Autologous Pancreatic Islet Transplantation in Human Bone Marrow

The liver is the current site of choice for pancreatic islet transplantation, even though it is far from being ideal. We recently have shown in mice that the bone marrow (BM) may be a valid alternative to the liver, and here we report a pilot study to test feasibility and safety of BM as a site for islet transplantation in humans. Four patients who developed diabetes after total pancreatectomy were candidates for the autologous transplantation of pancreatic islet. Because the patients had contraindications for intraportal infusion, islets were infused in the BM. In all recipients, islets engrafted successfully as shown by measurable posttransplantation C-peptide levels and histopathological evidence of insulin-producing cells or molecular markers of endocrine tissue in BM biopsy samples analyzed during follow-up. Thus far, we have recorded no adverse events related to the infusion procedure or the presence of islets in the BM. Islet function was sustained for the maximum follow-up of 944 days. The encouraging results of this pilot study provide new perspectives in identifying alternative sites for islet infusion in patients with type 1 diabetes. Moreover, this is the first unequivocal example of successful engraftment of endocrine tissue in the BM in humans.Islet transplantation represents an important therapeutic option for adults with unstable type 1 diabetes (T1D) who, despite their best efforts, have wide and unpredictable fluctuations of glucose levels or who are no longer able to sense hypoglycemia with an increased risk of acute and chronic complications of diabetes and a significant worsening of quality of life (1). The liver is the current site of choice for pancreatic islet transplantation, even though it is far from being ideal because of immunologic (2–4), anatomic (5), and metabolic (6–8) factors leading to significant early graft loss. Along with preexisting and transplant-induced autospecific and allospecific immune responses (9), a nonspecific response, predominantly mediated by innate inflammatory processes related to mechanics and site, plays a major role in the loss of islets and islet function after transplantation in the liver (4,10–13). As reported by many studies, an estimated 60–80% of the transplanted islet mass is lost within hours or days after intrahepatic islet infusion (12,14,15), mainly because of immediate blood-mediated inflammatory reaction (16), thrombosis (11,17), and hepatic tissue ischemia (18,19) with release of liver enzymes (20,21). Furthermore, from a clinical point of view, the process of islet infusion in the liver is associated with an increase of portal pressure proportional to the islet mass (22), thus limiting the total islet mass to be transplanted (23). Recognizing these problems has increased the interest in the search for alternative sites for islet transplantation to avoid liver-specific problems (24). Despite the success of experimental islet transplantation in mouse models using different sites, the results of only a few of those studies were applied in large animal models and none was applied in human models.Bone marrow (BM) may be an alternative site for pancreatic islet transplantation because it offers a protected and extravascular, although well-vascularized, microenvironment (25). Because of BM broad distribution and easy access, islet infusion in the BM may overcome technical limitations and reduce complications of islet infusion in the liver through the portal vein (24). In a recent preclinical study, we tested whether syngeneic pancreatic islets could engraft in the BM of diabetic mice by comparing survival, function, and morphology of syngeneic islets infused in the BM or in the liver (26). Islets engrafted efficiently in the BM of diabetic mice and for >1 year posttransplantation, the glucose metabolism of those animals was similar to that of nondiabetic mice. Furthermore, mice with islets infused in the BM were more likely to reach euglycemia than mice with islets infused in liver. Islets in the BM showed a compact morphology with a preserved ratio between α-cells and β-cells, with only marginal effects on bone structure. Moreover, the presence of islets in the BM did not affect hematopoietic activity, even when this function was strongly upregulated in response to virus-induced BM aplasia. Based on these results, we were granted approval to use this approach in humans, and we performed a pilot study in which patients with diabetes and hepatic contraindications for liver islet autotransplantation (IAT) received a single intra-BM islet infusion in the iliac crest.     

Has Time Come for New Goals in Human Islet Transplantation?

The enthusiasm regarding clinical islet transplantation has been dampened by the long-term results. Concerns about the associated risks of life-long immunosuppression and the striking imbalance between potential recipients and available donor pancreata warrant changes in some of the current goals.
Islet transplantation will never be a cure of type 1 diabetes in the majority of patients with no secondary complications, but is a valid option for a limited number of patients with brittle diabetes waiting for an organ or after organ transplantation. Furthermore, insulin independence should not be the main goal of islet transplantation, but avoidance of severe hypoglycemia and good glycemic control, which can be achieved with a relatively small functional beta-cell mass. Therefore, initially one islet infusion is sufficient. Retransplantation at a later time point remains an option, if glucose control deteriorates.
Efforts to improve islet transplantation should no longer focus on islet isolation and immunosuppression, but rather on the low posttransplant survival rate of islets caused by activation of the coagulation pathway and the limited oxygen delivery to the islets. Transplantation of smaller islets be it naturally small or size tailored reaggregated islets has the potential to facilitate these processes.     

Rescue Purification Maximizes the Use of Human Islet Preparations for Transplantation

The relative inefficiency of the islet purification process may hamper obtaining enough islets for transplantation even with adequate pre-purification counts. In this study, we determined the effect of an additional purification step on total islet yields and pancreas utilization at our center. Twenty-five pancreata were processed using the automated method followed by continuous gradient purification (CGP), and the less pure islet fractions were subjected to additional rescue gradient purification (RGP). CGP and RGP islets were combined and transplanted into patients with type 1 diabetes. CGP and RGP islets showed no significant differences in cell viability, insulin secretion in vitro and function when transplanted into chemically diabetic mice. Mean RGP contribution to the final preparation was 27.9 +/- 19.9%. In 12 of 25 preparations, CGP yielded <5000 IEQ/kg of recipient body weight, and inclusion of RGP islets to the final preparation allowed to obtain the minimal islet number required for transplantation. Transplanted islets resulted in sustained C-peptide production, HbA1(C) normalization and insulin-independence or reduced insulin requirements. Taken together, our data suggest that RGP islets are comparable in terms of viability and potency to CGP islets. RGP may be of assistance in maximizing the number of islet preparations successfully used in transplant protocols.     

Gastrointestinal Transplantation: An Update

Gastrointestinal transplantation is a life-saving option for patients who have chronic intestinal failure and cannot tolerate total parenteral nutrition (TPN). Early referral is important because of the scarcity of donors and the increased risk of complications in debilitated recipients. One-year patient survival rates range from 50% to 70%. Despite the use of intense immune suppression, most patients experience at least 1 episode of graft rejection. More than 80% of the survivors are able to stop TPN and resume an unrestricted oral diet. Patients with functioning grafts have a good quality of life. (Liver Transpl 2000;6:515-519.)     

Islet Transplantation and Glucose Regulation

Transplantation of isolated islets of Langerhans for treatment of diabetes has been developed through experimental research in several species and is now being applied to humans with some success albeit limited. A significant problem for human islet allotransplantation or autotransplantation (following pancreatectomy) is the relatively poor yield of islets available for transplantation. The metabolic function of islet transplant recipients that have achieved insulin independence reflects the relatively small mass of insulin-secreting tissue implanted and the fact that only the intraportal site of transplantation appears to allow sufficient graft function to achieve insulin independence. The long-term function of such grafts has been poor, with most grafts showing deterioration in function within 5 years. Studies of islet transplantation in other species showed a similar result, although other sites for islet graft implantation, such as the spleen or kidney capsule, may be associated with a better outcome. These studies, however, also suffer from problems of relatively limited islet mass. Only in the rodent model where isogeneic strains are available is it possible to transplant sufficient numbers of islets to obtain an equivalent functional islet mass similar to that found in the normal pancreas; and in this case near-normal glucose metabolism is obtained and is maintained for the life-span of the animal.     

Antiangiogenic and Immunomodulatory Effects of Rapamycin on Islet Endothelium: Relevance for Islet Transplantation

Donor intra-islet endothelial cells contribute to neovascularization after transplantation. Several factors may interfere with this process and ultimately influence islet engraftment. Rapamycin, a central immunosuppressant in islet transplantation, is an mTOR inhibitor that has been shown to inhibit cancer angiogenesis. The aim of this study was to evaluate the effects of rapamycin on islet endothelium. Rapamycin inhibited the outgrowth of endothelial cells from freshly purified human islets and the formation of capillary-like structures in vitro and in vivo after subcutaneous injection within Matrigel plugs into SCID mice. Rapamycin decreased migration, proliferation and angiogenic properties of human and mouse islet-derived endothelial cell lines with appearance of apoptosis. The expression of angiogenesis-related factors VEGF, alphaVbeta3 integrin and thrombospondin-1 on islet endothelium was altered in the presence of rapamycin. On the other hand, rapamycin decreased the surface expression of molecules involved in immune processes such as ICAM-1 and CD40 and reduced the adhesion of T cells to islet endothelium. Our results suggest that rapamycin exerts dual effects on islet endothelium inducing a simultaneous inhibition of angiogenesis and a down-regulation of receptors involved in lymphocyte adhesion and activation.     

The Clinical Impact of Islet Transplantation

Islet cell transplantation has recently emerged as one of the most promising therapeutic approaches to improving glycometabolic control in diabetic patients and, in many cases, achieving insulin independence. Unfortunately, many persistent flaws still prevent islet transplantation from becoming the gold standard treatment for type 1 diabetic patients. We review the state of the art of islet transplantation, outcomes, immunosuppression and—most important—the impact on patients' survival and long‐term diabetic complications and eventual alternative options. Finally, we review the many problems in the field and the challenges to islet survival after transplantation. The rate of insulin independence 1 year after islet cell transplantation has significantly improved in recent years (60% at 1 year posttransplantation compared with 15% previously). Recent data indicate that restoration of insulin secretion after islet cell transplantation is associated with an improvement in quality of life, with a reduction in hypoglycemic episodes and potentially with a reduction in long‐term diabetic complications. Once clinical islet transplantation has been successfully established, this treatment could even be offered to diabetic patients long before the onset of diabetic complications.     

Islet Transplantation in Genetically Determined Diabetes

Hyperglycemia induced in animals by beta cell toxins or by pancreatectomy can be reversed by pancreatic islet transplantation. Abnormal carbohydrate metabolism in juvenile onset human diabetics has also been corrected, albeit temporarily because of graft rejection, by pancreatic transplantation. It does not necessarily follow that naturally occurring diabetes in animals or adult onset diabetes in man would respond to similar treatment. Islet transplantation was studied in mice with chemically induced or genetically determined diabetes. Streptozotocin-induced diabetic mice were permanently cured by syngeneic islets and, when immunosuppressed, were rendered normoglycemic for six weeks after receiving xenogeneic rat islets. In contrast, histocompatible islets from normoglycemic coisogenic donors were ineffective in hyperglycemic db/db recipients as were xenogeneic rat islets in immunosuppressed db/db hosts. However, when islets were isolated from db/db donors and transplated to genetically normal coisogenic mice, which had been rendered hyperglycemic with streptozotocin, they became normoglycemic. Apparently the metabolic defect in the db/db mice, which is similar in some ways to human maturity onset diabetes, does not reside in their islets as these cells can function normally if transplanted to genetically nondiabetic hosts. In two other types of genetic diabetes (ob/ob and NZO) islet transplantation was more effective. Pancreatic transplantation is unlikely to be the proper treatment for all types of diabetes even if technical and immunological problems are overcome.     

胰岛细胞移植的现状与进展

胰岛细胞移植是目前糖尿病治疗中的一个研究热点,也是有望从根本上治疗糖尿病的一种方法。自2000年加拿大Edmonton报到了对7例1型糖尿病患者成功施行胰岛细胞移植术以来,国内外掀起了胰岛细胞移植的研究热潮,在动物实验和临床研究方面取得了一定的突破。本文就胰岛细胞移植的现状与进展进行简要的综述。     

Quality of Life After Islet Transplantation

This study analyzed quality of life in patients with type 1 diabetes that received islet transplantation. Twenty-three subjects were followed over 3 years. In addition to an interview, patients self-completed two standardized psychometric questionnaires, HSQ 2.0 and DQOL, before and after transplant, and scores were compared. Analysis was also adjusted for potential "confounders" such as graft dysfunction, insulin therapy and adverse events. DQOL: the Impact score significantly improved at all time points of the follow-up; satisfaction and worry scales also significantly improved at selected time points. Longitudinal analysis demonstrated that reintroduction of insulin had a negative effect on all three scales, but significant improvement in Impact scale persisted even after adjusting for this factor. HSQ 2.0: only the Health Perception scale preliminarily showed significant improvement at most time points. Longitudinal analysis showed loss of significance when insulin therapy was considered. Other scores were improved only at selected time points or not affected. Bodily pain scale showed deterioration at selected times. Interview: glucose control stability, not insulin independence, was reported as the main beneficial factor influencing QOL. In conclusion, islet transplantation has a positive influence on patients' QOL, despite chronic immunosuppression side effects. Re-introduction of insulin modifies QOL outcomes.     

胰岛移植的分子影像示踪与监测

目的总结目前胰岛移植分子影像示踪与监测的不同方法。方法对当前国内、外有关胰岛移植分子影像示踪与监测的研究报道进行综述和分析。结果 MRI具有高敏感性,高空间分辨率,可应用于临床,可在MR实时引导下进行药物注射,无放射性损伤等优点,但缺点是无法区分肝细胞以及未存活的移植胰岛细胞,在肝铁离子过量的患者中无法进行监测。核素分子显像的优点是特异性高,只有存活的细胞产生信号,可应用于临床,但具有离子辐射,有可能会篡改基因,无解剖成像,空间分辨率低,放射性示踪半衰期较短。光学成像特异性高,只有存活的细胞产生信号,获取途径多样,无放射性损伤,但有可能会篡改基因,空间分辨率低,尚无法应用于临床。结论磁共振、核素影像、光学成像以及基于微囊胰岛的多模态影像,都可以对胰岛移植物进行分子影像的示踪与监测,基于微囊胰岛的多模态影像可能会是未来最佳的方法。     

Islet Transplantation: Lessons Learned Since the Edmonton Breakthrough

This work sought to summarize the main issues of the last decade in the field of clinical islet transplantation. Ten years ago in Edmonton, a new protocol initiated for islet transplantation brought a breakthrough to the field. The earlier, rather poor results were in a sharp contrast to the first published results of 100% insulin freedom at 1 year. However, later it became clear that the promising initial results decline with time; at around 5 years, only about 10% of the patients maintain freedom from external insulin. Despite that fact, a milestone was set and intensive research started worldwide. New hopes were raised for patients. Modifications of the original protocol have been implemented to improve clinical results; however, islet transplantation remains an experimental procedure to date.     

Positron Emission Tomography in Clinical Islet Transplantation

The fate of islets in clinical transplantation is unclear. To elude on this positron emission tomography combined with computed tomography (PET/CT) was performed for 60 min during islet transplantation in five patients receiving six transplants. A fraction of the islets (23%) were labeled with 18F‐fluorodeoxyglucose ([¹⁸F]FDG) and carefully mixed with unlabeled islets just prior to intraportal transplantation. The peak radioactivity concentration in the liver was found at 19 min after start of islet infusion and corresponded to only 75% of what was expected, indicating that islets are lost during the transplantation procedure. No accumulation of radioactivity was found in the lungs. A nonphysiological peak of C‐peptide was found in plasma during and immediately after transplantation in all subjects. Distribution in the liver was heterogeneous with wide variations in location and concentration. Islets found in areas with concentrations of >400 IEQ/cc liver tissue varied between 1% and 32% of the graft in different subjects. No side effects attributed to the PET/CT procedure were found. Clinical outcome in all patients was comparable to that previously observed indicating that the [¹⁸F]FDG labeling procedure did not harm the islets. The technique has potential to be used to assess approaches to enhance islet survival and engraftment in clinical transplantation.