Institute of Allergy and Clinical Immunology, Seoul National University Medical Research Center, Seoul 03080, Korea.
Department of Translational Medicine, Seoul National University College of Medicine, Seoul 03080, Korea.
These authors contributed equally to this work.
Department of Internal Medicine, Gachon University Gil Medical Center, Incheon 21565, Korea.
Department of Internal Medicine, Seoul National University College of Medicine, Seoul 03080, Korea.
Department of Pharmacy, Jeju National University College of Pharmacy, Jeju 63243, Korea.
Department of Medical Science, Seoul National University College of Medicine, Seoul 03080, Korea.
Abstract
Although asthma is a common chronic airway disease that responds well to anti-inflammatory agents, some patients with asthma are unresponsive to conventional treatment. Mesenchymal stem cells (MSCs) have therapeutic potential for the treatment of inflammatory diseases owing to their immunomodulatory properties. However, the target cells of MSCs are not yet clearly known. This study aimed to determine the effect of human umbilical cord-derived MSCs (hUC-MSCs) on asthmatic lungs by modulating innate immune cells and effector T cells using a murine asthmatic model. Intravenously administered hUC-MSCs reduced airway resistance, mucus production, and inflammation in the murine asthma model. hUC-MSCs attenuated not only T helper (Th) 2 cells and Th17 cells but also augmented regulatory T cells (Tregs). As for innate lymphoid cells (ILC), hUC-MSCs effectively suppressed ILC2s by downregulating master regulators of ILC2s, such as Gata3 and Tcf7. Finally, regarding lung macrophages, hUC-MSCs reduced the total number of macrophages, particularly the proportion of the enhanced monocyte-derived macrophage population. In a closer examination of monocyte-derived macrophages, hUC-MSCs reduced the M2a and M2c populations. In conclusion, hUC-MSCs can be considered as a potential anti- asthmatic treatment given their therapeutic effect on the asthmatic airway inflammation in a murine asthma model by modulating innate immune cells, such as ILC2s, M2a, and M2c macrophages, as well as affecting Tregs and effector T cells.
Fig. 1 Effect of hUC-MSCs on AHR and lung inflammation in a murine asthma model. (A) Establishment of a murine asthma model with OVA and Alum administration. (B) AHR was measured 24 h after the last challenge. (C) The number of total inflammatory cells, including macrophages, neutrophils, eosinophils, and lymphocytes in BAL fluid. (D) Proportion of eosinophils and neutrophils to CD45+ cells in the lungs. (E) IL-5, IL-13, Il-17, and IFN-γ cytokine expression analysis by ELISA in BAL fluid. (F) H&E (100×) and PAS staining (100×) for lung histology. (G) Lung inflammation score and PAS score in a murine asthma model. (H) Muc5ac expression in lungs. n = 4 for each group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. All results are representative of at least three independent experiments. Alum, aluminum hydroxide; RL, resistance of lung; BAL, bronchoalveolar lavage; WT, wild type.Fig. 2 Effect of hUC-MSCs on effector T cell activation in vivo and ex vivo . The number of (A) IL-5+ CD4+ T cells, IL-13+ CD4+ T cells, (B) IL-17A+ CD4+ T cells and IFN-γ+ CD4+ T cells in the lungs and their dot plots. (C and D) Foxp3+CD25+CD4+ T cells among CD4+ T cells in the lungs and the spleen, and the mRNA expression of Foxp3 and Il10 with and without hUC-MSCs treatment in the lungs. (E) Experimental scheme and dot plots showing Tregs induced by hUC-MSCs during differentiation from naïve CD4+ T cells isolated from the spleen. (F) The effect of hUC-MSCs on ex vivo Tregs differentiated from naïve CD4+ T cells isolated from the spleen (control). n = 4 for each group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. All results are representative of three independent experiments.Fig. 3 Effect of hUC-MSCs on ILCs in vivo and ex vivo. (A) Dot plot and the number of IL-5+ ILCs, IL-13+ ILCs, (B) IL-17A+ ILCs and IFN-γ+ ILCs in the lung homogenate. (C) Protocol for differentiation of ILCs isolated from the lungs and co-culturing of hUC-MSCs in vitro, and mechanism-dependent changes in their activation. (D) The number of IL-5+ ILCs and IL-13+ ILCs due to hUC-MSCs treatment ex vivo. (E) Il5, Il13, Il10, (F) Gata3, Tcf7, (G) Il1rl1 and Il7r gene expression in ILC2s. Naïve ILCs isolated from the mouse lungs were used as the control. n = 4 for each group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. All results are representative of at least three independent experiments.Fig. 4 Effect of hUC-MSCs on lung mRNA expression in a murine asthma model. (A) Changes in Il5, Il13, Gata3, and Rora gene expression in the lungs following hUC-MSC treatment. (B) Changes in Il17a, Rorc, (C) Ifng and Tbx21 gene expression in the lungs following hUC-MSC treatment. (D and E) Mrc1, H2-ab, Ym1, Relma, Cd163, Arg1, Pparg, and Cd86 gene expression in the lungs following hUC-MSC treatment. (F) Mcp1 and Itgam gene expression in the lungs following hUC-MSC treatment. n = 4 for each group, **P < 0.01, ***P < 0.001, ****P < 0.0001. All results are representative of at least three independent experiments.Fig. 5 Effect of hUC-MSCs on macrophages in a murine asthma model. (A) Number of lung macrophages. (B) Dot plot showing lung AMs and MoMs population in vivo. (C) Change of SiglecF+CD11c+CD11b+− resident AMs and SiglecF+CD11c+CD11b+ transient AMs in the lungs. (D) Change of MoMs and their subtype in the lungs. All macrophage subtypes represent the ratio to total macrophages. n = 4 for each group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001; ns, not significant. All results are representative of at least three independent experiments. MoM, monocyte derived macrophage.Fig. 6 Effect of hUC-MSCs on ex vivo and in vitro macrophage mRNA expression. (A) Change of M1 and M2 MoMs in the lungs. (B) M2 macrophage subtype changes in the lungs induced by hUC-MSCs. (C) Changes in M2 activation of mouse AMs MH-S (CRL-2019) in vitro by hUC-MSCs. Unstimulated mouse AMs were used as controls. (D) Changes in M2 activation of human PBMC-derived macrophage direct by hUC-MSC treatment. M1 and M2 MoM subtypes represent the ratio to total MoMs. The unstimulated human PBMC-derived macrophages were used as the control. n = 4 for each group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. All results are representative of at least three independent experiments.
그림 출처: PubMed Central 공개 논문 (PMC9676992)
Conflict of interest statement
CONFLICT OF INTEREST . The authors have no potential conflicts of interest to disclose.
