Abstract
BackgroundBone marrow mesenchymal stem cell (BM-MSC) has been shown to treat pulmonary arterial hypertension (PAH). However, excessive reactive oxygen species (ROS) increases the apoptosis of BM-MSCs, leading to poor survival and engraft efficiency. Thus, improving the ability of BM-MSCs to scavenge ROS may considerably enhance the effectiveness of transplantation therapy. Mammalian Ste20-like kinase 1 (Mst1) is a pro-apoptotic molecule which increases ROS production. The aim of this study is to uncover the underlying mechanisms the effect of Mst1 inhibition on the tolerance of BM-MSCs under H2O2 condition.MethodsMst1 expression in BM-MSCs was inhibited via transfection with adenoviruses expressing a short hairpin (sh) RNA directed against Mst1 (Ad-sh-Mst1) and exposure to H2O2. Cell viability was detected by Cell Counting Kit 8 (CCK-8) assay, and cell apoptosis was analyzed by Annexin V-FITC/PI, Caspase 3 Activity Assay kits, and pro caspase 3 expression. ROS level was evaluated by the ROS probe DCFH-DA, mitochondrial membrane potential (ΔΨm) assay, SOD1/2, CAT, and GPx expression. Autophagy was assessed using transmission electron microscopy, stubRFP-sensGFP-LC3 lentivirus, and autophagy-related protein expression. The autophagy/Keap1/Nrf2 signal in H2O2-treated BM-MSC/sh-Mst1 was also measured.ResultsMst1 inhibition reduced ROS production; increased antioxidant enzyme SOD1/2, CAT, and GPx expression; maintained ΔΨm; and alleviated cell apoptosis in H2O2-treated BM-MSCs. In addition, this phenomenon was closely correlated with the autophagy/Keap1/Nrf2 signal pathway. Moreover, the antioxidant pathway Keap1/Nrf2 was also blocked when autophagy was inhibited by the autophagy inhibitor 3-MA. However, Keap1 or Nrf2 knockout via siRNA had no effect on autophagy activation or suppression.ConclusionMst1 inhibition mediated the cytoprotective action of mBM-MSCs against H2O2-induced oxidative stress injury. The underlying mechanisms involve autophagy activation and the Keap1/Nrf2 signal pathway.Graphical abstract
Highlights
Mesenchymal stem cell (MSC)-based therapies have been investigated for pulmonary arterial hypertension (PAH) treatment due to the “homing” ability [1, 2]
We investigate the modulation of mouse Bone marrow mesenchymal stem cell (BM-MSC) via Mammalian Ste20-like kinase 1 (Mst1) expression downregulation under H2O2 conditions and determine its underlying mechanisms
The results indicated that the isolated mBM-MSCs can be used for the subsequent experiments
Summary
Mesenchymal stem cell (MSC)-based therapies have been investigated for pulmonary arterial hypertension (PAH) treatment due to the “homing” ability [1, 2]. Increasing evidence supports the enhanced generation of pathological levels of reactive oxygen species (ROS) at the injured pulmonary vasculature in PAH [3]. Excessive ROS induces MSC apoptosis, leading to poor survival and engraft efficiency [7, 8]. Improving the ROS scavenging ability of MSCs is essential to promote MSC engraftment and enhance tissue repair. Bone marrow mesenchymal stem cell (BM-MSC) has been shown to treat pulmonary arterial hypertension (PAH). Excessive reactive oxygen species (ROS) increases the apoptosis of BM-MSCs, leading to poor survival and engraft efficiency. Improving the ability of BM-MSCs to scavenge ROS may considerably enhance the effectiveness of transplantation therapy. The aim of this study is to uncover the underlying mechanisms the effect of Mst inhibition on the tolerance of BM-MSCs under H2O2 condition
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