Ex vivo Pretreatment of Islets with Mitomycin C

Abstract

Strategies to reduce the immunogenicity of pancreatic islets and to prevent the activation of proinflammatory events are essential for successful islet engraftment. Pretransplant islet culture presents an opportunity for preconditioning to improve outcomes of islet transplantation. We previously demonstrated that ex vivo mitomycin C (MMC) pretreatment and subsequent culture significantly prolonged graft survival. Fully understanding the biological process of pretreatment could result in the development of a protocol to improve the survival of islet grafts. Microarrays were employed to conduct a comprehensive analysis of genes expressed in untreated or MMC-treated rat islets that were subsequently cultured for 3 d. A bioinformatics software was used to identify biological processes that were most affected by MMC pretreatment, and validation studies, including in vivo and in vitro assay, were performed. The gene expression analysis identified significant downregulation of annotated functions associated with cellular movement and revealed significant downregulation of multiple genes encoding proinflammatory mediators with chemotactic activity. Validation studies revealed significantly decreased levels of interleukin 6 (IL-6), monocyte chemoattractant protein 3 (MCP-3), and matrix metallopeptidase 2 (MMP2) in culture supernatants of MMC-treated islets compared with controls. Moreover, we showed the suppression of leukocyte chemotactic activity of MMC-treated islets in vitro. We also showed that MMC-treated islets secreted lower levels of chemoattractants that synergistically reduced the immunogenic potential of islets. Histological and immunohistochemical analyses of the implant site revealed that infiltration of monocytes, CD3-positive T cells, and B cells was decreased in MMC-treated islets. In conclusion, the ex vivo pretreatment of islets with MMC and subsequent culture can reduce the immunogenic potential and prolong the survival of islet grafts by inducing the suppression of multiple leukocyte chemotactic factors.

Mitomycin C treatment improves pancreatic islet graft longevity in intraportal islet transplantation by suppressing proinflammatory response

“…Sato et al reported contrasting changes in gene expression between MMC-preconditioned islets and those subjected to culture only. Specifically, the expression of genes involved in various aspects of cell activation, such as cellular movement, immune cell trafficking and inflammatory responses, was downregulated in MMC-conditioned islets 25 . Moreover, we demonstrated that MMC preconditioning significantly suppressed the expression of IL-1β, MCP-1, IL-6 and TNF-α in islet allografts.…”

Comprehensive analysis of gene expression of isolated pancreatic islets during pretransplant culture

“…Short-term culture of isolated islets could provide an opportunity for preconditioning to improve the outcome of islet transplantation. Our previous examination of islets treated with mitomycin-C (MMC) prior to transplantation revealed a decrease in multiple cytokines from MMC-treated islets, including IL-6, which could lead to prolonged engraftment of MMC treated islets without immunosuppression 31 - 33) . These findings suggest that preconditioning during islet culture as a strategy against inflammatory responses could be promising for islet transplantation.…”