Establishment of a prolonged pancreas preservation model for islet isolation research in mice

Establishing a prolonged pancreas preservation model in a small animal is important for islet isolation research. Use of a rat pancreas model has been reported, but no published reports have used a mouse pancreas for prolonged cold preservation prior to islet isolation. For the model, a mouse is preferred over a rat because of its small size, well-known immune characterization and variety of gene-modulated models. In the present study, we established a prolonged pancreas preservation model in a mouse for islet isolation research. The collagenase solution was injected successfully after 24 and 48 h cold preservations in University of Wisconsin solution, and islets could be isolated from both groups of preserved pancreata. The islet yields from the control, 24 h preserved, and 48 h preserved pancreata were 183.9 ± 13.9, 128.5 ± 15.5, and 24.6 ± 12.9 per pancreas, respectively. The propidium iodide–positive area assay was significantly increased in both preserved groups, and insulin secretion levels in response to 20.0 mM glucose and stimulation indices were significantly decreased in the 48 h preserved group. Inflammation-related genes mRNA levels were significantly upregulated in the 24 h preserved group, as previously shown in the human model. Thus, this model might be useful for prehuman islet isolation screening research, reserving research using human pancreata for the most promising approaches.     

Establishment of a prolonged pancreas preservation model for islet isolation research in mice

Establishing a prolonged pancreas preservation model in a small animal is important for islet isolation research. Use of a rat pancreas model has been reported, but no published reports have used a mouse pancreas for prolonged cold preservation prior to islet isolation. For the model, a mouse is preferred over a rat because of its small size, well-known immune characterization, and variety of gene-modulated models. In the present study, we established a prolonged pancreas preservation model in a mouse for islet isolation research. The collagenase solution was injected successfully after 24 and 48 h cold preservations in University of Wisconsin solution, and islets could be isolated from both groups of preserved pancreata. The islet yields from the control, 24 h preserved, and 48 h preserved pancreata were 183.9 ± 13.9, 128.5 ± 15.5, and 24.6 ± 12.9 per pancreas, respectively. The propidium iodide-positive area assay was significantly increased in both preserved groups, and insulin secretion levels in response to 20.0 mM glucose and stimulation indices were significantly decreased in the 48 h preserved group. Inflammation-related genes mRNA levels were significantly upregulated in the 24 h preserved group, as previously shown in the human model. Thus, this model might be useful for prehuman islet isolation screening research, reserving research using human pancreata for the most promising approaches.     

Clinical islet isolation outcomes with a highly purified neutral protease for pancreas dissociation

Background: Pancreas dissociation is a critical initial component of the islet isolation procedure and introduces high variability based on factors including the enzyme type, specificity and potency. Product refinement and alterations to the application strategies have improved isolation outcomes over time; however, islet utilization from donor organs remains low. In this study we evaluate a low endotoxin-high activity grade neutral protease in clinical islet isolation.

Materials and Methods: The use of a non-collagenolytic enzyme, either thermolysin or high active neutral protease, was randomized in clinical islet isolations to evaluate efficacy. Additionally a retrospective comparison to neutral protease NB was conducted.

Results:The thermolysin group had lower trapped islet population and increased purity and post-culture islet mass in comparison to high active grade neutral protease. Comparison of neutral protease NB GMP grade to high active neutral protease displayed no measurable difference in islet mass or viability and transplantation outcomes at 1 mo post-transplant were favorable for both groups.

Conclusions: High activity neutral protease can generate clinical grade islets and may prove beneficial to islet function and viability based on a reduced endotoxin load but dosing of neutral protease requires ongoing optimization.     

An analysis of the role of collagenase and protease in the enzymatic dissociation of the rat pancreas for islet isolation

Summary Crude Clostridium histolyticum collagenase is widely used for the enzymatic degradation of pancreatic extracellular matrix in order to isolate the islets of Langerhans. The variable enzymatic composition of crude collagenases is a critical issue which contributes to the poor reproducibility of islet isolation procedures. In this study, the separate contributions of collagenase and protease to the islet isolation process were analysed by testing various combinations of purified collagenase and purified protease in rat pancreas dissociations under conditions which eliminated all other proteolytic activity. Under these conditions, complete tissue dissociation by purified collagenase required 99±10 min, whereas increasing amounts of protease progressively reduced this time to a minimum of 36±1 min. Histochemical analysis of the dissociation process showed that protease enhanced the degradation of all four major components of the extracellular matrix: collagen was degraded more completely, while proteoglycans, glycoproteins and elastin were degraded at a higher rate. Pancreas dissociation under the present, strictly controlled conditions resulted in a high yield of viable islets: 4.2–5.0 l islet tissue volume (3,300–3,800 islets) were isolated per g pancreas in the presence of a high or low protease concentration, respectively. Prolonged dissociation in the presence of protease resulted in a dramatic decrease in islet yield which correlated with the observation that the enzyme accelerated islet disintegration. It is concluded that the collagenase-induced dissociation of the extracellular matrix is facilitated by protease. Our study shows that high yields of viable islets can be obtained under controlled enzymatic conditions, provided that the exposure of islets to protease is limited.     

Donor insulin use predicts beta-cell function after islet transplantation

Insulin is routinely used to manage hyperglycaemia in organ donors and during the peri-transplant period in islet transplant recipients. However, it is unknown whether donor insulin use (DIU) predicts beta-cell dysfunction after islet transplantation. We reviewed data from the UK Transplant Registry and the UK Islet Transplant Consortium; all first-time transplants during 2008-2016 were included. Linear regression models determined associations between DIU, median and coefficient of variation (CV) peri-transplant glucose levels and 3-month islet graft function. In 91 islet cell transplant recipients, DIU was associated with lower islet function assessed by BETA-2 scores (β [SE] -3.5 [1.5], P = .02), higher 3-month post-transplant HbA1c levels (5.4 [2.6] mmol/mol, P = .04) and lower fasting C-peptide levels (−107.9 [46.1] pmol/l, P = .02). Glucose at 10 512 time points was recorded during the first 5 days peri-transplant: the median (IQR) daily glucose level was 7.9 (7.0-8.9) mmol/L and glucose CV was 28% (21%-35%). Neither median glucose levels nor glucose CV predicted outcomes post-transplantation. Data on DIU predicts beta-cell dysfunction 3 months after islet transplantation and could help improve donor selection and transplant outcomes.     

An effective purification method using large bottles for human pancreatic islet isolation

The purification process is one of the most difficult procedures in pancreatic islet isolation. It was demonstrated that the standard purification method using a COBE 2991 cell processor with Ficoll density gradient solution harmed islets mechanically by high shear force. We reported that purification using large bottles with a lower viscosity gradient solution could improve the efficacy of porcine islet purification. In this study, we examined whether the new bottle purification method could improve the purification of human islets. Nine human pancreata from brain-dead donors were used. After pancreas digestion, the digested tissue was divided into three groups. Each group was purified by continuous density gradient using ET-Kyoto and iodixanol gradient solution with either the standard COBE method (COBE group) or the top loading (top group) or bottom loading (bottom group) bottle purification methods. Islet yield, purity, recovery rate after purification, and in vitro and in vivo viability were compared. Islet yield per pancreas weight (IE/g) and the recovery rate in the top group were significantly higher than in the COBE and bottom groups. Furthermore, the average size of purified islets in the top group was significantly larger than in the COBE group, which indicated that the bottle method could reduce the shear force to the islets. In vivo viability was also significantly higher in the top group compared with the COBE group. In conclusion, the top-loading bottle method could improve the quality and quantity of human islets after purification.     

An effective purification method using large bottles for human pancreatic islet isolation

The purification process is one of the most difficult procedures in pancreatic islet isolation. It was demonstrated that the standard purification method using a COBE 2991 cell processor with Ficoll density gradient solution harmed islets mechanically by high shear force. We reported that purification using large bottles with a lower viscosity gradient solution could improve the efficacy of porcine islet purification. In this study, we examined whether the new bottle purification method could improve the purification of human islets. Nine human pancreata from brain-dead donors were used. After pancreas digestion, the digested tissue was divided into three groups. Each group was purified by continuous density gradient using ET-Kyoto and iodixanol gradient solution with either the standard COBE method (COBE group) or the top loading (top group) or bottom loading (bottom group) bottle purification methods. Islet yield, purity, recovery rate after purification, and in vitro and in vivo viability were compared. Islet yield per pancreas weight (IE/g) and the recovery rate in the top group were significantly higher than in the COBE and bottom groups. Furthermore, the average size of purified islets in the top group was significantly larger than in the COBE group, which indicated that the bottle method could reduce the shear force to the islets. In vivo viability was also significantly higher in the top group compared with the COBE group. In conclusion, the top-loading bottle method could improve the quality and quantity of human islets after purification.     

Improvement of collagenase distribution with the ductal preservation for human islet isolation

A delivery of collagenase at the islet-exocrine interface is crucial for successful human islet isolation. In this study, we investigated how the ductal preservation method at the procurement site affected collagenase distribution. At first, we analyzed human islet isolation data among groups using Serva collagenase with or without ductal injection (DI) or using new Liberase MTF with DI. Then, to assess the distribution of collagenase, human pancreata were classified into two groups: without DI (no DI, n = 5) and with DI at the procurement site (DI, n = 5). Collagenase with 1% marking dye was perfused in the same manner as in our clinical isolation. The distension of the pancreas and the microscopic distribution of the dyed collagenase in pancreas sections were examined. For microscopic analysis, islets were counted and classified into three criteria: unreached, dye didn't reach the islet surface; surface, dye resided on the surface of the islet but not inside; and inside, dye was found inside the islet. As a result, DI groups substantially improved islet yields. In addition, Liberase MTF with DI significantly improved efficacy of pancreas digestion. All pancreata were well distended macroscopically. However, microscopically, the majority of islets in the no DI group were untouched by the dyed collagenase. Ductal preservation substantially improved dyed collagenase delivery on the surface of islets. In conclusion, delivery of collagenase on the surface of islets was unexpectedly insufficient without DI, which was substantially improved by DI. Thus, ductal preservation is a potent method to improve collagenase delivery and islet yields.     

Improvement of collagenase distribution with the ductal preservation for human islet isolation

A delivery of collagenase at the islet-exocrine interface is crucial for successful human islet isolation. In this study, we investigated how the ductal preservation method at the procurement site affected collagenase distribution. At first, we analyzed human islet isolation data among groups using Serva collagenase with or without ductal injection (DI) or using new Liberase MTF with DI. Then, to assess the distribution of collagenase, human pancreata were classified into two groups: without DI (no DI, n = 5) and with DI at the procurement site (DI, n = 5). Collagenase with 1% marking dye was perfused in the same manner as in our clinical isolation. The distension of the pancreas and the microscopic distribution of the dyed collagenase in pancreas sections were examined. For microscopic analysis, islets were counted and classified into three criteria: unreached, dye didn’t reach the islet surface; surface, dye resided on the surface of the islet but not inside; and inside, dye was found inside the islet. As a result, DI groups substantially improved islet yields. In addition, Liberase MTF with DI significantly improved efficacy of pancreas digestion. All pancreata were well distended macroscopically. However, microscopically, the majority of islets in the no DI group were untouched by the dyed collagenase. Ductal preservation substantially improved dyed collagenase delivery on the surface of islets. In conclusion, delivery of collagenase on the surface of islets was unexpectedly insufficient without DI, which was substantially improved by DI. Thus, ductal preservation is a potent method to improve collagenase delivery and islet yields.     

Quantification of islet size and architecture

Human islets exhibit distinct islet architecture particularly in large islets that comprise of a relatively abundant fraction of α-cells intermingled with β-cells, whereas mouse islets show largely similar architecture of a β-cell core with α-cells in the periphery. In humans, islet architecture is islet-size dependent. Changes in endocrine cell mass preferentially occurred in large islets as demonstrated in our recent study on pathological changes of the pancreas in patients with type 2 diabetes.¹ Kilimnik G, Zhao B, Jo J, Periwal V, Witkowski P, Misawa R, et al. Altered islet composition and disproportionate loss of large islets in patients with type 2 diabetes. PLoS One 2011; 6:e27445; http://dx.doi.org/10.1371/journal.pone.0027445; PMID: 22102895 [Crossref], [PubMed], [Web of Science ®] , [Google Scholar] The size dependency of human islets in morphological changes prompted us to develop a method to capture the representative islet distribution in the whole pancreas section combined with a semi-automated analysis to quantify changes in islet architecture. The computer-assisted quantification allows detailed examination of endocrine cell composition in individual islets and minimizes sampling bias. The standard immunohistochemistry based method is widely applicable to various specimens, which is particularly useful for large animal studies but is also applied to a large-scale analysis of the whole organ section from mice. In this article, we describe the method of image capture, parameters measured, data analysis and interpretation of the data.     

Quantification of islet size and architecture

Human islets exhibit distinct islet architecture particularly in large islets that comprise of a relatively abundant fraction of α-cells intermingled with β-cells, whereas mouse islets show largely similar architecture of a β-cell core with α-cells in the periphery. In humans, islet architecture is islet-size dependent. Changes in endocrine cell mass preferentially occurred in large islets as demonstrated in our recent study on pathological changes of the pancreas in patients with type 2 diabetes. ( 1) The size dependency of human islets in morphological changes prompted us to develop a method to capture the representative islet distribution in the whole pancreas section combined with a semi-automated analysis to quantify changes in islet architecture. The computer-assisted quantification allows detailed examination of endocrine cell composition in individual islets and minimizes sampling bias. The standard immunohistochemistry based method is widely applicable to various specimens, which is particularly useful for large animal studies but is also applied to a large-scale analysis of the whole organ section from mice. In this article, we describe the method of image capture, parameters measured, data analysis and interpretation of the data.     

Pancreatic islet isolation variables in non-human primates (rhesus macaques)

BACKGROUND: Non-human primates (NHPs) are important preclinical models for pancreatic islet transplantation (PIT) because of their close phylogenetic and immunological relationship with humans. However, low availability of NHP tissue, long learning curves and prohibitive expenses constrain the consistency of isolated NHP islets for PIT studies. To advance preclinical studies, we attempted to identify key variables that consistently influence the quantity and quality of NHP islets. METHODS: Seventy-two consecutive pancreatic islet isolations from rhesus macaques were reviewed retrospectively. A scaled down, semi-automated islet isolation method was used, and monkeys with streptozotocin-induced diabetes, weighing 3-7 kg, served as recipients for allotransplantation. We analysed the effects of 22 independent variables grouped as donor factors, surgical factors and isolation technique factors. Islet yields, success of isolation and transplantation results were used as quantitative and qualitative outcomes. RESULTS: In the multivariate analysis, variables that significantly affected islet yield were the type of monkey, pancreas preservation, enzyme lot and volume of enzyme delivered. The variables associated with successful isolation were the enzyme lot and volume delivered. The transplant result was correlated with pancreas preservation, enzyme lot, endotoxin levels and COBE collection method. CONCLUSIONS: Islet quantity and quality are highly variable between isolations. The data reviewed suggest that future NHP isolations should use bilayer preservation, infuse more than 80 ml of Liberase into the pancreas, collect non-fractioned tissue from the COBE, and strictly monitor for infection.     

Islet vs. pancreas transplantation in Brazil: Defining criteria for pancreas allocation decision

Background: Many studies have evaluated whether there are characteristics related to pancreas donors and the islet isolation process that can influence in pancreatic islet yield. However, this analysis has not yet been performed in Brazil, one of the world leaders in whole pancreas organ transplantation (WOPT), where pancreas allocation for pancreatic islet transplantation (PIT) has no officially defined criteria. Definition of parameters that would predict the outcome of islet isolation from local pancreas donors would be useful for defining allocation priority in Brazil. Objective: To analyze the relationship between multiple donor-related and islet isolation variables with the total number of isolated pancreatic islet equivalents (IEQ) in a Brazilian sample of pancreas donors. Methods: Several variables were analyzed in 74 pancreata relative to the outcome of total IEQs obtained at the end of the process. Results: In univariate analysis, body mass index (BMI) (p = 0.003), the presence of fatty infiltrates in the pancreas as observed during harvesting (p = 0.042) and pancreas digestion time (p = 0.046) were identified as variables related to a greater IEQ yield. In a multivariate analysis a statistically significant contribution to the variability of islet yield was found only for the BMI (p = 0.017). A ROC curve defined a BMI = 30 as a cut-off point, with pancreata from donors with BMI > 30 yielding more islets than donors with BMI < 30 (p < 0.001). Conclusion: These data reinforces the importance of the donor BMI as a defining parameter for successful islet isolation and establishes this variable as a potential pancreas allocation criterion in Brazil, where there is an unequal competition for good quality organs between WOPT and PIT.     

Clinical islet transplantation in Japan

Introduction

The results of clinical islet transplantation in Japan are, here in, reported and discussed its efficacy and problems.

Methods

Since the first islet transplantation was performed in 2004, 65 islet isolations and 34 islet transplantations to 18 type 1 diabetic patients have been performed in Japan.

Results

Following islet transplantation, patients experienced decreased insulin requirements and lower hemoglobin A1C levels, and positive serum C-peptide levels. All patients achieved stabilized blood glucose levels and the disappearance of hypoglycemic unawareness. Although three patients achieved insulin independency for a limited period, persistent islet graft function was difficult to maintain. Overall islet graft survival was 86.5% at 6 months, 78.7% at 1 year, and 62.9% at 2 years after the first islet transplantation. In our institution, we carried out 23 islet isolations and six islet transplantations to four patients. Although insulin independency was not achieved, all patients showed a disappearance of hypoglycemic unawareness.

Conclusions

Using data from the Japanese Trial of Islet Transplantation, the effectiveness of islet transplantation was shown even when using the pancreata from non-heart-beating donors. Although there are a number of problems to be solved and further improvement is needed, we can state that the introduction of clinical islet transplantation offers hope for type 1 diabetic patients.     

Transgenic mice overproducing islet amyloid polypeptide have increased insulin storage and secretion in vitro

Summary To determine whether chronic overproduction of islet amyloid polypeptide alters beta-cell function, we studied a line of transgenic mice which overexpress islet amyloid polypeptide in their beta-cells. At 3 months of age, these transgenic mice had greater pancreatic content of both islet amyloid polypeptide and insulin. Further, basal and glucose-stimulated secretion of both islet amyloid polypeptide and insulin were also elevated in the perfused pancreas of the transgenic animals. These findings demonstrate that chronic overproduction and secretion of islet amyloid polypeptide are associated with increased insulin storage and enhanced secretion of insulin in vitro. This increase in insulin storage and secretion may be due to a direct effect of islet amyloid polypeptide on the beta-cell or a betacell adaptation to islet amyloid polypeptide-induced insulin resistance.Abbreviations IAPP Islet amyloid polypeptide - bp base pair - TFA trifluoroacetic acid - IRI immunoreactive insulin - SLI somatostatin-like immunoreactivity - IAPP-LI IAPP-like immunoreactivity     

Rupture of a giant non-functioning pancreatic islet cell tumor following minimal trauma

The interesting occurrence of a previously asymptomatic giant non-functioning islet cell tumor of the pancreas identified following rupture secondary to relatively insignificant abdominal trauma is reported. Despite the rarity of the condition, this circumstance underscores the need to be aware of the possibility of pre-existing pathology in evaluating and treating patients for blunt abdominal trauma. To our knowledge, this is the first reported instance of this rare disease entity presented secondary to blunt abdominal trauma.     

C-peptide constricts pancreatic islet arterioles in diabetic, but not normoglycaemic mice

BACKGROUND: Pancreatic islet blood flow is regulated separately from that of the exocrine pancreas, and a consistent finding during impaired glucose tolerance is an increased blood perfusion. The aim of the present study was to investigate whether C-peptide affects pancreatic islet arterioles in normal and diabetic mice. MATERIALS AND METHODS: Control and diabetic C57-Bl mice were studied after 2 weeks of alloxan-induced diabetes. Islet arterioles were dissected and microperfused with Dulbecco's modified Eagle medium (DMEM) solution. The effect of luminal application of mouse C-peptide was investigated. RESULTS: C-peptide reduced the diameter of islet arterioles from diabetic mice (-10+/-4%, P<0.05) compared to base-line values, whilst arterioles from normoglycaemic animals did not respond to C-peptide (P=0.2). CONCLUSION: These findings suggest a role for C-peptide in the regulation of islet blood flow, especially during conditions with impaired glucose tolerance.     

Different digestion enzymes used for human pancreatic islet isolation: A mixed treatment comparison (MTC) meta-analysis

Collagenases are critical reagents determining yield and quality of isolated human pancreatic islets and may affect islet transplantation outcome. Some islet transplantation centers have compared 2 or more collagenase blends; however, the results regarding differences in quantity and quality of islets are conflicting. Thus, for the first time, a mixed treatment comparison (MTC) meta-analysis was carried out to compile data about the effect of different collagenases used for human pancreas digestion on islet yield, purity, viability and stimulation index (SI). Pubmed, Embase and Cochrane libraries were searched. Of 755 articles retrieved, a total of 15 articles fulfilled the eligibility criteria and were included in the MTC meta-analysis. Our results revealed that Vitacyte and Liberase MTF were associated with a small increase in islet yield (islet equivalent number/g pancreas) when compared with Sevac enzyme [standardized mean difference (95% credible interval – CrI) = −2.19 (−4.25 to −0.21) and −2.28 (−4.49 to −0.23), respectively]. However, all other enzyme comparisons did not show any significant difference regarding islet yield. Purity and viability percentages were not significantly different among any of the analyzed digestion enzymes. Interestingly, Vitacyte and Serva NB1 were associated with increased SI when compared with Liberase MTF enzyme [unstandardized weighted mean difference (95% CrI) = −1.69 (−2.87 to −0.51) and −1.07 (−1.79 to −0.39), respectively]. In conclusion, our MTC meta-analysis suggests that the digestion enzymes currently being used for islet isolation works with similar efficiency regarding islet yield, purity and viability; however, Vitacyte and Serva NB1 enzymes seem to be associated with an improved SI as compared with Liberase MTF.     

Different digestion enzymes used for human pancreatic islet isolation: A mixed treatment comparison (MTC) meta-analysis

Collagenases are critical reagents determining yield and quality of isolated human pancreatic islets and may affect islet transplantation outcome. Some islet transplantation centers have compared 2 or more collagenase blends; however, the results regarding differences in quantity and quality of islets are conflicting. Thus, for the first time, a mixed treatment comparison (MTC) meta-analysis was carried out to compile data about the effect of different collagenases used for human pancreas digestion on islet yield, purity, viability and stimulation index (SI). Pubmed, Embase and Cochrane libraries were searched. Of 755 articles retrieved, a total of 15 articles fulfilled the eligibility criteria and were included in the MTC meta-analysis. Our results revealed that Vitacyte and Liberase MTF were associated with a small increase in islet yield (islet equivalent number/g pancreas) when compared with Sevac enzyme [standardized mean difference (95% credible interval – CrI) = ?2.19 (?4.25 to ?0.21) and ?2.28 (?4.49 to ?0.23), respectively]. However, all other enzyme comparisons did not show any significant difference regarding islet yield. Purity and viability percentages were not significantly different among any of the analyzed digestion enzymes. Interestingly, Vitacyte and Serva NB1 were associated with increased SI when compared with Liberase MTF enzyme [unstandardized weighted mean difference (95% CrI) = ?1.69 (?2.87 to ?0.51) and ?1.07 (?1.79 to ?0.39), respectively]. In conclusion, our MTC meta-analysis suggests that the digestion enzymes currently being used for islet isolation works with similar efficiency regarding islet yield, purity and viability; however, Vitacyte and Serva NB1 enzymes seem to be associated with an improved SI as compared with Liberase MTF.     

The IKEM pancreas and islet transplant program as part of healthcare for type 1 diabetes patients: retrospective analysis of outcome from 1983 to 2010

Currently, 25-30 pancreas transplantations per year are carried out in type 1 diabetes (T1D) recipients residing in Czech Republic. Most of the recipients are transplanted together with kidney allografts, but pancreas is also transplanted alone in selected patients with brittle diabetes. Since 2005, the Institute for Clinical and Experimental Medicine (IKEM) islet transplant program was initiated as complementary therapeutic modality. The aim of this paper was to analyze the transplant program at our clinical center, and to examine the survival of recipients, and their pancreas, kidney, and islet grafts. Patient and graft survival rates were evaluated in the following three categories using Kaplan-Meier test: simultaneous pancreas and kidney transplantation (SPKTx), pancreas transplantation alone (PTA), and islet transplantation (ITx). Three hundred and ninety SPKTx, 34 PTA and 44 ITx were carried out between 1983 and 2010. One- and 5-year patient survival rates were 92 % and 81% in SPKTx, respectively. In SPKTx, the 1-year survival rate of pancreas grafts was 78%, and the 5-year rate was 66%. Kidney graft survival rates were 89% and 79%, respectively, after the same follow-up periods. In the PTA category, recipient survivals were 100% after 1 year, and 92% after 3 years. 70% and 65% of pancreatic grafts were working properly at 1 and 3-year follow-ups, respectively. To date, we have carried out 44 islet transplantations in 31 recipients. Islet function (C-peptide ≥ 0.2 ng/ml) was documented in 60% of recipients after 12 months. So far, only 3 patients remained free of exogenous insulin. While SPKTx is a well established treatment for uremic T1D patients, ITx represents an emerging complementary treatment modality. The latter is especially suitable for high-risk recipients, but routine clinical application is still hampered by the limited availability of usable organ transplants and viability of transplanted islets.