Anna Casu

Long‐Term Controlled Normoglycemia in Diabetic Non‐Human Primates After Transplantation with hCD46 Transgenic Porcine Islets

Xenotransplantation of porcine islets into diabetic non‐human primates is characterized by (i) an initial massive graft loss possibly due to the instant blood‐mediated inflammatory reaction and (ii) the requirement of intensive, clinically unfriendly immunosuppressive therapy. We investigated whether the transgenic expression of a human complement‐regulatory protein (hCD46) on porcine islets would improve the outcome of islet xenotransplantation in streptozotocin‐induced diabetic Cynomolgus monkeys. Immunosuppression consisted of thymoglobulin, anti‐CD154 mAb for costimulation blockade, and mycophenolate mofetil. Following the transplantation of islets from wild‐type pigs (n = 2) or from 1,3‐galactosyltransferase gene‐knockout pigs (n = 2), islets survived for a maximum of only 46 days, as evidenced by return to hyperglycemia and the need for exogenous insulin therapy. The transplantation of islets from hCD46 pigs resulted in graft survival and insulin‐independent normoglycemia in four of five monkeys for the 3 months follow‐up of the experiment. One normalized recipient, selected at random, was followed for >12 months. Inhibition of complement activation by the expression of hCD46 on the pig islets did not substantially reduce the initial loss of islet mass, rather was effective in limiting antibody‐mediated rejection. This resulted in a reduced need for immunosuppression to preserve a sufficient islet mass to maintain normoglycemia long‐term.

Rapid loss of intraportally transplanted islets: an overview of pathophysiology and preventive strategies

Abstract:  Islets isolated from multiple pancreas donors are often necessary to achieve euglycemia in type 1 diabetic patients treated by islet allotransplantation. This increases the burden on the limited pool of donor organs. After infusion into the portal vein, a substantial percentage of islets are lost in the immediate post‐transplant period through an inflammatory response termed the instant blood‐mediated inflammatory reaction (IBMIR). IBMIR is equally, if not more of a problem after islet xenotransplantation, e.g., using pig islets in non‐human primates. Coagulation, platelet aggregation, complement activation, and neutrophil and monocyte infiltration play roles in this reaction. IBMIR is potentially triggered by islet surface molecules, such as tissue factor and collagen residues that are normally not in direct contact with the blood. Also, stress during the islet isolation process results in the expression and production of several inflammatory mediators by the islets themselves. The potential mechanisms involved in this rapid graft loss and treatment options to reduce this loss are reviewed. Preventive strategies for IBMIR can include systemic treatment of the recipient, pre‐conditioning of the isolated islets, or, in the case of xenotransplantation, genetic modification of the organ‐source pig. Pre‐conditioning of islets in culture by exposure to anti‐inflammatory agents or by genetic modification harbors fewer risks of systemic complications in the recipient. The future of clinical islet transplantation will, at least in part, depend on the success of efforts made to reduce rapid graft loss, and thus allow islet transplantation to become a more efficient therapy by the use of single donors.

Metabolic aspects of pig-to-monkey (Macaca fascicularis) islet transplantation : implications for translation into clinical practice

Aims/hypothesisAttempts to use an alternative source of islets to restore glucose homeostasis in diabetic patients require preclinical islet xenotransplantation models to be tested. These models raise questions about metabolic compatibility between species and the most appropriate metabolic parameters to be used to monitor graft function. The present study investigated and compared relevant gluco-metabolic parameters in pigs, monkeys and the pig-to-monkey islet transplantation model to gain insight into the potential clinical outcome of pig-to-human islet transplantation.MethodsBasal and IVGTT-stimulated blood glucose, C-peptide, insulin and glucagon levels were assessed in non-diabetic pigs and monkeys. The same parameters were used to evaluate the performance of porcine islet xenografts in diabetic monkeys.ResultsNon-diabetic cynomolgus monkeys showed lower levels of fasting and stimulated blood glucose but higher levels of C-peptide and insulin than non-diabetic pigs. The reported levels in humans lie between those of monkeys and pigs, and differences in metabolic parameters between pigs and humans appear to be smaller than those between pigs and cynomolgus monkeys. The transplantation data indicated that the degree of graft function (evaluated by the measurement of C-peptide levels) necessary to normalise blood glucose in the recipient was determined by the recipient levels rather than by the donor levels.Conclusions/interpretationThe differences between donor and recipient species may affect the transplantation outcome and need to be considered when assessing graft function in xenotransplantation models. Given the differences between monkeys and humans as potential recipients of pig islets, it should be easier to reach glucose homeostasis in pig-to-human than in pig-to-non-human primate islet xenotransplantation.

Clinical islet xenotransplantation: how close are we?

Type 1 diabetes (T1D) is a major health problem throughout the world. In the U.S., it is estimated that about 1.5 million people suffer from T1D. Even when well controlled—by frequent monitoring of blood glucose and administration of insulin, the long-term complications of the disease are significant and include cardiovascular disease, nephropathy, retinopathy, and neuropathy (1). Here we review recent progress in preclinical models of pig islet xenotransplantation and discuss the remaining challenges that need to be addressed before the application of this form of therapy can be established in patients with T1D. During the past decade, islet allotransplantation alone (without previous kidney transplantation) using deceased human donor pancreata has been indicated mainly in patients who have had T1D for >5 years with life-threatening hypoglycemic episodes and wide fluctuations in blood glucose levels. Although the initial long-term results were rather disappointing (2), the results of islet allotransplantation have improved significantly in recent years, with 5-year insulin-independent normoglycemia achieved in >50% of patients at experienced centers (3). There is increasing evidence that successful islet allotransplantation greatly reduces the incidence of hypoglycemic episodes (2) and reduces or slows the incidence of late complications of T1D (4). This may extend the indications for islet transplantation to patients with progressive complications. For example, islet transplantation in a patient with preterminal renal failure may prevent disease progression, possibly avoiding the need for hemodialysis and kidney transplantation, provided that nonnephrotoxic immunosuppressive drug therapy is administered. Currently, in the U.S., the median waiting time for a kidney allograft from a deceased human donor is >4 years (5). However, islets from two deceased human donor pancreata are frequently required to achieve normoglycemia in a diabetic patient. Because of the limited number of suitable deceased donor pancreata, the overall number of …

Chapter 4: Pre-clinical efficacy and complication data required to justify a clinical trial

Abstract:  Pre‐clinical efficacy and complication data required to justify a porcine islet transplantation clinical trial have been considered. To be relevant to clinical islet transplantation, pre‐clinical data should be obtained in a pig‐to‐diabetic non‐human primate (NHP) model. In view of the difficulties of maintaining immunosuppressed NHPs for long periods of time, flexibility must be allowed with regard to the following recommendations. A pre‐clinical trial would be deemed a success if five of eight consecutive experiments in NHPs with proven diabetes meet the following guidelines.

Insulin secretion and glucose metabolism in alpha 1,3‐galactosyltransferase knock‐out pigs compared to wild‐type pigs

Casu A, Echeverri GJ, Bottino R, van der Windt DJ, He J, Ekser B, Ball S, Ayares D, Cooper DKC. Insulin secretion and glucose metabolism in alpha 1,3‐galactosyltransferase knock‐out pigs compared to wild‐type pigs. Xenotransplantation 2010; 17: 131–139.

Seasonality of birth in children (0-14 years) and young adults (0-29 years) with type 1 diabetes mellitus in Sardinia differs from that in the general population

A cohort of 1,118 children (0-14 years) and 810 adolescents and young adults (15-29 years) with type 1 diabetes mellitus (DM) diagnosed in Sardinia between 1989 and 1998 were analyzed for seasonality of month of birth, and compared to the pattern registered in 314,084 live births. Patients with DM of both age groups had a statistically significant different seasonality pattern from the general population, revealing an increased birth rate during the summer months, a mirror image of the seasonality of onset of disease.

Recovery of Endogenous β-Cell Function in Nonhuman Primates After Chemical Diabetes Induction and Islet Transplantation

OBJECTIVE—To describe the ability of nonhuman primate endocrine pancreata to reestablish endogenous insulin production after chemical β-cell destruction. RESEARCH DESIGN AND METHODS—Eleven monkeys (Macaca fascicularis) were rendered diabetic with streptozotocin. Eight diabetic monkeys received intraportal porcine islet transplantation. RESULTS—Two monkeys transplanted after 75 days of type 1 diabetes showed recovery of endogenous C-peptide production a few weeks after transplantation, concomitant with graft failure. Histological analysis of the pancreas of these monkeys showed insulin-positive cells, single or in small aggregates, scattered in the pancreas and adjacent to ducts. Interestingly, numerous CK19+ cells costained with proinsulin and PDX-1 antibodies. Furthermore, the peculiar double phenotype glucagon-positive/GLUT2+ was observed. In these monkeys as well as in all others, the original islets showed no insulin staining. CONCLUSIONS—Our data provide evidence that, in nonhuman primates, the pancreas can reestablish endogenous insulin production after chemical β-cell destruction. This seems to be a nongeneralizable event with only 2 out of 11 monkeys recovering β-cell function. In these two monkeys, younger age and islet graft behavior might have played a role in triggering endogenous β-cell recovery.

Bayesian approach to study the temporal trend and the geographical variation in the risk of type 1 diabetes. The Sardinian Conscript Type 1 Diabetes Registry

Background:  A previous analysis of the Sardinian Conscript Type 1 Diabetes Registry indicated an abrupt increase in the risk for type 1 diabetes (T1D) among Sardinian male cohorts starting from the 1946 one.

Acute gastric dilatation after porcine islet transplantation in a cynomolgus monkey – case history and review of the literature

Abstract:  We report a streptozotocin‐induced diabetic cynomolgus monkey that developed acute gastric dilatation within the first week after intraportal porcine islet transplantation. The monkey presented with increasing anorexia and somnolence, associated with persistent metabolic acidosis. On physical examination, a diffuse mass was palpable in the abdomen. Because of its deteriorating clinical condition, the monkey was euthanized. Necropsy revealed acute gastric dilatation. Potential etiological factors, possible methods of prevention, and guidelines for management are reviewed, with special emphasis on the incidence and management of acute gastric dilatation in non‐human primates.