Immunosuppressive effect of mesenchymal stem cell-derived exosomes on a concanavalin A-induced liver injury model

Author: Ryo Tamura, Shinji Uemoto and Yasuhiko Tabata

Affiliation: Department of Surgery, Graduate School of Medicine, Kyoto University,Kyoto, Japan.

Department of Biomaterials, Field of Tissue Engineering,Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan.

Inflammation and Regeneration201636:26DOI: 10.1186/s41232-016-0030-5

©  The Author(s) 2016Received: 24 August 2016

Accepted: 1 November 2016Published: 17 November 2016

Immunosuppressive effect of mesenchymal stem cell-derived exosomes on a

concanavalin A-induced liver injury modelCourse Title: Stem Cells and Tissue Engineering

Course Code: GEB427Semester: Spring-2017

Course instructor: Roushney Fatima MuktiLecturer

Department of Genetic Engineering & Biotechnology, East West University

 Presented by:1. Zannat Mahal2. Sumaiya Binte Salam3. Md. Shabab Mehebub

Department of Genetic Engineering & Biotechnology, East West University

Key-wordsMSCs = Mesenchymal Stem Cells

ALT = Alanine Aminotransferase (used to detect liver injury)

Con A = Concavalin A (directly injecting it into mice, induces T-cell mediated liver injury)

NPC = Non-parenchymal liver cell

Cytometry = a process for cell sorting

PKH- Fluorescent label of live cells over an extended period of time

KI-67- a cellular marker for proliferation to observe cell growth.

INTRODUCTION

In case of a successful organ transplantation, it is crucial to control organ rejection by the host body.

Many immunosuppressive drugs have been developed to prevent the rejection but they require long term follow-up of transplant recipient.

Mesenchymal stem cell can be applied as promising immune modulator to minimize the use of immunosuppressants.

As MSC functions are mediated through the secretion of trophic factors in exosomes, MSC-derived exosomes have been reported as a non-cellular alternative to MSC for transplantation therapy.

Objective In this study, the suppressive effect of exosomes was investigated on

an immune-induced liver injury model.

The liver injury was induced by injection of concanavalin A (conA) .

The suppressive effects of MSC-derived exosomes and the MSCs in this model were indentified by the level of plasma ALT, histopathological examination, the mRNA expression of pro- and anti-inflammatory cytokines and the population alteration of Treg among NPCs.

Additionally, in vivo tracing with fluorescent lebeled exosomes and MSCs was performed in liver tissue.

MethodsAnimals and cultivation of MSCs and fibroblasts

Characterization of MSC and NPC

Collection of exosomes

Characterization of exosomes collected

Fluorescent labelling of exosomes and MSCs with PKH26

Preparation of the mouse liver injury model and administration of exosomes and MSCs

Sample collection and immunohistochemical staining of liver specimens

mRNA expression analysis of inflammatory cytokines

Statistical analysis

Results

Characterization of MSCs and exosomes

In vivo accumulation of PKH26-labeled exosomes and MSCs and co-localization of PKH

In vivo effect of exosomes derived from MSCs and fibroblasts

mRNA expression of pro- and anti-inflammatory cytokines and flow cytometric analysis of NPCs

Fig. Characterization of MSCs and exosomes. Differentiation ofMSCs into three cellular lineages. i) Microscopy images of MSCs, ii)

MSCs stained with oil red O at 7 days after adipogenic differentiationculture, iii) MSCs stained with alizarin red staining at 30 days afterosteogenic differentiation culture, and iv) MSCs stained with alcianblue staining at 18 days after chondrogenic differentiation culture.

Results

Fig. Characterization of exosomes collected. Collection procedure for exosomes (left) & Size distribution of exosomes (right)

Results

Fig. In vivo tracing of PKH26 fluorescent dye-labeled MSCs and exosomes. a Localization of PKH26 dye after the injection of PKH26- labeled MSC (arrow). White box showed the magnified image of the place where PKH was observed in the lung. b Presence of PKH26 dye in the liver specimen and their co-localization with FITC-stained F4/80-positive cells after the injection of PKH26-labeled exosomes (arrow). i) PKH26 after the injection of PKH26 labeled-exosomes, ii) FITC-stained F4/80-positive cells

Results

Discussion • Even though, MSC transplantation is considered to be a promising

adjunctive therapy, reports of unintended differentiation of cells have been reported.

• Exosomes, being non-cellular, bear no such problem.• Pediatric recipients of organ transplantation require long-term

immune control, and it is strongly desired to develop a sustainable and adjustable treatment free from any adverse effects. The use of MSC-derived exosomes has major advantages to meet these requirements and may become a promising noncellular alternative to MSCs for transplantation therapy.

Conclusion• MSC-derived exosomes have been shown to demonstrate

suppressive effect on an immune-mediated liver injury model and have some advantages over MSCs and might become a promising alternative to MSCs for transplantation therapy.

Thank You for your

Patience!