Islet Engraftment Research Articles

Methacrylic acid copolymer coating of polypropylene mesh chamber improves subcutaneous islet engraftment

Subcutaneous devices can be used to house therapeutic cells such as pancreatic islets so that the cells can be retrieved. However, a high number of cells may be required to reverse diabetes, since a portion of the graft can be lost after transplantation due to ischemia and therefore the right device design is important. Increasing the vascularity of the subcutaneous space prior to cell transplantation is a strategic goal for cell transplantation, as it promotes islet survival, glucose-sensing and insulin secretion. In this study, a porous cell transplantation device was coated with 40% methacrylic acid-co-isodecyl acrylate (MAA-co-IDA), a biomaterial which promotes a vascular response without additional biologics. Three weeks after device implantation, the vessel density surrounding the device was double that of an uncoated device. The vasculature was mature and connected to the host bloodstream, as demonstrated by perfusion studies and histology. The tissue response to coated devices demonstrated lower levels of inflammation, measured by reduced gene expression of i-NOS and IL1β, and increased expression of IL4. Syngeneic islets (300 islet equivalents) transplanted into the prevascularized coated device were able to return diabetic animals to normoglycemia for up to 11 weeks and resolve a glucose bolus similarly to non-diabetic mice by 3 weeks post-transplantation. We expect that the vessels and microenvironment resulting from the device coating are permissive to islet survival and thus enabled islets to reverse diabetes.

Mechanism of Transplanted Islet Engraftment in Visceral White Adipose Tissue.

White adipose tissue (WAT) is a candidate transplantation site for islets. However, the mechanism of islet engraftment in WAT has not been fully investigated. In this study, we attempted to clarify the therapeutic effect and mechanism of islet transplantation into visceral WAT. Two hundred mouse islets were transplanted into epididymal WAT of syngeneic diabetic mice by wrapping islets with the tissue (fat-covered group). Mice that received intraperitoneal and renal subcapsular islet transplantations were used as negative and positive control groups, respectively. The transplant efficacy, including improvements in blood glucose and plasma insulin levels and in glucose tolerance tests, of the fat-covered group was superior to the negative control group and almost equal to the positive control group. Vessel density of engrafted islets in the fat-covered group was higher than that in the positive control group. It was speculated that the mechanism of islet engraftment in WAT might consist of trapping islets in WAT, adhesion of islets via a combination of adhesion factors (fibronectin and integrin β1), and promotion of angiogenesis in islets by expression of angiogenic factors induced by adiponectin. Visceral WAT is an important candidate for islet transplantation. Adhesion factors and adiponectin might contribute to islet engraftment into WAT. Further studies to elucidate the detailed mechanism are necessary.

1686-P: Brown Adipose Tissue as a Novel Site for Islet Transplantation

Type 1 diabetes (T1D) results in hyperglycemia, increased risk for secondary complications, and life-long dependence on exogenous insulin. Islet transplantation is a promising alternative to insulin therapy for T1D treatment. However, long-term function of islets transplanted into the FDA approved hepatic portal vein remains low, with much of the islet mass being lost early after transplantation due to immune responses and drug toxicity. These challenges have prompted a search for alternative transplantation sites that are readily accessible, immune-privileged, offer graft retrievability, and promote islet survival. A novel site that exhibits these traits is brown adipose tissue (BAT), which is highly vascularized, innervated, and contains immunomodulatory immune cells. BAT functions to maintain thermogenesis and can regulate metabolic homeostasis. We hypothesize that islet engraftment in BAT can restore euglycemia in diabetic mice, promote revascularization, and delay immune rejection. Syngeneic transplantation of 250 NOD.scid islets into the BAT of streptozotocin (STZ)-treated diabetic NOD.scid mice restored euglycemia for over two months, similar to islets transplanted under the pre-clinical kidney capsule transplant site. Islet function was also comparable across transplant groups as measured by a glucose tolerance test. BAT thermogenic function during a cold challenge showed no difference in cold tolerance after transplantation. Histology and mRNA analysis revealed BAT engrafted islets expressed islet hormones, functional islet transcription factors, and CD31+ vascularization at the islet graft site. Overall, our data suggest BAT is a novel and efficacious site for islet transplantation. Future islet transplant studies will determine if the inherent properties of BAT can extend the longevity of transplanted islets by investigating the immunomodulatory capacity of BAT-resident mesenchymal stem cells and immune cells, compared to current transplantation sites. Disclosure J. Barra: None. J. Kepple: None. H.M. Tse: None. C.S. Hunter: None. Funding National Institute of Diabetes and Digestive and Kidney Diseases (R01DK099550); JDRF (SRA-2016-270-S-B, 2-SRA-2019-692-S-B); National Institute of General Medical Sciences (T32GM008111)

Long-term reversal of diabetes by subcutaneous transplantation of pancreatic islet cells and adipose-derived stem cell sheet using surface-immobilized heparin and engineered collagen scaffold

ObjectiveEsterified collagen (EC) can be functionalized with heparin to enhance islet graft stability. Growth factors secreted by human adipose-derived stem cells (hADSCs) can bind efficiently to EC-heparin (EC-Hep), which enhances...

Immune checkpoint CD47 molecule engineered islets mitigate instant blood-mediated inflammatory reaction and show improved engraftment following intraportal transplantation.

Instant blood-mediated inflammatory reaction (IBMIR) causes significant destruction of islets transplanted intraportally. Myeloid cells are a major culprit of IBMIR. Given the critical role of CD47 as a negative checkpoint for myeloid cells, we hypothesized that the presence of CD47 on islets will minimize graft loss by mitigating IBMIR. We herein report the generation of a chimeric construct, SA-CD47, encompassing the extracellular domain of CD47 modified to include core streptavidin (SA). SA-CD47 protein was expressed in insect cells and efficiently displayed on biotin-modified mouse islet surface without a negative impact on their viability and function. Rat cells engineered with SA-CD47 were refractory to phagocytosis by mouse macrophages. SA-CD47-engineered islets showed intact structure and minimal infiltration by CD11b+ granulocytes/macrophages as compared with SA-engineered controls in an in vitro loop assay mitigating IBMIR. In a syngeneic marginal mass model of intraportal transplantation, SA-CD47-engineered islets showed better engraftment and function as compared with the SA-control group (87.5% vs 14.3%). Engraftment was associated with low levels of intrahepatic inflammatory cells and mediators of islet destruction, including high-mobility group box-1, tissue factor, and IL-1β. These findings support the use of CD47 as an innate immune checkpoint to mitigate IBMIR for enhanced islet engraftment with translational potential.

Key role of macrophages in tolerance induction via T regulatory type 1 (Tr1) cells.

T regulatory type 1 (Tr1) cells are a class of regulatory T cells (Tregs ) participating in peripheral tolerance, hence the rationale behind their testing in clinical trials in different disease settings. One of their applications is tolerance induction to allogeneic islets for long-term diabetes-free survival. Currently the cellular and molecular mechanisms that promote Tr1-cell induction in vivo remain poorly understood. We employed a mouse model of transplant tolerance where treatment with granulocyte colony-stimulating factor (G-CSF)/rapamycin induces permanent engraftment of allogeneic pancreatic islets in C57BL/6 mice via Tr1 cells. The innate composition of graft and spleen cells in tolerant mice was analyzed by flow cytometry. Graft phagocytic cells were co-cultured with CD4+ T cells in vitro to test their ability to induce Tr1-cell induction. Graft phagocytic cells were depleted in vivo at different time-points during G-CSF/rapamycin treatment, to identify their role in Tr1-cell induction and consequently in graft survival. In the spleen, the site of Tr1-cell induction, no differences in the frequencies of macrophages or dendritic cells (DC) were observed. In the graft, the site of antigen uptake, a high proportion of macrophages and not DC was detected in tolerant but not in rejecting mice. Graft-infiltrating macrophages of G-CSF/rapamycin-treated mice had an M2 phenotype, characterized by higher CD206 expression and interleukin (IL)-10 production, whereas splenic macrophages only had an increased CD206 expression. Graft-infiltrating cells from G-CSF/rapamycin-treated mice-induced Tr1-cell expansion in vitro. Furthermore, Tr1-cell induction was perturbed upon in-vivo depletion of phagocytic cells, early and not late during treatment, leading to graft loss suggesting that macrophages play a key role in tolerance induction mediated by Tr1 cells. Taken together, in this mouse model of Tr1-cell induced tolerance to allogeneic islets, M2 macrophages infiltrating the graft upon G-CSF/rapamycin treatment are key for Tr1-cell induction. This work provides mechanistic insight into pharmacologically induced Tr1-cell expansion in vivo in this stringent model of allogeneic transplantation.

Shielding islets with human amniotic epithelial cells enhances islet engraftment and revascularization in a murine diabetes model.

Hypoxia is a major cause of considerable islet loss during the early posttransplant period. Here, we investigate whether shielding islets with human amniotic epithelial cells (hAECs), which possess anti-inflammatory and regenerative properties, improves islet engraftment and survival. Shielded islets were generated on agarose microwells by mixing rat islets (RIs) or human islets (HI) and hAECs (100 hAECs/IEQ). Islet secretory function and viability were assessed after culture in hypoxia (1% O2 ) or normoxia (21% O2 ) in vitro. In vivo function was evaluated after transplant under the kidney capsule of diabetic immunodeficient mice. Graft morphology and vascularization were evaluated by immunohistochemistry. Both shielded RIs and HIs show higher viability and increased glucose-stimulated insulin secretion after exposure to hypoxia in vitro compared with control islets. Transplant of shielded islets results in considerably earlier normoglycemia and vascularization, an enhanced glucose tolerance, and a higher β cell mass. Our results show that hAECs have a clear cytoprotective effect against hypoxic damages in vitro. This strategy improves β cell mass engraftment and islet revascularization, leading to an improved capacity of islets to reverse hyperglycemia, and could be rapidly applicable in the clinical situation seeing that the modification to HIs are minor.

Human Amniotic Epithelial Cells and Human Amniotic Membrane as a Vehicle for Islet Cell Transplantation

Immunologic and nonimmunologic mechanisms contribute to islet loss on transplantation in the liver. A site outside of the vascular bed may act as a sanctuary site avoiding immediate inflammatory response. We developed a unique approach by using a human amniotic membrane (HAM) for islet transplant onto the liver surface by combining it with human amniotic epithelial cells (HAECs). Decellularized 1.0 × 1.0-cm HAM was placed on the surface of the liver; 2000 islet equivalent of human islets mixed with 0.4 × 106 HAECs were infused into the space between the membrane and surface. Two pieces of the decellularized HAM were placed under the subcutaneous space to study the angiogenic potential. After cotransplant 57.1% of recipient mice became normoglycemic by the third day after transplant, and 100% attained euglycemia 2 weeks post-transplant. Bodyweight of 85.7% of mice gradually increased over time. The HAM and islets were identified in histologic section in all. Neovascularization was observed to be dramatically increased. In conclusion, HAM is a very versatile biological membrane; HAECs help angiogenesis and better engraftment of islets on the liver surface, eliminating the need for vascular deployment of islet cells and avoiding the risk of portal vein thrombosis and instant blood-mediated inflammatory reaction–related islet loss in the liver.

Local release of NECA (5′-(N-ethylcarboxamido)adenosine) from implantable polymeric sheets for enhanced islet revascularization in extrahepatic transplantation site

Clinical intraportal pancreatic islet infusion is popular for treating type I diabetes. However, multiple doses of islets and anti-rejection protocols are needed to compensate for early large cell losses post-infusion due to the harsh hepatic environment. Thus, extrahepatic sites are utilized to enable efficient islet engraftment and reduce islet mass. Here, we reported an effective islet revascularization protocol that was based on the co-implantation of islet/fibrin gel construct with poly(lactic-co-glycolic) acid sheet releasing NECA (5′-(N-ethylcarboxamido) adenosine; a potent agonist of adenosine) into mouse epididymal fat pad. Thin, flexible sheets (d = 4 mm) prepared by simple casting exhibited sustained NECA release for up to 21 days, which effectively improved early islet engraftment with a median diabetic reversal time of 18.5 days. Western blotting revealed the facilitative effect of NECA on VEGF expression from islets in vitro and from grafts in vivo. In addition, NECA directly promoted the angiogenic activities of islet-derived endothelial cells by enhancing their proliferation and vessel-like tube formation. As a result, neovasculatures were effectively formed in the engrafted islet vicinity, as evidenced by vasculature imaging and immunofluorescence. Taken together, we suggest NECA-releasing PLGA sheets offer a safe and effective drug delivery system that enhances islet engraftment while reducing islet mass at extrahepatic sites for clinical relevance.

Co-transplantation of human adipose-derived mesenchymal stem cells with neonatal porcine islets within a prevascularized subcutaneous space augments the xenograft function.

Cell transplantation has been widely recognized as a curative treatment strategy for variety of diseases including type I diabetes (T1D). Broader patient inclusion for this therapeutic option is restricted by a limited supply of healthy human islet donors and significant loss of islets immediately postintrahepatic transplant due to immune activation. Neonatal porcine islets (NPIs) are a potential ubiquitous β-cell source for treating T1D. Mesenchymal stem cells (MSCs) have the inherent capacity to secrete immunoregulatory, anti-inflammatory, and proangiogenic factors and, thus, have the potential to improve islet engraftment, survival, and function. Herein, we assessed the effect of human adipose-derived MSCs (AdMSCs) on NPI metabolic outcomes in diabetic mice when co-transplanted within the prevascularized subcutaneous deviceless (DL) space or kidney capsule (KC). Graft function has been evaluated by weekly blood glucose, stimulated porcine insulin, glucose tolerance, and total cellular graft insulin content. Compared with NPI alone, co-transplantation of NPIs and AdMSCs resulted in significantly earlier normoglycemia (*P<.05), improved glucose tolerance (*P<.05), superior stimulated serum porcine insulin (**P<.01), and increased graft insulin content (*P<.05) in the DL site and not the KC. Thus, our study demonstrates that co-transplantation of human AdMSCs with NPIs is an effective tactic to augment islet xenograft function in a clinically relevant extrahepatic site.

Pancreatic Tissue-Derived Extracellular Matrix Bioink for Printing 3D Cell-Laden Pancreatic Tissue Constructs.

The transplantation of pancreatic islets is a promising treatment for patients who suffer from type 1 diabetes accompanied by hypoglycemia and secondary complications. However, islet transplantation still has several limitations such as the low viability of transplanted islets due to poor islet engraftment and hostile environments. In addition, the insulin-producing cells differentiated from human pluripotent stem cells have limited ability to secrete sufficient hormones that can regulate the blood glucose level; therefore, improving the maturation by culturing cells with proper microenvironmental cues is strongly required. In this article, we elucidate protocols for preparing a pancreatic tissue-derived decellularized extracellular matrix (pdECM) bioink to provide a beneficial microenvironment that can increase glucose sensitivity of pancreatic islets, followed by describing the processes for generating 3D pancreatic tissue constructs using a microextrusion-based bioprinting technique.

Transplantation of PEGylated islets enhances therapeutic efficacy in a diabetic nonhuman primate model.

Islet cell transplantation can lead to insulin independence, reduced hypoglycemia, and amelioration of diabetes complications in patients with type 1 diabetes. The systemic delivery of anti-inflammatory agents, while considered crucial to limit the early loss of islets associated with intrahepatic infusion, increases the burden of immunosuppression. In an effort to decrease the pharmaceutical load to the patient, we modified the pancreatic islet surface with long-chain poly(ethylene glycol) (PEG) to mitigate detrimental host-implant interactions. The effect of PEGylation on islet engraftment and long-term survival was examined in a robust nonhuman primate model via three paired transplants of dosages 4300, 8300, and 10000islet equivalents per kg body weight. A reduced immunosuppressive regimen of anti-thymocyte globulin induction plus tacrolimus in the first posttransplant month followed by maintenance with sirolimus monotherapy was employed. To limit transplant variability, two of the three pairs were closely MHC-matched recipients and received MHC-disparate PEGylated or untreated islets isolated from the same donors. Recipients of PEGylated islets exhibited significantly improved early c-peptide levels, reduced exogenous insulin requirements, and superior glycemic control, as compared to recipients of untreated islets. These results indicate that this simple islet modification procedure may improve islet engraftment and survival in the setting of reduced immunosuppression.

Recruited fibroblasts reconstitute the peri-islet membrane: a longitudinal imaging study of human islet grafting and revascularisation

Aims/hypothesisRapid and adequate islet revascularisation and restoration of the islet–extracellular matrix (ECM) interaction are significant factors influencing islet survival and function of the transplanted islets in individuals with type 1 diabetes. Because the ECM encapsulating the islets is degraded during islet isolation, understanding the process of revascularisation and engraftment after transplantation is essential and needs further investigation.MethodsHere we apply a longitudinal and high-resolution imaging approach to investigate the dynamics of the pancreatic islet engraftment process up to 11 months after transplantation. Human and mouse islet grafts were inserted into the anterior chamber of the mouse eye, using a NOD.ROSA-tomato.Rag2−/− or B6.ROSA-tomato host allowing the investigation of the expansion of host vs donor cells and the contribution of host cells to aspects such as promoting the encapsulation and vascularisation of the graft.ResultsA fibroblast-like stromal cell population of host origin rapidly migrates to ensheath the transplanted islet and aid in the formation of a basement membrane-like structure. Moreover, we show that the vessel network, while reconstituted by host endothelial cells, still retains the overall architecture of the donor islets.Conclusions/interpretationIn this transplantation situation the fibroblast-like stromal cells appear to take over as main producers of ECM or act as a scaffold for other ECM-producing cells to reconstitute a peri-islet-like basement membrane. This may have implications for our understanding of long-term graft rejection and for the design of novel strategies to interfere with this process.

Improving the Function and Engraftment of Transplanted Pancreatic Islets Using Pulsed Focused Ultrasound Therapy

This study demonstrates that pulsed focused ultrasound (pFUS) therapy can non-invasively enhance the function and engraftment of pancreatic islets following transplantation. In vitro, we show that islets treated with pFUS at low (peak negative pressure (PNP): 106kPa, spatial peak temporal peak intensity (Isptp): 0.71 W/cm2), medium (PNP: 150kPa, Isptp: 1.43 W/cm2) or high (PNP: 212kPa, Isptp: 2.86 W/cm2) acoustic intensities were stimulated resulting in an increase in their function (i.e. insulin secretion at low-intensity: 1.15 ± 0.17, medium-intensity: 2.02 ± 0.25, and high-intensity: 2.54 ± 0.38 fold increase when compared to control untreated islets; P < 0.05). Furthermore, we have shown that this improvement in islet function is a result of pFUS increasing the intracellular concentration of calcium (Ca2+) within islets which was also linked to pFUS increasing the resting membrane potential (Vm) of islets. Following syngeneic renal sub-capsule islet transplantation in C57/B6 mice, pFUS (PNP: 2.9 MPa, Isptp: 895 W/cm2) improved the function of transplanted islets with diabetic animals rapidly re-establishing glycemic control. In addition, pFUS was able to enhance the engraftment by facilitating islet revascularization and reducing inflammation. Given a significant number of islets are lost immediately following transplantation, pFUS has the potential to be used in humans as a novel non-invasive therapy to facilitate islet function and engraftment, thereby improving the outcome of diabetic patients undergoing islet transplantation.

Nanotechnology Approaches to Modulate Immune Responses to Cell-based Therapies for Type 1 Diabetes.

Islet transplantation is a promising curative treatment option for type 1 diabetes (T1D) as it can provide physiological blood glucose control. The widespread utilization of islet transplantation is limited due to systemic immunosuppression requirements, persisting graft immunodestruction, and poor islet engraftment. Traditional macro- and micropolymeric encapsulation strategies can alleviate the need for antirejection immunosuppression, yet the increased graft volume and diffusional distances imparted by these coatings can be detrimental to graft viability and glucose control. Additionally, systemic administration of pro-engraftment and antirejection therapeutics leaves patients vulnerable to adverse off-target side effects. Nanoscale engineering techniques can be used to immunocamouflage islets, modulate the transplant microenvironment, and provide localized pro-engraftment cues. In this review, we discuss the applications of nanotechnology to advance the clinical potential of islet transplantation, with a focus on cell surface engineering, bioactive functionalization, and use of nanoparticles in T1D cell-based treatments.

48: Islets Loaded in Hydrogel Derived from Human Amniotic Membrane Reverse Diabetes in Mice

Background Development of macroencapsulation, neovascularized devices and biopolymer scaffolds that could be easily loaded with islets and implanted, with the aim of constructing a bioartificial pancreas may help to reduce the risks and improve the success rate of islet transplantation. Human amniotic membrane (HAM) is inexpensive and attractive as a biomaterial due to its structural similarities to islet extracellular matrix (ECM), and its immunomodulatory, anti-inflammatory and antifibrotic properties. The aim of our study was to develop hydrogel derived from HAM and assess whether it could support islet function in vitro and in vivo. Methods/ Materials The hydrogels were generated from HAM and accessed for porosity and ECM content. The protein content in HAM derived hydrogels and native HAM lysates were measured. To assess hydrogel impact on islet viability and function isolated rat islets were incorporated into the hydrogels and cultured for one week. The cell viability was evaluated by FDA/PI staining. To demonstrate islet function the glucose stimulated insulin secretion (GSIS) tests were performed using standard ELISA. Next, we assessed whether incorporation of islets into hydrogel could enhance engraftment and lead to better glycemic control in diabetic SCID mice. For this purpose 350 rat islets (IEQ) loaded into the hydrogels or islets alone (control) were transplanted into the epididymal fat of diabetic SCID mice. Blood glucose levels were monitored daily and intraperitoneal glucose tolerance tests (IPGTTs) were carried out. Grafts and serum were harvested at 1, 2, 6 and 12 weeks after transplantation to assess outcome. Results The ECM concentration in the hydrogel affected the pore size. Insulin and glucagon expression and viability of islets incorporated into hydrogel was significantly higher than that of islets in free-floating culture. In addition, significant enhancement of GSIS was observed from islets embedded in hydrogel as compared to controls. In vivo experiments showed that, transplantation of 350 IEQ embedded in hydrogel lead to enhanced engraftment, vascularization, viability and better glycaemic control compared to control mice transplanted with islets alone. Conclusions Incorporation of pancreatic islet into amnion-derived hydrogels enhances islet engraftment and is a valuable approach to improve islet transplantation outcomes. This work was supported by a grant from the Swiss National Science Foundation (Grant #310030_173138, to TB, EB, DB) and a grant from the European Foundation for Study of Diabetes (to EB and DB).

49: Shielding Islets with Human Amniotic Epithelial Cells Protects Islets Against Hypoxia and Enhances Islet Engraftment and Revascularization after Transplantation in a Murine Diabetic Model

49: Shielding Islets with Human Amniotic Epithelial Cells Protects Islets Against Hypoxia and Enhances Islet Engraftment and Revascularization after Transplantation in a Murine Diabetic Model

Selective local irradiation improves islet engraftment and survival in intra–bone marrow islet transplantation

BackgroundBone marrow (BM) is as an alternative site for islet transplantation, but it is not an immunoprotected microenvironment and allogeneic islets are rejected. However, the BM, for its structure and anatomic position, offers the possibility to modulate microenvironment by local interventions. We here investigate whether local irradiation is able to improve islet engraftment and prevent rejection in BM in the absence of immunosuppression. MethodsA model of BM local irradiation was set up. Islets were transplanted in syngeneic and fully major histocompatibility complex–mismatched recipients in control and locally irradiated BM; gain of normoglycemia and time to rejection were evaluated. ResultsBM local irradiation proved to be a selective and safe procedure. Syngeneic islet transplantation into locally irradiated BM had better outcome compared with not irradiated recipients in terms of capacity to gain normoglycemia (100% versus 56% in irradiated versus not irradiated mice). In the allogenic setting, glycemia was significantly lower in the first days after transplantation in the group of irradiated mice and local irradiation also delayed time to graft rejection (from 4 ± 1 days for not irradiated to 11 ± 1 days for locally irradiated mice). DiscussionThese data indicate that local immunosuppression by irradiation before islet transplantation in BM favors islet engraftment and delays time to rejection.

CCL2/MCP-1 and CXCL12/SDF-1 blockade by L-aptamers improve pancreatic islet engraftment and survival in mouse.

The blockade of pro-inflammatory mediators is a successful approach to improve the engraftment after islet transplantation. L-aptamers are chemically synthesized, nonimmunogenic bio-stable oligonucleotides that bind and inhibit target molecules conceptually similar to antibodies. We aimed to evaluate if blockade-aptamer-based inhibitors of C-C Motif Chemokine Ligand 2/monocyte chemoattractant protein-1 (CCL2/MCP-1) and C-X-C Motif Chemokine Ligand 12/stromal cell-derived factor-1 (CXCL12/SDF-1) are able to favor islet survival in mouse models for islet transplantation and for type 1 diabetes. We evaluated the efficacy of the CCL2-specific mNOX-E36 and the CXCL12-specific NOX-A12 on islet survival in a syngeneic mouse model of intraportal islet transplantation and in a multiple low doses of streptozotocin (MLD-STZ) diabetes induction model. Moreover, we characterized intrahepatic infiltrated leukocytes by flow cytometry before and 3 days after islet infusion in presence or absence of these inhibitors. The administration for 14days of mNOX-E36 and NOX-A12 significantly improved islet engraftment, either compound alone or in combination. Intrahepatic islet transplantation recruited CD45+ leucocytes and more specifically CD45+/CD11b+ mono/macrophages; mNOX-E36 and NOX-A12 treatments significantly decreased the recruitment of inflammatory monocytes, CD11b+ /Ly6Chigh /CCR2+ and CD11b+ /Ly6Chigh /CXCR4+ cells, respectively. Additionally, both L-aptamers significantly attenuated diabetes progression in the MLD-STZ model. In conclusion, CCL2/MCP-1 and CXCL12/SDF-1 blockade by L-aptamers is an efficient strategy to improve islet engraftment and survival.

The role of human CD46 in early xenoislet engraftment in a dual transplant model.

Membrane cofactor protein CD46 attenuates the complement cascade by facilitating cleavage of C3b and C4b. In solid organ xenotransplantation, organs expressing CD46 have been shown to resist hyperacute rejection. However, the incremental value of human CD46 expression for islet xenotransplantation remains poorly defined. This study attempted to delineate the role of CD46 in early neonatal porcine islet engraftment by comparing Gal-knocked out (GKO) and hCD46-transgenic (GKO/CD46) islets in a dual transplant model. Seven rhesus macaques underwent dual transplant and were sacrificed at 1hour (n=4) or 24hours (n=3). Both hemilivers were recovered and fixed for immunohistochemistry (CD46, insulin, neutrophil elastase, platelet, IgM, IgG, C3d, C4d, CD68, Caspase 3). Quantitative immunohistochemical analysis was performed using the Aperio Imagescope. Within 1hour of intraportal infusion of xenografts, no differences were observed between the two types of islets in terms of platelet, antibody, or complement deposition. Cellular infiltration and islet apoptotic activity were also similar at 1hour. At 24hours, GKO/CD46 islets demonstrated significantly less platelet deposition (P=0.01) and neutrophil infiltration (P=0.01) compared to GKO islets. In contrast, C3d (P=0.38) and C4d (P=0.45) deposition was equal between the two genotypes. Our findings suggest that expression of hCD46 on NPIs potentially provides a measurable incremental survival advantage in vivo by reducing early thrombo-inflammatory events associated with instant blood-mediated inflammatory reaction (IBMIR) following intraportal islet infusion.