[Show abstract][Hide abstract] ABSTRACT: Although encapsulation has been demonstrated to prolong islet survival, insufficient oxygen delivery via the surrounding microcirculation after subcutaneous transplantation results in suboptimal islet function and early graft death as early as the first week after transplantation. We employ the mouse dorsal window model, Laser Speckle Imaging (LSI) and Wide-field Functional Imaging (WiFI) to noninvasively quantify changes in relative blood flow (RBF) and hemoglobin oxygen saturation (HOS) to study the vascular changes following subcutaneous transplantation of alginate-encapsulated xenogeneic islets over a seven day period post transplantation.
Subcutaneous dorsal window chambers were installed on C57BL/6 albino mice and implanted with a blank alginate sheet, a sheet containing young porcine islets isolated from pre-weaned Landrace pigs (18-22 days old) or no implant (negative control). Images obtained using LSI and WiFI were analyzed using algorithms to quantify the changes in RBF and HOS respectively. At the conclusion of the experiment, the implants were extracted and analyzed for islet viability and function.
Arterial and venous dilatation, peri-implant neovascularization and increase in the RBF and arterial HOS were noted in both groups by day 7. On further quantification, the increase in HOS from 90.03+0.69 to 98.77+0.40 and RBF from 0.88 to 1.62 and by day 7 in the porcine islet group was statistically significant (p=0.03) while the increase in the control group (96.09+1.05 to 99.02+0.43;HOS, 0.97 to 0.98; RBF) was not (p=0.67). The transplanted islets maintained viability (79.08±3.28; Day 0, 79.35±0.84; Day7) and remained functional (SI = 2.16±0.09; Day 0, 1.98±0.08; Day7).
Our results suggest that implanted islets induce vascular changes, while remaining functional and maintaining viability. Future experiments will attempt to study the role played by islet secreted VEGF in increasing blood flow and arterial oxygenation in the implant-adjacent vasculature while also extending the study period for up to 2 weeks post transplantation.
International Pancreas and Islet Transplantation Association, 2013, Monterey, California; 09/2013
[Show abstract][Hide abstract] ABSTRACT: Cell encapsulation is a method of encasing cells in a semipermeable matrix that provides a permeable gradient for the passage of oxygen and nutrients, but effectively blocks immune regulating cells from reaching the graft, preventing rejection. This concept has been described as early as the 1930's but it has exhibited substantial achievements over the last decade. Several advances in encapsulation engineering, chemical purification, applications, and cell viability promises to make this a revolutionary technology. Several obstacles still need to be overcome before this process become a reality, including developing a reliable source of islets or insulin-producing cells, determining the ideal biomaterial to promote graft function, reducing the host response to the encapsulation device, and ultimately a streamlined, scaled up process for industry to be able to efficiently and safely produce encapsulated cells for clinical use. This paper provides a comprehensive review of cell encapsulation of islets for the treatment of type 1 diabetes including a historical perspective, current research findings, and future studies.
[Show abstract][Hide abstract] ABSTRACT: Background: Xenotransplantion provides a solution to human islet donor scarcity. However, the rapid, immune-mediated destruction of transplanted islets in the absence of potent and prolonged immunosuppression make it an unattractive solution. Studies have demonstrated that biomaterial encapsulation of xenograft islets prolong their survival and obviates the need for immunosuppression. However with encapsulation, islet survival is compromised due to insufficient oxygen delivery via the microcirculation. Thus, a brisk vascular response is vital to reduce the risk of hypoxia in the transplanted islets. We aim to compare the responses to isograft and xenograft islets by employing a combination of the mouse dorsal window-chamber model and Wide-field Functional Imaging (WiFI) and Laser Speckle Imaging(LSI).
Methods: Dorsal window chambers were installed on C57BL/6 albino mice. The implants were placed in intimate contact with the underside of the dermis of the opposing skin, within the viewing port of the model. We studied the host vascular response to the introduction of either a sheet containing islets isolated from C57Bl/6 mice or one containing islets isolated from 3 week old Yorkshire pigs. Two modes of WiFI were used to monitor hemodynamics over the ensuing 7-day period. Laser speckle imaging enables mapping of blood flow, and multispectral imaging enables mapping of hemoglobin oxygen saturation.
Results: Our data demonstrates that the dorsal window chamber enables longitudinal monitoring and an easy comparison of the host response to transplanted islets isolated from various donors over a one week period. Vascular changes that are readily noted are arteriolar, venular and venous dilatation and the formation of nascent arteriovenous connections. Peri-implant neovascularization, while noted in both groups was significantly more in the porcine islet group. We hypothesize that this is presumably due to a greater metabolic demand by porcine islets when compared to syngeneic murine islets. An increase in the relative rate of blood flow(Fig.1C,G and Fig.2 C,G) and hemoglobin oxygen saturation (Fig 1D,H and Fig 2D,H.) can also be observed. Future experiments will compare islet viability and the host response between the two groups upto two weeks after transplantation.
Fig 1. With the mouse dorsal window chamber and WiFI, the host vascular response is monitored over a 7-day period following implantation of syngeneic murine islets encapsulated in a high guluronate alginate sheet. (A,B,C,D) Brightfield, blood-flow, and hemoglobin oxygen saturation images, respectively, taken immediately following surgery. (E,F,G,H) Corresponding images seven days following surgery. Numerous islets( ) are visualized within the chamber. Peri-implant neovascularization( ) is clearly visible on Day 7(F). An increase in blood flow and hemoglobin oxygen saturation in the peri-implant microvasculature is also noted (G,H).
24th International Meeting of the Transplantation Society, 2012, Berlin, Germany; 07/2012
[Show abstract][Hide abstract] ABSTRACT: Islet encapsulation offers an immune system barrier for islet transplantation, and encapsulation within an alginate sheetlike structure offers the ability to be retrievable after transplanted. This study aims to show that human islets encapsulated into islet sheets remain functional and viable after 8 weeks in culture or when transplanted into the subcutaneous space of rats. Human islets were isolated from cadaveric organs. Dissociation and purification were done using enzymatic digestion and a continuous Ficoll-UWD gradient. Purified human islets were encapsulated in alginate sheets. Human Islet sheets were either kept in culture, at 37°C and 5% CO(2), or transplanted subcutaneously into Lewis rats. After 1, 2, 4, and 8 weeks, the human islet sheets were retrieved from the rats and assessed. The viability of the sheets was measured using fluorescein diacetate (FDA)/propidium iodide (PI), and function was measured through glucose-stimulated insulin release, in which the sheets were incubated for an hour in low-glucose concentration (2.8 mmol/L) and then high (28 mmol/L), then high (28 mmol/L) plus 3-isobutyl-1-methylxanthine (50 μm). Human islet sheets remained both viable, above 70%, and functional, with a stimulation index (insulin secretion in high glucose divided by insulin secretion in low glucose) above 1.5, over 8 weeks of culture or subcutaneous transplantation. Islet transplantation continues to make advances in the treatment of type 1 diabetes. These preliminary results suggest that encapsulated islets sheets can survive and maintain islet viability and function in vivo, within the subcutaneous region.
[Show abstract][Hide abstract] ABSTRACT: The Islet Sheet is a thin planar bioartificial endocrine pancreas fabricated by gelling highly purified alginate and islets of Langerhans. Acellular alginate layers form a uniform immunoprotective barrier to host rejection of the encapsulated cells, with the tissue nourished by passive diffusion from adjacent host tissue. The overall thickness of the Islet Sheet, 250 microm, is chosen to maximize nutrient diffusion. In this paper we describe the early development of the Islet Sheet, including purification and fractionation of the alginates used, difficulties in maintaining sheet planarity, and preliminary metabolic studies in pancreatectomized dogs. In a key experiment, approximately 75,000 allogeneic islet equivalents in six Islet Sheets were sutured to the omentum of a 7-kg female beagle dog at the time of pancreatectomy. Fasting euglycemia was maintained for 84 days. Fed blood sugars were usually below 150 mg/dL. A single injection of 2 U insulin was administered on day 9, and antibiotics were administered for two weeks. No other drugs were used. IVGTT post implant was not normal, but seemed to improve between 30 and 60 days. Upon omentectomy and sheet removal the metabolic parameters deteriorated to a frankly diabetic state within seven days. The sheets did not remain flat, but fragments were recovered within hard, mostly acellular capsules. Dithizone staining showed islets within alginate sheets recovered from the interior of these capsules, suggesting that allogeneic islet tissue survived 84 days and was responsible for maintaining fasting euglycemia.
Annals of the New York Academy of Sciences 12/2001; 944:252-66. · 4.38 Impact Factor