Adipose-Derived Mesenchymal Stem Cells Protect Endothelial Cells from Hypoxic Injury by Suppressing Terminal UPR In Vivo and In Vitro

Michael Keese; Jiaxing Zheng; Kaixuan Yan; Karen Bieback; Benito A Yard; Prama Pallavi; Christoph Reissfelder; Mark Andreas Kluth; Martin Sigl; Vugar Yugublu

doi:10.3390/ijms242417197

Adipose-Derived Mesenchymal Stem Cells Protect Endothelial Cells from Hypoxic Injury by Suppressing Terminal UPR In Vivo and In Vitro

Int J Mol Sci. 2023 Dec 6;24(24):17197. doi: 10.3390/ijms242417197.

Authors

Michael Keese^{1

2

3}, Jiaxing Zheng^{1

2}, Kaixuan Yan¹, Karen Bieback⁴, Benito A Yard⁵, Prama Pallavi^{1

2}, Christoph Reissfelder^{1

6}, Mark Andreas Kluth⁷, Martin Sigl⁸, Vugar Yugublu¹

Affiliations

¹ Department of Surgery, Medical Centre Mannheim, Medical Faculty Manheim, Heidelberg University, 68167 Mannheim, Germany.
² European Center of Angioscience (ECAS), Medical Faculty Manheim, Heidelberg University, 68167 Mannheim, Germany.
³ Department for Vascular Surgery, Theresienkrankenhaus Mannheim, 68165 Mannheim, Germany.
⁴ Institute of Transfusion Medicine and Immunology, Medical Faculty Manheim, Heidelberg University, 68167 Mannheim, Germany.
⁵ V Department of Medicine, Medical Faculty Manheim, Heidelberg University, 68167 Mannheim, Germany.
⁶ DKFZ-Hector Cancer Institute, Medical Faculty Mannheim, Heidelberg University, 68167 Mannheim, Germany.
⁷ RHEACELL GmbH & Co. KG, Im Neuenheimer Feld 517, 69120 Heidelberg, Germany.
⁸ Department of Cardiology, Angiology, Haemostaseology and Medical Intensive Care, University Medical Centre Mannheim, Medical Faculty Mannheim, Heidelberg University, 68167 Mannheim, Germany.

Abstract

Adipose-derived stem cells (ASCs) have been used as a therapeutic intervention for peripheral artery disease (PAD) in clinical trials. To further explore the therapeutic mechanism of these mesenchymal multipotent stromal/stem cells in PAD, this study was designed to test the effect of xenogeneic ASCs extracted from human adipose tissue on hypoxic endothelial cells (ECs) and terminal unfolded protein response (UPR) in vitro and in an atherosclerosis-prone apolipoprotein E-deficient mice (ApoE^-/- mice) hindlimb ischemia model in vivo. ASCs were added to Cobalt (II) chloride-treated ECs; then, metabolic activity, cell migration, and tube formation were evaluated. Fluorescence-based sensors were used to assess dynamic changes in Ca²⁺ levels in the cytosolic- and endoplasmic reticulum (ER) as well as changes in reactive oxygen species. Western blotting was used to observe the UPR pathway. To simulate an acute-on-chronic model of PAD, ApoE^-/- mice were subjected to a double ligation of the femoral artery (DLFA). An assessment of functional recovery after DFLA was conducted, as well as histology of gastrocnemius. Hypoxia caused ER stress in ECs, but ASCs reduced it, thereby promoting cell survival. Treatment with ASCs ameliorated the effects of ischemia on muscle tissue in the ApoE^-/- mice hindlimb ischemia model. Animals showed less muscle necrosis, less inflammation, and lower levels of muscle enzymes after ASC injection. In vitro and in vivo results revealed that all ER stress sensors (BIP, ATF6, CHOP, and XBP1) were activated. We also observed that the expression of these proteins was reduced in the ASCs treatment group. ASCs effectively alleviated endothelial dysfunction under hypoxic conditions by strengthening ATF6 and initiating a transcriptional program to restore ER homeostasis. In general, our data suggest that ASCs may be a meaningful treatment option for patients with PAD who do not have traditional revascularization options.

Keywords: adipose-derived stem cells; critical limb ischemia; endoplasmic reticulum; hypoxia; peripheral artery disease; unfolded protein response.

MeSH terms

Adipose Tissue / metabolism
Animals
Apolipoproteins E / genetics
Apolipoproteins E / metabolism
Endothelial Cells* / metabolism
Humans
Hypoxia / metabolism
Ischemia / metabolism
Mesenchymal Stem Cells* / metabolism
Mice
Neovascularization, Physiologic / physiology
Unfolded Protein Response

Substances

Apolipoproteins E

Grants and funding

This research was financed by TICEBA GmbH.