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Pancreatic islet transplantation is a promising strategy for the treatment of type I diabetes. High-mobility group box-1 (HMGB1), highly expressed in islet cells, is a potent immune stimulator in immune rejection. Heme oxygenase-1 (HO1) gene therapy can modulate the release of HMGB1 by altering intracellular molecules for successful cell transplantation. After delivery of the heme oxygenase-1 (HO1) gene to islet cells using an adeno-associated viral vector (AAV), it was evaluated the changes in cytoplasmic Ca

Intermittent subcutaneous (S.C.) injection of teriparatide [PTH (1-34)] is one of the effective therapies to cure osteoporosis. However, a long-term repeated administration of teriparatide by S.C. to the patients is highly challenging. Herein, a triple padlock nanocarrier prepared by a taurocholic acid-conjugated chondroitin sulfate A (TCSA) is designed to develop an oral dosage form of recombinant human teriparatide (rhPTH). Oral administration of TCSA/rhPTH to the bilateral ovariectomized (OVX) rats resulted in the recovery of the bone marrow density and healthy serum bone parameters from the severe osteoporotic conditions. Also, it enhanced new bone formation in the osteoporotic tibias. This triple padlock oral delivery platform overcame the current barriers associated with teriparatide administration and exhibited a promising therapeutic effect against osteoporosis.

Type 2 diabetes mellitus (T2DM) is a chronic and progressive hyperglycemic condition. Glucagon-like peptide-1 (GLP1) is an incretin secreted from pancreatic β-cells and helps to produce insulin to balance the blood glucose level without the risk of hypoglycemia. However, the therapeutic application of GLP1 is limited by its intrinsic short half-life and rapid metabolic clearance in the body. To enhance the antidiabetic effect of GLP1, we designed a human cysteine-modified IgG1-Fc antibody-mediated oral gene delivery vehicle, which helps to produce GLP1 sustainably in the target site with the help of increased half-life of the Fc-conjugated nanocarrier, protects GLP1 from acidic and enzymatic degradation in the gastrointestinal (GI) tract, uptakes and transports the GLP1 formulation through the neonatal Fc receptor (FcRn), and helps to release the GLP1 gene in the intestine. Our formulation could reduce the blood glucose from about an average of 320 mg/dL (hyperglycemic) to 150 mg/dL (normal blood glucose concentration) in diabetic mice, which is about 50% reduction of the total blood glucose concentration. GLP1 (500 μg) complexed with the IgG1-Fc carrier was proven to be the optimal dose for a complete reduction of hyperglycemic conditions in diabetic mice. A significant amount of insulin production and the presence of GLP1 peptide were observed in the pancreatic islets of oral GLP1 formulation-treated diabetic mice in immunohistochemistry analysis compared to nontreated diabetic mice. The orally given formulation was completely nontoxic according to the histopathology analysis of mice organ tissues, and no mice death was observed. Our antibody-mediated oral gene delivery system is a promising tool for various oral therapeutic gene delivery applications to treat diseases like diabetes.

Intermittent subcutaneous (S.C.) injection of teriparatide [PTH (1-34)] is one of the effective therapies to cure osteoporosis. However, a long-term repeated administration of teriparatide by S.C. to the patients is highly challenging. Herein, a triple padlock nanocarrier prepared by a taurocholic acid-conjugated chondroitin sulfate A (TCSA) is designed to develop an oral dosage form of recombinant human teriparatide (rhPTH). Oral administration of TCSA/rhPTH to the bilateral ovariectomized (OVX) rats resulted in the recovery of the bone marrow density and healthy serum bone parameters from the severe osteoporotic conditions. Also, it enhanced new bone formation in the osteoporotic tibias. This triple padlock oral delivery platform overcame the current barriers associated with teriparatide administration and exhibited a promising therapeutic effect against osteoporosis.

Pancreatic β cells are responsible for insulin secretion and are important for glucose regulation in a healthy body and diabetic disease patient without prelabeling of islets. While the conventional biomarkers for diabetes have been glucose and insulin concentrations in the blood, the direct determination of the pancreatic β cell mass would provide critical information for the disease status and progression. By combining fluorination and diversity-oriented fluorescence library strategy, we have developed a multimodal pancreatic β cell probe PiF for both fluorescence and for PET (positron emission tomography). By simple tail vein injection, PiF stains pancreatic β cells specifically and allows intraoperative fluorescent imaging of pancreatic islets. PiF-injected pancreatic tissue even facilitated an antibody-free islet analysis within 2 h, dramatically accelerating the day-long histological procedure without any fixing and dehydration step. Not only islets in the pancreas but also the low background of PiF in the liver allowed us to monitor the intraportal transplanted islets, which is the first in vivo visualization of transplanted human islets without a prelabeling of the islets. Finally, we could replace the built-in fluorine atom in PiF with radioactive 18F and successfully demonstrate in situ PET imaging for pancreatic islets.

Monitoring the intensity of immune reactions is very useful to diagnose the fate of transplanted cells. For directly monitoring immune reactions after pancreatic islet cell transplantation, here graphene oxide (GO) quencher was chemically conjugated with fluorescence dye via Granzyme B (GrB)-specific peptide, so called to GO-based immunosensor. GO-based immunosensor was stably immobilized onto surface of living cells without damage of viability and functionality. The cell-surface camouflaged GO-based immunosensor could detect GrB (>2 unit/ml) secreted from immune cells, thereby dequenching fluorescence. Collectively, we anticipated that the GO-based immunosensor could be utilized to monitor cell graft rejection after cell therapy in vivo.

Pancreatic islet transplantation is a promising strategy for diabetic patients. Unfortunately, host’s immune cells rapidly reject transplanted islets. Cytoprotective gene therapy has been tried to protect them. However, the transduce yield of ex vivo gene delivery into islets is very rare due to the anatomical structure of them. Therefore, we newly designed an Arg-Gly-Asp (RGD) peptide-incorporated adenovirus vector expressing cytoprotective heme oxygenase-1 (HO1) gene (RGD-Adv-HO1) for effective gene therapy to islets via RGD–integrin interaction between adenovirus and islet. After exposure to islets, the transduced RGD-Adv-HO1 did not affect the viability and insulin secretion of islets with higher transduction efficacy. Moreover, HO1 showed cytoprotective effect from the presence of paraquat-induced reactive oxygen species (ROS). When the transduced islets were xenotransplanted into streptozotocin-induced diabetic Balb/c mice, their survival time in vivo was significantly prolonged with curing blood glucose level, which was attributed to the stable expression of HO1. Additionally, we found that RGD-Adv-HO1 showed a good synergistic effect in xenotransplanted islets when accompanied with low dose of immunosuppressants tacrolimus and anti-CD154 monoclonal antibody. Collectively, this new combinatorial remedy of RGD-Adv-HO1 transduction with low dose of immunosuppressive agents could be an effective therapy for successful outcome of islet transplantation.

Although pancreatic islet implantation is an attractive strategy for curing diabetes mellitus, implanted cells are immunologically eliminated due to early islet graft loss. One of main issues in early islet graft loss is the secretion of high-mobility group-box-1 (HMGB1) protein from the damaged islet cells, which is known as a cytokine-like factor. Therefore, regulating the activity of HMGB1 protein offers an alternative strategy for improving outcomes of islet cell therapy. To this end, we first demonstrated that HMGB1 protein could be bound to its A-box fragment (HMGB1 A-box) with higher binding affinity, resembling anti-HMGB1 antibody. To be used as a pharmaceutical protein ex vivo, TAT-labeled HMGB1 A-box-His6 (TAT-HMGB1A) was structurally modified for cellular membrane penetration. TAT-HMGB1A significantly reduced secretion of endogenous HMGB1 protein through interaction in the cytosol without any damage to the viability or functionality of the islets. When TAT-HMGB1A-treated islets were implanted into diabetic nude mice, they completely cured diabetes, as evidenced by stable blood glucose level. TAT-HMGB1A treatment could also reduce the marginal islet mass needed to cure diabetes. Furthermore, TAT-HMGB1A positively protected xenotransplanted islets from xenogeneic immune reactions. Collectively, cell-penetrable TAT-HMGB1A could be used to modulate HMGB1 activity to increase successful outcomes of ex vivo pancreatic islet cell therapy.

Pancreatic islet transplantation is a promising treatment for treatment of type 1 diabetes; however, transplantation outcomes have been disappointing due to early graft loss that is mediated by many immune responses. Immune cells not only directly damaged islet but also produced reactive oxygen species (ROS), which is highly toxic to islet cells. Metallothionein (MT) can provide protection against oxidative stress by scavenging various ranges of ROS including superoxide, hydroxyl radical, hydrogen peroxide and nitric oxide. For scavenging immune response-induced ROS, cell-penetrating Tat peptide-metallothionein (Tat-MT) was delivered into islets. The viability of Tat-MT-treated islets was not damaged during co-culture with macrophages or ROS-generating paraquat. When Tat-MT-treated islets were xenotransplanted, ROS production was significantly attenuated at the islets. Eventually, the survival time of Tat-MT-treated islets was significantly enhanced without any immunosuppressant medicine. Additionally, we confirmed that the survival time of Tat-MT-treated islets in all animals was dramatically improved when accompanied with low dose immunosuppressive agents (tacrolimus and anti-CD154 monoclonal antibody), indicating that Tat-MT delivery could have synergistic effect with immunosuppressants. Collectively, this new combination therapy of Tat-MT delivery with low dose immunosuppressant would be a powerful remedy for successful outcome of islet xenotransplantation.

The number of people suffering from insulin-independent type 2 diabetes mellitus (T2DM) is ever increasing on a yearly basis. Current anti-diabetic medications often result in adverse weight gain and hypoglycemic episodes. Hypoglycemia can be avoided with glucagon-like peptide (GLP)-1 receptor agonists, which are expensive and require daily injections that may result immune activation. This study demonstrates the use of non-viral vector based oral delivery of GLP-1 gene through enterohepatic recycling pathways of bile acids. Oral administration of the plasmid DNA (pDNA) encoding GLP-1 decreased diabetic glucose levels to the normoglycemic range with significant weight reduction in a high-fat diet (HFD) induced diabetic mouse model and a genetically engineered T2DM rat model. This novel oral GLP1 delivery system is an attractive alternative to treat late-stage T2DM conditions that require repeated insulin injection and can potentially minimize the occurrence of hypoglycemic anomalies.

Pancreatic islet implantation in the liver is a promising approach for diabetes therapy. However, 70% of the islet mass fails to be engrafted in the liver due to the instant blood-mediated inflammatory reactions (IBMIR) resulting from direct contact between islet cells and the bloodstream. To overcome this issue, direct monitoring is very important for establishing prognosis after islet cell therapy. Here we established a new type of MR contrast agent with anticoagulant activity via heparin-immobilized superparamagnetic iron oxide (HSPIO). The HSPIO was chemically conjugated onto islet surface ex vivo without damage of their viability and functionality. The conjugated HSPIO nanoparticles onto islet surface could attenuate IBMIR in vitro and in vivo. The HSPIO-conjugated islets could cure the blood glucose levels of diabetes animals after implantation. In addition, the HSPIO nanoparticles were well maintained on the transplanted islets for a long time during modulation of inflammation. Also, they allowed for stable visualization of the implanted islet cells for more than 150 days without reduction of the MRI signal. Furthermore, when HSPIO itself was intraportally injected, it was rapidly eliminated without accumulation in the liver, suggesting that HSPIO nanoparticles could only track the immobilized islet. Collectively, this HSPIO nanoparticle having MRI sensitivity and anticoagulant activity could be utilized for successful islet implantation.

An approach is presented to create arrays of quasi-spherical microdroplets by surfaces called "spheroform" inspired by micro-/nanostructures found in the back of the Namib desert beetle. Spheroform is fabricated by microarray formation of catecholamine polymers on the superhydrophobic background. Spheroform is able to precisely control spheroid sizes of therapeutic islet cells and mesenchymal stem cells. The therapeutic spheroids exhibit improved biochemical activities.

Various types of particle-based drug delivery systems have been explored for the treatment of pulmonary diseases; however, bio-distribution and elimination of the particles should be monitored for better understanding of their therapeutic efficacy and safety. This study aimed to characterize the biological properties of micro-sized discoidal polymeric particles (DPPs) as lung-targeted drug delivery carriers. DPPs were prepared using a top-down fabrication approach and characterized by assessing size and zeta potential. They were labeled with zirconium-89 ( 89 Zr), and bio-distribution studies and PET imaging were performed for 7 days after intravenous administration. Their hydrodynamic size was 2.8 ± 6.1 μm and average zeta potential was −39.9 ± 5.39 mV. At doses of 5, 12.5, and 25 mg/kg, they showed no acute toxicity in nude mice. Desferrioxamine (DFO)-functionalized 89 Zr-labeled DPPs gave a decay-corrected radiochemical yield of 82.1 ± 0.2%. Furthermore, 89 Zr-DPPs, from chelate-free labeling methods, showed a yield of 48.5 ± 0.9%. Bio-distribution studies and PET imaging showed 89 Zr-DFO-DPPs to be mainly accumulated in the lungs and degraded within 3 d of injection. However, 89 Zr-DFO-DPPs showed significantly low uptake in the bone. Overall, our results suggested micro-sized DPPs as promising drug delivery carriers for the targeted treatment of various pulmonary diseases.

Pancreatic islet transplantation is a promising method for treatment of type 1 diabetes mellitus. However, transplanted islets can be destroyed due to host immune reactions. To immunologically protect transplanted islets, here an immunoprotective microfiber including islets by using a polydimethylsiloxane (PDMS)-based microfluidic device is newly designed. A cylindrical-flow channel in the microfluidic platform is used for producing collagen-alginate composite (CAC) fibers. This enables mass production and uniform diameter distribution (

Graphene oxide (GO) has received increasing attention in bioengineering fields due to its unique biophysical and electrical properties, along with excellent biocompatibility. The application of GO nanoparticles (GO-NPs) to engineer self-renewal and differentiation of human fetal neural stem cells (hfNSCs) is reported. GO-NPs added to hfNSC culture during neurosphere formation substantially promote cell-to-cell and cell-to-matrix interactions in neurospheres. Accordingly, GO-NP-treated hfNSCs show enhanced self-renewal ability and accelerated differentiation compared to untreated cells, indicating the utility of GO in developing stem cell therapies for neurogenesis.

In the pancreatic islet transplantation for curing diabetes mellitus, reaggregation of pancreatic islet single cells for making ‘islet spheroid’ is considered as a useful strategy in terms of islet viability and gene delivery into islet. However, during aggregation of islet spheroid, their insulin secretion pattern was not recovered to that of intact islets. We have developed a new cell-culture system using poly(dimethylsiloxane) (PDMS)-based concave-patterned film for aggregating islet spheroid. Here we optimized the effective formation of islets spheroid by using the concave-patterned film with different microwell diameter (300, 500, and 700-μm). Aggregated islet spheroids in different concave microwells showed same morphology with a limited size range and diameter dependency of microwell, which was similar to that of intact islet. However, islet spheroid in flat-bottomed dish (control) showed random aggregation with broad size distribution. Interestingly, insulin secretion pattern of islet spheroids in 300 and 500-μm concave patterned-film was same to that of intact islets, which was caused by the improved cell-to-cell interaction that was confirmed by electron microscope and strong phalloidin immunostain. However, islet spheroids in concave patterned-film with 700-μm diameter and flat-dish did not show normal insulin secretion in accordance with different glucose concentration, which might be caused by weak cell-to-cell interaction. On the other hand, the viability of islet spheroids in concave patterned-film was not affected during cultivation due to biocompatibility of PDMS polymer. Collectively, we demonstrated that PDMS-based concave-patterned film could be used for designing islet spheroids with improved cellular functionality and size uniformity to cure diabetes mellitus. Open image in new window

Radioactivity of high concentrations have existed in the handling nuclear materials in hot cell of PIEF(Post Irradiation Examination Facility). The exhaust filtration system was enabled to process cylindrical filters by using a manipulator in the hot cell. By establishing a double filtration system with two filters, backup protection against leakage or failure of the first is provided by the second filter. Additionally, this a arrangement is arrange intended to increase the total filtration efficiency. The result of the pressure drop changing in the air flow of the cylindrical and HEPA filters is observed by a curved line. A filtering efficiency of more than 99.99% to particle appears in the upstream and downstream during the efficiency test on the HEPA filters. The V-pleats type had a lower pressure drop than the separator type. There was no damage during usage and was found to be suitable with high capacity of air volume. Therefore, by carrying out performance tests of the exhaust filtration system, efficiency and safety can be achieved.

Transplantation of insulin-secreting pancreatic islets can provide real-time regulation of blood glucose in patients with type 1 diabetes mellitus. Currently, noninvasive and repetitive monitoring of islet engraftment and function is an emerging and promising modality for successful islet transplantation. Here we report a new technique for highly sensitive in vivo magnetic resonance (MR) imaging of transplanted pancreatic islets. To this end, heparinized superparamagnetic iron oxide (heparin-SPIO) nanoparticle was newly synthesized for chemical conjugation onto islet surface. Compared to typical cellular labeling of Feridex® via random endocytosis, chemical conjugation of heparin-SPIO was stable and improved the hypointensity of transplanted islets due to surface modification of every islet. These heparin-SPIO-conjugated islets showed normal viability and insulin secretion, and were quantified by spin echo T2-weighted MR imaging with linear correlation depending on transplanted islet mass in vitro and in vivo for 30 days. Also, from the immunohistochemistry, we confirmed the existence of heparin-SPIO and insulin biosynthesis in transplanted islets. However, Feridex-uptake islets showed late glucose responsiveness according to changing glucose concentration although they could normally control the blood glucose levels in diabetic mouse. Thus, we anticipate that this surface labeling with heparin-SPIO can be directly applicable for MR imaging of transplanted islets.

Pancreatic islet transplantation is an ideal therapy for type I diabetes. Interestingly, small size of islets may exhibit superior performance in transplantation in terms of function and hypoxia. To this end, large size of islets was dissociated to generate islet single cells and they were re-aggregated to make small size of islet-cell-cluster (ICC). However, the function of insulin secretion from ICCs was damaged due to weak cell-to-cell interaction. To overcome this limitation, here the dispersed islet single cells were re-aggregated at the concave microwell array that was fabricated with poly(dimethylsiloxane) (PDMS) by using soft-lithography and mold-replication technology. The aggregated ICCs had uniform size, and their insulin secretion in response to the change of low and high glucose concentration was similar to that of intact islets since the cell-cell interaction of islet single cells was strongly induced due to the shape of concave microwell. Also, after cultivation for 2 weeks, the expression of hypoxia-induced factor- 1α(HIF-1α; a hypoxia marker protein) in the ICCs was rare; meaning that hypoxic state in the central area of ICCs was attenuated. On the other hand, the ICCs generated in flat-bottomed wells had abnormal insulin secretion caused by weak cell-to-cell interaction. In addition, after cultivation for 2 weeks, the generated ICCs in flat-bottomed well had weak signal of HIF-1α, while intact islets had its strong signal. Therefore, new culture system using concave microwell array may be a promising alternative to success fully prepare functional ICCs. Open image in new window

Irradiation behavior of the reactor pressure vessel SA508 class 3 steel weld metals was examined by Charpy V Notch impact specimens. The specimens were exposed to a fluence of 2.8x1019 neutrons(n)/cm 2 (E>1 MeV) at 288℃. The irradiation damage of weld metal was evaluated by comparison between unirradiated and irradiated specimens in terms of absorbed energy and lateral expansion. The specimens for neutron irradiation were welded by submerged arc welding process at a heat input of 3.2 kJ/mm which showed good toughness in terms of weld microstructure, absorbed energy and lateral expansion. The post-irradiation Charpy V notch 41J and 68J transition temperature elevation were 65℃ and 70℃, respectively. This elevation was accompanied by a 20% reduction in Charpy V notch upper shelf energy level. The lateral expansion at 0.9㎜ irradiated Charpy specimens showed temperature elevation of 65℃ and was greatly decreased due to radiation damage. Key Words : Radiation damage, SA508 class 3, Submerged arc welding, Impact test, Absorbed energy, lateral expansion, Transition temperature elevation

While clinical islet transplantation is being investigated as a useful method to cure diabetes mellitus (DM), the outcome of this therapy remains imperfect. This is mainly due to early graft rejection of transplanted islets by the instant blood-mediated inflammatory reaction (IBMIR) or islet ischemia. Therefore, it is important to develop imaging tools for transplanted islets, so that real-time treatment can be used to protect the islets from host immune reactions if required. Recently, synthetic nanoparticles are emerging in the area of biomedical imaging. Innovative nanoparticle developments have significantly enhanced the versatility of different imaging modalities. Nanoparticles represent potential probes to monitor transplanted islets through magnetic resonance imaging (MRI), positron emission tomography (PET), and optical imaging. Nanoparticles can be delivered to transplanted islets by conjugation onto the cell membrane, intracellular delivery, cellular membrane receptor or transporter targeting. Visualizing the survival, function, and biodistribution of transplanted islets may suggest an appropriate treatment, which will enhance islet graft survival. This chapter focuses on recent advances and applications of various nanoparticle-based imaging techniques used in molecular imaging to monitor transplanted islets.

The hot cell facility for research activities related to the electrolytic reduction of spent fuel, which is designed to permit safe handling of radioactive materials up to 1,385 TBq, is planned to be built. The primary radiological safety design goal for the facility is to keep the radiation exposures to workers as-low-as-reasonably-achievable (ALARA). To meet this goal, the facility is designed to keep gamma and neutron radiation lower than the recommended dose-rate in normally occupied areas.The calculations by QAD-CGGP and MCNP-4C are used to evaluate the proposed engineering design concepts that would provide acceptable dose-rates during the normal operation in hot cell facility. The calculated maximum effective gamma dose-rates by QAD-CGGP on the surfaces of the facility at operation area and at service area are 2.97×10-3 and l.0l×l0-1 mSv/h, respectively. Those calculated by MCNP-4C are 2.99×10-3 and 7.88×10-2 mSv/h, respectively. The dose-rates contributed by neutrons are one order of magnitude les...

There is a clinical need for an alternative labeling agent for magnetic resonance imaging (MRI) in islet transplantation. We aimed to evaluate the feasibility of islet MRI using ferumoxytol, which is the only clinically-available ultrasmall superparamagnetic iron oxide. We compared islet function and viability of control islets and islets labeled with ferumoxytol and/or a heparin-protamine complex (HPF). Efficacy of ferumoxytol labeling was assessed in both ex vivo and in vivo models. Labeling for 48 h with HPF, but not up to 800 μg/mL ferumoxytol, deranged ex vivo islet viability and function. The T2∗ relaxation time was optimal when islets were labeled with 800 μg/mL of ferumoxytol for 48 h. Prussian blue stain, iron content assay, transmission electron microscopy (TEM) supported internalization of ferumoxytol particles. However, the labeling intensity in the ex vivo MRI of islets labeled with ferumoxytol was much weaker than that of islets labeled with ferucarbotran. In syngeneic intraportal islet transplantation, there was a correlation between the total area of visualized islets and the transplanted islet mass. In conclusion, islet MRI using ferumoxytol was feasible in terms of in vitro and in vivo efficacy and safety. However, the weak labeling efficacy is still a hurdle for the clinical application.

While subcutaneous tissue has been proposed as a potential site for pancreatic islet transplantation, concern remains that the microvasculature of subcutaneous tissue is too poor to support transplanted islets. In an effort to overcome this limitation, we evaluated whether fibrin gel with human adipose-derived stem cells (hADSCs) and rat pancreatic islets could cure diabetes mellitus when transplanted into the subcutaneous space of diabetic mice. Subcutaneously co-transplanted islets and hADSCs showed normalization of the diabetic recipient's blood glucose levels. The result was enhanced by co-treatment of fibroblast growth factor-2 (FGF2) in the fibrin gel. The hADSCs enhanced islet viability after transplantation by secreting various growth factors that can protect islets from hypoxic damage. Afterward, hADSCs could maintain islet viability by recruiting new microvasculature nearby the transplanted islets via overexpression of vascular endothelial growth factor (VEGF). The hADSCs did not directly differentiate into endothelial cells (no detection of biomarkers of human endothelial cells), but showed evidence of differentiation toward insulin-secreting cells (detection of human insulin). Mice receiving islet transplantation alone did not become normoglycemic. Collectively, co-transplantation of fibrin gel with islets and hADSCs will expand the indications for islet transplant therapy and the potential clinical application of cell-based therapy.

Subcutaneous site is ideal for clinical islet transplantation because it has the advantage of simple operation procedure under local anesthesia and can be biopsied when needed. However, the transplantation outcomes at subcutaneous site have been disappointing due to hypoxia-induced oxidative stress by poor vascularization. We hypothesized that subcutaneously transplanted islets would have hypoxia resistance by using internalization of metallothionein (MT), an antioxidant scavenging enzyme, which was mediated by fusion between MT and cell penetrating Tat peptide. The Tat-MT was dose-dependently transduced into islets without any damage. Tat-MT-treated islets could be protected from oxidative stress induced by intracellular nitric oxide donor, sodium nitroprusside (SNP). When Tat-MT-treated islets were subcutaneously transplanted into diabetic nude mice, they normally controlled the blood glucose levels without severe fluctuation (median survival time (MST): > 30 days), whereas most untreated islets were rejected (MST 17 days). From the intraperitoneal glucose tolerance test 5 days after posttransplantation, glucose responsiveness of Tat-MT-treated islets was similar to that of normal healthy mice, while untreated islets had delayed glucose responsiveness. From the results of immunohistochemical stain, Tat-MT-treated islets had strong anti-insulin positive cells and lower anti-HIF-1α positive cells. However, untreated islets had rare anti-insulin positive cells and strong anti-HIF-1α-positive cells. Collectively, these findings demonstrated that Tat-MT delivery into islet could offer a new strategy for successful islet transplantation under subcutaneous space.

Islet cell transplantation is a promising option for the restoration of normal glucose homeostasis in patients with type 1 diabetes. Because graft volume is a crucial issue in islet transplantations for patients with diabetes, we evaluated a new method for increasing functional tissue yield in xenogeneic grafts of encapsulated islets. Islets were labeled with three different superparamagnetic iron oxide nano particles (SPIONs; dextran-coated SPION, siloxane-coated SPION, and heparin-coated SPION). Magnetic separation was performed to separate encapsulated islets from the empty capsules, and cell viability and function were tested. Islets labeled with 1000 μg Fe/ml dextran-coated SPIONs experienced a 69.9% reduction in graft volume, with a 33.2% loss of islet-containing capsules. Islets labeled with 100 μg Fe/ml heparin-coated SPIONs showed a 46.4% reduction in graft volume, with a 4.5% loss of capsules containing islets. No purification could be achieved using siloxane-coated SPIONs due to its toxicity to the primary islets. SPION labeling of islets is useful for transplant purification during islet separation as well as in vivo imaging after transplantation. Furthermore, purification of encapsulated islets can also reduce the volume of the encapsulated islets without impairing their function by removing empty capsules.

Magnetic resonance imaging (MRI) is a tremendous modality for noninvasive cell tracking. To this end, superparamagnetic iron oxide (SPIO), one of the MRI contrast agents, should be labeled to the cells before transplantation. Currently, cellular labelling with SPIOs such as Feridex and Resovist is generally carried out through their engulfment into cytosol via endocytosis. However, the labelling efficacy via endocytosis is relatively low due to their non-specific random engulfment and degradation in the cytosol. To overcome these limitations, transfection agents such as poly-L-lysine and lipofectamine are complexed with SPIOs and treated to the cells. However, these strategies should be optimized due to the cytotoxicity of transfection agents themselves. Recently, there were developments of chemical conjugation of SPIOs onto cellular membrane. To this end, the surface of SPIOs was coated with heparin polysaccharide and chemically conjugated with collagen matrix layer of cell surface by using linker polymer, which was stably maintained in vivo. This new remedy can overcome the limitations of cell labelling via endocytosis. Collectively, these strategies could be applied for noninvasive imaging of MRI after cell transplantation in vivo.

Human mesenchymal stem cells (hMSCs) offer significant therapeutic potential in the field of regenerative medicine and high-resolution magnetic resonance imaging (MRI) is useful modality to visualize in vivo kinetics of transplanted stem cells. For successful MR imaging, there is a great need for effective contrast agents for stem cell labeling with high uptake yield and low toxicity. Here, we present superparamagnetic iron oxide (SPIO) nanoparticles coated with unfractionated heparin (UFH-SPIO) as a new negative contrast agent for in vivo MR imaging of hMSCs. The uptake of UFH-SPIO by hMSCs was effective without the aid of transfection agents, which was dependent on the concentration and exposure time. The uptake efficiency of UFH-SPIO was greater than that of DEX-SPIO (SPIO coated with dextran) by approximately 3 folds when treated for 1 h. TEM and Prussian blue staining confirmed that UFH-SPIO nanoparticles were internalized into the cytosol of hMSCs which existed during in vitro subculture for 28 days. Low temperature endocytosis inhibition assay demonstrated that the incorporation of UFH-SPIO into hMSCs was likely to be mediated by endocytosis. When the phantom of UFH-SPIO-labeled hMSCs was visualized with 3-T T 2 -weighted MRI, the hypointensity signals of UFH-SPIO-labeled hMSCs were linearly correlated with the concentration of the nanoparticles. The cellular labeling using UFH-SPIO did not reduce the viability, proliferation or differentiation potential to osteogenic and adipogenic lineages of hMSCs. When the UFH-SPIO-labeled hMSCs were transplanted into the left renal subcapsular membranes of nude mice, they were successfully visualized and detected by T 2 and T 2 ∗ -weighted MRI for a month. Collectively, these results suggest that UFH-SPIO nanoparticles are promising as a new MRI contrast agent for in vivo long-term tracking of hMSCs.

In this study, we suggest in situ islet spheroid formation and encapsulation on a single platform without replating as a method for producing mono-disperse spheroids and minimizing damage to spheroids during encapsulation. Using this approach, the size of spheroid can be controlled by modulating the size of the concave well. Here, we used 300 μm concave wells to reduce spheroid size and thereby eliminating the central necrosis caused by large volume. As the encapsulation material, we used alginate and collagen-alginate composite (CAC), and evaluated their suitability through diverse in vitro tests, including measurements of viability, oxygen consumption rate (OCR), hypoxic damage to encapsulated spheroids, and insulin secretion. For in situ encapsulation, alginate or CAC was spread over a concave microwell array containing spheroids, and CaCl(2) solution was diffused through a nano-porous dialysis membrane to achieve uniform polymerization, forming convex structures. By this process, the formation of uniform-size islet spheroids and their encapsulation without an intervening replating step was successfully performed. As a control, intact islets were evaluated concurrently. The in vitro test demonstrated excellent performance of CAC-encapsulated spheroids, and on the basis of these results, we transplanted the islet spheroids-encapsulated with CAC into the intraperitoneal cavity of mice with induced diabetes for 4 weeks, and evaluated subsequent glucose control. Intact islets were also transplanted as control to investigate the effect of encapsulation. Transplanted CAC-encapsulated islet spheroids maintained glucose levels below 200 mg/dL for 4 weeks, at which they were still active. At the end of the implantation experiment, we carried out intraperitoneal glucose tolerance test (IPGTT) in mice to investigate whether the implanted islets remained responsive to glucose. The glucose level in mice with CAC-encapsulated islet spheroids dropped below 200 mg/dL 60 min after glucose injection and was stably maintained. In conclusion, the proposed encapsulation method enhances the viability and function of islet spheroids, and protects these spheroids from immune attack.

Background Pancreatic islet encapsulation is one way to address the disadvantages of islet transplantation. Not only does encapsulation involve bidirectional diffusion of nutrients, oxygen, and glucose, but also it protects the graft from the recipient’s immune reaction. The high mobility group box 1 (HMGB1), one of higher expression proteins in islet, can be secreted from transplanted islets and induce the inflammation. Therefore, the regulation of HMGB1-mediated inflammation is very important for successful islet transplantation. In this study, we used the HMGB1 A box, an antagonist of HMGB1 receptor in the immune cells, in the encapsulation of isolated islets as a new strategy. Result For co-encapsulation of HMGB1 A box protein with islets, we evaluated the distribution of alginate bead diameter. The average diameter of empty alginate bead was similar to that of alginate bead with islets. When different concentrations of HMGB1 A box protein was co-encapsulated with islets, it did not affect the viability and insulin secretion function of the islets. When the alginate beads with islets plus HMGB1 A box protein were cultured with macrophage, the amount of TNF-α secreted from the macrophages was significantly attenuated when compared to cultivation of unencapsulated islets or encapsulated islets. When the alginate beads with islets plus HMGB1 A box protein were intraperitoneally xenotransplanted into the diabetic mice, the survival rate of the islets was strongly improved with 2-fold. Conclusion Collectively, these results suggested that the encapsulation of HMGB1 A box protein might offer a protective effect in islet transplantation.