Liyang Dong1, Ying Wang2, Tingting Zheng3, Yanan Pu4, Yongbin Ma4,5, Xin Qi4, Wenzhe Zhang6, Fei Xue6, Zirui Shan6, Jiameng Liu3, Xuefeng Wang7,8, Chaoming Mao9. 1. Department of Nuclear Medicine, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, 212000, People's Republic of China. dongliyang0829@hotmail.com. 2. Department of Respiratory Diseases, The Affiliated Huai'an Hospital of Xuzhou Medical University, Huai'an, Jiangsu, 223002, People's Republic of China. 3. Department of Nuclear Medicine, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, 212000, People's Republic of China. 4. Jiangsu Key Laboratory of Pathogen Biology, Department of Pathogen Biology and Immunology, Nanjing Medical University, Nanjing, Jiangsu, 211166, People's Republic of China. 5. Department of Neurology Laboratory, The Affiliated Jintan Hospital of Jiangsu University, Jintan, Jiangsu, 213200, People's Republic of China. 6. Department of Central Laboratory, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, 212000, People's Republic of China. 7. Department of Nuclear Medicine, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, 212000, People's Republic of China. xuefengwang@ujs.edu.cn. 8. Department of Central Laboratory, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, 212000, People's Republic of China. xuefengwang@ujs.edu.cn. 9. Department of Nuclear Medicine, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, 212000, People's Republic of China. jq1001@ujs.edu.cn.
Abstract
BACKGROUND: As one of the main functional forms of mesenchymal stem cells (MSCs), MSC-derived extracellular vesicles (MSC-EVs) have shown an alternative therapeutic option in experimental models of allergic asthma. Oxygen concentration plays an important role in the self-renewal, proliferation, and EV release of MSCs and a recent study found that the anti-asthma effect of MSCs was enhanced by culture in hypoxic conditions. However, the potential of hypoxic MSC-derived EVs (Hypo-EVs) in asthma is still unknown. METHODS: BALB/c female mice were sensitized and challenged with ovalbumin (OVA), and each group received PBS, normoxic human umbilical cord MSC-EVs (Nor-EVs), or Hypo-EVs weekly. After treatment, the animals were euthanized, and their lungs and bronchoalveolar lavage fluid (BALF) were collected. With the use of hematoxylin and eosin (HE), periodic acid-Schiff (PAS) and Masson's trichrome staining, enzyme-linked immune sorbent assay (ELISA), Western blot analysis, and real-time PCR, the inflammation and collagen fiber content of airways and lung parenchyma were investigated. RESULTS: Hypoxic environment can promote human umbilical cord MSCs (hUCMSCs) to release more EVs. In OVA animals, the administration of Nor-EVs or Hypo-EVs significantly ameliorated the BALF total cells, eosinophils, and pro-inflammatory mediators (IL-4 and IL-13) in asthmatic mice. Moreover, Hypo-EVs were generally more potent than Nor-EVs in suppressing airway inflammation in asthmatic mice. Compared with Nor-EVs, Hypo-EVs further prevented mouse chronic allergic airway remodeling, concomitant with the decreased expression of pro-fibrogenic markers α-smooth muscle actin (α-SMA), collagen-1, and TGF-β1-p-smad2/3 signaling pathway. In vitro, Hypo-EVs decreased the expression of p-smad2/3, α-SMA, and collagen-1 in HLF-1 cells (human lung fibroblasts) stimulated by TGF-β1. In addition, we showed that miR-146a-5p was enriched in Hypo-EVs compared with that in Nor-EVs, and Hypo-EV administration unregulated the miR-146a-5p expression both in asthma mice lung tissues and in TGF-β1-treated HLF-1. More importantly, decreased miR-146a-5p expression in Hypo-EVs impaired Hypo-EV-mediated lung protection in OVA mice. CONCLUSION: Our findings provided the first evidence that hypoxic hUCMSC-derived EVs attenuated allergic airway inflammation and airway remodeling in chronic asthma mice, potentially creating new avenues for the treatment of asthma.
BACKGROUND: As one of the main functional forms of mesenchymal stem cells (MSCs), MSC-derived extracellular vesicles (MSC-EVs) have shown an alternative therapeutic option in experimental models of allergic asthma. Oxygen concentration plays an important role in the self-renewal, proliferation, and EV release of MSCs and a recent study found that the anti-asthma effect of MSCs was enhanced by culture in hypoxic conditions. However, the potential of hypoxic MSC-derived EVs (Hypo-EVs) in asthma is still unknown. METHODS: BALB/c female mice were sensitized and challenged with ovalbumin (OVA), and each group received PBS, normoxic human umbilical cord MSC-EVs (Nor-EVs), or Hypo-EVs weekly. After treatment, the animals were euthanized, and their lungs and bronchoalveolar lavage fluid (BALF) were collected. With the use of hematoxylin and eosin (HE), periodic acid-Schiff (PAS) and Masson's trichrome staining, enzyme-linked immune sorbent assay (ELISA), Western blot analysis, and real-time PCR, the inflammation and collagen fiber content of airways and lung parenchyma were investigated. RESULTS: Hypoxic environment can promote human umbilical cord MSCs (hUCMSCs) to release more EVs. In OVA animals, the administration of Nor-EVs or Hypo-EVs significantly ameliorated the BALF total cells, eosinophils, and pro-inflammatory mediators (IL-4 and IL-13) in asthmatic mice. Moreover, Hypo-EVs were generally more potent than Nor-EVs in suppressing airway inflammation in asthmatic mice. Compared with Nor-EVs, Hypo-EVs further prevented mouse chronic allergic airway remodeling, concomitant with the decreased expression of pro-fibrogenic markers α-smooth muscle actin (α-SMA), collagen-1, and TGF-β1-p-smad2/3 signaling pathway. In vitro, Hypo-EVs decreased the expression of p-smad2/3, α-SMA, and collagen-1 in HLF-1 cells (human lung fibroblasts) stimulated by TGF-β1. In addition, we showed that miR-146a-5p was enriched in Hypo-EVs compared with that in Nor-EVs, and Hypo-EV administration unregulated the miR-146a-5p expression both in asthmamice lung tissues and in TGF-β1-treated HLF-1. More importantly, decreased miR-146a-5p expression in Hypo-EVs impaired Hypo-EV-mediated lung protection in OVAmice. CONCLUSION: Our findings provided the first evidence that hypoxic hUCMSC-derived EVs attenuated allergic airway inflammation and airway remodeling in chronic asthmamice, potentially creating new avenues for the treatment of asthma.
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