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七甲川花菁类荧光小分子IR-61改善大鼠肺纤维化
Fluorescent small molecule IR-61 improves pulmonary fibrosis induced by intratracheal bleomycin in rats
- 第三军医大学学报 2020年42卷第4期 页码：350-358
- 作者机构：
1.550025贵阳, 贵州医科大学公共卫生学院，环境污染与疾病监控教育部重点实验室
2.400038 重庆, 陆军军医大学(第三军医大学)军事预防医学系火箭军医学教研室，创伤、烧伤与复合伤国家重点实验室
3.646000 四川 泸州，西南医科大学附属医院肿瘤科
- 作者简介：
[张爱华，E-mail: aihuagzykd@163.com ZHANG Aihua, E-mail: aihuagzykd@163.com
[史春梦，E-mail: shicm@sina.com SHI Chunmeng, E-mail: shicm@sina.com
- 基金信息：国家重点研究发展计划项目(2016YFC1000805);重庆市高校创新团队资助项目(CXTDG201602020);军队后勤科研重大项目(AWS17J007)
DOI：10.16016/j.1000-5404.201909108
中图分类号： R563.13; R966; R977.6收稿：2019-09-15，
修回：2019-12-11，
纸质出版：2020-02-28
- 稿件说明：
移动端阅览
黄玉, 王子文, 王会兰, 等. 七甲川花菁类荧光小分子IR-61改善大鼠肺纤维化[J]. 第三军医大学学报, 2020,42(4):350-358.
Yu HUANG, Ziwen WANG, Huilan WANG, et al. Fluorescent small molecule IR-61 improves pulmonary fibrosis induced by intratracheal bleomycin in rats[J]. Journal of Third Military Medical University, 2020, 42(4): 350-358.
黄玉, 王子文, 王会兰, 等. 七甲川花菁类荧光小分子IR-61改善大鼠肺纤维化[J]. 第三军医大学学报, 2020,42(4):350-358. DOI： 10.16016/j.1000-5404.201909108.
Yu HUANG, Ziwen WANG, Huilan WANG, et al. Fluorescent small molecule IR-61 improves pulmonary fibrosis induced by intratracheal bleomycin in rats[J]. Journal of Third Military Medical University, 2020, 42(4): 350-358. DOI： 10.16016/j.1000-5404.201909108.
摘要
目的
2
探索七甲川花菁类荧光小分子IR-61在肺纤维化发生过程中的作用以及对肺成纤维细胞转分化、增殖和迁移的影响。
方法
2
采用气管滴注博来霉素(bleomycin
BLM，2.5 mg/kg)建立大鼠肺纤维化模型；腹腔注射PBS或IR-61(2.0 mg/kg，每周2次)；BLM处理10 d后，尾静脉注射IR-61(2.0 mg/kg)，于24 h后取各器官行近红外荧光成像(near infrared fluorescence imaging，NIR)，观察IR-61在各器官的蓄积情况；治疗28 d后，取肺组织制作切片，行HE和Masson染色，显微镜下观察；采用RT-PCR、Western blot测定人肺成纤维细胞(human fetal lung fibroblast
HFL1)与原代大鼠肺成纤维细胞转分化及增殖相关基因Collagen-1、Collagen-3、α-SMA、Fibronectin、FOXM1、CCNB1、CDC25b和AURKB的mRNA及蛋白表达；用细胞划痕实验进行迁移检测。
结果
2
① NIR显示:IR-61可靶向至博来霉素诱导的受损伤肺组织。②形态学观察显示:BLM+IR-61组较博来霉素组形态改善，肺塌陷少，纤维结节少。经组织学鉴定，BLM+IR-61处理组较BLM处理组肺纤维化程度低[(0.103 3±0.0145 3)
vs
(0.460 0±0.0378 6)，
P
<
0.05
]
。IR-61预处理HFL1与原代大鼠肺成纤维细胞后，用TGF-β1刺激，Western blot和RT-PCR分析结果显示:TGF-β1+IR-61处理组较TGF-β1处理组转分化特异性基因Collagen-1、Collagen-3、α-SMA、Fibronectin的mRNA及蛋白表达降低(
P
<
0.05)，同时，增殖特异性FOXM1、CCNB1、CDC25b和AURKB基因的表达也降低(
P
<
0.05)，说明IR-61可以抑制TGF-β1诱导的肺成纤维细胞的转分化与增殖的作用。③细胞划痕实验结果显示:TGF-β1诱导肺成纤维细胞迁移活性增强，IR-61能有效抑制肺成纤维细胞的迁移能力。
结论
2
IR-61通过靶向到受损伤的肺组织并通过抑制肺成纤维细胞的转分化、增殖和迁移来减轻博来霉素诱导的肺纤维化。
Abstract
Objective
2
To evaluate the effects of IR-61 on the development of pulmonary fibrosis and on the proliferation and migration of lung fibroblasts in rats.
Methods
2
Forty male SD rats (6 to 8 weeks old) were randomized equally into normal control group
IR-61 group
bleomycin group and bleomycin+IR-61 group. In the latter 2 groups
rat models of
pulmonary fibrosis were established by intratracheal infusion of bleomycin (2.5 mg/kg). In the 2 groups with IR-61 treatment
IR-61 was intraperitoneally injected at the dose of 2.0 mg/kg 30 min following intratracheal bleomycin infusion and was then administered twice a week. At 10 d after bleomycin treatment
3 rats from each group were selected for IR-61 injection (2 mg/kg)
via
the tail vein to observe the accumulation of IR-61 in each organ at 24 h using near infrared fluorescence imaging (NIR). After 28 d of treatment
the rats were euthanized and the lung tissue was collected for HE and Masson staining. RT-PCR and Western blotting were performed to detect the changes of the expression of the genes related with lung fibroblast transdifferentiation and proliferation (including Collagen-1
Collagen-3
α-SMA
fibronectin
FOXM1
CCNB1
CDC25b and AURKB) in human lung fibroblasts (HFL1) and primary rat lung fibroblasts in response to IR-61 and transforming growth factor-β1 (TGF-β1) treatments; the migration of the cells was assessed using cell scratch test.
Results
2
NIR imaging showed that IR-61 could target the injured lung tissue induced by bleomycin. Morphological observation showed that compared with bleomycin-treated rats
the rats with IR-61 treatment presented with better structural integrity of the lungs with less obvious lung collapse
fewer fibrotic nodules and a significantly milder of pulmonary fibrosis (0.103 3±0.014 53
vs
0.460 0±0.037 86
P
<
0.05). In HFL1 cells and primary rat lung fibroblasts
IR-61 pretreatment prior to TGF-β1 stimulation resulted in significantly lowered mRNA and protein expression of collagen-1
collagen-3
α-SMA
and fibronectin as well as the proliferation-specific genes FOXM1
CCNB1
CDC25b and AURKB (
P
<
0.05). Cell scratch test showed that TGF-β1 treatment significantly increased the cell migration activity
which was effectively suppressed by IR-61 treatment.
Conclusion
2
IR-61 ameliorates bleomycin-induced pulmonary fibrosis in rats by targeting the injured lung tissue and inhibiting the transdifferentiation
proliferation and migration of lung fibroblasts.
关键词
Keywords
references
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