Inhibitory Effect of Thrombomodulin on HMGB1-Stimulated IFN-γ Production of Hepatic NKT and Gr-1+ Cells, Facilitating to Prevent Early Loss of Transplanted Islets in the Liver of Mice

Abstract

Currently, insulin independence in patients with IDDM has been hardly achieved after pancreatic islet transplantation (tx) from a single donor mainly due to early loss of transplanted islets. Previously, we have shown in mice that pancreatic islets contain abundant HMGB1, released into circulation and triggering NKT cell-dependent IFN-γ production of Gr-1+ cells (neutrophils) in the liver receiving islets (JCI 2010) which is an essential component of early loss of transplanted islets (JEM 2005). In the present study, we hypothesize that the beneficial effect of thrombomodulin (TM) on engraftments of islets in the liver might be mediated through inhibition of HMGB1-NKT-Gr-1+cell pathways since TM has been reported to produce sequestration of HMGB1 (JCI 2005). Hyperglycemia of STZ-diabetic mice (C57BL/6) receiving 200 syngenic islets from a single donor into the liver via the portal vein was ameliorated when TM was administered IV for 3 times (0, 12 and 24hrs, 200μg/injection/mouse, n=5), while those of mice (n=5) treated with saline did not. Morphologically, intact islet grafts with well granulated β cells were seen in the liver of normoglycemic recipients treated with TM, while in contrast, degenerated islets with de-granulated β cells were seen in hyperglycemic control mice. IPGTT (1g/kg glucose) at 60 days after tx revealed that the glucose tolerance of TM-treated mice receiving 200 islets (n=5) was superior to that of normoglycemic mice receiving 400 islets without TM treatment (n=5). FACS analysis showed that IFN-γ production of NKT cells and Gr-1+ cells accumulated in the liver of mice receiving islets and treated with saline was up-regulated at 6 hours after tx as reported previously, while in contrast, that in mice receiving 200 islets and treated with TM was down-regulated with reduction in number of infiltrating Gr-1+ cells. IFN-γ production of NKT cells and Gr-1+ cells accumulated in the liver of mice at 2 hours after the IV injection of HMGB1 (100 μg/injection/mouse) without islet transplantation was up-regulated, while in marked contrast, that in mice treated with TM (500μg, iv) prior to the HMGB1 injection was down-regulated with reduced number of infiltrated Gr-1+ cells. These findings indicate that TM prevents the early loss of transplanted islets in the liver of mice through inhibition of stimulatory effects of HMGB1. Importantly, recombinant TM has already been used in clinics with great impact on its efficacy for the treatment of sepsis with disseminated intravascular coagulation in Japan. Thus, the safety issue regarding the clinical use of TM has been cleared and it seems ready to apply this to clinical islet transplantation to improve the efficiency of intraportal islet transplantation.