Exosomes from high glucose-treated renal tubular epithelial cells induce macrophage-to-myofibroblast transformation
10.19405/j.cnki.issn1000-1492.2022.06.002
- Author:
Dong Wang
1
;
Yonggui Wu
2
;
Xiangming Qi
2
Author Information
1. Dept of Nephropathy , The First Afiliated Hospital , Anhui Medical University , Hefei 230022 ; Dept of Nephropathy , Hubei Suizhou Center Hospital , Suizhou 441300
2. Dept of Nephropathy , The First Afiliated Hospital , Anhui Medical University , Hefei 230022
- Publication Type:Journal Article
- Keywords:
exosomes;
macrophage;
myofibroblast
- From:
Acta Universitatis Medicinalis Anhui
2022;57(6):847-856
- CountryChina
- Language:Chinese
-
Abstract:
Abstract:To investigate the role and mechanism of macrophage-to-myofibroblast transition(MMT) induced by exosomes from high glucose-treated renal tubular epithelial cells.
Methods:Human renal tubular epithelial cells(HK2) were divided into glucose control group(5.5 mmol/L D-glucose) and high glucose group(30.0 mmol/L D-glucose) and cultured for 48 hours. The supernatant was collected by ultracentrifugation and identified by transmission electron microscopy and Western blot. PKH67 labelled exosomes were used to stimulate THP-1 macrophages. Laser scanning confocal microscopy was used to observe the phagocytosis process. The expression of inducible nitric oxide synthase(iNOS), α-SMA and CD206 was detected by flow cytometry and Western blot to determine the best concentration and time point. Laser scanning confocal microscopy was used to detect the fluorescence co-expression of rabbit polyclonal to mannose receptor(CD206), α-smooth muscle actin(α-SMA), collage type-Ⅳ(Col-IV) and fibronectin(FN). The expression of tumor growth factor-beta 1(TGF-β1), interleukin(IL)-6,IL-10 was separately detected by real-time quantitative PCR(qRT-PCR) and enzyme-linked immunosorbent assay(ELISA). The expression of TGF-β1, mothers against decapentaplegic homolog 3(Smad3), phosphorylated-smad 3(p-Smad3) in THP-1 macrophages was detected by Western blot.
Results:The expression of cluster of differentiation 63(CD63)and tumor susceptibility gene 101 protein(TSG101)was positive, while the Calnexin protein was negative in the supernatant, confirming that the specimen was exosomes with high purity. THP-1 macrophages could internalize each group of exosomes. 40 mg/L exosomes for 96 hours were the best experimental condition. After being stimulated by high-glucose-exosomes, the percentage of M1 macrophages would climax in 24 hours, and the rate of M2 macrophages would climax in 96 hours. Immunofluorescence staining showed that exosomes released by HG-treated HK2 induced CD206, α-SMA, Col-IV and FN accumulation in cultured THP-1 macrophages. Compared with normal-glucose-exosomes, high-glucose-exosomes increased the expression of TGF-β1, IL-10 in M2 macrophages and decreased the expression of IL-6(allP<0.05). Moreover, TGF-β1 and p-Smad3 proteins expression also increased significantly(allP<0.05).
Conclusion:Exosomes secreted by renal tubular epithelial cells in high glucose environment can induce M2 macrophages transdifferentiate into myofibroblasts, and its mechanism may be related to activation of TGF-β1/Smad3 signal pathway.
- Full text:2025022109062284245高糖环境下肾小管上皮细胞来...巨噬细胞向肌成纤维细胞转化_王东.pdf
