Islet transplantation outcomes in type 1 diabetes and transplantation of HLA-DQ8/DR4: results of a single-centre retrospective cohort in Canada.

Background: In solid organ transplantation, HLA matching between donor and recipient is associated with superior outcomes. In islet transplantation, an intervention for Type 1 diabetes, HLA matching between donor and recipient is not performed as part of allocation. Susceptibility to Type 1 diabetes is associated with the presence of certain HLA types. This study was conducted to determine the impact of these susceptibility antigens on islet allograft survival. Methods: This is a single-centre retrospective cohort study. This cohort of transplant recipients (n = 268) received islets from 661 donor pancreases between March 11th, 1999 and August 29th, 2018 at the University of Alberta Hospital (Edmonton, AB, Canada). The frequency of the Type 1 diabetes susceptibility HLA antigens (HLA-A24, -B39, -DQ8, -DQ2 and-DQ2-DQA1∗05) in recipients and donors were determined. Recipient and donor HLA antigens were examined in relation to time to first C-peptide negative status/graft failure or last observation point. Taking into account multiple transplants per patient, we fitted a Gaussian frailty survival analysis model with baseline hazard function stratified by transplant number, adjusted for cumulative islet dose and other confounders. Findings: Across all transplants recipients of donors positive for HLA-DQ8 had significantly better graft survival (adjusted HRs 0.33 95% CI 0.17-0.66; p = 0.002). At first transplant only, donors positive for HLA-DQ2-DQA1∗05 had inferior graft survival (adjusted HR 1.96 95% CI 1.10-3.46); p = 0.02), although this was not significant in the frailty analysis taking multiple transplants into account (adjusted HR 1.46 95% CI 0.77-2.78; p = 0.25). Other HLA antigens were not associated with graft survival after adjustment for confounders. Interpretation: Our findings suggest islet transplantation from HLA-DQ8 donors is associated with superior graft outcomes. A donor positive for HLA-DQ2-DQA1∗05 at first transplant was associated with inferior graft survival but not when taking into account multiple transplants per recipient. The relevance of HLA-antigens on organ allocation needs further evaluation and inclusion in islet transplant registries and additional observational and interventional studies to evaluate the role of HLA-DQ8 in islet graft survival are required. Funding: None.

Encapsulated human islets in alginate fiber maintain long-term functionality.

Maintenance of islet function after in vitro culture is crucial for both transplantation and research. Here we evaluated the effects of encapsulation in alginate fiber on the function of human islets which were distributed by the Alberta Islet Distribution Program. Encapsulated human islets from 15 deceased donors were cultured under 5.5 or 25 mM glucose conditions in vitro. The amounts of C-peptide and glucagon secreted from encapsulated islets into the culture media were measured periodically, and immunohistochemical studies were performed. Encapsulated islets maintained C-peptide and glucagon secretion for more than 75 days in 5 cases; in two cases, their secretion was also successfully detected even on day 180. α- and ß-cell composition and ß-cell survival in islets were unaltered in the fiber after 75 or 180 days of culture. The encapsulated islets cultured with 5.5 mM glucose, but not those with 25 mM glucose, exhibited glucose responsiveness of C-peptide secretion until day 180. We demonstrate that alginate encapsulation enabled human islets to maintain their viability and glucose responsiveness of C-peptide secretion after long-term in vitro culture, potentially for more than for 180 days.

Pancreas and Islet Transplantation: Comparative Outcome Analysis of a Single-centre Cohort Over 20-years.

OBJECTIVE: To provide the largest single-center analysis of islet (ITx) and pancreas (PTx) transplantation. SUMMARY BACKGROUND DATA: Studies describing long-term outcomes with ITx and PTx are scarce. METHODS: We included adults undergoing ITx (n=266) and PTx (n=146) at the University of Alberta from January 1999 to October 2019. Outcomes include patient and graft survival, insulin independence, glycemic control, procedure-related complications, and hospital readmissions. Data are presented as medians (interquartile ranges, IQR) and absolute numbers (percentages, %) and compared using Mann-Whitney and χ2 tests. Kaplan-Meier estimates, Cox proportional hazard models and mixed main effects models were implemented. RESULTS: Crude mortality was 9.4% and 14.4% after ITx and PTx, respectively ( P= 0.141). Sex-adjusted and age-adjusted hazard-ratio for mortality was 2.08 (95% CI, 1.04-4.17, P= 0.038) for PTx versus ITx. Insulin independence occurred in 78.6% and 92.5% in ITx and PTx recipients, respectively ( P= 0.0003), while the total duration of insulin independence was 2.1 (IQR 0.8-4.6) and 6.7 (IQR 2.9-12.4) year for ITx and PTx, respectively ( P= 2.2×10 -22 ). Graft failure ensued in 34.2% and 19.9% after ITx and PTx, respectively ( P =0.002). Glycemic control improved for up to 20-years post-transplant, particularly for PTx recipients (group, P= 7.4×10 -7 , time, P =4.8×10 -6 , group*time, P= 1.2×10 -7 ). Procedure-related complications and hospital readmissions were higher after PTx ( P =2.5×10 -32 and P= 6.4×10 -112 , respectively). CONCLUSIONS: PTx shows higher sex-adjusted and age-adjusted mortality, procedure-related complications and readmissions compared with ITx. Conversely, insulin independence, graft survival and glycemic control are better with PTx. This study provides data to balance risks and benefits with ITx and PTx, which could improve shared decision-making.

Evaluating the Potential for ABO-incompatible Islet Transplantation: Expression of ABH Antigens on Human Pancreata, Isolated Islets, and Embryonic Stem Cell-derived Islets.

BACKGROUND: ABO-incompatible transplantation has improved accessibility of kidney, heart, and liver transplantation. Pancreatic islet transplantation continues to be ABO-matched, yet ABH antigen expression within isolated human islets or novel human embryonic stem cell (hESC)-derived islets remain uncharacterized. METHODS: We evaluated ABH glycans within human pancreata, isolated islets, hESC-derived pancreatic progenitors, and the ensuing in vivo mature islets following kidney subcapsular transplantation in rats. Analyses include fluorescence immunohistochemistry and single-cell analysis using flow cytometry. RESULTS: Within the pancreas, endocrine and ductal cells do not express ABH antigens. Conversely, pancreatic acinar tissues strongly express these antigens. Acinar tissues are present in a substantial portion of cells within islet preparations obtained for clinical transplantation. The hESC-derived pancreatic progenitors and their ensuing in vivo-matured islet-like clusters do not express ABH antigens. CONCLUSIONS: Clinical pancreatic islet transplantation should remain ABO-matched because of contaminant acinar tissue within islet preparations that express ABH glycans. Alternatively, hESC-derived pancreatic progenitors and the resulting in vivo-matured hESC-derived islets do not express ABH antigens. These findings introduce the potential for ABO-incompatible cell replacement treatment and offer evidence to support scalability of hESC-derived cell therapies in type 1 diabetes.

C-peptide Targets and Patient-centered Outcomes of Relevance to Cellular Transplantation for Diabetes.

BACKGROUND: C-peptide levels are a key measure of beta-cell mass following islet transplantation, but threshold values required to achieve clinically relevant patient-centered outcomes are not yet established. METHODS: We conducted a cross-sectional retrospective cohort study evaluating patients undergoing islet transplantation at a single center from 1999 to 2018. Cohorts included patients achieving insulin independence without hypoglycemia, those with insulin dependence without hypoglycemia, and those with recurrent symptomatic hypoglycemia. Primary outcome was fasting C-peptide levels at 6 to 12 mo postfirst transplant; secondary outcomes included stimulated C-peptide levels and BETA-2 scores. Fasting and stimulated C-peptide and BETA-2 cutoff values for determination of hypoglycemic freedom and insulin independence were evaluated using receiver operating characteristic curves. RESULTS: We analyzed 192 patients, with 122 (63.5%) being insulin independent without hypoglycemia, 61 (31.8%) being insulin dependent without hypoglycemia, and 9 (4.7%) experiencing recurrent symptomatic hypoglycemia. Patients with insulin independence had a median (interquartile range) fasting C-peptide level of 0.66 nmol/L (0.34 nmol/L), compared with 0.49 nmol/L (0.25 nmol/L) for those being insulin dependent without hypoglycemia and 0.07 nmol/L (0.05 nmol/L) for patients experiencing hypoglycemia ( P < 0.001). Optimal fasting C-peptide cutoffs for insulin independence and hypoglycemia were ≥0.50 nmol/L and ≥0.12 nmol/L, respectively. Cutoffs for insulin independence and freedom of hypoglycemia using stimulated C-peptide were ≥1.2 nmol/L and ≥0.68 nmol/L, respectively, whereas optimal cutoff BETA-2 scores were ≥16.4 and ≥5.2. CONCLUSIONS: We define C-peptide levels and BETA-2 scores associated with patient-centered outcomes. Characterizing these values will enable evaluation of ongoing clinical trials with islet or stem cell therapies.

A predictive computational platform for optimizing the design of bioartificial pancreas devices.

The delivery of encapsulated islets or stem cell-derived insulin-producing cells (i.e., bioartificial pancreas devices) may achieve a functional cure for type 1 diabetes, but their efficacy is limited by mass transport constraints. Modeling such constraints is thus desirable, but previous efforts invoke simplifications which limit the utility of their insights. Herein, we present a computational platform for investigating the therapeutic capacity of generic and user-programmable bioartificial pancreas devices, which accounts for highly influential stochastic properties including the size distribution and random localization of the cells. We first apply the platform in a study which finds that endogenous islet size distribution variance significantly influences device potency. Then we pursue optimizations, determining ideal device structures and estimates of the curative cell dose. Finally, we propose a new, device-specific islet equivalence conversion table, and develop a surrogate machine learning model, hosted on a web application, to rapidly produce these coefficients for user-defined devices.

E2F1 transcription factor mediates a link between fat and islets to promote ß cell proliferation in response to acute insulin resistance.

Prevention or amelioration of declining ß cell mass is a potential strategy to cure diabetes. Here, we report the pathways utilized by ß cells to robustly replicate in response to acute insulin resistance induced by S961, a pharmacological insulin receptor antagonist. Interestingly, pathways that include CENP-A and the transcription factor E2F1 that are independent of insulin signaling and its substrates appeared to mediate S961-induced ß cell multiplication. Consistently, pharmacological inhibition of E2F1 blocks ß-cell proliferation in S961-injected mice. Serum from S961-treated mice recapitulates replication of ß cells in mouse and human islets in an E2F1-dependent manner. Co-culture of islets with adipocytes isolated from S961-treated mice enables ß cells to duplicate, while E2F1 inhibition limits their growth even in the presence of adipocytes. These data suggest insulin resistance-induced proliferative signals from adipocytes activate E2F1, a potential therapeutic target, to promote ß cell compensation.

Uncoupling protein 2 and aldolase B impact insulin release by modulating mitochondrial function and Ca2+ release from the ER.

Uncoupling protein 2 (UCP2), a mitochondrial protein, is known to be upregulated in pancreatic islets of patients with type 2 diabetes (T2DM); however, the pathological significance of this increase in UCP2 expression is unclear. In this study, we highlight the molecular link between the increase in UCP2 expression in ß-cells and ß-cell failure by using genetically engineered mice and human islets. ß-cell-specific UCP2-overexpressing transgenic mice (ßUCP2Tg) exhibited glucose intolerance and a reduction in insulin secretion. Decreased mitochondrial function and increased aldolase B (AldB) expression through oxidative-stress-mediated pathway were observed in ßUCP2Tg islets. AldB, a glycolytic enzyme, was associated with reduced insulin secretion via mitochondrial dysfunction and impaired calcium release from the endoplasmic reticulum (ER). Taken together, our findings provide a new mechanism of ß-cell dysfunction by UCP2 and AldB. Targeting the UCP2/AldB axis is a promising approach for the recovery of ß-cell function.

Outcomes Following Extrahepatic and Intraportal Pancreatic Islet Transplantation: A Comparative Cohort Study.

BACKGROUND: Preliminary studies show promise for extrahepatic islet transplantation (ITx). However, clinical comparisons with intraportal ITx outcomes remain limited. METHODS: This single-center cohort study evaluates patients receiving extrahepatic or intraportal ITx between 1999 and 2018. Primary outcome was stimulated C-peptide level. Secondary outcomes were fasting plasma glucose, BETA-2 scores, and fasting C-peptide level. Multivariable logistic modeling evaluated factors independently associated with a composite variable of early graft failure and primary nonfunction within 60 d of ITx. RESULTS: Of 264 patients, 9 (3.5%) received extrahepatic ITx (gastric submucosal = 2, subcutaneous = 3, omental = 4). Group demographics were similar at baseline (age, body mass index, diabetes duration, and glycemic control). At 1-3 mo post-first infusion, patients receiving extrahepatic ITx had significantly lower stimulated C-peptide (0.05 nmol/L versus 1.2 nmol/L, P < 0.001), higher fasting plasma glucose (9.3 mmol/L versus 7.3 mmol/L, P < 0.001), and lower BETA-2 scores (0 versus 11.6, P < 0.001) and SUITO indices (1.5 versus 39.6, P < 0.001) compared with those receiving intraportal ITx. Subjects receiving extrahepatic grafts failed to produce median C-peptide ≥0.2 nmol/L within the first 60 d after transplant. Subsequent intraportal infusion following extrahepatic transplants achieved equivalent outcomes compared with patients receiving intraportal transplant alone. Extrahepatic ITx was independently associated with early graft failure/primary non-function (odds ratio 1.709, confidence interval 73.8-39 616.0, P < 0.001), whereas no other factors were independently predictive. CONCLUSIONS: Using current techniques, intraportal islet infusion remains the gold standard for clinical ITx, with superior engraftment, graft function, and glycemic outcomes compared with extrahepatic transplantation of human islets.

Pancreatic islet transplantation in type 1 diabetes: 20-year experience from a single-centre cohort in Canada.

BACKGROUND: Islet transplantation offers an effective treatment for selected people with type 1 diabetes and intractable hypoglycaemia. Long-term experience, however, remains limited. We report outcomes from a single-centre cohort up to 20 years after islet transplantation. METHODS: This cohort study included patients older than 18 years with type 1 diabetes undergoing allogeneic islet transplantation between March 11, 1999, and Oct 1, 2019, at the University of Alberta Hospital (Edmonton, AB, Canada). Patients who underwent islet-after-kidney transplantation and islet transplantation alone or islet transplantation before whole-pancreas transplantation (follow-up was censored at the time of whole-pancreas transplantation) were included. Patient survival, graft survival (fasting plasma C-peptide >0·1 nmol/L), insulin independence, glycaemic control, and adverse events are reported. To identify factors associated with prolonged graft survival, recipients with sustained graft survival (≥90% of patient follow-up duration) were compared with those who had non-sustained graft survival (<90% of follow-up duration). Multivariate binary logistic regression analyses were done to determine predictors of sustained graft survival. FINDINGS: Between March 11, 1999, and Oct 1, 2019, 255 patients underwent islet transplantation and were included in the analyses (149 [58%] were female and 218 [85%] were White). Over a median follow-up of 7·4 years (IQR 4·4-12·2), 230 (90%) patients survived. Median graft survival was 5·9 years (IQR 3·0-9·5), and graft failure occurred in 91 (36%) patients. 178 (70%) recipients had sustained graft survival, and 77 (30%) had non-sustained graft survival. At baseline, compared with patients with non-sustained graft survival, those with sustained graft survival had longer median type 1 diabetes duration (33·5 years [IQR 24·3-41·7] vs 26·2 years [17·0-35·5]; p=0·0003), median older age (49·4 years [43·5-56·1] vs 44·2 years [35·4-54·2]; p=0·0011), and lower median insulin requirements (0·53 units/kg per day [0·45-0·67] vs 0·59 units/kg per day [0·48-0·70]; p=0·032), but median HbA1c concentrations were similar (8·2% [7·5-9·0] vs 8·5% [7·8-9·2]; p=0·23). 201 (79%) recipients had insulin independence, with a Kaplan-Meier estimate of 61% (95% CI 54-67) at 1 year, 32% (25-39) at 5 years, 20% (14-27) at 10 years, 11% (6-18) at 15 years, and 8% (2-17) at 20 years. Patients with sustained graft survival had significantly higher rates of insulin independence (160 [90%] of 178 vs 41 [53%] of 77; p<0·0001) and sustained improvements in glycaemic control mixed-main-effects model group effect, p<0·0001) compared with those with non-sustained graft survival. Multivariate analyses identified the combined use of anakinra plus etanercept (adjusted odds ratio 7·5 [95% CI 2·7-21·0], p<0·0001) and the BETA-2 score of 15 or higher (4·1 [1·5-11·4], p=0·0066) as factors associated with sustained graft survival. In recipients with sustained graft survival, the incidence of procedural complications was lower (23 [5%] of 443 infusions vs 17 [10%] of 167 infusions; p=0·027), whereas the incidence of cancer was higher (29 of [16%] of 178 vs four [5%] of 77; p=0·015) than in those with non-sustained graft survival; most were skin cancers (22 [67%] of 33). End-stage renal disease and severe infections were similar between groups. INTERPRETATION: We present the largest single-centre cohort study of long-term outcomes following islet transplantation. Although some limitations with our study remain, such as the retrospective component, a relatively small sample size, and the absence of non-transplant controls, we found that the combined use of anakinra plus etanercept and the BETA-2 score were associated with improved outcomes, and therefore these factors could inform clinical practice. FUNDING: None.

Imeglimin Ameliorates ß-Cell Apoptosis by Modulating the Endoplasmic Reticulum Homeostasis Pathway.

The effects of imeglimin, a novel antidiabetes agent, on ß-cell function remain unclear. Here, we unveiled the impact of imeglimin on ß-cell survival. Treatment with imeglimin augmented mitochondrial function, enhanced insulin secretion, promoted ß-cell proliferation, and improved ß-cell survival in mouse islets. Imeglimin upregulated the expression of endoplasmic reticulum (ER)-related molecules, including Chop (Ddit3), Gadd34 (Ppp1r15a), Atf3, and Sdf2l1, and decreased eIF2α phosphorylation after treatment with thapsigargin and restored global protein synthesis in ß-cells under ER stress. Imeglimin failed to protect against ER stress-induced ß-cell apoptosis in CHOP-deficient islets or in the presence of GADD34 inhibitor. Treatment with imeglimin showed a significant decrease in the number of apoptotic ß-cells and increased ß-cell mass in Akita mice. Imeglimin also protected against ß-cell apoptosis in both human islets and human pluripotent stem cell-derived ß-like cells. Taken together, imeglimin modulates the ER homeostasis pathway, which results in the prevention of ß-cell apoptosis both in vitro and in vivo.

Glucose metabolism and pyruvate carboxylase enhance glutathione synthesis and restrict oxidative stress in pancreatic islets.

Glucose metabolism modulates the islet ß cell responses to diabetogenic stress, including inflammation. Here, we probed the metabolic mechanisms that underlie the protective effect of glucose in inflammation by interrogating the metabolite profiles of primary islets from human donors and identified de novo glutathione synthesis as a prominent glucose-driven pro-survival pathway. We find that pyruvate carboxylase is required for glutathione synthesis in islets and promotes their antioxidant capacity to counter inflammation and nitrosative stress. Loss- and gain-of-function studies indicate that pyruvate carboxylase is necessary and sufficient to mediate the metabolic input from glucose into glutathione synthesis and the oxidative stress response. Altered redox metabolism and cellular capacity to replenish glutathione pools are relevant in multiple pathologies beyond obesity and diabetes. Our findings reveal a direct interplay between glucose metabolism and glutathione biosynthesis via pyruvate carboxylase. This metabolic axis may also have implications in other settings where sustaining glutathione is essential.

Silencing the G-protein coupled receptor 3-salt inducible kinase 2 pathway promotes human ß cell proliferation.

Loss of pancreatic ß cells is the hallmark of type 1 diabetes, for which provision of insulin is the standard of care. While regenerative and stem cell therapies hold the promise of generating single-source or host-matched tissue to obviate immune-mediated complications, these will still require surgical intervention and immunosuppression. Here we report the development of a high-throughput RNAi screening approach to identify upstream pathways that regulate adult human ß cell quiescence and demonstrate in a screen of the GPCRome that silencing G-protein coupled receptor 3 (GPR3) leads to human pancreatic ß cell proliferation. Loss of GPR3 leads to activation of Salt Inducible Kinase 2 (SIK2), which is necessary and sufficient to drive cell cycle entry, increase ß cell mass, and enhance insulin secretion in mice. Taken together, our data show that targeting the GPR3-SIK2 pathway is a potential strategy to stimulate the regeneration of ß cells.

Total pancreatectomy with islet cell autotransplantation in a 2-year-old child with hereditary pancreatitis due to a PRSS1 mutation.

Acute recurrent and chronic pancreatitis in children carries high morbidity and burden. Compared to adults, ~75% of the cases of chronic pancreatitis in children are associated with underlying genetic mutations. The decision to intervene and the optimal timing poses unique challenges. Total pancreatectomy and islet cell autotransplantation (TPIAT) provides definitive therapy to relieve pain and improve quality of life while minimizing the risk of pancreatogenic diabetes. Substantial clinical data are available for adults; however, information on clinical outcomes in children remains scarce, particularly for very young children. Herein, we present an unusual, complex case of a 2-year-old child that underwent a successful TPIAT due to hereditary pancreatitis with an underlying mutation in PRSS1 gene, complicated by unremitting pancreatic ascites, hemorrhage, and sepsis. This is the youngest case to be reported in the literature. We provide a comprehensive report of the course and procedures implemented in this patient to guide other teams when considering these extraordinary measures in similar cases.

CD82 is a marker to isolate ß cell precursors from human iPS cells and plays a role for the maturation of ß cells.

Generation of pancreatic ß cells from pluripotent stem cells is a key technology to develop cell therapy for insulin-dependent diabetes and considerable efforts have been made to produce ß cells. However, due to multiple and lengthy differentiation steps, production of ß cells is often unstable. It is also desirable to eliminate undifferentiated cells to avoid potential risks of tumorigenesis. To isolate ß cell precursors from late stage pancreatic endocrine progenitor (EP) cells derived from iPS cells, we have identified CD82, a member of the tetraspanin family. CD82+ cells at the EP stage differentiated into endocrine cells more efficiently than CD82- EP stage cells. We also show that CD82+ cells in human islets secreted insulin more efficiently than CD82- cells. Furthermore, knockdown of CD82 expression by siRNA or inhibition of CD82 by monoclonal antibodies in NGN3+ cells suppressed the function of ß cells with glucose-stimulated insulin secretion, suggesting that CD82 plays a role in maturation of EP cells to ß cells.

Clinical islet transplantation: Current progress and new frontiers.

Islet transplantation (IT) is now a robust treatment for selected patients with type 1 diabetes suffering from recurrent hypoglycemia and impaired awareness of hypoglycemia. A global soar of clinical islet transplant programs attests to the commitment of many institutions and researchers to advance IT as a potential cure for this devastating disease. However, many challenges limiting the widespread applicability of clinical IT remain. In this review, we will touch on the milestones in the history of IT and its path to clinical success, discuss the current challenges around IT, propose some possible solutions, and elaborate on the frontiers envisioned in the future of clinical IT.

Frequency of Obliteration of the Dorsal and Ventral Ducts of the Pancreas in Islet Transplantation.

BACKGROUND: Islet isolation is an essential process in every human islet transplantation protocol. Intraductal enzyme delivery followed by adequate distention of the pancreas is the most critical step in islet isolation. Anomalies of the pancreatic duct system can significantly affect this process. Thus, identification and characterization of ductal patency is of paramount importance to achieve optimal islet isolation. AIMS: To investigate the frequency of duct obliteration in the human pancreas and explore donor/patient characteristics associated with specific ductal variations. METHODS: We examined ductal patency of pancreata allocated for islet allotransplantation (n = 597) and autotransplantation (n = 21) after removal of the duodenum during islet isolation procedure. Donor/patient factors were reviewed from the batch files. RESULTS: Among 559 deceased donor pancreata without pancreas divisum (n = 38, 6.4%), both ducts were patent in 50.1%, only ventral duct was patent in 46.7%, and only dorsal duct was patent in 3.2%. Donor age was not associated with the frequency of obliterated dorsal duct. Black race tended to have the higher frequency of patent dorsal duct. As expected, pancreas divisum was more frequent in chronic pancreatitis cases (n = 6, 28.6%). Within 7 cases of chronic pancreatitis with unknown etiology, we encountered one case of ventral duct obliteration. CONCLUSIONS: The minor duodenal papilla and aging do not likely play an important role in the occurrence of dorsal duct obliteration. Although frequency of obliterated ventral duct was low in our population, physicians, including gastroenterologists and endoscopists, as well as islet transplantation researchers should be aware of this possibility.

Glucose-dependent partitioning of arginine to the urea cycle protects ß-cells from inflammation.

Chronic inflammation is linked to diverse disease processes, but the intrinsic mechanisms that determine cellular sensitivity to inflammation are incompletely understood. Here, we show the contribution of glucose metabolism to inflammation-induced changes in the survival of pancreatic islet ß-cells. Using metabolomic, biochemical and functional analyses, we investigate the protective versus non-protective effects of glucose in the presence of pro-inflammatory cytokines. When protective, glucose metabolism augments anaplerotic input into the TCA cycle via pyruvate carboxylase (PC) activity, leading to increased aspartate levels. This metabolic mechanism supports the argininosuccinate shunt, which fuels ureagenesis from arginine and conversely diminishes arginine utilization for production of nitric oxide (NO), a chief mediator of inflammatory cytotoxicity. Activation of the PC-urea cycle axis is sufficient to suppress NO synthesis and shield cells from death in the context of inflammation and other stress paradigms. Overall, these studies uncover a previously unappreciated link between glucose metabolism and arginine-utilizing pathways via PC-directed ureagenesis as a protective mechanism.

Luseogliflozin increases beta cell proliferation through humoral factors that activate an insulin receptor- and IGF-1 receptor-independent pathway.

AIMS/HYPOTHESIS: Sodium-glucose cotransporter 2 (SGLT2) inhibitors, which prevent the renal reabsorption of glucose, decrease blood glucose levels in an insulin-independent manner. We previously reported creating a mouse model of systemic inhibition of target receptors for both insulin and IGF-1 by treating animals with OSI-906, a dual insulin/IGF-1 receptor inhibitor, for 7 days. The OSI-906-treated mice exhibited an increased beta cell mass, hepatic steatosis and adipose tissue atrophy, accompanied by hyperglycaemia and hyperinsulinaemia. In the present study, we investigated the effects of an SGLT2 inhibitor, luseogliflozin, on these changes in OSI-906-treated mice. METHODS: We treated C57BL/6J male mice either with vehicle, luseogliflozin, OSI-906 or OSI-906 plus luseogliflozin for 7 days, and phenotyping was performed to determine beta cell mass and proliferation. Subsequently, we tested whether serum-derived factors have an effect on beta cell proliferation in genetically engineered beta cells, mouse islets or human islets. RESULTS: SGLT2 inhibition with luseogliflozin significantly ameliorated hyperglycaemia, but not hyperinsulinaemia, in the OSI-906-treated mice. Liver steatosis and adipose tissue atrophy induced by OSI-906 were not altered by treatment with luseogliflozin. Beta cell mass and proliferation were further increased by SGLT2 inhibition with luseogliflozin in the OSI-906-treated mice. Luseogliflozin upregulated gene expression related to the forkhead box M1 (FoxM1)/polo-like kinase 1 (PLK1)/centromere protein A (CENP-A) pathway in the islets of OSI-906-treated mice. The increase in beta cell proliferation was recapitulated in a co-culture of Irs2 knockout and Insr/IR knockout (ßIRKO) beta cells with serum from both luseogliflozin- and OSI-906-treated mice, but not after SGLT2 inhibition in beta cells. Circulating factors in both luseogliflozin- and OSI-906-treated mice promoted beta cell proliferation in both mouse islets and cadaveric human islets. CONCLUSIONS/INTERPRETATION: These results suggest that luseogliflozin can increase beta cell proliferation through the activation of the FoxM1/PLK1/CENP-A pathway via humoral factors that act in an insulin/IGF-1 receptor-independent manner.