Publications by authors named "Matthias Hebrok"

111 Publications

Selective deletion of human leukocyte antigens protects stem cell-derived islets from immune rejection.

Cell Rep 2021 Aug;36(7):109538

Diabetes Center, Department of Medicine, University of California, San Francisco, San Francisco, CA 94143, USA. Electronic address:

Stem cell-based replacement therapies hold the promise to restore function of damaged or degenerated tissue such as the pancreatic islets in people with type 1 diabetes. Wide application of these therapies requires overcoming the fundamental roadblock of immune rejection. To address this issue, we use genetic engineering to create human pluripotent stem cells (hPSCs) in which the majority of the polymorphic human leukocyte antigens (HLAs), the main drivers of allogeneic rejection, are deleted. We retain the common HLA class I allele HLA-A2 and less polymorphic HLA-E/F/G to allow immune surveillance and inhibition of natural killer (NK) cells. We employ a combination of in vitro assays and humanized mouse models to demonstrate that these gene manipulations significantly reduce NK cell activity and T-cell-mediated alloimmune response against hPSC-derived islet cells. In summary, our approach produces hypoimmunogenic hPSCs that can be readily matched with recipients to avoid alloimmune rejection.
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http://dx.doi.org/10.1016/j.celrep.2021.109538DOI Listing
August 2021

Superporous agarose scaffolds for encapsulation of adult human islets and human stem-cell-derived β cells for intravascular bioartificial pancreas applications.

J Biomed Mater Res A 2021 Jun 30. Epub 2021 Jun 30.

Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, California, USA.

Type 1 diabetic patients with severe hypoglycemia unawareness have benefitted from cellular therapies, such as pancreas or islet transplantation; however, donor shortage and the need for immunosuppression limits widespread clinical application. We previously developed an intravascular bioartificial pancreas (iBAP) using silicon nanopore membranes (SNM) for immunoprotection. To ensure ample nutrient delivery, the iBAP will need a cell scaffold with high hydraulic permeability to provide mechanical support and maintain islet viability and function. Here, we examine the feasibility of superporous agarose (SPA) as a potential cell scaffold in the iBAP. SPA exhibits 66-fold greater hydraulic permeability than the SNM along with a short (<10 μm) diffusion distance to the nearest islet. SPA also supports short-term functionality of both encapsulated human islets and stem-cell-derived enriched β-clusters in a convection-based system, demonstrated by high viability (>95%) and biphasic insulin responses to dynamic glucose stimulus. These findings suggest that the SPA scaffold will not limit nutrient delivery in a convection-based bioartificial pancreas and merits continued investigation.
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http://dx.doi.org/10.1002/jbm.a.37236DOI Listing
June 2021

Single-cell transcriptome analysis defines heterogeneity of the murine pancreatic ductal tree.

Elife 2021 May 19;10. Epub 2021 May 19.

Diabetes Center, University of California, San Francisco, San Francisco, United States.

To study disease development, an inventory of an organ's cell types and understanding of physiologic function is paramount. Here, we performed single-cell RNA-sequencing to examine heterogeneity of murine pancreatic duct cells, pancreatobiliary cells, and intrapancreatic bile duct cells. We describe an epithelial-mesenchymal transitory axis in our three pancreatic duct subpopulations and identify osteopontin as a regulator of this fate decision as well as human duct cell dedifferentiation. Our results further identify functional heterogeneity within pancreatic duct subpopulations by elucidating a role for geminin in accumulation of DNA damage in the setting of chronic pancreatitis. Our findings implicate diverse functional roles for subpopulations of pancreatic duct cells in maintenance of duct cell identity and disease progression and establish a comprehensive road map of murine pancreatic duct cell, pancreatobiliary cell, and intrapancreatic bile duct cell homeostasis.
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http://dx.doi.org/10.7554/eLife.67776DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8184217PMC
May 2021

Stem Cell-Based Clinical Trials for Diabetes Mellitus.

Front Endocrinol (Lausanne) 2021 26;12:631463. Epub 2021 Feb 26.

Diabetes Center, University of California San Francisco, San Francisco, CA, United States.

Since its introduction more than twenty years ago, intraportal allogeneic cadaveric islet transplantation has been shown to be a promising therapy for patients with Type I Diabetes (T1D). Despite its positive outcome, the impact of islet transplantation has been limited due to a number of confounding issues, including the limited availability of cadaveric islets, the typically lifelong dependence of immunosuppressive drugs, and the lack of coverage of transplant costs by health insurance companies in some countries. Despite improvements in the immunosuppressive regimen, the number of required islets remains high, with two or more donors per patient often needed. Insulin independence is typically achieved upon islet transplantation, but on average just 25% of patients do not require exogenous insulin injections five years after. For these reasons, implementation of islet transplantation has been restricted almost exclusively to patients with brittle T1D who cannot avoid hypoglycemic events despite optimized insulin therapy. To improve C-peptide levels in patients with both T1 and T2 Diabetes, numerous clinical trials have explored the efficacy of mesenchymal stem cells (MSCs), both as supporting cells to protect existing β cells, and as source for newly generated β cells. Transplantation of MSCs is found to be effective for T2D patients, but its efficacy in T1D is controversial, as the ability of MSCs to differentiate into functional β cells is poor, and transdifferentiation does not seem to occur. Instead, to address limitations related to supply, human embryonic stem cell (hESC)-derived β cells are being explored as surrogates for cadaveric islets. Transplantation of allogeneic hESC-derived insulin-producing organoids has recently entered Phase I and Phase II clinical trials. Stem cell replacement therapies overcome the barrier of finite availability, but they still face immune rejection. Immune protective strategies, including coupling hESC-derived insulin-producing organoids with macroencapsulation devices and microencapsulation technologies, are being tested to balance the necessity of immune protection with the need for vascularization. Here, we compare the diverse human stem cell approaches and outcomes of recently completed and ongoing clinical trials, and discuss innovative strategies developed to overcome the most significant challenges remaining for transplanting stem cell-derived β cells.
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http://dx.doi.org/10.3389/fendo.2021.631463DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7953062PMC
February 2021

Non-xenogeneic expansion and definitive endoderm differentiation of human pluripotent stem cells in an automated bioreactor.

Biotechnol Bioeng 2021 02 16;118(2):979-991. Epub 2020 Dec 16.

Department of Chemical and Biological Engineering, Tufts University, Medford, Massachusetts, USA.

Scalable processes are requisite for the robust biomanufacturing of human pluripotent stem cell (hPSC)-derived therapeutics. Toward this end, we demonstrate the xeno-free expansion and directed differentiation of human embryonic and induced pluripotent stem cells to definitive endoderm (DE) in a controlled stirred suspension bioreactor (SSB). Based on previous work on converting hPSCs to insulin-producing progeny, differentiation of two hPSC lines was optimized in planar cultures yielding up to 87% FOXA2 /SOX17 cells. Next, hPSCs were propagated in an SSB with controlled pH and dissolved oxygen. Cultures displayed a 10- to 12-fold increase in cell number over 5-6 days with the maintenance of pluripotency (>85% OCT4 ) and viability (>85%). For differentiation, SSB cultures yielded up to 89% FOXA2 /SOX17 cells or ~ 8 DE cells per seeded hPSC. Specification to DE cell fate was consistently more efficient in the bioreactor compared to planar cultures. Hence, a tunable strategy is established that is suitable for the xeno-free manufacturing of DE cells from different hPSC lines in scalable SSBs. This study advances bioprocess development for producing a wide gamut of human DE cell-derived therapeutics.
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http://dx.doi.org/10.1002/bit.27629DOI Listing
February 2021

Emerging routes to the generation of functional β-cells for diabetes mellitus cell therapy.

Nat Rev Endocrinol 2020 09 25;16(9):506-518. Epub 2020 Jun 25.

Diabetes Center, University of California San Francisco, San Francisco, CA, USA.

Diabetes mellitus, which affects more than 463 million people globally, is caused by the autoimmune ablation or functional loss of insulin-producing β-cells, and prevalence is projected to continue rising over the next decades. Generating β-cells to mitigate the aberrant glucose homeostasis manifested in the disease has remained elusive. Substantial advances have been made in producing mature β-cells from human pluripotent stem cells that respond appropriately to dynamic changes in glucose concentrations in vitro and rapidly function in vivo following transplantation in mice. Other potential avenues to produce functional β-cells include: transdifferentiation of closely related cell types (for example, other pancreatic islet cells such as α-cells, or other cells derived from endoderm); the engineering of non-β-cells that are capable of modulating blood sugar; and the construction of synthetic 'cells' or particles mimicking functional aspects of β-cells. This Review focuses on the current status of generating β-cells via these diverse routes, highlighting the unique advantages and challenges of each approach. Given the remarkable progress in this field, scalable bioengineering processes are also discussed for the realization of the therapeutic potential of derived β-cells.
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http://dx.doi.org/10.1038/s41574-020-0375-3DOI Listing
September 2020

Loss of the transcription factor MAFB limits β-cell derivation from human PSCs.

Nat Commun 2020 06 2;11(1):2742. Epub 2020 Jun 2.

UCSF Diabetes Center, University of California San Francisco, San Francisco, CA, 94143, USA.

Next generation sequencing studies have highlighted discrepancies in β-cells which exist between mice and men. Numerous reports have identified MAF BZIP Transcription Factor B (MAFB) to be present in human β-cells postnatally, while its expression is restricted to embryonic and neo-natal β-cells in mice. Using CRISPR/Cas9-mediated gene editing, coupled with endocrine cell differentiation strategies, we dissect the contribution of MAFB to β-cell development and function specifically in humans. Here we report that MAFB knockout hPSCs have normal pancreatic differentiation capacity up to the progenitor stage, but favor somatostatin- and pancreatic polypeptide-positive cells at the expense of insulin- and glucagon-producing cells during endocrine cell development. Our results describe a requirement for MAFB late in the human pancreatic developmental program and identify it as a distinguishing transcription factor within islet cell subtype specification. We propose that hPSCs represent a powerful tool to model human pancreatic endocrine development and associated disease pathophysiology.
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http://dx.doi.org/10.1038/s41467-020-16550-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7265500PMC
June 2020

LIN28B Impairs the Transition of hESC-Derived β Cells from the Juvenile to Adult State.

Stem Cell Reports 2020 01 26;14(1):9-20. Epub 2019 Dec 26.

The Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, CA 94143, USA; Department of Urology, University of California, San Francisco, CA 94143, USA. Electronic address:

Differentiation of human embryonic stem cells into pancreatic β cells holds great promise for the treatment of diabetes. Recent advances have led to the production of glucose-responsive insulin-secreting cells in vitro, but resulting cells remain less mature than their adult primary β cell counterparts. The barrier(s) to in vitro β cell maturation are unclear. Here, we evaluated a potential role for microRNAs. MicroRNA profiling showed high expression of let-7 family microRNAs in vivo, but not in in vitro differentiated β cells. Reduced levels of let-7 in vitro were associated with increased levels of the RNA binding protein LIN28B, a negative regulator of let-7 biogenesis. Ablation of LIN28B during human embryonic stem cell (hESC) differentiation toward β cells led to a more mature glucose-stimulated insulin secretion profile and the suppression of juvenile-specific genes. However, let-7 overexpression had little effect. These results uncover LIN28B as a modulator of β cell maturation in vitro.
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http://dx.doi.org/10.1016/j.stemcr.2019.11.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6962644PMC
January 2020

Lipid Droplet Accumulation in Human Pancreatic Islets Is Dependent On Both Donor Age and Health.

Diabetes 2020 03 13;69(3):342-354. Epub 2019 Dec 13.

Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN

Human but not mouse islets transplanted into immunodeficient NSG mice effectively accumulate lipid droplets (LDs). Because chronic lipid exposure is associated with islet β-cell dysfunction, we investigated LD accumulation in the intact human and mouse pancreas over a range of ages and states of diabetes. Very few LDs were found in normal human juvenile pancreatic acinar and islet cells, with numbers subsequently increasing throughout adulthood. While accumulation appeared evenly distributed in postjuvenile acinar and islet cells in donors without diabetes, LDs were enriched in islet α- and β-cells from donors with type 2 diabetes (T2D). LDs were also found in the islet β-like cells produced from human embryonic cell-derived β-cell clusters. In contrast, LD accumulation was nearly undetectable in the adult rodent pancreas, even in hyperglycemic and hyperlipidemic models or 1.5-year-old mice. Taken together, there appear to be significant differences in pancreas islet cell lipid handling between species, and the human juvenile and adult cell populations. Moreover, our results suggest that LD enrichment could be impactful to T2D islet cell function.
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http://dx.doi.org/10.2337/db19-0281DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7034188PMC
March 2020

Using a barcoded AAV capsid library to select for clinically relevant gene therapy vectors.

JCI Insight 2019 11 14;4(22). Epub 2019 Nov 14.

Departments of Pediatrics and Genetics, Stanford University, Stanford, California, USA.

While gene transfer using recombinant adeno-associated viral (rAAV) vectors has shown success in some clinical trials, there remain many tissues that are not well transduced. Because of the recent success in reprogramming islet-derived cells into functional β cells in animal models, we constructed 2 highly complex barcoded replication competent capsid shuffled libraries and selected for high-transducing variants on primary human islets. We describe the generation of a chimeric AAV capsid (AAV-KP1) that facilitates transduction of primary human islet cells and human embryonic stem cell-derived β cells with up to 10-fold higher efficiency compared with previously studied best-in-class AAV vectors. Remarkably, this chimeric capsid also enabled transduction of both mouse and human hepatocytes at very high levels in a humanized chimeric mouse model, thus providing a versatile vector that has the potential to be used in both preclinical testing and human clinical trials for liver-based diseases and diabetes.
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http://dx.doi.org/10.1172/jci.insight.131610DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6948855PMC
November 2019

Supporting Survival of Transplanted Stem-Cell-Derived Insulin-Producing Cells in an Encapsulation Device Augmented with Controlled Release of Amino Acids.

Adv Biosyst 2019 Sep 9;3(9). Epub 2019 Aug 9.

Department of Bioengineering and Therapeutic Sciences, University of California, 1700 4th Street, Byers Hall, Box 2520, San Francisco, CA 94158, USA.

Pancreatic islet transplantation is a promising treatment for type I diabetes, which is a chronic autoimmune disease in which the host immune cells attack insulin-producing beta cells. The impact of this therapy is limited due to tissue availability and dependence on immunosuppressive drugs that prevent immune rejection of the transplanted cells. These issues can be solved by encapsulating stem cell-derived insulin-producing cells in an immunoprotective device. However, encapsulation exacerbates ischemia, and the lack of vasculature at the implantation site post-transplantation worsens graft survival. Here, an encapsulation device that supplements nutrients to the cells is developed to improve the survival of encapsulated stem cell-derived insulin-producing cells in the poorly vascularized subcutaneous space. An internal compartment in the device is fabricated to provide zero-order release of alanine and glutamine for several weeks. The amino acid reservoir sustains viability of insulin-producing cells in nutrient limiting conditions in vitro. Moreover, the reservoir also increases cell survival by 30% after transplanting the graft in the subcutaneous space.
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http://dx.doi.org/10.1002/adbi.201900086DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6800235PMC
September 2019

mTORC1 to AMPK switching underlies β-cell metabolic plasticity during maturation and diabetes.

J Clin Invest 2019 07 2;129(10):4124-4137. Epub 2019 Jul 2.

The Diabetes Center, UCSF, San Francisco, California, USA.

Pancreatic beta cells (β-cells) differentiate during fetal life, but only postnatally acquire the capacity for glucose-stimulated insulin secretion (GSIS). How this happens is not clear. In exploring what molecular mechanisms drive the maturation of β-cell function, we found that the control of cellular signaling in β-cells fundamentally switched from the nutrient sensor target of rapamycin (mTORC1) to the energy sensor 5'-adenosine monophosphate-activated protein kinase (AMPK), and that this was critical for functional maturation. Moreover, AMPK was activated by the dietary transition taking place during weaning, and this in turn inhibited mTORC1 activity to drive the adult β-cell phenotype. While forcing constitutive mTORC1 signaling in adult β-cells relegated them to a functionally immature phenotype with characteristic transcriptional and metabolic profiles, engineering the switch from mTORC1 to AMPK signaling was sufficient to promote β-cell mitochondrial biogenesis, a shift to oxidative metabolism, and functional maturation. We also found that type 2 diabetes, a condition marked by both mitochondrial degeneration and dysregulated GSIS, was associated with a remarkable reversion of the normal AMPK-dependent adult β-cell signature to a more neonatal one characterized by mTORC1 activation. Manipulating the way in which cellular nutrient signaling pathways regulate β-cell metabolism may thus offer new targets to improve β-cell function in diabetes.
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http://dx.doi.org/10.1172/JCI127021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6763225PMC
July 2019

The Pdx1-Bound Swi/Snf Chromatin Remodeling Complex Regulates Pancreatic Progenitor Cell Proliferation and Mature Islet β-Cell Function.

Diabetes 2019 09 14;68(9):1806-1818. Epub 2019 Jun 14.

Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN

Transcription factors positively and/or negatively impact gene expression by recruiting coregulatory factors, which interact through protein-protein binding. Here we demonstrate that mouse pancreas size and islet β-cell function are controlled by the ATP-dependent Swi/Snf chromatin remodeling coregulatory complex that physically associates with Pdx1, a diabetes-linked transcription factor essential to pancreatic morphogenesis and adult islet cell function and maintenance. Early embryonic deletion of just the Swi/Snf Brg1 ATPase subunit reduced multipotent pancreatic progenitor cell proliferation and resulted in pancreas hypoplasia. In contrast, removal of both Swi/Snf ATPase subunits, Brg1 and Brm, was necessary to compromise adult islet β-cell activity, which included whole-animal glucose intolerance, hyperglycemia, and impaired insulin secretion. Notably, lineage-tracing analysis revealed Swi/Snf-deficient β-cells lost the ability to produce the mRNAs for and other key metabolic genes without effecting the expression of many essential islet-enriched transcription factors. Swi/Snf was necessary for Pdx1 to bind to the gene enhancer, demonstrating the importance of this association in mediating chromatin accessibility. These results illustrate how fundamental the Pdx1:Swi/Snf coregulator complex is in the pancreas, and we discuss how disrupting their association could influence type 1 and type 2 diabetes susceptibility.
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http://dx.doi.org/10.2337/db19-0349DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6702633PMC
September 2019

Author Correction: Recapitulating endocrine cell clustering in culture promotes maturation of human stem-cell-derived β cells.

Nat Cell Biol 2019 06;21(6):792

Diabetes Center, University of California San Francisco, San Francisco, CA, USA.

In the version of this article originally published, the Gene Expression Omnibus (GEO) accession number listed in the data availability section was incorrectly given as GSE10979 instead of GSE109795. The sentence should read "RNA-seq data that support the findings of this study have been deposited in the Gene Expression Omnibus (GEO) under accession code GSE109795," and the code should link to https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE109795. The error has been corrected in the HTML and PDF versions of the paper.
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http://dx.doi.org/10.1038/s41556-019-0316-3DOI Listing
June 2019

Pancreatic pericytes originate from the embryonic pancreatic mesenchyme.

Dev Biol 2019 05 13;449(1):14-20. Epub 2019 Feb 13.

Department of Cell and Developmental Biology, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv 69978, Israel. Electronic address:

The embryonic origin of pericytes is heterogeneous, both between and within organs. While pericytes of coelomic organs were proposed to differentiate from the mesothelium, a single-layer squamous epithelium, the embryonic origin of pancreatic pericytes has yet to be reported. Here, we show that adult pancreatic pericytes originate from the embryonic pancreatic mesenchyme. Our analysis indicates that pericytes of the adult mouse pancreas originate from cells expressing the transcription factor Nkx3.2. In the embryonic pancreas, Nkx3.2-expressing cells constitute the multilayered mesenchyme, which surrounds the pancreatic epithelium and supports multiple events in its development. Thus, we traced the fate of the pancreatic mesenchyme. Our analysis reveals that pancreatic mesenchymal cells acquire various pericyte characteristics, including gene expression, typical morphology, and periendothelial location, during embryogenesis. Importantly, we show that the vast majority of pancreatic mesenchymal cells differentiate into pericytes already at embryonic day 13.5 and progressively acquires a more mature pericyte phenotype during later stages of pancreas organogenesis. Thus, our study indicates the embryonic pancreatic mesenchyme as the primary origin to adult pancreatic pericytes. As pericytes of other coelomic organs were suggested to differentiate from the mesothelium, our findings point to a distinct origin of these cells in the pancreas. Thus, our study proposes a complex ontogeny of pericytes of coelomic organs.
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http://dx.doi.org/10.1016/j.ydbio.2019.01.020DOI Listing
May 2019

Recapitulating endocrine cell clustering in culture promotes maturation of human stem-cell-derived β cells.

Nat Cell Biol 2019 02 1;21(2):263-274. Epub 2019 Feb 1.

Diabetes Center, University of California San Francisco, San Francisco, CA, USA.

Despite advances in the differentiation of insulin-producing cells from human embryonic stem cells, the generation of mature functional β cells in vitro has remained elusive. To accomplish this goal, we have developed cell culture conditions to closely mimic events occurring during pancreatic islet organogenesis and β cell maturation. In particular, we have focused on recapitulating endocrine cell clustering by isolating and reaggregating immature β-like cells to form islet-sized enriched β-clusters (eBCs). eBCs display physiological properties analogous to primary human β cells, including robust dynamic insulin secretion, increased calcium signalling in response to secretagogues, and improved mitochondrial energization. Notably, endocrine cell clustering induces metabolic maturation by driving mitochondrial oxidative respiration, a process central to stimulus-secretion coupling in mature β cells. eBCs display glucose-stimulated insulin secretion as early as three days after transplantation in mice. In summary, replicating aspects of endocrine cell clustering permits the generation of stem-cell-derived β cells that resemble their endogenous counterparts.
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http://dx.doi.org/10.1038/s41556-018-0271-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6746427PMC
February 2019

The BRG1/SOX9 axis is critical for acinar cell-derived pancreatic tumorigenesis.

J Clin Invest 2018 08 16;128(8):3475-3489. Epub 2018 Jul 16.

Department of Gastroenterology and Hepatology, Kyoto University Graduate School of Medicine, Kyoto, Japan.

Chromatin remodeler Brahma related gene 1 (BRG1) is silenced in approximately 10% of human pancreatic ductal adenocarcinomas (PDAs). We previously showed that BRG1 inhibits the formation of intraductal pancreatic mucinous neoplasm (IPMN) and that IPMN-derived PDA originated from ductal cells. However, the role of BRG1 in pancreatic intraepithelial neoplasia-derived (PanIN-derived) PDA that originated from acinar cells remains elusive. Here, we found that exclusive elimination of Brg1 in acinar cells of Ptf1a-CreER; KrasG12D; Brg1fl/fl mice impaired the formation of acinar-to-ductal metaplasia (ADM) and PanIN independently of p53 mutation, while PDA formation was inhibited in the presence of p53 mutation. BRG1 bound to regions of the Sox9 promoter to regulate its expression and was critical for recruitment of upstream regulators, including PDX1, to the Sox9 promoter and enhancer in acinar cells. SOX9 expression was downregulated in BRG1-depleted ADMs/PanINs. Notably, Sox9 overexpression canceled this PanIN-attenuated phenotype in KBC mice. Furthermore, Brg1 deletion in established PanIN by using a dual recombinase system resulted in regression of the lesions in mice. Finally, BRG1 expression correlated with SOX9 expression in human PDAs. In summary, BRG1 is critical for PanIN initiation and progression through positive regulation of SOX9. Thus, the BRG1/SOX9 axis is a potential target for PanIN-derived PDA.
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http://dx.doi.org/10.1172/JCI94287DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6063489PMC
August 2018

Stem Cell Therapies for Treating Diabetes: Progress and Remaining Challenges.

Cell Stem Cell 2018 Jun;22(6):810-823

Diabetes Center, University of California, San Francisco, San Francisco, CA 94143, USA; Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, CA 94143, USA. Electronic address:

Restoration of insulin independence and normoglycemia has been the overarching goal in diabetes research and therapy. While whole-organ and islet transplantation have become gold-standard procedures in achieving glucose control in diabetic patients, the profound lack of suitable donor tissues severely hampers the broad application of these therapies. Here, we describe current efforts aimed at generating a sustainable source of functional human stem cell-derived insulin-producing islet cells for cell transplantation and present state-of-the-art efforts to protect such cells via immune modulation and encapsulation strategies.
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http://dx.doi.org/10.1016/j.stem.2018.05.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6007036PMC
June 2018

Synaptotagmin 4 Regulates Pancreatic β Cell Maturation by Modulating the Ca Sensitivity of Insulin Secretion Vesicles.

Dev Cell 2018 05 12;45(3):347-361.e5. Epub 2018 Apr 12.

Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Department of Veterans Affairs, Tennessee Valley Health Authority, Nashville, TN 37232, USA; Center for Stem Cell Biology, Vanderbilt University School of Medicine, Department of Veterans Affairs, Tennessee Valley Health Authority, Nashville, TN 37232, USA; The Program of Developmental Biology, Vanderbilt University School of Medicine, Department of Veterans Affairs, Tennessee Valley Health Authority, Nashville, TN 37232, USA. Electronic address:

Islet β cells from newborn mammals exhibit high basal insulin secretion and poor glucose-stimulated insulin secretion (GSIS). Here we show that β cells of newborns secrete more insulin than adults in response to similar intracellular Ca concentrations, suggesting differences in the Ca sensitivity of insulin secretion. Synaptotagmin 4 (Syt4), a non-Ca binding paralog of the β cell Ca sensor Syt7, increased by ∼8-fold during β cell maturation. Syt4 ablation increased basal insulin secretion and compromised GSIS. Precocious Syt4 expression repressed basal insulin secretion but also impaired islet morphogenesis and GSIS. Syt4 was localized on insulin granules and Syt4 levels inversely related to the number of readily releasable vesicles. Thus, transcriptional regulation of Syt4 affects insulin secretion; Syt4 expression is regulated in part by Myt transcription factors, which repress Syt4 transcription. Finally, human SYT4 regulated GSIS in EndoC-βH1 cells, a human β cell line. These findings reveal the role that altered Ca sensing plays in regulating β cell maturation.
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http://dx.doi.org/10.1016/j.devcel.2018.03.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5962294PMC
May 2018

Replication confers β cell immaturity.

Nat Commun 2018 02 2;9(1):485. Epub 2018 Feb 2.

Diabetes Center, Department of Medicine, University of California, San Francisco, CA, USA.

Pancreatic β cells are highly specialized to regulate systemic glucose levels by secreting insulin. In adults, increase in β-cell mass is limited due to brakes on cell replication. In contrast, proliferation is robust in neonatal β cells that are functionally immature as defined by a lower set point for glucose-stimulated insulin secretion. Here we show that β-cell proliferation and immaturity are linked by tuning expression of physiologically relevant, non-oncogenic levels of c-Myc. Adult β cells induced to replicate adopt gene expression and metabolic profiles resembling those of immature neonatal β that proliferate readily. We directly demonstrate that priming insulin-producing cells to enter the cell cycle promotes a functionally immature phenotype. We suggest that there exists a balance between mature functionality and the ability to expand, as the phenotypic state of the β cell reverts to a less functional one in response to proliferative cues.
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http://dx.doi.org/10.1038/s41467-018-02939-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5797102PMC
February 2018

Mitigating Ischemic Injury of Stem Cell-Derived Insulin-Producing Cells after Transplant.

Stem Cell Reports 2017 09 10;9(3):807-819. Epub 2017 Aug 10.

Department of Surgery, University of California San Francisco, San Francisco, CA 94143, USA; UCSF Diabetes Center, University of California San Francisco, San Francisco, CA 94143, USA. Electronic address:

The advent of large-scale in vitro differentiation of human stem cell-derived insulin-producing cells (SCIPC) has brought us closer to treating diabetes using stem cell technology. However, decades of experiences from islet transplantation show that ischemia-induced islet cell death after transplant severely limits the efficacy of the therapy. It is unclear to what extent human SCIPC are susceptible to ischemia. In this study, we show that more than half of SCIPC die shortly after transplantation. Nutrient deprivation and hypoxia acted synergistically to kill SCIPC in vitro. Amino acid supplementation rescued SCIPC from nutrient deprivation, likely by providing cellular energy. Generating SCIPC under physiological oxygen tension of 5% conferred hypoxia resistance without affecting their differentiation or function. A two-pronged strategy of physiological oxygen acclimatization during differentiation and amino acid supplementation during transplantation significantly improved SCIPC survival after transplant.
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http://dx.doi.org/10.1016/j.stemcr.2017.07.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5599226PMC
September 2017

Nanoporous Immunoprotective Device for Stem-Cell-Derived β-Cell Replacement Therapy.

ACS Nano 2017 08 7;11(8):7747-7757. Epub 2017 Aug 7.

UCSF-UC Berkeley Joint PhD Program in Bioengineering , San Francisco, California 94143, United States.

Encapsulation of human embryonic stem-cell-differentiated beta cell clusters (hES-βC) holds great promise for cell replacement therapy for the treatment of diabetics without the need for chronic systemic immune suppression. Here, we demonstrate a nanoporous immunoprotective polymer thin film cell encapsulation device that can exclude immune molecules while allowing exchange of oxygen and nutrients necessary for in vitro and in vivo stem cell viability and function. Biocompatibility studies show the device promotes neovascular formation with limited foreign body response in vivo. The device also successfully prevented teratoma escape into the peritoneal cavity of mice. Long-term animal studies demonstrate evidence of engraftment, viability, and function of cells encapsulated in the device after 6 months. Finally, in vivo study confirms that the device was able to effectively immuno-isolate cells from the host immune system.
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http://dx.doi.org/10.1021/acsnano.7b01239DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5667644PMC
August 2017

Atypical flat lesions derive from pancreatic acinar cells.

Pancreatology 2017 May - Jun;17(3):350-353. Epub 2017 Apr 25.

Institute of Pathology, University Clinic Duesseldorf, Heinrich-Heine University, Duesseldorf, Germany.

Objectives: Pancreatic ductal adenocarcinoma (PDAC) is thought to derive from different precursor lesions including the recently identified atypical flat lesions (AFL). While all precursor lesions and PDAC share ductal characteristics, there is an ongoing debate about the cellular origin of the different PDAC precursor lesions. In particular, pancreatic acinar cells have previously been shown to display a remarkable plasticity being able to undergo ductal dedifferentiation in the context of oncogenic stimuli.

Methods: Histological analyses were performed in a murine PDAC model that specifically expresses oncogenic Kras in adult pancreatic acinar cells. Occurrence, characterization, and lineage tracing of AFLs were investigated.

Results: Upon expression of oncogenic Kras in adult pancreatic acinar cells, AFLs with typical morphology and expression profile arise. Lineage tracing confirmed that the AFLs were of acinar origin.

Conclusions: Using a murine PDAC model, this study identifies pancreatic acinar cells as a cellular source for AFLs.
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http://dx.doi.org/10.1016/j.pan.2017.04.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5770228PMC
March 2018

Global Protease Activity Profiling Provides Differential Diagnosis of Pancreatic Cysts.

Clin Cancer Res 2017 Aug 19;23(16):4865-4874. Epub 2017 Apr 19.

Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, California.

Pancreatic cysts are estimated to be present in 2%-3% of the adult population. Unfortunately, current diagnostics do not accurately distinguish benign cysts from those that can progress into invasive cancer. Misregulated pericellular proteolysis is a hallmark of malignancy, and therefore, we used a global approach to discover protease activities that differentiate benign nonmucinous cysts from premalignant mucinous cysts. We employed an unbiased and global protease profiling approach to discover protease activities in 23 cyst fluid samples. The distinguishing activities of select proteases was confirmed in 110 samples using specific fluorogenic substrates and required less than 5 μL of cyst fluid. We determined that the activities of the aspartyl proteases gastricsin and cathepsin E are highly increased in fluid from mucinous cysts. IHC analysis revealed that gastricsin expression was associated with regions of low-grade dysplasia, whereas cathepsin E expression was independent of dysplasia grade. Gastricsin activity differentiated mucinous from nonmucinous cysts with a specificity of 100% and a sensitivity of 93%, whereas cathepsin E activity was 92% specific and 70% sensitive. Gastricsin significantly outperformed the most widely used molecular biomarker, carcinoembryonic antigen (CEA), which demonstrated 94% specificity and 65% sensitivity. Combined analysis of gastricsin and CEA resulted in a near perfect classifier with 100% specificity and 98% sensitivity. Quantitation of gastricsin and cathepsin E activities accurately distinguished mucinous from nonmucinous pancreatic cysts and has the potential to replace current diagnostics for analysis of these highly prevalent lesions. .
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http://dx.doi.org/10.1158/1078-0432.CCR-16-2987DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5712228PMC
August 2017

All mixed up: defining roles for β-cell subtypes in mature islets.

Genes Dev 2017 02;31(3):228-240

Diabetes Center, Department of Medicine, University of California at San Francisco, San Francisco, California 94143, USA.

Following differentiation during fetal development, β cells further adapt to their postnatal role through functional maturation. While adult islets are thought to contain functionally mature β cells, recent analyses of transgenic rodent and human pancreata reveal a number of novel heterogeneity markers in mammalian β cells. The marked heterogeneity long after maturation raises the prospect that diverse populations harbor distinct roles aside from glucose-stimulated insulin secretion. In this review, we outline our current understanding of the β-cell maturation process, emphasize recent literature on novel heterogeneity markers, and offer perspectives on reconciling the findings from these two areas.
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http://dx.doi.org/10.1101/gad.294389.116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5358720PMC
February 2017

Designing β Cells.

Authors:
Matthias Hebrok

Cell Metab 2017 02;25(2):223-224

Diabetes Center, Department of Medicine, University of California, San Francisco, San Francisco, CA 94143, USA. Electronic address:

Diabetes is a growing epidemic, and many patients depend on insulin injections to control the disease and minimize long-term complications. In a recent manuscript published in Science, Xie et al. (2016) generate insulin-producing cells from a somatic embryonic kidney cell line through minimal genetic modification capable of regulating glucose levels in diabetic mice.
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http://dx.doi.org/10.1016/j.cmet.2017.01.018DOI Listing
February 2017

PDX1 dynamically regulates pancreatic ductal adenocarcinoma initiation and maintenance.

Genes Dev 2016 12;30(24):2669-2683

Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan 48109, USA.

Aberrant activation of embryonic signaling pathways is frequent in pancreatic ductal adenocarcinoma (PDA), making developmental regulators therapeutically attractive. Here we demonstrate diverse functions for pancreatic and duodenal homeobox 1 (PDX1), a transcription factor indispensable for pancreas development, in the progression from normal exocrine cells to metastatic PDA. We identify a critical role for PDX1 in maintaining acinar cell identity, thus resisting the formation of pancreatic intraepithelial neoplasia (PanIN)-derived PDA. Upon neoplastic transformation, the role of PDX1 changes from tumor-suppressive to oncogenic. Interestingly, subsets of malignant cells lose PDX1 expression while undergoing epithelial-to-mesenchymal transition (EMT), and PDX1 loss is associated with poor outcome. This stage-specific functionality arises from profound shifts in PDX1 chromatin occupancy from acinar cells to PDA. In summary, we report distinct roles of PDX1 at different stages of PDA, suggesting that therapeutic approaches against this potential target need to account for its changing functions at different stages of carcinogenesis. These findings provide insight into the complexity of PDA pathogenesis and advocate a rigorous investigation of therapeutically tractable targets at distinct phases of PDA development and progression.
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http://dx.doi.org/10.1101/gad.291021.116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5238727PMC
December 2016

Rebranding asymptomatic type 1 diabetes: the case for autoimmune beta cell disorder as a pathological and diagnostic entity.

Diabetologia 2017 01 26;60(1):35-38. Epub 2016 Oct 26.

Department of Pathology, Immunology, and Laboratory Medicine, College of Medicine, University of Florida, Gainesville, FL, USA.

The asymptomatic phase of type 1 diabetes is recognised by the presence of beta cell autoantibodies in the absence of hyperglycaemia. We propose that an accurate description of this stage is provided by the name 'Autoimmune Beta Cell Disorder' (ABCD). Specifically, we suggest that this nomenclature and diagnosis will, in a proactive manner, shift the paradigm towards type 1 diabetes being first and foremost an immune-mediated disease and only later a metabolic disease, presaging more active therapeutic intervention in the asymptomatic stage of disease, before end-stage beta cell failure. Furthermore, we argue that accepting ABCD as a diagnosis will be critical in order to accelerate pharmaceutical, academic and public activities leading to clinical trials that could reverse beta cell autoimmunity and halt progression to symptomatic insulin-requiring type 1 diabetes. We recognize that there are both opportunities and challenges in the implementation of the ABCD concept but hope that the notion of 'asymptomatic autoimmune disease' as a disorder will be widely discussed and eventually accepted.
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http://dx.doi.org/10.1007/s00125-016-4144-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5516264PMC
January 2017

Loss of Pancreas upon Activated Wnt Signaling Is Concomitant with Emergence of Gastrointestinal Identity.

PLoS One 2016 13;11(10):e0164714. Epub 2016 Oct 13.

Unidad de Gestión Clínica de Endocrinología y Nutrición, Hospital Universitario Virgen del Rocío, Sevilla, Spain.

Organ formation is achieved through the complex interplay between signaling pathways and transcriptional cascades. The canonical Wnt signaling pathway plays multiple roles during embryonic development including patterning, proliferation and differentiation in distinct tissues. Previous studies have established the importance of this pathway at multiple stages of pancreas formation as well as in postnatal organ function and homeostasis. In mice, gain-of-function experiments have demonstrated that activation of the canonical Wnt pathway results in pancreatic hypoplasia, a phenomenon whose underlying mechanisms remains to be elucidated. Here, we show that ectopic activation of epithelial canonical Wnt signaling causes aberrant induction of gastric and intestinal markers both in the pancreatic epithelium and mesenchyme, leading to the development of gut-like features. Furthermore, we provide evidence that β -catenin-induced impairment of pancreas formation depends on Hedgehog signaling. Together, our data emphasize the developmental plasticity of pancreatic progenitors and further underscore the key role of precise regulation of signaling pathways to maintain appropriate organ boundaries.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0164714PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5063371PMC
May 2017

Human islets contain four distinct subtypes of β cells.

Nat Commun 2016 07 11;7:11756. Epub 2016 Jul 11.

Oregon Stem Cell Center, Papé Family Pediatric Research Institute, Department of Pediatrics, Oregon Health and Science University, 3181 SW Sam Jackson Park Road, Portland, Oregon 97239, USA.

Human pancreatic islets of Langerhans contain five distinct endocrine cell types, each producing a characteristic hormone. The dysfunction or loss of the insulin-producing β cells causes diabetes mellitus, a disease that harms millions. Until now, β cells were generally regarded as a single, homogenous cell population. Here we identify four antigenically distinct subtypes of human β cells, which we refer to as β1-4, and which are distinguished by differential expression of ST8SIA1 and CD9. These subpopulations are always present in normal adult islets and have diverse gene expression profiles and distinct basal and glucose-stimulated insulin secretion. Importantly, the β cell subtype distribution is profoundly altered in type 2 diabetes. These data suggest that this antigenically defined β cell heterogeneity is functionally and likely medically relevant.
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http://dx.doi.org/10.1038/ncomms11756DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4942571PMC
July 2016
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