Human mesenchymal stem cells-derived conditioned medium inhibits hypoxia-induced death of neonatal porcine islets by inducing autophagy

Yuzhi Xu; Mengqun Tan; Xiaoqian Ma; Hongde Li; Xuesong He; Zeyi Chen; Yixiong Tan; Wei Nie; Pengfei Rong; Wei Wang

doi:10.1111/xen.12556

Human mesenchymal stem cells-derived conditioned medium inhibits hypoxia-induced death of neonatal porcine islets by inducing autophagy

Xenotransplantation. 2020 Jan;27(1):e12556. doi: 10.1111/xen.12556. Epub 2019 Oct 2.

Authors

Yuzhi Xu^{1

2}, Mengqun Tan³, Xiaoqian Ma^{1

3}, Hongde Li^{1

2}, Xuesong He³, Zeyi Chen³, Yixiong Tan^{1

2}, Wei Nie^{1

3}, Pengfei Rong^{1

2}, Wei Wang^{1

3}

Affiliations

¹ Cell Transplantation and Gene Therapy Institute, The Third Xiang Ya Hospital, Central South University, Changsha, China.
² Department of Radiology, The Third Xiang Ya Hospital of Central South University, Changsha, China.
³ Engineering and Technology Research Center for Xenotransplantation of Human Province, Changsha, China.

PMID: 31578787
DOI: 10.1111/xen.12556

Abstract

Background: The dysfunction of islet grafts is generally attributed to hypoxia-induced damage. Mesenchymal stem cells (MSCs) are currently thought to effectively protect cells from various risk factors via regulating autophagy. In our study, we investigated if human umbilical cord-derived MSCs could ameliorate hypoxia-induced apoptosis in porcine islets by modulating autophagy, and we explored the underlying mechanisms.

Methods: Neonatal porcine islet cell clusters (NICCs) were cultured with human umbilical cord-derived MSC conditioned medium (huc-MSC-CM) and RPMI-1640 medium (control) under hypoxic conditions (1% O₂ ) in vitro. NICCs were treated with 3-methyladenine (3-MA) and chloroquine (CQ) to examine the role of huc-MSC-CM in regulating autophagy. Finally, the levels of several cytokines secreted by huc-MSCs were detected by ELISAs, and the corresponding inhibitors were applied to investigate which cytokine mediates the protective effects of huc-MSC-CM. The effects of huc-MSC-CM on NICCs viability and autophagy were examined using AO/PI staining, flow cytometry analysis, transmission electron microscopy (TEM) and confocal fluorescence microscopy analysis. The insulin secretion of NICCs was tested with an insulin immunoradiometric assay kit.

Results: Compared to the control, the huc-MSC-CM treatment improved the viability of NICCs, inhibited apoptosis, increased autophagic activity and the levels of PI3K class III and phosphorylated Akt, while the ratio of phosphorylated mTOR/mTOR was reduced. These changes were reversed by CQ and 3-MA treatments. High concentrations of IL-6 were detected in hu-MSC-CM. Furthermore, recombinant IL-6 pre-treatment exerted similar effects as huc-MSC-CM, and these effects were reversed by a specific inhibitor of IL-6 (Sarilumab).

Conclusions: Our results demonstrated that huc-MSC-CM improved islet viability and function by increasing autophagy through the PI3K/Akt/mTOR pathway under hypoxic conditions. Additionally, IL-6 plays an important role in the function of huc-MSC-CM.

Keywords: apoptosis; autophagy; hypoxia; mesenchymal stem cells; neonatal porcine islet cell clusters.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Animals, Newborn
Autophagy
Cell Death
Cells, Cultured
Culture Media, Conditioned
Humans
Hypoxia / metabolism*
Islets of Langerhans / physiology*
Mesenchymal Stem Cells / physiology*
Phosphatidylinositol 3-Kinases / metabolism
Proto-Oncogene Proteins c-akt / metabolism
Signal Transduction
Swine
TOR Serine-Threonine Kinases / metabolism

Substances

Culture Media, Conditioned
Proto-Oncogene Proteins c-akt
TOR Serine-Threonine Kinases