Local release of dexamethasone from macroporous scaffolds accelerates islet transplant engraftment by promotion of anti-inflammatory M2 macrophages
Graphical abstract
Introduction
Type 1 diabetes (T1D) is an autoimmune disease resulting in the targeted destruction of insulin-secreting beta cells. Clinical islet transplantation, where allogeneic pancreatic islets are infused into the portal vein of the liver, has achieved insulin independence for individuals suffering from labile diabetes with severe hypoglycemic unawareness [1]. Graft efficacy is unstable, however, as multiple transplant-related factors, such as poor engraftment and host inflammatory and immunological responses, result in the significant loss of islets [2]. In an effort to improve graft duration, our laboratory developed a three-dimensional macroporous polydimethylsiloxane (PDMS) scaffold for housing transplanted islets [3]. Placement of islet-loaded scaffolds within an extrahepatic site (i.e. the omentum) results in stable efficacy in syngeneic and allogeneic diabetic rodent and nonhuman primate models [4]. The highly interconnected and porous structure of the PDMS scaffold facilitates positive interactions between graft and host tissue, resulting in a supportive microenvironment [3]. Further, the localization of islets within a defined implant permits ease of monitoring and retrieval, as well as the integration of beneficial factors, such as proangiogenic hydrogels [5].
With the potential of this 3-D platform to locally instruct host responses to the islet graft, efforts were focused on decreasing inflammatory events instigated post-transplant. In clinical studies, inflammation has been implicated as a prominent player in decreasing islet viability and graft efficacy [6]. Activation of the inflammatory cascade starts early, whereby pro-inflammatory mediators, such as tissue factor and high-mobility group box-1 protein, are upregulated by stressed islets following pancreatic isolation. The transplantation of islets subsequently triggers thrombosis, complement cascade, and an instant blood-mediated inflammatory reaction (IBMIR), which results in early islet loss (∼60% of the islet mass) and the promotion of adaptive immune responses [7], [8]. When biomaterials are added to the transplant microenvironment, activation of inflammatory processes can be further amplified, due to foreign body responses (FBR) [9]. To mitigate inflammation in clinical islet transplantation, supplemental administration of TNFα and IL-1β blockers (etanercept and anakinra) has been shown to dampen pro-inflammatory effectors and lead to improved allograft survival [10]. The intravenous delivery of these agents, however, results in detrimental systemic impacts such as an increased risk of serious infections.
The engineering of a localized drug delivery platform could avoid the side effects of systemic anti-inflammatory regimens, while also precisely modulating inflammatory responses at the transplant site. An optimal drug and dosage would suppress active inflammatory pathways and instruct local host responses towards desirable phenotypes. Directing macrophage phenotype is of particular interest, as macrophages are key contributors in determining the overall outcome of the host response to an implant [11]. During normal activation and resolution of inflammatory processes, the macrophage population covers a balanced spectrum of phenotypes. On one end of this spectrum are classically activated macrophages (termed M1), which generate a microenvironment rich in reactive oxygen species, degradative enzymes, and pro-inflammatory cytokines (e.g. IL-1β, IL-6, and TNFα) [12]. On the other end are the alternatively activated subtype (termed M2) that is associated with a different portfolio of cytokines (e.g. IL-10 and TGFβ) that facilitate healthy tissue remodeling [13]. Local pro-inflammatory instigators and undesirable implants tip this balance towards M1, which can result in detrimental downstream events such as the generation of a fibrotic capsule, the promotion of detrimental Th1 responses, and the destruction of the graft [14]. Alternatively, modest promotion of the M2 phenotype has been shown to direct positive tissue remodeling, resulting in functional engraftment of the implant [15]. These M2 macrophages impart additional benefits to islet transplants, as they have been shown to promote islet function and beta cell proliferation [16].
A strong drug candidate for this approach is dexamethasone (Dex). Dex, an immunosuppressive glucocorticoid capable of potent suppression of inflammatory pathways, has been shown to polarize human blood-derived monocytes toward the anti-inflammatory (M2) phenotype, while retaining their migratory function [17], [18]. The Dex dosage must be carefully tailored, however, to prevent overloading at the implant site, as elevated glucocorticoids can severely impair cell mobility, resulting in compromised engraftment and vascularization [19]. Further, Dex has also been shown to impair the glucose responsiveness of beta cells at high concentrations [20]. In this study, we utilized our macroporous PMDS scaffold platform to locally deliver dexamethasone in a controlled manner within the desired therapeutic range (Fig. 1). This streamlined approach eliminated the need for disparate drug depots, as previously explored [21]. Evaluation of kinetic release of Dex from the 3-D scaffold permitted tailoring of drug loading to desired doses. Following optimization, the impact of this drug-release platform on the efficiency of islet engraftment and subsequent host cell responses were investigated.
Section snippets
Fabrication of Dex-PDMS scaffolds and characterization of Dex-releasing kinetics
Macroporous PDMS scaffolds of 85% total porosity were fabricated by particle leaching and solvent casting, as described previously [3]. In brief, the PDMS matrix was prepared by mixing PDMS part A with cross-linker part B (RTV 615 A/B kit, Momentive) at a ratio of 4:1 (A:B). Dexamethasone (Enzo life sciences) was subsequently added to the PDMS mixture at the desired percentage (1, 0.5, 0.25, and 0.1% weight-to-weight, which results in a total mass of 182, 90.5, 45.1, and 18 μg Dex per scaffold
Characterization of Dex release profiles from PDMS scaffolds
The macroporous PDMS scaffold serves as a supportive 3-D platform for housing islets following transplantation (Fig. 2A–C). For simplicity of engineering design, dexamethasone was incorporated directly into the PDMS liquid prior to the fabrication of the resulting scaffold. To ensure appropriate Dex release kinetics from the 3-D implant, varying concentrations of Dex (1.0, 0.5, 0.25, and 0.1% w/w) were screened. Despite the unpredictable surface area of the scaffold, drug release profiles were
Discussion
Biomaterial delivery systems are attractive platforms over direct daily injections, as they facilitate the localized delivery of therapeutic agents in a controllable fashion over extended time periods, while eliminating the complications of patient compliance. Via the manipulation of biomaterial characteristics (e.g. architecture, degradation, and drug loading), customized constructs can meet distinctive physiological demands. By incorporating Dex directly into the 3-D platform used to house
Conclusion
A Dex-PDMS scaffold provides a feasible platform to locally deliver immunomodulatory dexamethasone in a controllable manner and thereby foster a protective microenvironment for engrafted islets. In this study, we characterized the impact of Dex-PDMS scaffolds on modulating inflammatory response in the transplant milieu. The local, low-dose delivery of Dex via a macroporous scaffold demonstrated macrophage polarization towards an anti-inflammatory phenotype and significantly improved islet
Acknowledgements
This work was supported by a partnership grant from the JDRF and Leona M. and Harry B. Helmsley Charitable Trust (3-SRA-2015-38-Q-R) and the University of Florida Preeminence funding. We thank Irayme Labrada for her excellent technical assistant during animal transplants. We thank the Diabetes Research Institute Preclinical and Translational Models Core lab for providing the rodent islets used in this project. We thank Kevin Johnson from the Diabetes Research Institute for his consistent
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