This study investigated the immunosuppressive effects of mesenchymal stem cell (MSC)-derived exosomes in a mouse model of liver injury induced by concanavalin A (Con A). Exosomes were isolated from MSCs and characterized. Mice received injections of MSC exosomes or MSCs labeled with fluorescent dye after Con A injection. Results showed MSC exosomes localized to injured livers and reduced liver damage and inflammation similar to MSCs. The study suggests MSC exosomes may be a promising non-cellular alternative to MSCs for transplantation therapy by controlling immune responses with fewer safety concerns than live MSCs.

1. 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 Regeneration201636:26
DOI: 10.1186/s41232-016-0030-5
© The Author(s) 2016
Received: 24 August 2016
Accepted: 1 November 2016
Published: 17 November 2016

2. Immunosuppressive effect of mesenchymal stemcell-derived exosomes on a
concanavalin A-induced liver injury model
Course Title: Stem Cells and Tissue Engineering
Course Code: GEB427
Semester: Spring-2017
Course instructor: Roushney Fatima Mukti
Lecturer
Department of Genetic Engineering & Biotechnology,
East West University
Presented by:
1. Zannat Mahal
2. Sumaiya Binte Salam
3. Md. Shabab Mehebub
Department of Genetic Engineering & Biotechnology,
East West University

4. Key-words
MSCs = 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.

5. 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.

6. 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.

7. Methods
Animals 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

8. 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

9. Fig. Characterization of MSCs and exosomes. Differentiation of
MSCs into three cellular lineages. i) Microscopy images of MSCs, ii)
MSCs stained with oil red O at 7 days after adipogenic differentiation
culture, iii) MSCs stained with alizarin red staining at 30 days after
osteogenic differentiation culture, and iv) MSCs stained with alcian
blue staining at 18 days after chondrogenic differentiation culture.
Results

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

11. 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

12. 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.

13. 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.