[Isolation and cryopreservation of human islets of Langerhans]. / Isolement et cryopréservation d'îlots de Langerhans humains.

Islet transplantation represents an alternative to whole pancreas transplantation for the treatment of patients suffering from diabetes type I. The transplantation of a sufficient number of islets is an essential condition for successful allograft. Islet cryopreservation allows the storage of islet preparations for subsequent pooling, at the time of transplantation, of cryopreserved islets with a fresh preparation in order to increase the mass of transplanted pancreatic endocrine tissue. From May 1994 to April 1995, islets were isolated from 22 human pancreases using a modified automated method, and 19 preparations were cryopreserved. The function of cryopreserved islets was tested in vitro (static incubation and perifusion). The results of static incubation experiments confirmed that the insulin secretion of cryopreserved human islets in response to glucose stimulation was comparable to the response of islets that have not been frozen. In static incubation experiments, the mean (+/- SEM) insulin secretion of islets, prior to cryopreservation, was 239.3 (+/- 58.9) and 479.5 (+/- 59.5) pg/islet/15 min at 2.8 mM glucose and 16.7 mM glucose respectively. The mean (+/- SEM) insulin secretion of cryopreserved islets was 274 (+/- 103.2) and 468.5 (+/- 191.9) pg/islet/15 min at 2.8 mM and 16.7 mM glucose respectively. The perifusion experiments also demonstrated a significant increase of insulin secretion from cryopreserved islets perifused with a stimulating glucose concentration. Our experiments allow us to envisage the use of cryopreserved islet preparations for allotransplantation in diabetic patients.

[Islets of Langerhans allografts in the pig: initial results]. / Allogreffes d'îlots de Langerhans chez le porc: premiers résultats.

Islet of Langerhans transplantation represents a promising treatment of diabetes. Use of porcine islets for xenotransplantation could offer a solution to shortage of organ donors. However, isolation of pig islets remains a real challenge because of their marked fragility. Using a modified automated method for islet isolation, we performed 10 intraportal islet allografts in pigs. Surgically pancreatectomized pigs were transplanted with purified islet preparations pooled from pancreases of 3 donors (slaughter-house pigs, age 5-8 months). Six recipients were not immunosuppressed and four received an immunosuppressive treatment of Cyclosporine and Azathioprine. In the first group (non-immunosuppressed recipients), insulin secretion was observed for a mean of 4.8 days after transplantation. In the second group (with immunosuppressive treatment) the recipients sustained an insulin secretion for 6-9 days. However, in both groups liver biopsies showed signs of acute rejection and destruction of the transplanted islets. Pig islet allotransplantation, using as recipients surgically pancreatectomized pigs, can be considered as a suitable model to assess the functional results of mass islet isolation. We were able to reverse diabetes transitory in a large animal, by transplantation of purified pig islet preparations. However, even when an immunosuppressive treatment was administered to the recipients, rejection seemed to represent an important factor in the functional outcome of the islet grafts.

[Mechanism of rejection of islet of Langerhans xenografts]. / Les mécanismes de rejet dans les xénogreffes d'îlots de Langerhans.

For immediately vascularised xenografts, the presence of preformed antibodies associated with the activation of the complement induces a hyperacute humoral rejection in discordant combinations. For neovascularised xenografts, such as the islets of Langerhans, the absence of an initial vascularisation prevents the acute humoral rejection and there is some evidence to suggest that the rejection is cell mediated. To analyse the influence of species response to cellular transplantation, we compared the functional outcome and the immunohistochemical analysis of concordant and discordant xenotransplated islets with allotransplanted islets of Langerhans. In the concordant xenotransplantation experiment, rat (Sprague Dawley) purified islets (N = 600) were transplanted under the kidney capsule of streptozotocin diabetic mice. In the discordant experiment, purified human islets (N = 1000), in the allotransplantation model Balb mouse islets (N = 600) were transplanted under the kidney capsule of C57BL/6 mice. Daily blood glucose levels were monitored until rejection occurred (blood glucose > 11 mMol/L) and kidney biopsies were taken every second day for immunohistological examination. In allotransplanted mice, rejection occurred after a mean of 15 +/- 4 days, in concordant xenotransplanted mice after a mean of 10 +/- 3 days and in the discordant xenotransplanted mice after a mean of 8 +/- 1 days. The immunohistological analysis showed in both xenotransplantation models a deposit of IgM and C3 surrounding the grafted cells, starting at day 1 and persisting until rejection, associated with the presence of Mac-1 positive cells. The intensity of the humoral response was moderate to important in the concordant group, but very strong and unchanged until rejection in the disconcordant group. An invasion with CD4+ and CD8+ cells started between day 4 and 6 in both groups, progressively increasing until rejection. In the allotransplantation model, no humoral response was detectable and the cellular response started between day 10 and 12 after transplantation. We conclude, that a cellular response is involved in the rejection process of both concordant and discordant xenotransplanted islets of Langerhans. Compared to an allotransplantation model, the humoral response does not destroy the function of transplanted islets, but could trigger or accelerate the cell-mediated response.