Implantation of standardized beta-cell grafts in a liver segment of IDDM patients: graft and recipient characteristics in two cases of insulin-independence under maintenance immunosuppression for prior kidney graft

Abstract

Islet transplantation can restore insulin production in insulin-dependent diabetic (IDDM) patients in whom this capacity had been lost for many years [1±7]. This observation has been made in kidney and liver recipients, where advantage is taken of the need for a continuous immune suppression. Survival of the grafts is variable, but generally less than 1 year. A prolonged beta-cell function with a state of insulin-independence after one year was achieved in 7 % of the cases recorded by the Islet Transplant Reg- Diabetologia (1998) Summary Islet allografts in insulin-dependent diabetic (IDDM) patients exhibit variable survival lengths and low rates of insulin-independence despite treatment with anti-T-cell antibodies and maintenance immunosuppression. Use of poorly characterized freshly isolated preparations makes it difficult to determine whether failures are caused by variations in donor tissue. This study assesses survival of standardized beta-cell allografts in C-peptide negative IDDM patients on maintenance immunosuppression following kidney transplantation and without receiving anti-T-cell antibodies or additional immunosuppression. Human islets were isolated from pancreatic segments after maximal 20 h cold-preservation. During culture, preparations were selected according to quality control tests and combined with grafts with standardized cell composition (³ 50 % beta cells), viability ( ³ 90 % ), total beta-cell number (1 to 2 × 10 6 /kg body weight) and insulin-producing capacity (2 to 4 nmol × graft ±1 × h ±1 ). Grafts were injected in a liver segment through the repermeabilized umbilical vein. After 2 weeks C-peptide positivity, four out of seven recipients became C-peptide negative; two of them were initially GAD 65 -antibody positive and exhibited a rise in titre during graft destruction. The other three patients remained C-peptide positive for more than 1 year, two of them becoming insulin-independent with near-normal fasting glycaemia and HbA 1 c ; they remained GAD 65 -and islet cell antibody negative. The three patients with surviving grafts presented a history of anti-thymocyte globulin therapy at kidney transplantation. Long-term surviving grafts increased C-peptide release following intravenous glucagon or oral glucose but not following intravenous glucose. Thus, cultured human beta-cells can survive for more than 1 year in IDDM patients on maintenance anti-rejection therapy for a prior kidney graft and without the need for an increased immunosuppression at the time of implantation. The use of functionally standardized beta-cell grafts helps to identify recipient and graft factors which influence their survival and metabolic effects. Insulin-independence can be achieved by injection of 1.5 million beta-cells per kg body weight in a liver segment. These beta-cell implants respond well to adenylcyclase activators but poorly to glucose. [Diabetologia (1998) 41: 452±459]

Formation of insulin-positive cells in implants of human pancreatic duct cell preparations from young donors

“…We examined whether isolated postnatal human duct-cell preparations have this property when transplanted under the kidney capsule of normal mice. A duct cell-containing fraction was obtained during the islet-cell isolation procedure that is conducted for our clinical trial on beta-cell transplantation, and is further enriched during suspension culture [19,20,21,22,23]. Grafts are then prepared that contain approximately 0.5 million cells and mainly consist of CK 19 marked duct cells (80%) with acinar cells as major contaminant (18%); endocrine and insulin-positive cells were rare (<2%) but consistently present.…”

“…We used duct-cell preparations from 14 donors of age 1 to 63 years. The steps of the isolation procedure were similar to those described previously [19,20] as were the culture conditions [22,23]. Briefly, after collagenase digestion of the pancreas and Ficoll gradient centrifugation, the residual cell fraction that remained after islet isolation was harvested, washed and cultured in suspension for 7±3 days (mean ± SD) at 37°C in Ham's F10 medium (BioWhittaker, Md., USA) containing 0.5% bovine serum albumin.…”

“…It has been proposed that addition of an extracellular matrix and selected agents might induce differentiation of duct cells to islet cells [18]. It is important to know whether a similar process can occur in human islet grafts which are known to be heavily contaminated by pancreatic duct cells [19,20,21]. To investigate this possibility we prepared human pancreatic duct cells [22,23] and followed their cellular composition and cell population volumes over 10 weeks after transplantation in nude mice.…”

IL-1β and IFN-γ induce the expression of diverse chemokines and IL-15 in human and rat pancreatic islet cells, and in islets from pre-diabetic NOD mice

“…Human islet cells were isolated from pancreases obtained from five adult heart-beating organ donors [8]. The mean donor age (± SEM) was 21±6 years and the islet preparations contained 70±2% beta cells and 15% alpha cells, as assessed by immunocytochemistry and electron microscopy [33,35]. For the RT-PCR experiments, human islets cells (10 5 ) were cultured in Ham's F10 medium [36], and exposed for 6 h or 24 h to recombinant human IL-1β (50 U/ml), recombinant human IFN-γ (1000 U/ml, Genzyme, Cambridge, USA) and the iNOS blocker N G -methyl-L-arginine (L-MA, 1.0 mmol/l; Sigma, Bornem, Belgium), alone or in combinations.…”

CD40 expression on human pancreatic duct cells: role in nuclear factor-kappa B activation and production of pro-inflammatory cytokines

“…Duct cells were also shown to produce nitric oxide [4] and to express MHC class II [3], leading to proposals that they might contribute to beta cell damage in Type 1 diabetes mellitus [8]. In the transplantation setting, islet suspensions prepared from human pancreases contain a high proportion of non-endocrine duct cells [9,10], implying that these cells could affect graft acceptance. Defining the path-ways of duct cell activation is therefore relevant to understanding both the pathogenesis of Type 1 diabetes and islet transplantation.…”