Engraftment and tolerance induction in cellular therapy : focus on mesenchymal stromal cells

Different cell types have been employed in the search for a cellular therapy that is applicable for the treatment of heart failure. Remuscularization strategies using stem-cell-derived cardiomyocytes have been prone to arrhythmias. Mesenchymal stromal cells (MSCs) are one of the most promising cell types for the treatment of heart failure due to their diverse regenerative properties. We have isolated MSCs from the human fetal heart in the form of human fetal cardiac MSCs (hfcMSCs), which might comprise a more relevant cell source than bone marrow-derived MSCs, for the treatment of heart failure. Although the cell source is of importance for successful cell therapy, the efficacy of the cells also depends on avoiding anchorage-dependent apoptosis (anoikis) and immune rejection. Immune rejection may be counteracted with immunosuppressive drugs; however, these drugs generally have serious side effects that limit their suitability in cell therapy. We have used a transient blockade of T cell costimulation with the aim to induce immunologic tolerance through the generation of Regulatory T cells (Tregs) specifically for the antigens of the transplant. Anoikis may be diminished by the simultaneous administration of the exogenous extracellular matrix (ECM), which provides anchorage possibilities for the transplanted cells.

In Paper I and II the use of costimulation blockade in xenogeneic and allogeneic mouse models was explored and we showed that Foxp3+ Tregs are associated with tolerated and viable grafts as well as reduced reactivity in mixed lymphocyte reaction tests. MSCs, known for their immunomodulatory properties, were shown to synergize with costimulation blockade to facilitate immunological tolerance to allogeneic insulin-producing islets in mice. In Paper III we showed that Matrigel synergizes with costimulation blockade to enable enhanced cell retention of hfcMSCs after subcutaneous implantation in mice. The hfcMSCs displayed vasculogenic potential in vivo, shown as the expression of CD31, and also the formation of vessel-like structures staining positive for laminin a4. In Paper IV, hfcMSCs were cultured on a 3D scaffold composed of a novel type of artificial spider silk, NT2RepCT, to conserve the cell connections and ECM at the time of implantation. The hfcMSCs displayed vasculogenic potential in the 3D cultures on NT2RepCT as well, with the formation of an ECM that stained positive for laminin a4 and fibronectin and contained vessel-like structures expressing CD31. After in vivo implantation, the hfcMSCs on the NT2RepCT were subjected to immune rejection regardless of costimulation blockade or isotype control treatment.

In this thesis hfcMSCs were shown to have a vasculogenic potential as they also deposit laminins that are important for vascular structures and thereby may have the potential to prevent ischemic heart failure. We also demonstrated a synergistic effect between costimulation blockade and MSCs or Matrigel for enhanced cell retention. Artificial spider silk showed great potential for advanced in vitro studies with 3D cultures, but could not be used in vivo in its current form.

List of scientific papers

I. Costimulation Blockade Induces FoxP3+ Regulatory T Cells to Human Embryonic Stem Cells. Karin Ljung, Oscar E. Simonson, Ulrika Felldin, Ewa Wärdell, Cristian Ibarra, Liselotte Antonsson, Makiko Kumagai-Braesch, Outi Hovatta, Riina Lampela, Karl-Henrik Grinnemo, and Matthias Corbascio. Bioresearch Open Access. 2013; 2(6):455-458.
https://doi.org/10.1089/biores.2013.0039

II. Multipotent Mesenchymal Stromal Cells Synergize with Costimulation Blockade in the Inhibition of the Immune Responses and the Induction of FoxP3+ Regulatory T Cells. Tohru Takahashi, Annika Tibell, Karin Ljung, Yu Saito, Anna Grönlund, Cecilia Österholm, Jan Holgersson, Torbjörn Lundgren, Bo-Göran Ericzon, Matthias Corbacio, Makiko Kumagai-Braesch. Stem Cells Translational Medicine. 2014; 3(12): 1484-1494.
https://doi.org/10.5966/sctm.2014-0012

III. Human Fetal Cardiac Mesenchymal Stromal Cells Differentiate in vivo into Endothelial Cells and Contribute to Vaculogenesis in Immunologically Competent Mice. Karin Ljung, Anna Grönlund, Ulrika Felldin, Sergey Rodin, Matthias Corbascio, Cecilia Österholm, Karl-Henrik Grinnemo. [Submitted]

IV. Artificial Spider Silk Provides a 3D Scaffold for the Growth, Vasculogenesis and Matrix Formation of Human Fetal Cardiac Mesenchymal Stromal Cells. Karin Ljung, Marlene Andersson, Lotta Floderus, Kerstin Nordling, Matthias Corbascio, Jan Johansson, Cecilia Österholm, Karl-Henrik Grinnemo, Anna Rising. [Manuscript]