摘要
目的通过腺病毒载体介导的小鼠γ干扰素(m IFN-γ)在小鼠气道上皮细胞转基因表达,探讨其对博莱霉素诱导小鼠肺纤维化模型治疗的作用机制。方法42只雄性ICR小鼠,随机分为阴性对照组(NC组)、纤维化模型组(FMA、FMB、FMC组)和转基因治疗组(GTA、GTB、GTC组),除NC组外,其余各组经鼻滴入博莱霉素建立纤维化模型。治疗各组在第14天经鼻给予带有m IFN-γ基因的复制缺陷型腺病毒(AdCMVm IFN-γ)悬液。经流式细胞仪测定外周血Th1/Th2和Tc1/Tc2,计数支气管肺泡灌洗液(BALF)细胞分类,右肺行HE及M asson染色,测定左肺组织中羟脯氨酸(HYP)含量,半定量RT-PCR方法了解肺组织中多种细胞因子的表达。测定小鼠潮气量(Vt)、0.1秒率(FEV0.1/FVC%)和静态胸肺顺应性(Cst)。结果AdCMVm IFN-γ治疗各组BALF中的细胞总数与同期模型组相比显著增高(P<0.01),但其肺组织中HYP含量均显著降低(P<0.01)。在呼吸生理上,GTC组Vt和Cst较FMC组明显改善(P<0.05)。AdCMVm IFN-γ局部转基因表达可抑制Th2、Tc2优势表达状态。与模型组比较,治疗各组中IL-12、MMP-2和MMP-9显著升高;IL-13、TGF-β1和TIMP-1显著降低。结论m IFN-γ转基因表达可减轻肺组织纤维化的程度,除了直接作用,还可能通过调节肺组织中多种细胞因子表达实现。
Objective To investigate the transgene expresscon of adenoviral vector mediated mufine interferon-γ （AdCMVmIFN-γ） and its effect on bleomycin-induced pulmonary fibrosis model. Methods 42 ICR male mice were randomly divided into one negative control group （ NC）, three pulmonary fibrosis model groups （ FMA, FMB, FMC） and three gene therapy groups （GTA, GTB, GTC）. Except for NC group, mice in the other groups were administrated by nasal instillation with bleomycin. On day 14th, AdCMVmIFN-γ solution was administrated by nasal instillation in gene therapy groups. T lymphocytes were quantified for Th1/Th2 and Tc1/Tc2 by flow cytometry. Accumulation of inflammatory cells in bronchioalveolar lavage （BALF） was quantified by cell count. The fight lung was stained with either hematoxylin-eosin or Masson trichrome. Left lung was weighed and its hydroxyproline （HYP） content was assayed by HCl acid hydrolysis. Multi-cytokine expression in lung tissue was tested by RT-PCR. The tidal volume （V1 ）, FEV0.1/FVC% and static compliance （Cst） of mice were measured before they were sacrificed. Results The total cell number in BALF in gene therapy groups were significantly higher than that in pulmonary fibrosis model groups at the same time point （P 〈0.01 ）. However, HYP content in lung tissue in gene therapy groups were significantly lower than that in pulmonary fibrosis model groups （P 〈0.01 ）. Vt and Cst in GTC group were significantly improved than those in FMC group （P 〈0.05）. Th2 and Tc2 were showed as dominant expression at the fibrosis forming stage, which could be abrogated by AdCMVmIFN-γ locally transgenic expression. Compared with the cytokine expression in the lung in FM groups, the expression of IL-12, MMP-2 and MMP-9 mRNA were significantly higher in GT groups; the expression of IL- 13, TGF-β1 and TIMP-1 mRNA were significantly lower in GT groups. Conclusions The molecular mechanisms of suppressing pulmonary fibrosis by mIFN-γ transgenic expression involve the indirect action by the regulation of cytokine expression locally in the lungs, in addition to the direct effects of locally mIFN-γ ovcrexpression.
出处
《山东医药》
CAS
北大核心
2009年第28期14-17,共4页
Shandong Medical Journal
基金
北京市自然科学基金资助项目(7042031)。
关键词
博莱霉素
肺间质纤维化
基因治疗
肺功能
细胞因子
干扰素-γ重组
bleomycin
pulmonary interstitial fibrosis
gene therapy
lung function
cytokine
interferongamma, re-combinant
