意义(P < 0.05),CCN1组脂肪细胞完整性更高,囊泡化与空泡化程度、炎症细胞聚集程度和纤维化程度更低,差异均有统计学意义(均为P < 0.000 1)。免疫荧光染色结果显示,CCN1组中活性脂肪细胞比例更高且形态均一,对照组存在较少的活性脂肪细胞且大小不一,伴有囊泡化现象。与对照组比较,CCN1组新生血管数量更多,血小板衍生生长因子(PDGF)和成纤维细胞生长因子(FGF)mRNA表达水平均升高,差异均有统计学意义(均为P < 0.05)。高通量测序结果显示两组的转录组水平差异有统计学意义,CCN1组中与M1型巨噬细胞相关的细胞表面标志物、炎症因子及趋化因子基因表达呈下调趋势,实时荧光定量聚合酶链反应(RT-qPCR)结果显示CCN1组白细胞介素(IL)-8、IL-1和Toll样受体(TLR)2 mRNA表达水平较对照组均降低,差异均有统计学意义(P < 0.01~0.05)。  结论  在自体脂肪移植过程中添加外源性CCN1能有效促进脂肪组织新生血管化,提高移植脂肪存活率,其机制可能是通过下调TLR2表达介导巨噬细胞向M2型转化。

     

  • 图  1  两组大鼠自体移植脂肪存活情况比较

    注:A图为自体脂肪移植术后3个月,移植脂肪组织大体观察; B图为两组大鼠移植脂肪组织质量保留率比较。

    Figure  1.  Comparison of the survival of autologous fat grafts of rats between two groups

    图  2  两组大鼠自体移植脂肪组织形态比较

    注:A图示HE染色结果,CCN1组可见更多完整脂肪细胞,而对照组可见更多囊泡结构(黑色星号),较多炎症细胞聚集(黑色箭头),同时纤维化(黑色三角)程度更高; B图对HE染色结果进行半定量分析,与对照组比较,aP < 0.000 1。

    Figure  2.  Comparison of morphology of autologous fat graft tissues of rats between two groups

    图  3  两组大鼠自体移植脂肪组织中活性脂肪细胞比例比较

    注:A图示脂肪移植术后3个月移植脂肪组织免疫荧光染色(×200)结果,CCN1组可见更多大小均一、结构完整的活性脂肪细胞,对照组存在较少的活性脂肪细胞且大小不一,伴有囊泡化现象; B图为perilipin染色区域的定量分析结果,与对照组比较,aP < 0.01。

    Figure  3.  Comparison of active adipocytes proportion in autologous fat graft tissues of rats between two groups

    图  4  两组大鼠自体移植脂肪组织新生血管数量比较

    注:A图示脂肪移植术后3个月移植脂肪组织新生血管内皮细胞免疫荧光染色(×800)结果,可见CD31阳性内皮细胞(箭头所示); B图为CD31染色区域的定量分析结果; C图为RT-qPCR检测两组PDGF、VEGF、FGF mRNA表达水平; 与对照组比较,aP < 0.05。

    Figure  4.  Comparison of neovascular number of autologous fat graft tissues of rats between two groups

    图  5  两组大鼠自体移植脂肪组织促炎因子mRNA表达水平比较

    注:与对照组比较,aP < 0.05,bP < 0.01。

    Figure  5.  Comparison of the mRNA expression levels of proinflammatory cytokines of autologous fat graft tissuesof rats between two groups

    表  1  高通量测序分析两组mRNA表达水平差异

    Table  1.   High-throughput sequencing analysis of differenceof mRNA expression levels between two groups

    M1型巨噬细胞相关基因 log2(差异倍数) 表达量改变
    细胞表面标志物
      SOCS3 -1.963 36 下调
      TLR2 -1.364 11 下调
      TLR7 -1.486 99 下调
      TLR9 -1.599 46 下调
      IL-18RAP -1.669 3 下调
    炎症因子和趋化因子
      IL-1β -4.319 61 下调
      TNF-αIP3 -1.654 89 下调
      IL-23α -1.700 33 下调
      IL-6 -2.539 59 下调
      IL-18 -1.477 76 下调
      CCL3 -4.207 43 下调
      CCL4 -1.771 42 下调
      CXCL1 -3.184 19 下调
      CXCL2 -6.822 16 下调
      CXCL6 -3.915 91 下调
    下载: 导出CSV
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  • 收稿日期:  2021-04-28
  • 网络出版日期:  2021-07-13
  • 刊出日期:  2021-07-15

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