Lu, Y. et al. Bioluminescent monitoring of islet graft survival after transplantation. Mol. Ther. 9, 428-435

Stanford University, Stanford, California, United States
Molecular Therapy (Impact Factor: 6.23). 04/2004; 9(3):428-35. DOI: 10.1016/j.ymthe.2004.01.008
Source: PubMed


Islet transplantation offers a potential therapy to restore glucose homeostasis in type 1 diabetes patients. A method to image transplanted islets noninvasively and repeatedly would greatly assist studies of islet transplantation. Using recombinant adenovirus, we show that isolated rodent and human islets can be genetically engineered to express luciferase and then imaged after implantation into NOD-scid mice using a cooled charge-coupled device. The magnitude of the signal was dependent on the islet dose. Adenovirus-directed luciferase expression, however, rapidly attenuated. We next tested lentivirus vectors that should direct the long-term expression of reporter genes in transduced islets. Transplanted lentivirus-transduced islets restored euglycemia long term in streptozotocin-treated NOD-scid mice. The signal from implanted lentivirus-transduced islets was related directly to the implanted islet mass, and the signal did not attenuate over the observation period. Viral transduction, luciferase expression, and repeated imaging had no apparent long-term deleterious effects on islet function after implantation. These data demonstrate that the introduction of reporter genes into an isolated tissue allows the long-term monitoring of its survival following implantation. Such imaging technologies may allow earlier detection of graft rejection and the adjustment of therapies to prolong graft survival posttransplantation.

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Available from: Martha Campbell-Thompson
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    • "In islet studies, BLI capability is incorporated by isolating islets from luciferase-expressing transgenic animals (Chen et al., 2006, 2008) or by virally transducing islets from wild-type animals with the luciferase gene (Fowler et al., 2005; Lu et al., 2004). Viral luciferase incorporation in primary cells such as islets would require genetic manipulation of the graft in every transplant procedure, making routine BLI application costly and time-consuming . "
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    ABSTRACT: Cell-based therapies to treat loss-of-function hormonal disorders such as diabetes and Parkinson's disease are routinely coupled with encapsulation strategies, but an understanding of when and why grafts fail in vivo is lacking. Consequently, investigators cannot clearly define the key factors that influence graft success. Although bioluminescence is a popular method to track the survival of free cells transplanted in preclinical models, little is known of the ability to use bioluminescence for real-time tracking of microencapsulated cells. Furthermore, the impact that dynamic imaging distances may have, due to freely-floating microcapsules in vivo, on cell survival monitoring is unknown. This work addresses these questions by applying bioluminescence to a pancreatic substitute based on microencapsulated cells. Recombinant insulin-secreting cells were transduced with a luciferase lentivirus and microencapsulated in Ba(2+) crosslinked alginate for in vitro and in vivo studies. In vitro quantitative bioluminescence monitoring was possible and viable microencapsulated cells were followed in real time under both normoxic and anoxic conditions. Although in vivo dispersion of freely-floating microcapsules in the peritoneal cavity limited the analysis to a qualitative bioluminescence evaluation, signals consistently four orders of magnitude above background were clear indicators of temporal cell survival. Strong agreement between in vivo and in vitro cell proliferation over time was discovered by making direct bioluminescence comparisons between explanted microcapsules and parallel in vitro cultures. Broader application of this bioluminescence approach to retrievable transplants, in supplement to currently used end-point physiological tests, could improve understanding and accelerate development of cell-based therapies for critical clinical applications. Copyright © 2014 John Wiley & Sons, Ltd.
    Full-text · Article · Jul 2014 · Journal of Tissue Engineering and Regenerative Medicine
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    • "Finally, these studies provided new information about islet fate following transplantation. Luciferase signal emanating from the graft remained stable for at least 140 days, indicating graft stability in this model of transplantation [62]. The feasibility of monitoring transplanted islets by BLI was also demonstrated in the intrahepatic transplantation model. "
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    ABSTRACT: Replacement of insulin production by pancreatic islet transplantation has great potential as a therapy for type 1 diabetes mellitus. At present, the lack of an effective approach to islet grafts assessment limits the success of this treatment. The development of molecular imaging techniques has the potential to fulfill the goal of real-time noninvasive monitoring of the functional status and viability of the islet grafts. We review the application of a variety of imaging modalities for detecting endogenous and transplanted beta-cell mass. The review also explores the various molecular imaging strategies for assessing islet delivery, the metabolic effects on the islet grafts as well as detection of immunorejection. Here, we highlight the use of combined imaging and therapeutic interventions in islet transplantation and the in vivo monitoring of stem cells differentiation into insulin-producing cells.
    Full-text · Article · Oct 2011 · Journal of Transplantation
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    • "Bioluminescent reporters, such as firefly luciferase, emit photons that can be detected with high sensitivity using a cooled charge-coupled device (CCD). We and others have demonstrated human and rodent islets can be genetically engineered ex vivo to express luciferase so that the islets can be monitored following their transplantation by CCD and that CCD signal corresponded with the implanted β-cell mass (1,2). Several laboratories have generated transgenic mice that express luciferase in their β-cells and shown that the CCD signal from the pancreas is correlated with β-cell mass (3–5). "
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    ABSTRACT: β-Cells that express an imaging reporter have provided powerful tools for studying β-cell development, islet transplantation, and β-cell autoimmunity. To further expedite diabetes research, we generated transgenic C57BL/6 "MIP-TF" mice that have a mouse insulin promoter (MIP) driving the expression of a trifusion (TF) protein of three imaging reporters (luciferase/enhanced green fluorescent protein/HSV1-sr39 thymidine kinase) in their β-cells. This should enable the noninvasive imaging of β-cells by charge-coupled device (CCD) and micro-positron emission tomography (PET), as well as the identification of β-cells at the cellular level by fluorescent microscopy. MIP-TF mouse β-cells were multimodality imaged in models of type 1 and type 2 diabetes. MIP-TF mouse β-cells were readily identified in pancreatic tissue sections using fluorescent microscopy. We show that MIP-TF β-cells can be noninvasively imaged using microPET. There was a correlation between CCD and microPET signals from the pancreas region of individual mice. After low-dose streptozotocin administration to induce type 1 diabetes, we observed a progressive reduction in bioluminescence from the pancreas region before the appearance of hyperglycemia. Although there have been reports of hyperglycemia inducing proinsulin expression in extrapancreatic tissues, we did not observe bioluminescent signals from extrapancreatic tissues of diabetic MIP-TF mice. Because MIP-TF mouse β-cells express a viral thymidine kinase, ganciclovir treatment induced hyperglycemia, providing a new experimental model of type 1 diabetes. Mice fed a high-fat diet to model early type 2 diabetes displayed a progressive increase in their pancreatic bioluminescent signals, which were positively correlated with area under the curve-intraperitoneal glucose tolerance test (AUC-IPGTT). MIP-TF mice provide a new tool for monitoring β-cells from the single cell level to noninvasive assessments of β-cells in models of type 1 diabetes and type 2 diabetes.
    Full-text · Article · Mar 2011 · Diabetes
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