Anti-inflammatory and immunomodulatory effects of human induced pluripotent stem cells-derived mesenchymal stem/stromal cells and their extracellular vesicles

IF 4 3区环境科学与生态学 Q3 CELL & TISSUE ENGINEERING

Regenerative Therapy Pub Date : 2026-03-01 Epub Date: 2026-02-26 DOI:10.1016/j.reth.2026.101081

Yusaku Komoike , Daisuke Kamiya , Haruka Sasaki-Ban , Atsushi Matsuda , Hirokazu Matsumoto , Makoto Ikeya

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引用次数: 0

Abstract

Introduction

Mesenchymal stem/stromal cells (MSCs) are multipotent cells that secrete multiple factors for tissue regeneration. These tissue-derived MSCs can self-renew, but their numbers are limited and decrease with age. The previously established xeno-free MSCs (XF-iMSCs), derived from human induced pluripotent stem cells (hiPSCs) of the neural crest cell (NCC) lineage, can regenerate damaged bone and skeletal muscle. However, the anti-inflammatory and immunomodulatory effects of XF-iMSCs have not been elucidated. Here, we aimed to elucidate the effects of XF-iMSCs and their extracellular vesicles (XF-iEv) on inflammation and immunomodulation.

Methods

XF-iMSCs were generated from hiPSCs using NCCs. Mouse PBMCs and splenocytes were obtained from male C57BL/6 mice. The MSCs were characterized using flow cytometry. Cytokine secretion stimulated by lipopolysaccharides (LPS) or Dynabeads CD3/CD28 was measured by ELISA. The proliferation of CellTrace Violet-labeled effector T cells (Teff) cultured with MSCs or their EVs was analyzed using a suppression assay. EVs were purified from the MSC culture medium using a MagCapture Exosome Isolation Kit. Proteome analysis of the EVs was performed using non-labeled liquid chromatography-tandem mass spectrometry.

Results

XF-iMSCs expressed representative MSC cell surface markers, including CD44, CD73, and CD105, but not CD45 or HLA-DR. XF-iMSCs and human adipocyte-derived MSCs (hAC-MSCs) suppressed LPS-stimulated IL-6 and TNF-α secretion in mouse PBMCs. Suppression of LPS-induced IL-6 and TNF-α secretion by XF-iEv and hAC-MSCs (hAC-Ev) was concentration-dependent. Fifty times-concentrated EVs from both XF-iMSCs and hAC-MSCs strongly suppressed IL-2, IFN-γ, and IL-17 secretion induced by dynabeads CD3/CD28 in mouse splenocytes. The effect of XF-iMSCs against inflammatory and anti-inflammatory cytokine production induced by LPS in hPBMCs was comparable to that of primary human adipocytes, bone marrow, and umbilical cord-derived MSCs. XF-iEv (x50) strongly inhibited TNF-α secretion from LPS-stimulated human PBMCs, although there was no effect on IL-6 secretion. Condition medium (CM) from XF-iMSCs promoted IL-10 secretion but not concentrated XF-iEv (x50). XF-iMSCs suppressed Teff proliferation to a level comparable to that of hAC-MSCs. XF-iEv had 1217 proteins with unique components compared to hAC-Ev.

Conclusions

XF-iMSCs and their EVs exert anti-inflammatory and immunomodulatory effects in human and mouse cell-based assays and have promising therapeutic applications for autoimmune diseases.

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人诱导多能干细胞来源的间充质干细胞/基质细胞及其细胞外囊泡的抗炎和免疫调节作用。

间充质干细胞（Mesenchymal stem/stromal cells, MSCs）是一种多能细胞，可分泌多种组织再生因子。这些组织来源的MSCs可以自我更新，但它们的数量有限，并随着年龄的增长而减少。先前建立的无xeno间充质干细胞（XF-iMSCs）来源于神经嵴细胞（NCC）谱系的人诱导多能干细胞（hiPSCs），可以再生受损的骨骼和骨骼肌。然而，XF-iMSCs的抗炎和免疫调节作用尚未被阐明。在这里，我们旨在阐明XF-iMSCs及其细胞外囊泡（XF-iEv）在炎症和免疫调节中的作用。方法：利用NCCs从hiPSCs中获得XF-iMSCs。从雄性C57BL/6小鼠获得小鼠pbmc和脾细胞。流式细胞术对MSCs进行表征。采用ELISA法检测脂多糖（LPS）或Dynabeads CD3/CD28刺激细胞因子的分泌。用抑制实验分析了细胞微量紫标记的效应T细胞（Teff）与间充质干细胞或其ev培养的增殖。使用MagCapture外泌体分离试剂盒从MSC培养基中纯化ev。采用非标记液相色谱-串联质谱法对ev进行蛋白质组学分析。结果：XF-iMSCs表达具有代表性的MSC细胞表面标记物，包括CD44、CD73和CD105，但不表达CD45或HLA-DR。XF-iMSCs和人脂肪细胞来源的MSCs （hAC-MSCs）抑制lps刺激的小鼠PBMCs中IL-6和TNF-α的分泌。XF-iEv和hAC-MSCs （hAC-Ev）对lps诱导的IL-6和TNF-α分泌的抑制呈浓度依赖性。50倍浓缩的XF-iMSCs和hAC-MSCs均能强烈抑制dynabeads CD3/CD28诱导的小鼠脾细胞IL-2、IFN-γ和IL-17的分泌。XF-iMSCs对LPS诱导的hPBMCs炎症和抗炎细胞因子产生的影响与原代人脂肪细胞、骨髓和脐带来源的MSCs相当。XF-iEv （x50）强烈抑制lps刺激的人外周血单核细胞分泌TNF-α，但对IL-6分泌没有影响。来自XF-iMSCs的条件培养基（CM）促进了IL-10的分泌，但没有浓缩XF-iEv （x50）。XF-iMSCs抑制Teff增殖的水平与hAC-MSCs相当。与hAC-Ev相比，XF-iEv具有1217种独特成分的蛋白。结论：XF-iMSCs及其EVs具有抗炎和免疫调节作用，在人类和小鼠细胞检测中具有良好的治疗自身免疫性疾病的应用前景。

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来源期刊

Regenerative Therapy Engineering-Biomedical Engineering

CiteScore

6.00

自引率

2.30%

发文量

106

审稿时长

49 days

期刊介绍： Regenerative Therapy is the official peer-reviewed online journal of the Japanese Society for Regenerative Medicine. Regenerative Therapy is a multidisciplinary journal that publishes original articles and reviews of basic research, clinical translation, industrial development, and regulatory issues focusing on stem cell biology, tissue engineering, and regenerative medicine.