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Generation of pancreatic β cells from human pluripotent stem cells (hPSCs) holds promise as a cell replacement therapy for diabetes. In this study, we establish a link between the state of the actin cytoskeleton and the expression of pancreatic transcription factors that drive pancreatic lineage specification. Bulk and single-cell RNA sequencing demonstrated that different degrees of actin polymerization biased cells toward various endodermal lineages and that conditions favoring a polymerized cytoskeleton strongly inhibited neurogenin 3-induced endocrine differentiation. Using latrunculin A to depolymerize the cytoskeleton during endocrine induction, we developed a two-dimensional differentiation protocol for generating human pluripotent stem-cell-derived β (SC-β) cells with improved in vitro and in vivo function. SC-β cells differentiated from four hPSC lines exhibited first- and second-phase dynamic glucose-stimulated insulin secretion. Transplantation of islet-sized aggregates of these cells rapidly reversed severe preexisting diabetes in mice at a rate close to that of human islets and maintained normoglycemia for at least 9 months. Generation of pancreatic β cells from stem cells is enhanced by manipulating the cytoskeleton.

When encapsulated with alginate derivatives that resist the foreign-body response, human embryonic stem cell–derived beta cells restore long-term normoglycemia in immunocompetent mice without the need for immunosuppression. The transplantation of glucose-responsive, insulin-producing cells offers the potential for restoring glycemic control in individuals with diabetes 1 . Pancreas transplantation and the infusion of cadaveric islets are currently implemented clinically 2 , but these approaches are limited by the adverse effects of immunosuppressive therapy over the lifetime of the recipient and the limited supply of donor tissue 3 . The latter concern may be addressed by recently described glucose-responsive mature beta cells that are derived from human embryonic stem cells (referred to as SC-β cells), which may represent an unlimited source of human cells for pancreas replacement therapy 4 . Strategies to address the immunosuppression concerns include immunoisolation of insulin-producing cells with porous biomaterials that function as an immune barrier 5 , 6 . However, clinical implementation has been challenging because of host immune responses to the implant materials 7 . Here we report the first long-term glycemic correction of a diabetic, immunocompetent animal model using human SC-β cells. SC-β cells were encapsulated with alginate derivatives capable of mitigating foreign-body responses in vivo and implanted into the intraperitoneal space of C57BL/6J mice treated with streptozotocin, which is an animal model for chemically induced type 1 diabetes. These implants induced glycemic correction without any immunosuppression until their removal at 174 d after implantation. Human C-peptide concentrations and in vivo glucose responsiveness demonstrated therapeutically relevant glycemic control. Implants retrieved after 174 d contained viable insulin-producing cells.

This paper describes a new protocol for producing insulin‐producing cells in vitro that represents another potential cell source for a diabetes cell therapy. These cells can be loaded into a protective device that is implanted under the skin. The device is designed to protect the cells from immune rejection by the implant recipient. The implant can engraft and respond to glucose by secreting insulin, thus potentially replacing the β cells lost in patients with type 1 diabetes. The PEC‐01 cell population, differentiated from human embryonic stem cells (hESCs), contains pancreatic progenitors (PPs) that, when loaded into macroencapsulation devices (to produce the VC‐01 candidate product) and transplanted into mice, can mature into glucose‐responsive insulin‐secreting cells and other pancreatic endocrine cells involved in glucose metabolism. We modified the protocol for making PEC‐01 cells such that 73%–80% of the cell population consisted of PDX1‐positive (PDX1+) and NKX6.1+ PPs. The PPs were further differentiated to islet‐like cells (ICs) that reproducibly contained 73%–89% endocrine cells, of which approximately 40%–50% expressed insulin. A large fraction of these insulin‐positive cells were single hormone‐positive and expressed the transcription factors PDX1 and NKX6.1. To preclude a significant contribution of progenitors to the in vivo function of ICs, we used a simple enrichment process to remove remaining PPs, yielding aggregates that contained 93%–98% endocrine cells and 1%–3% progenitors. Enriched ICs, when encapsulated and implanted into mice, functioned similarly to the VC‐01 candidate product, demonstrating conclusively that in vitro‐produced hESC‐derived insulin‐producing cells can mature and function in vivo in devices. A scaled version of our suspension culture was used, and the endocrine aggregates could be cryopreserved and retain functionality. Although ICs expressed multiple important β cell genes, the cells contained relatively low levels of several maturity‐associated markers. Correlating with this, the time to function of ICs was similar to PEC‐01 cells, indicating that ICs required cell‐autonomous maturation after delivery in vivo, which would occur concurrently with graft integration into the host. Significance Type 1 diabetes (T1D) affects approximately 1.25 million people in the U.S. alone and is deadly if not managed with insulin injections. This paper describes the production of insulin‐producing cells in vitro and a new protocol for producing the cells, representing another potential cell source for a diabetes cell therapy. These cells can be loaded into a protective device that is implanted under the skin. The device is designed to protect the cells from immune rejection by the implant recipient. The implant can engraft and respond to glucose by secreting insulin, thus potentially replacing the β cells lost in patients with T1D.

The main goal of treatment for type 1 diabetes is to control glycaemia with insulin therapy to reduce disease complications. For some patients, technological approaches to insulin delivery are inadequate, and allogeneic islet transplantation is a safe alternative for those patients who have had severe hypoglycaemia complicated by impaired hypoglycaemia awareness or glycaemic lability, or who already receive immunosuppressive drugs for a kidney transplant. Since 2000, intrahepatic islet transplantation has proven efficacious in alleviating the burden of labile diabetes and preventing complications related to diabetes, whether or not a previous kidney transplant is present. Age, body-mass index, renal status, and cardiopulmonary status affect the choice between pancreas or islet transplantation. Access to transplantation is limited by the number of deceased donors and the necessity of immunosuppression. Future approaches might include alternative sources of islets (eg, xenogeneic tissue or human stem cells), extrahepatic sites of implantation (eg, omental, subcutaneous, or intramuscular), and induction of immune tolerance or encapsulation of islets.

The need to suppress a patient’s immune system after the transplantation of allogeneic cells is associated with wide-ranging side effects. We report the outcomes of transplantation of genetically modified allogeneic donor islet cells into a man with long-standing type 1 diabetes. We used clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated protein 12b (Cas12b) editing and lentiviral transduction to genetically edit the cells to avoid rejection; the cells were then transplanted into the participant’s forearm muscle. He did not receive any immunosuppressive drugs and, at 12 weeks after transplantation, showed no immune response against the gene-edited cells. C-peptide measurements showed stable and glucose-responsive insulin secretion. A total of four adverse events occurred, none of which were serious or related to the study drug. (Funded by the Leona M. and Harry B. Helmsley Charitable Trust; EudraCT number, 2023-507988-19-00 ; ClinicalTrials.gov number, NCT06239636 .) A patient with type 1 diabetes underwent transplantation of genetically modified allogeneic islet cells with no immunosuppression. The results of this study support the concept of immune evasion by these modified cells.

A seven-stage protocol enables differentiation of human embryonic stem cells into cells similar to pancreatic beta cells. Transplantation of pancreatic progenitors or insulin-secreting cells derived from human embryonic stem cells (hESCs) has been proposed as a therapy for diabetes. We describe a seven-stage protocol that efficiently converts hESCs into insulin-producing cells. Stage (S) 7 cells expressed key markers of mature pancreatic beta cells, including MAFA, and displayed glucose-stimulated insulin secretion similar to that of human islets during static incubations in vitro . Additional characterization using single-cell imaging and dynamic glucose stimulation assays revealed similarities but also notable differences between S7 insulin-secreting cells and primary human beta cells. Nevertheless, S7 cells rapidly reversed diabetes in mice within 40 days, roughly four times faster than pancreatic progenitors. Therefore, although S7 cells are not fully equivalent to mature beta cells, their capacity for glucose-responsive insulin secretion and rapid reversal of diabetes in vivo makes them a promising alternative to pancreatic progenitor cells or cadaveric islets for the treatment of diabetes.

Development of a cell therapy for diabetes would be greatly aided by a renewable supply of human β-cells. Here we show that pancreatic endoderm derived from human embryonic stem (hES) cells efficiently generates glucose-responsive endocrine cells after implantation into mice. Upon glucose stimulation of the implanted mice, human insulin and C-peptide are detected in sera at levels similar to those of mice transplanted with ∼3,000 human islets. Moreover, the insulin-expressing cells generated after engraftment exhibit many properties of functional β-cells, including expression of critical β-cell transcription factors, appropriate processing of proinsulin and the presence of mature endocrine secretory granules. Finally, in a test of therapeutic potential, we demonstrate that implantation of hES cell–derived pancreatic endoderm protects against streptozotocin-induced hyperglycemia. Together, these data provide definitive evidence that hES cells are competent to generate glucose-responsive, insulin-secreting cells.

Chronically deregulated blood-glucose concentrations in diabetes mellitus result from a loss of pancreatic insulin-producing β cells (type 1 diabetes, T1D) or from impaired insulin sensitivity of body cells and glucose-stimulated insulin release (type 2 diabetes, T2D). Here, we show that therapeutically applicable β-cell-mimetic designer cells can be established by minimal engineering of human cells. We achieved glucose responsiveness by a synthetic circuit that couples glycolysis-mediated calcium entry to an excitation-transcription system controlling therapeutic transgene expression. Implanted circuit-carrying cells corrected insulin deficiency and self-sufficiently abolished persistent hyperglycemia in T1D mice. Similarly, glucoseinducible glucagon-like peptide 1 transcription improved endogenous glucose-stimulated insulin release and glucose tolerance in T2D mice. These systems may enable a combination of diagnosis and treatment for diabetes mellitus therapy.

Loss and functional failure of pancreatic β-cells results in disruption of glucose homeostasis and progression of diabetes. Although whole pancreas or pancreatic islet transplantation serves as a promising approach for β-cell replenishment and diabetes therapy, the severe scarcity of donor islets makes it unattainable for most diabetic patients. Stem cells, particularly induced pluripotent stem cells (iPSCs), are promising for the treatment of diabetes owing to their self-renewal capacity and ability to differentiate into functional β-cells. In this review, we first introduce the development of functional β-cells and their heterogeneity and then turn to highlight recent advances in the generation of β-cells from stem cells and their potential applications in disease modeling, drug discovery and clinical therapy. Finally, we have discussed the current challenges in developing stem cell-based therapeutic strategies for improving the treatment of diabetes. Although some significant technical hurdles remain, stem cells offer great hope for patients with diabetes and will certainly transform future clinical practice.

Extracellular matrix (ECM) molecules are responsible for structural and biochemical support, as well as for regulation of molecular signalling and tissue repair in many organ structures, including the pancreas. In pancreatic islets, collagen type IV and VI, and laminins are the most abundant molecules, but other ECM molecules are also present. The ECM interacts with specific combinations of integrin α/β heterodimers on islet cells and guides many cellular processes. More specifically, some ECM molecules are involved in beta cell survival, function and insulin production, while others can fine tune the susceptibility of islet cells to cytokines. Further, some ECM induce release of growth factors to facilitate tissue repair. During enzymatic isolation of islets for transplantation, the ECM is damaged, impacting islet function. However, restoration of the ECM in human islets (for example by adding ECM to the interior of immunoprotective capsules) has been shown to enhance islet function. Here, we provide current insight into the role of ECM molecules in islet function and discuss the clinical potential of ECM manipulation to enhance pancreatic islet function and survival.