Mesenchymal stromal/stem cells promote intestinal epithelium regeneration after chemotherapy-induced damage

Abstract

Background Allogeneic hematopoietic stem cell transplantation (HSCT) is a curative treatment for leukemia and a range of non-malignant disorders. The success of the therapy is hampered by occurrence of acute graft-versus-host disease (aGvHD); an inflammatory response damaging recipient organs, with gut, liver, and skin being the most susceptible. Intestinal GvHD injury is often a life-threatening complication in patients unresponsive to steroid treatment. Allogeneic mesenchymal stromal/stem cell (MSC) infusions are a promising potential treatment for steroid-resistant aGvHD. Data from our institution and others demonstrate rescue of approximately 40–50% of aGvHD patients with MSCs in Phase I, II studies and minor side effects. Although promising, better understanding of MSC mode of action and patient response to MSC-based therapy is essential to improve this lifesaving treatment. Methods Single cell human small intestine organoids were embedded in Matrigel, grown for 5 days and treated with busulfan for 48 h. Organoids damaged by treatment with busulfan or control organoids were co-cultured with 5000, 10,000, and 50,000 MSCs for 24 h, 48 h or 7 days and the analyses such as surface area determination, proliferation and apoptosis assessment, RNA sequencing and proteomics were performed. Results Here, we developed a 3D co-culture model of human small intestinal organoids and MSCs, which allows to study the regenerative effects of MSCs on intestinal epithelium in a more physiologically relevant setting than existing in vitro systems. Using this model we mimicked chemotherapy-mediated damage of the intestinal epithelium. The treatment with busulfan, the chemotherapeutic commonly used as conditioning regiment before the HSCT, affected pathways regulating epithelial to mesenchymal transition, proliferation, and apoptosis in small intestinal organoids, as shown by transcriptomic and proteomic analysis. The co-culture of busulfan-treated intestinal organoids with MSCs reversed the effects of busulfan on the transcriptome and proteome of intestinal epithelium, which we also confirmed by functional evaluation of proliferation and apoptosis. Conclusions Collectively, we demonstrate that our in vitro co-culture system is a new valuable tool to facilitate the investigation of the molecular mechanisms behind the therapeutic effects of MSCs on damaged intestinal epithelium. This could benefit further optimization of the use of MSCs in HSCT patients.

1. Introduction

Allogeneic hematopoietic stem cell transplantation (HSCT) is used as a treatment for a variety of acquired and inherited disorders of the hematopoietic system, including inborn errors of metabolism, disorders of the immune system, and hematologic malignancies (e.g. leukemia and lymphoma). An essential component of HSCT is the conditioning regimen, consisting of chemotherapy and/or total body irradiation, administrated prior to hematopoietic cell infusion. The purpose of the conditioning is to target any residual leukemic disease, to provide sufficient myeloablation for engraftment and enough immunoablation to prevent rejection against the transplanted graft. However, the conditioning also contributes to the development of a major complication of allogeneic HSCT, acute graft-versus-host-disease (aGvHD), in which immune cells from the donor attack healthy recipient tissue, including liver, skin, and gut [1–3]. Especially intestinal aGvHD is a life-threatening complication. Recent findings suggest that GvHD development in the gut is enhanced by prior chemotherapy-mediated epithelial damage [4]. This was confirmed in in vitro experiments whereby chemotherapy-mediated epithelial damage increased T cell proliferation and activation [4].

First-line treatment of aGvHD is based on systemic corticosteroids that cause immunosuppression to prevent fatal disease progression [5]. However, approximately 50% of aGvHD patients become refractory to corticosteroid treatment, resulting in high morbidity and mortality rates, and low quality of life in these patients [6–8]. Many strategies have been used for a second-line treatment of steroid refractory aGvHD patients.