Explore the vast range of titles available.

Aims/hypothesis Increased circulating levels of incompletely processed insulin (i.e. proinsulin) are observed clinically in type 1 and type 2 diabetes. Previous studies have suggested that Ca 2+ signalling within beta cells regulates insulin processing and secretion; however, the mechanisms that link impaired Ca 2+ signalling with defective insulin maturation remain incompletely understood. Methods We generated mice with beta cell-specific sarcoendoplasmic reticulum Ca 2+ ATPase-2 (SERCA2) deletion (βS2KO mice) and used an INS-1 cell line model of SERCA2 deficiency. Whole-body metabolic phenotyping, Ca 2+ imaging, RNA-seq and protein processing assays were used to determine how loss of SERCA2 impacts beta cell function. To test key findings in human model systems, cadaveric islets were treated with diabetogenic stressors and prohormone convertase expression patterns were characterised. Results βS2KO mice exhibited age-dependent glucose intolerance and increased plasma and pancreatic levels of proinsulin, while endoplasmic reticulum (ER) Ca 2+ levels and glucose-stimulated Ca 2+ synchronicity were reduced in βS2KO islets. Islets isolated from βS2KO mice and SERCA2-deficient INS-1 cells showed decreased expression of the active forms of the proinsulin processing enzymes PC1/3 and PC2. Additionally, immunofluorescence staining revealed mis-location and abnormal accumulation of proinsulin and proPC2 in the intermediate region between the ER and the Golgi (i.e. the ERGIC) and in the cis-Golgi in beta cells of βS2KO mice. Treatment of islets from human donors without diabetes with high glucose and palmitate concentrations led to reduced expression of the active forms of the proinsulin processing enzymes, thus phenocopying the findings observed in βS2KO islets and SERCA2-deficient INS-1 cells. Similar findings were observed in wild-type mouse islets treated with brefeldin A, a compound that perturbs ER-to-Golgi trafficking. Conclusions/interpretation Taken together, these data highlight an important link between ER Ca 2+ homeostasis and proinsulin processing in beta cells. Our findings suggest a model whereby chronic ER Ca 2+ depletion due to SERCA2 deficiency impairs the spatial regulation of prohormone trafficking, processing and maturation within the secretory pathway. Data availability RNA-seq data have been deposited in the Gene Expression Omnibus (GEO; accession no.: GSE207498). Graphical Abstract

The non-β endocrine cells in pancreatic islets play an essential counterpart and regulatory role to the insulin-producing β-cells in the regulation of blood-glucose homeostasis. While significant progress has been made towards the understanding of β-cell regeneration in adults, very little is known about the regeneration of the non-β endocrine cells such as glucagon-producing α-cells and somatostatin producing δ-cells. Previous studies have noted the increase of α-cell composition in diabetes patients and in animal models. It is thus our hypothesis that non-β-cells such as α-cells and δ-cells in adults can regenerate, and that the regeneration accelerates in diabetic conditions. To test this hypothesis, we examined islet cell composition in a streptozotocin (STZ)-induced diabetes mouse model in detail. Our data showed the number of α-cells in each islet increased following STZ-mediated β-cell destruction, peaked at Day 6, which was about 3 times that of normal islets. In addition, we found δ-cell numbers doubled by Day 6 following STZ treatment. These data suggest α- and δ-cell regeneration occurred rapidly following a single diabetes-inducing dose of STZ in mice. Using in vivo BrdU labeling techniques, we demonstrated α- and δ-cell regeneration involved cell proliferation. Co-staining of the islets with the proliferating cell marker Ki67 showed α- and δ-cells could replicate, suggesting self-duplication played a role in their regeneration. Furthermore, Pdx1(+)/Insulin(-) cells were detected following STZ treatment, indicating the involvement of endocrine progenitor cells in the regeneration of these non-β cells. This is further confirmed by the detection of Pdx1(+)/glucagon(+) cells and Pdx1(+)/somatostatin(+) cells following STZ treatment. Taken together, our study demonstrated adult α- and δ-cells could regenerate, and both self-duplication and regeneration from endocrine precursor cells were involved in their regeneration.

Introduction Initially approved for the fifth‐line or later therapeutic setting, the chimeric antigen receptor (CAR) T‐cell regimen ciltacabtagene autoleucel (cilta‐cel) was recently approved for second‐line (2L) treatment in relapsed/refractory multiple myeloma (RRMM). Oncology practitioners use clinical trials to inform treatment, but real‐world impressions and impact on practice are lacking. We aimed to determine whether presenting CARTITUDE‐4 clinical trial data would impact real‐world preferences/perceptions around CAR T‐cell therapy. Methods Recruiting from the Cardinal Health Oncology Provider Extended Network (OPEN), we surveyed hematologists/oncologists to investigate fourth‐line (4L) preferences in a hypothetical patient with triple‐class–refractory MM. We posed the same questions and answers before and after the trial presentation and compared pre‐/post‐preferences toward cilta‐cel and sequencing relative to bispecific antibodies (BsAbs). Using the same methodology as described above, we also performed a secondary analysis comparing pre‐/post‐perceptions on the use of CAR T‐cell therapy in earlier lines for patients with triple‐class–refractory MM. Results Among 50 respondents, decision‐making factors before the trial presentation included CAR T‐cell center availability (58%), comorbidities (52%), and center locations (34%). Additionally, 48% of 46 respondents chose 4L cilta‐cel. Among 47, 40% wanted more real‐world/long‐term CAR T‐cell therapy outcomes in any line, 38% wanted more 2L data, and 34% favored 2L/third‐line (3L) use. After the presentation, the preference for cilta‐cel doubled from 48% to 88% (p < 0.001) among 50 respondents and rose from 34% to 55% (p = 0.001) for earlier‐line CAR T‐cell therapy among 49. Moreover, 55% of 49 respondents preferred CAR T‐cell therapy prior to BsAbs. Discussion We have shown that making oncology practitioners aware of trials precipitated decision‐making factors and led to notable, significant shifts in future intended practice patterns. Being aware of trial data enables practitioners to make more informed decisions, tailor therapies to individual patients, and ultimately improve outcomes.

Ongoing studies suggest an important role for iPLA2β in a multitude of biological processes and it has been implicated in neurodegenerative, skeletal and vascular smooth muscle disorders, bone formation, and cardiac arrhythmias. Thus, identifying an iPLA2βinhibitor that can be reliably and safely used in vivo is warranted. Currently, the mechanism-based inhibitor bromoenol lactone (BEL) is the most widely used to discern the role of iPLA2β in biological processes. While BEL is recognized as a more potent inhibitor of iPLA2 than of cPLA2 or sPLA2, leading to its designation as a \"specific\" inhibitor of iPLA2, it has been shown to also inhibit non-PLA2 enzymes. A potential complication of its use is that while the S and R enantiomers of BEL exhibit preference for cytosol-associated iPLA2β and membrane-associated iPLA2γ, respectively, the selectivity is only 10-fold for both. In addition, BEL is unstable in solution, promotes irreversible inhibition, and may be cytotoxic, making BEL not amenable for in vivo use. Recently, a fluoroketone (FK)-based compound (FKGK18) was described as a potent inhibitor of iPLA2β. Here we characterized its inhibitory profile in beta-cells and find that FKGK18: (a) inhibits iPLA2β with a greater potency (100-fold) than iPLA2γ, (b) inhibition of iPLA2β is reversible, (c) is an ineffective inhibitor of α-chymotrypsin, and (d) inhibits previously described outcomes of iPLA2β activation including (i) glucose-stimulated insulin secretion, (ii) arachidonic acid hydrolysis; as reflected by PGE2 release from human islets, (iii) ER stress-induced neutral sphingomyelinase 2 expression, and (iv) ER stress-induced beta-cell apoptosis. These findings suggest that FKGK18 is similar to BEL in its ability to inhibit iPLA2β. Because, in contrast to BEL, it is reversible and not a non-specific inhibitor of proteases, it is suggested that FKGK18 is more ideal for ex vivo and in vivo assessments of iPLA2β role in biological functions.

Type 1 diabetes (T1D) is a consequence of autoimmune β cell destruction, but the role of lipids in this process is unknown. We previously reported that activation of Ca2+-independent phospholipase A2β (iPLA2β) modulates polarization of macrophages (MΦ). Hydrolysis of the sn-2 substituent of glycerophospholipids by iPLA2β can lead to the generation of oxidized lipids (eicosanoids), pro- and antiinflammatory, which can initiate and amplify immune responses triggering β cell death. As MΦ are early triggers of immune responses in islets, we examined the impact of iPLA2β-derived lipids (iDLs) in spontaneous-T1D prone nonobese diabetic mice (NOD), in the context of MΦ production and plasma abundances of eicosanoids and sphingolipids. We find that (a) MΦNOD exhibit a proinflammatory lipid landscape during the prediabetic phase; (b) early inhibition or genetic reduction of iPLA2β reduces production of select proinflammatory lipids, promotes antiinflammatory MΦ phenotype, and reduces T1D incidence; (c) such lipid changes are reflected in NOD plasma during the prediabetic phase and at T1D onset; and (d) importantly, similar lipid signatures are evidenced in plasma of human subjects at high risk for developing T1D. These findings suggest that iDLs contribute to T1D onset and identify select lipids that could be targeted for therapeutics and, in conjunction with autoantibodies, serve as early biomarkers of pre-T1D.

Purpose of Review Type 1 diabetes (T1D) is an autoimmune disease marked by β-cell destruction. Immunotherapies for T1D have been investigated since the 1980s and have focused on restoration of tolerance, T cell or B cell inhibition, regulatory T cell (Treg) induction, suppression of innate immunity and inflammation, immune system reset, and islet transplantation. The purpose of this review is to provide an overview and lessons learned from single immunotherapy trials, describe recent and ongoing combination immunotherapy trials, and provide perspectives on strategies for future combination clinical interventions aimed at preserving insulin secretion in T1D. Recent Findings Combination immunotherapies have had mixed results in improving short-term glycemic control and insulin secretion in recent-onset T1D. Summary A handful of studies have successfully reached their primary end-point of improved insulin secretion in recent-onset T1D. However, long-term improvements glycemic control and the restoration of insulin independence remain elusive. Future interventions should focus on strategies that combine immunomodulation with efforts to alleviate β-cell stress and address the formation of antigens that activate autoimmunity.

A bidirectional and complex relationship exists between bone and glycemia. Persons with type 2 diabetes (T2D) are at risk for bone loss and fracture, however, heightened osteoanabolism may ameliorate T2D-induced deficits in glycemia as bone-forming osteoblasts contribute to energy metabolism via increased glucose uptake and cellular glycolysis. Mice globally lacking nuclear matrix protein 4 (Nmp4), a transcription factor expressed in all tissues and conserved between humans and rodents, are healthy and exhibit enhanced bone formation in response to anabolic osteoporosis therapies. To test whether loss of Nmp4 similarly impacted bone deficits caused by diet-induced obesity, male wild-type and Nmp4−/− mice (8 weeks) were fed either low-fat diet or high-fat diet (HFD) for 12 weeks. Endpoint parameters included bone architecture, structural and estimated tissue-level mechanical properties, body weight/composition, glucose-stimulated insulin secretion, glucose tolerance, insulin tolerance, and metabolic cage analysis. HFD diminished bone architecture and ultimate force and stiffness equally in both genotypes. Unexpectedly, the Nmp4−/− mice exhibited deficits in pancreatic β-cell function and were modestly glucose intolerant under normal diet conditions. Despite the β-cell deficits, the Nmp4−/− mice were less sensitive to HFD-induced weight gain, increases in % fat mass, and decreases in glucose tolerance and insulin sensitivity. We conclude that Nmp4 supports pancreatic β-cell function but suppresses peripheral glucose utilization, perhaps contributing to its suppression of induced skeletal anabolism. Selective disruption of Nmp4 in peripheral tissues may provide a strategy for improving both induced osteoanabolism and energy metabolism in comorbid patients.

Ongoing studies suggest an important role for iPLA.sub.2 [beta] in a multitude of biological processes and it has been implicated in neurodegenerative, skeletal and vascular smooth muscle disorders, bone formation, and cardiac arrhythmias. Thus, identifying an iPLA.sub.2 [beta]inhibitor that can be reliably and safely used in vivo is warranted. Currently, the mechanism-based inhibitor bromoenol lactone (BEL) is the most widely used to discern the role of iPLA.sub.2 [beta] in biological processes. While BEL is recognized as a more potent inhibitor of iPLA.sub.2 than of cPLA.sub.2 or sPLA.sub.2, leading to its designation as a \"specific\" inhibitor of iPLA.sub.2, it has been shown to also inhibit non-PLA.sub.2 enzymes. A potential complication of its use is that while the S and R enantiomers of BEL exhibit preference for cytosol-associated iPLA.sub.2 [beta] and membrane-associated iPLA.sub.2 [gamma], respectively, the selectivity is only 10-fold for both. In addition, BEL is unstable in solution, promotes irreversible inhibition, and may be cytotoxic, making BEL not amenable for in vivo use. Recently, a fluoroketone (FK)-based compound (FKGK18) was described as a potent inhibitor of iPLA.sub.2 [beta]. Here we characterized its inhibitory profile in beta-cells and find that FKGK18: (a) inhibits iPLA.sub.2 [beta] with a greater potency (100-fold) than iPLA.sub.2 [gamma], (b) inhibition of iPLA.sub.2 [beta] is reversible, (c) is an ineffective inhibitor of [alpha]-chymotrypsin, and (d) inhibits previously described outcomes of iPLA.sub.2 [beta] activation including (i) glucose-stimulated insulin secretion, (ii) arachidonic acid hydrolysis; as reflected by PGE2 release from human islets, (iii) ER stress-induced neutral sphingomyelinase 2 expression, and (iv) ER stress-induced beta-cell apoptosis. These findings suggest that FKGK18 is similar to BEL in its ability to inhibit iPLA.sub.2 [beta]. Because, in contrast to BEL, it is reversible and not a non-specific inhibitor of proteases, it is suggested that FKGK18 is more ideal for ex vivo and in vivo assessments of iPLA.sub.2 [beta] role in biological functions.

The non-[beta] endocrine cells in pancreatic islets play an essential counterpart and regulatory role to the insulin-producing [beta]-cells in the regulation of blood-glucose homeostasis. While significant progress has been made towards the understanding of [beta]-cell regeneration in adults, very little is known about the regeneration of the non-[beta] endocrine cells such as glucagon-producing [alpha]-cells and somatostatin producing [delta]-cells. Previous studies have noted the increase of [alpha]-cell composition in diabetes patients and in animal models. It is thus our hypothesis that non-[beta]-cells such as [alpha]-cells and [delta]-cells in adults can regenerate, and that the regeneration accelerates in diabetic conditions. To test this hypothesis, we examined islet cell composition in a streptozotocin (STZ)-induced diabetes mouse model in detail. Our data showed the number of [alpha]-cells in each islet increased following STZ-mediated [beta]-cell destruction, peaked at Day 6, which was about 3 times that of normal islets. In addition, we found [delta]-cell numbers doubled by Day 6 following STZ treatment. These data suggest [alpha]- and [delta]-cell regeneration occurred rapidly following a single diabetes-inducing dose of STZ in mice. Using in vivo BrdU labeling techniques, we demonstrated [alpha]- and [delta]-cell regeneration involved cell proliferation. Co-staining of the islets with the proliferating cell marker Ki67 showed [alpha]- and [delta]-cells could replicate, suggesting self-duplication played a role in their regeneration. Furthermore, Pdx1.sup.+ /Insulin.sup.- cells were detected following STZ treatment, indicating the involvement of endocrine progenitor cells in the regeneration of these non-[beta] cells. This is further confirmed by the detection of Pdx1.sup.+ /glucagon.sup.+ cells and Pdx1.sup.+ /somatostatin.sup.+ cells following STZ treatment. Taken together, our study demonstrated adult [alpha]- and [delta]-cells could regenerate, and both self-duplication and regeneration from endocrine precursor cells were involved in their regeneration.