In Vitro Assessment of Human Islet Vulnerability to Instant Blood-Mediated Inflammatory Reaction (IBMIR) and Its Use to Demonstrate a Beneficial Effect of Tissue Culture

Abstract

Culture of human pancreatic islets is now routinely carried out prior to clinical islet allotransplantation, using conditions that have been developed empirically. One of the major causes of early islet destruction after transplantation is the process termed instant blood-mediated inflammatory reaction (IBMIR). The aim of this study was to develop in vitro methods to investigate IBMIR and apply them to the culture conditions used routinely in our human islet isolation laboratory. Freshly isolated or precultured (24 h, 48 h) human islets were incubated in either ABO-compatible allogeneic human blood or Hank's buffered salt solution (HBSS) for 1 h at 37°C. Tissue factor (TF) expression and leukocyte migration were assessed by light microscopy. TF was also quantified by ELISA. To assess b-cell function, glucose-stimulated insulin secretion (GSIS) assay was carried out. The extent of islet b-cell damage was quantified using a proinsulin assay. Islets cultured for 24 h had higher GSIS when compared to freshly isolated or 48-h precultured islets. Freshly isolated islets had significantly higher TF content than 24-h and 48-h precultured islets. Incubation of freshly isolated human islets in allogeneic human blood released 6.5-fold higher level of proinsulin in comparison to freshly isolated human islets in HBSS. The high level of proinsulin released was significantly attenuated when precultured islets (24 h or 48 h) were exposed to fresh blood. Histological examination of fresh islets in blood clot showed that some islets were fragmented, showing signs of extraislet insulin leakage and extensive neutrophil infiltration and necrosis. These features were markedly reduced when the islets were cultured for 24 h. These results suggest that our standard 24-h islet culture is markedly beneficial in attenuating IBMIR, as evidenced by increased GSIS, lower content of TF, decrease islet fragmentation, and proinsulin release.

Does Islet Size Really Influence Graft Function After Clinical Islet Transplantation?

“…Changes in islet morphology (rounding up) during culture and the significant reduction in islet number after culture in the present study is evidence of just such a process. We have also recently shown that short-term tissue culture ameliorates the destruction of human islets by IBMIR [38]. Thus it is possible that islets cultured for short periods prior to transplant are effectively preconditioned and any correlation between islet size and function in grafts is lost or reduced.…”

Impact of Pancreatic Rat Islet Density on Cell Survival during Hypoxia

“…Based on clinical hepatic islet transplantations, 10,000 IEQ/kg is needed to reach normoglycaemia [ 1 ]. Unfortunately, it has been estimated that approximately 40% of the transplanted islets will die during the instant blood mediated inflammatory reaction (IBMIR) [ 23 ] and another 10 to 20% will die during vascularization [ 24 ]. These losses can be avoided when using a bioartificial pancreas.…”

Transplantation technologies for treatment of carbohydrate metabolism disorders

“…Наиболее изученным последствием внутрипортального введения островков является мгновенная кровь-опосредованная воспалительная реакция (The instant blood-mediated inflammatory reaction, IBMIR), представляющая собой иммунный ответ, развивающийся сразу после пересадки и реализующийся образованием тромбов и инфильтрацией тучными клетками и макрофагами [27,28]. Микротромбы, состоящие из тромбоцитов, нейтрофилов и моноцитов, появляются уже через 5 минут после инфузии островков [29,30]. Реакция инициируется коагуляционным каскадом, пик которого приходится на 6-12 часов после инфузии островков [31].…”