AP39, a Mitochondrial-Targeted H2S Donor, Improves Porcine Islet Survival in Culture.

The rapid deterioration of transplanted islets in culture is a well-established phenomenon. We recently reported that pancreas preservation with AP39 reduces reactive oxygen species (ROS) production and improves islet graft function. In this study, we investigated whether the addition of AP39 to the culture medium could reduce isolated islet deterioration and improve islet function. Isolated islets from porcine pancreata were cultured with 400 nM AP39 or without AP39 at 37 °C. After culturing for 6-72 h, the islet equivalents of porcine islets in the AP39(+) group were significantly higher than those in the AP39(-) group. The islets in the AP39(+) group exhibited significantly decreased levels of ROS production compared to the islets in the AP39(-) group. The islets in the AP39(+) group exhibited significantly increased mitochondrial membrane potential compared to the islets in the AP39(-) group. A marginal number (1500 IEs) of cultured islets from each group was then transplanted into streptozotocin-induced diabetic mice. Culturing isolated islets with AP39 improved islet transplantation outcomes in streptozotocin-induced diabetic mice. The addition of AP39 in culture medium reduces islet deterioration and furthers the advancements in ß-cell replacement therapy.

Pancreas Preservation in Modified Histidine-lactobionate Solution Is Superior to That in University of Wisconsin Solution for Porcine Islet Isolation.

BACKGROUND: We previously reported that modified extracellular-type trehalose-containing Kyoto (MK) solution, which contains a trypsin inhibitor (ulinastatin), significantly improved the islet yield compared with University of Wisconsin (UW) preservation, which is the gold standard for organ preservation for islet isolation. In this study, we evaluated the efficiency of a modified histidine-lactobionate (MHL) solution in addition to UW or MK solution. The MHL solution has a high sodium-low potassium composition with low viscosity compared with the UW solution. Moreover, similar to MK solution, MHL solution also contains ulinastatin. METHODS: Porcine pancreata were preserved in UW, MK, or MHL solution, followed by islet isolation. An optimized number (1500 IE) of isolated islets from each group were then transplanted into streptozotocin-induced diabetic mice. RESULTS: The islet yield before and after purification was significantly higher in the MHL group than in the UW group. On the contrary, the islet yield before and after purification was not significantly different between the MHL and MK groups. Preserving the porcine pancreata in MHL solution improved the outcome of islet transplantation in streptozotocin-induced diabetic mice compared with that in UW solution. CONCLUSIONS: Pancreas preservation with MHL solution preserves islet function better than UW solution. The effect of MHL solution is similar to that of MK solution, suggesting that MHL solution can be used as an alternative to MK solution for pancreatic islet transplantation.

Pancreas Preservation with a Neutrophil Elastase Inhibitor, Alvelestat, Contributes to Improvement of Porcine Islet Isolation and Transplantation.

For pancreatic islet transplantation, pancreas procurement, preservation, and islet isolation destroy cellular and non-cellular components and activate components such as resident neutrophils, which play an important role in the impairment of islet survival. It has been reported that inhibitors of neutrophil elastase (NE), such as sivelestat and α1-antitrypsin, could contribute to improvement of islet isolation and transplantation. In this study, we investigated whether pancreatic preservation with alvelestat, a novel NE inhibitor, improves porcine islet yield and function. Porcine pancreata were preserved with or without 5 µM alvelestat for 18 h, and islet isolation was performed. The islet yields before and after purification were significantly higher in the alvelestat (+) group than in the alvelestat (-) group. After islet transplantation into streptozotocin-induced diabetic mice, blood glucose levels reached the normoglycemic range in 55% and 5% of diabetic mice in the alvelestat (+) and alvelestat (-) groups, respectively. These results suggest that pancreas preservation with alvelestat improves islet yield and graft function and could thus serve as a novel clinical strategy for improving the outcome of islet transplantation.

Pancreas preservation with amphotericin B deteriorates islet yield for porcine islet isolation.

BACKGROUND: Amphotericin B is a crucial agent in the management of serious systemic fungal infections. It is also known to be cytotoxic. In this study, we evaluated the effect of amphotericin B added to the preservation solution on islet yield during islet isolation. METHODS: Porcine pancreata were preserved in the preservation solution with or without amphotericin B (0.25 µg/mL) for approximately 18 hours at 4°C, and then islet isolation was performed. An optimized number (1750 IE) of isolated islets from each group were transplanted into streptozotocin-induced diabetic mice. The culture of isolated islets and acinar tissue with amphotericin B was also evaluated. RESULTS: The islet yield before and after purification in the amphotericin B (-) group was significantly higher than that in the amphotericin B (+) group. After islet transplantation into diabetic mice, blood glucose levels reached the normoglycemic range, with 50% and 0% of that of the diabetic mice in the amphotericin B (-) and amphotericin B (+) groups, respectively. In the culture study, amphotericin B was found to be cytotoxic to porcine islets and acinar tissue. CONCLUSIONS: Amphotericin B added to the preservation solution deteriorates islet yield during porcine islet isolation. Thus, the use of amphotericin B should be considered carefully for the preservation of the pancreas for islet isolation and islet culture before islet transplantation.

Preservation of pancreas in the University of Wisconsin solution supplemented with AP39 reduces reactive oxygen species production and improves islet graft function.

Ischemia-reperfusion injury (IRI) results in increased rates of delayed graft function and early graft loss. It has recently been reported that hydrogen sulfide (H2 S) protects organ grafts against prolonged IRI. Here, we investigated whether the preservation of pancreas in University of Wisconsin (UW) solution supplemented with AP39, which is a mitochondrial-targeted H2 S donor, protected pancreatic islets against IRI and improved islet function. Porcine pancreata were preserved in the UW solution with AP39 (UW + AP39) or the vehicle (UW) for 18 h, followed by islet isolation. The islet yields before and after purification were significantly higher in the UW + AP39 group than in the UW group. The islets isolated from the pancreas preserved in UW + AP39 exhibited significantly decreased levels of reactive oxygen species (ROS) production and a significantly increased mitochondrial membrane potential as compared to the islets isolated from the pancreas preserved in the vehicle. We found that the pancreas preserved in UW + AP39 improved the outcome of islet transplantation in streptozotocin-induced diabetic mice. These results suggest that the preservation of pancreas in UW + AP39 protects the islet grafts against IRI and could thus serve as a novel clinical strategy for improving islet transplantation outcomes.

Pancreas preservation in extracellular-type p38 inhibitor-containing solution improves islet yield for porcine islet isolation.

BACKGROUND: For islet transplantation, pancreas preservation and islet isolation activate p38, which is a member of the stress-activated group of mitogen-activated protein kinases (MAPKs). In this study, we evaluated an extracellular-type p38 inhibitor-containing (EP) solution with University of Wisconsin (UW) solution, the gold standard for organ preservation. The EP solution has high sodium-low potassium composition with low viscosity compared to UW solution. Moreover, EP solution contains a recently developed p38 inhibitor (11R-p38I110 ) from our laboratory. METHODS: Porcine pancreata were preserved in UW, EP, or EP-P solution (EP solution without 11R-p38I110 ), and then islet isolation was performed. An optimized number (1500 IE) of isolated islets from each group were transplanted into streptozotocin-induced diabetic mice. RESULTS: The islet yield before and after purification was significantly higher in the EP group than in the UW group. The islet yield before and after purification was not significantly different between the EP and EP-P groups; however, the EP solution prevented a reduction in the number of islets during culture. Western blot analysis showed that p38 activation was attenuated by EP solution. For islet transplantation into streptozotocin-induced diabetic mice, pancreas preservation in EP solution improved the outcome of islet transplantation. CONCLUSIONS: Pancreas preservation with EP solution preserved islet function better than with UW solution. The advantages of EP solution over UW solution may include the inhibition of p38 activity as well as the composition of the solution.