TGF-β1-Licensed Murine MSCs Show Superior Therapeutic Efficacy in Modulating Corneal Allograft Immune Rejection In Vivo

Kevin Lynch,Oliver Treacy,Xizhe Chen,Nick Murphy,Paul Lohan,Md Nahidul Islam,Ellen Donohoe,Matthew D Griffin,Luke Watson,Steven Mcloughlin,Grace O’Malley,Aideen E Ryan,Thomas Ritter

doi:10.1016/j.ymthe.2020.05.023

Abstract

Mesenchymal stromal cells (MSCs) are a promising therapeutic option for multiple immune diseases/disorders; however, efficacy of MSC treatments can vary significantly. We present a novel licensing strategy to improve the immunosuppressive capacity of MSCs. Licensing murine MSCs with transforming growth factor-β1 (TGF-β MSCs) significantly improved their ability to modulate both the phenotype and secretome of inflammatory bone marrow-derived macrophages and significantly increased the numbers of regulatory T lymphocytes following co-culture assays. These TGF-β MSC-expanded regulatory T lymphocytes also expressed significantly higher levels of PD-L1 and CD73, indicating enhanced suppressive potential. Detailed analysis of T lymphocyte co-cultures revealed modulation of secreted factors, most notably elevated prostaglandin E2 (PGE2). Furthermore, TGF-β MSCs could significantly prolong rejection-free survival (69.2% acceptance rate compared to 21.4% for unlicensed MSC-treated recipients) in a murine corneal allograft model. Mechanistic studies revealed that (1) therapeutic efficacy of TGF-β MSCs is Smad2/3-dependent, (2) the enhanced immunosuppressive capacity of TGF-β MSCs is contact-dependent, and (3) enhanced secretion of PGE2 (via prostaglandin EP4 [E-type prostanoid 4] receptor) by TGF-β MSCs is the predominant mediator of Treg expansion and T cell activation and is associated with corneal allograft survival. Collectively, we provide compelling evidence for the use of TGF-β1 licensing as an unconventional strategy for enhancing MSC immunosuppressive capacity.

Highlights

Full-thickness corneal transplantation is a last resort for patients suffering from dystrophic, degenerative, infectious, or inflammatory corneal disorders.[1]
We observed a significant increase in CD73 expression following TGF-b licensing, which has particular relevance to the current study due to the reported role played by CD73 in immune tolerance.[41,42,43]
These data illustrate that TGF-b mesenchymal stromal cells (MSCs) have a particular phenotype that is distinguishable from unlicensed MSCs, and we sought to investigate how these cells would influence T lymphocytes in syngeneic co-culture assays

Summary

Introduction

Full-thickness corneal transplantation (penetrating keratoplasty) is a last resort for patients suffering from dystrophic, degenerative, infectious, or inflammatory corneal disorders.[1]. Despite the increased rate of uptake of these newer procedures, penetrating keratoplasty remains the most frequent keratoplasty procedure performed worldwide, for cases of deep-seated corneal infection and noninflammatory-associated deep stromal scars, with approximately 185,000 full-thickness corneal transplant procedures performed worldwide in 2012 alone.[1,2]. While topical corticosteroids with or without adjuvant immunosuppressant therapy remains the gold standard treatment option for prolongation of allograft survival and, tissue acceptance, patients are still highly susceptible to immune-mediated rejection.[3] Limbal stem cell (LSC) infusion therapy is a possible treatment option in some cases; widespread use is restricted due to limited availability of sufficient numbers of LSCs and is not possible for patients with bilateral LSC deficiency.[4] As a result, mesenchymal stromal cells (MSCs) have emerged as a viable therapeutic option due to the relative ease of isolating the cells and their high proliferation rates in vitro, generating high cell yields, as well as their immunomodulatory properties.[5]

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