scholarly journals Proteomic and Transcriptional Profiles of Human Stem Cell-Derived β Cells Following Enteroviral Challenge

2020 ◽  
Vol 8 (2) ◽  
pp. 295 ◽  
Author(s):  
Julius O. Nyalwidhe ◽  
Agata Jurczyk ◽  
Basanthi Satish ◽  
Sambra Redick ◽  
Natasha Qaisar ◽  
...  
Keyword(s):  
Stem Cell ◽  
Ex Vivo ◽  
Human Islets ◽  
Islet Autoimmunity ◽  
Type I ◽  
Human Stem Cell ◽  
Β Cells ◽  
Enteroviral Infection ◽  
Insulin Gene Expression ◽  
New Treatments

Enteroviral infections are implicated in islet autoimmunity and type 1 diabetes (T1D) pathogenesis. Significant β-cell stress and damage occur with viral infection, leading to cells that are dysfunctional and vulnerable to destruction. Human stem cell-derived β (SC-β) cells are insulin-producing cell clusters that closely resemble native β cells. To better understand the events precipitated by enteroviral infection of β cells, we investigated transcriptional and proteomic changes in SC-β cells challenged with coxsackie B virus (CVB). We confirmed infection by demonstrating that viral protein colocalized with insulin-positive SC-β cells by immunostaining. Transcriptome analysis showed a decrease in insulin gene expression following infection, and combined transcriptional and proteomic analysis revealed activation of innate immune pathways, including type I interferon (IFN), IFN-stimulated genes, nuclear factor-kappa B (NF-κB) and downstream inflammatory cytokines, and major histocompatibility complex (MHC) class I. Finally, insulin release by CVB4-infected SC-β cells was impaired. These transcriptional, proteomic, and functional findings are in agreement with responses in primary human islets infected with CVB ex vivo. Human SC-β cells may serve as a surrogate for primary human islets in virus-induced diabetes models. Because human SC-β cells are more genetically tractable and accessible than primary islets, they may provide a preferred platform for investigating T1D pathogenesis and developing new treatments.

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

2021 ◽  
Author(s):  
Rebecca Shaheen ◽  
Rachel E. Gurlin ◽  
Rebecca Gologorsky ◽  
Charles Blaha ◽  
Pujita Munnangi ◽  
...  
Keyword(s):  
Stem Cell ◽  
Human Islets ◽  
Human Stem Cell ◽  
Bioartificial Pancreas ◽  
Β Cells ◽  
Adult Human

scholarly journals Integration of Islet/Beta-Cell Transplants with Host Tissue Using Biomaterial Platforms

Endocrinology ◽  
2020 ◽  
Vol 161 (11) ◽  
Author(s):  
Daniel W Clough ◽  
Jessica L King ◽  
Feiran Li ◽  
Lonnie D Shea
Keyword(s):  
Stem Cell ◽  
Immune Responses ◽  
Cell Function ◽  
Type I Diabetes ◽  
Glucose Sensing ◽  
Trophic Factors ◽  
Design Parameters ◽  
Type I ◽  
Β Cells ◽  
Glucose Levels

Abstract Cell-based therapies are emerging for type I diabetes mellitus (T1D), an autoimmune disease characterized by the destruction of insulin-producing pancreatic β-cells, as a means to provide long-term restoration of glycemic control. Biomaterial scaffolds provide an opportunity to enhance the manufacturing and transplantation of islets or stem cell–derived β-cells. In contrast to encapsulation strategies that prevent host contact with the graft, recent approaches aim to integrate the transplant with the host to facilitate glucose sensing and insulin distribution, while also needing to modulate the immune response. Scaffolds can provide a supportive niche for cells either during the manufacturing process or following transplantation at extrahepatic sites. Scaffolds are being functionalized to deliver oxygen, angiogenic, anti-inflammatory, or trophic factors, and may facilitate cotransplantation of cells that can enhance engraftment or modulate immune responses. This local engineering of the transplant environment can complement systemic approaches for maximizing β-cell function or modulating immune responses leading to rejection. This review discusses the various scaffold platforms and design parameters that have been identified for the manufacture of human pluripotent stem cell–derived β-cells, and the transplantation of islets/β-cells to maintain normal blood glucose levels.

scholarly journals Characterization of an immunodeficient mouse model of mucopolysaccharidosis type I suitable for preclinical testing of human stem cell and gene therapy

2007 ◽  
Vol 74 (6) ◽  
pp. 429-438 ◽  
Author(s):  
Mayra F. Garcia-Rivera ◽  
Leah E. Colvin-Wanshura ◽  
Matthew S. Nelson ◽  
Zhenhong Nan ◽  
Shaukat A. Khan ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Stem Cell ◽  
Mouse Model ◽  
Type I ◽  
Mucopolysaccharidosis Type ◽  
Mucopolysaccharidosis Type I ◽  
Human Stem Cell ◽  
Preclinical Testing ◽  
Cell And Gene Therapy

scholarly journals Free Fatty Acids Induce a Proinflammatory Response in Islets via the Abundantly Expressed Interleukin-1 Receptor I

Endocrinology ◽  
2009 ◽  
Vol 150 (12) ◽  
pp. 5218-5229 ◽  
Author(s):  
Marianne Böni-Schnetzler ◽  
Simone Boller ◽  
Sarah Debray ◽  
Karim Bouzakri ◽  
Daniel T. Meier ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Free Fatty Acids ◽  
Interleukin 1 ◽  
Human Islets ◽  
Type I ◽  
Β Cells ◽  
Mouse Tissues ◽  
Mouse Islets ◽  
Human And Mouse ◽  
Proinflammatory Factors

Abstract Islets of patients with type 2 diabetes mellitus (T2DM) display features of an inflammatory process including elevated levels of the cytokine IL-1β, various chemokines, and macrophages. IL-1β is a master regulator of inflammation, and IL-1 receptor type I (IL-1RI) blockage improves glycemia and insulin secretion in humans with T2DM and in high-fat-fed mice pointing to a pivotal role of IL-1RI activity in intra-islet inflammation. Given the association of dyslipidemia and T2DM, we tested whether free fatty acids (FFA) promote the expression of proinflammatory factors in human and mouse islets and investigated a role for the IL-1RI in this response. A comparison of 22 mouse tissues revealed the highest IL-1RI expression levels in islets and MIN6 β-cells. FFA induced IL-1β, IL-6, and IL-8 in human islets and IL-1β and KC in mouse islets. Elevated glucose concentrations enhanced FFA-induced proinflammatory factors in human islets. Blocking the IL-1RI with the IL-1R antagonist (IL-1Ra) strongly inhibited FFA-mediated expression of proinflammatory factors in human and mouse islets. Antibody inhibition of IL-1β revealed that FFA stimulated IL-1RI activity via the induction of the receptor ligand. FFA-induced IL-1β and KC expression in mouse islets was completely dependent on the IL-1R/Toll-like receptor (TLR) docking protein Myd88 and partly dependent on TLR2 and -4. Activation of TLR2 in purified human β-cells and islets stimulated the expression of proinflammatory factors, and IL-1RI activity increased the TLR2 response in human islets. We conclude that FFA and TLR stimulation induce proinflammatory factors in islets and that IL-1RI engagement results in signal amplification.

scholarly journals A vascularized 3D model of the human pancreatic islet for ex vivo study of immune cell-islet interaction

2021 ◽  
Author(s):  
R. Hugh F. Bender ◽  
Benjamen T O'Donnell ◽  
Bhupinder Shergill ◽  
Brittany Q Pham ◽  
Damie J Juat ◽  
...  
Keyword(s):  
Immune Cell ◽  
Ex Vivo ◽  
Cell Damage ◽  
Three Dimensional ◽  
Human Islets ◽  
Culture Conditions ◽  
Β Cells ◽  
Standard Culture ◽  
Key Steps

Insulin is an essential regulator of blood glucose homeostasis that is produced exclusively by β cells within the pancreatic islets of healthy individuals. In those affected by diabetes, immune inflammation, damage, and destruction of islet β cells leads to insulin deficiency and hyperglycemia. Current efforts to understand the mechanisms underlying β cell damage in diabetes rely on in vitro-cultured cadaveric islets. However, isolation of these islets involves removal of crucial matrix and vasculature that supports islets in the intact pancreas. Unsurprisingly, these islets demonstrate reduced functionality over time in standard culture conditions, thereby limiting their value for understanding native islet biology. Leveraging a novel, vascularized micro-organ (VMO) approach, we have recapitulated elements of the native pancreas by incorporating isolated human islets within a three-dimensional matrix nourished by living, perfusable blood vessels. Importantly, these islets show long-term viability and maintain robust glucose-stimulated insulin responses. Furthermore, vessel-mediated delivery of immune cells to these tissues provides a model to assess islet-immune cell interactions and subsequent islet killing -- key steps in type 1 diabetes pathogenesis. Together, these results establish the islet-VMO as a novel, ex vivo platform for studying human islet biology in both health and disease.

scholarly journals Transcriptional dynamics of induced pluripotent stem cell differentiation into β cells reveals full endodermal commitment and homology with human islets

Cytotherapy ◽  
2020 ◽  
Author(s):  
Silvia Pellegrini ◽  
Raniero Chimienti ◽  
Giulia Maria Scotti ◽  
Francesca Giannese ◽  
Dejan Lazarevic ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Differentiation ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Stem Cell Differentiation ◽  
Human Islets ◽  
Β Cells ◽  
Transcriptional Dynamics ◽  
Induced Pluripotent

Nicotinamide, a Potent SIRT2 Inhibitor, Delays Differentiation of Hematopoietic Progenitor Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4142-4142
Author(s):  
Toni Peled ◽  
Sophie Adi ◽  
Elina Glukhman ◽  
Frida Grynspan ◽  
Arnon Nagler ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Fate ◽  
Cell Cultures ◽  
Ex Vivo ◽  
Cell Phenotype ◽  
Type I ◽  
Hematopoietic Stem ◽  
Additional Function ◽  
Base Exchange ◽  
Cell Fate Decisions

Abstract CD38, originally described as a differentiation marker, has emerged as an important multifunctional transmembrane protein. Its most intriguing and well-characterized function is its ability to catalyze the synthesis of cyclic ADP-ribose (cADPR) from NAD. Of particular interest is its presence on the inner membrane of the nucleus, suggesting that CD38/cADPR may play a direct role in mediating nuclear activation and gene expression. Our studies on ex vivo expansion of Hematopoietic Stem Cells (HSCs) have led us to test whether alteration of CD38 function carries the potential of affecting cell fate decisions of HSCs. Inhibition of CD38 enzymatic activity was achieved by treating CD34+ cell cultures with nicotinamide (NA), a well-known base-exchange inhibitor demonstrated to inhibit the synthesis of cADPR from NAD. We report here that exogenously added nicotinamide (5–10 mM) to CD34+ cell cultures supplemented with cytokines (SCF, TPO, IL-6, FLt3, +/− IL-3) resulted in significant enrichment of CD34+CD38− (79±9.3%, n=9) and CD34+CD38−Lin− (19±3%, n=8) cells, as compared with control cultures treated only with cytokines (6.3±1.8%, n=9, and 0.7±0.06%, n=8, respectively, p<0.01). The functionality of these early progenitor subsets was demonstrated using the extended LTC-CFC assay, performed in the absence of NA. These results raised the intriguing possibility that cADPR production may have a pivotal role in regulation of CD34+ cell fate. However, inhibition of cADPR downstream signal transduction pathways by its specific antagonist, 8-amino-cADPR did not yield any effect on CD34+ cell cultures, excluding the possibility that nicotinamide modulates CD34+ cell fate solely by inhibition of cADPR synthesis. Nicotinamide is also a well-known potent inhibitor of SIRT2, a unique NAD(+)-dependent type III histone deacetylase (HDAC) with mono-ADP-ribosyltransferase activity involved in gene silencing, metabolism, apoptosis and aging. NA blocks NAD(+) hydrolysis by binding to an adjacent conserved pocket, and is therefore suggested as the physiologically relevant regulator of SIRT2 enzymes. This additional function of nicotinamide raises the intriguing possibility that HSC enrichment achieved by nicotinamide treatment may be related to specific inhibition of SIRT2 deacetylase activity and modulation of chromatin architecture leading to re-activation of previously silenced genes. In line with this hypothesis, Milhem et all. recently reported that addition of trichostatin A, a specific HDAC (type I and II) inhibitor, along with a DNA hypomethylating agent, modulated HSC fate ex vivo resulting in the retention of stem cell phenotype, number, and function (Blood, 2004; 103; 4102). Ongoing work is aimed at elucidating whether inhibition of SIRT2 is specifically involved in NA mechanism of activity leading to modulation of hematopoietic stem cell fate in ex vivo conditions.

Characterization of an Immunodeficient Mucopolysaccharidosis Type I (MPS-I) Mouse Model Suitable for Preclinical Testing of Human Stem Cell and Gene Therapy.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1265-1265
Author(s):  
Mayra F. Quito-Rivera ◽  
Leah E. Colvin-Wanshura ◽  
Matthew S. Nelson ◽  
Zhenhong Nan ◽  
Shaukat A. Khan ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Type I ◽  
Mucopolysaccharidosis Type ◽  
Mucopolysaccharidosis Type I ◽  
Human Stem Cell ◽  
Therapeutic Approaches ◽  
Behavioral Abnormalities ◽  
Mps I

Abstract Mucopolysaccharidosis type I (MPS-I; Hurler syndrome) is an inborn metabolic disorder due to lack of the lysosomal glycosaminoglycan (GAG)-degrading enzyme alpha-L-iduronidase (IDUA). The resulting GAG accumulation causes progressive multi-system dysfunction and death in the first decade. We recently identified structural abnormalities in the accumulated GAGs that lead to defective heparin-binding cytokine signaling in MPS-I (Pan C et al, Blood 2005 in press: DOI 10.1182/blood-2005-02-0657), which may constitute a mechanism responsible for the diverse clinical manifestations of MPS-I and related mucopolysaccharidoses. Allogeneic hematopoietic stem cell transplantation (HSCT), if performed early in the disease, ameliorates many clinical features and extends life. However, HSCT is not available to all patients, and also does not adequately correct some of the most devastating features of MPS-I including mental retardation and skeletal deformities. Therefore, novel cellular, enzyme replacement and gene therapy approaches need to be developed for MPS-I. Such therapeutic strategies will best be tested in an immunodeficient host which is less likely to develop immune reactions to transplanted human or gene-corrected cells or their secreted IDUA enzyme. We have bred and characterized a homozygous immunodeficient MPS-I mouse (developed as a heterozygous MPS-I mouse on a NOD/SCID background by Jackson Labs) that is well suited for examining the efficacy of such therapeutic approaches. The phenotype of homozygous NOD/SCID/MPS-I mice closely mimicked the clinical features of human MPS-I including coarsening of facial features, skeletal deformities and diverse behavioral abnormalities. IDUA enzyme was completely undetectable in the liver, spleen, heart, lung, kidney and brain of homozygous animals (P<0.001) and was reduced to approximately 50% in heterozygous animals. Homozygous animals developed marked GAG accumulation (3 to 25 fold) in the same organs (P=0.03 to P<0.0001). Neuropathological examination showed accumulation of GM3 gangliosides in the cerebral peduncles, cerebellum and ventral brainstem of homozygous mice. Importantly, measurement of urinary GAG excretion (5-fold higher in homozygous animals; P<0.003) provided a non-invasive and reliable method that can be used to serially follow the biochemical efficacy of therapeutic interventions. We also identified and validated using rigorous biostatistical methods, a highly reproducible method for evaluating sensorimotor function and learning in this mouse model. This Rotarod Test revealed marked abnormalities in sensorimotor integration involving the cerebellum, niagro-striatal and proprioceptive pathways and motor cortex, as well as in learning. We believe that NOD/SCID/MPS-I mice will provide an extremely suitable animal model for assessing the systemic as well as neurological effects of human stem cell transplantation and gene therapeutic approaches, using the above techniques to measure efficacy. We have started using this model to assess the effect of intra-cerebroventricular implantation of human stem cells on the biochemical, pathological and behavioral abnormalities identified. Similar strategies may be invaluable for developing and studying animal models of other MPSs and related diseases.

scholarly journals Phenotypic analysis of c-Kit expression in epithelial monolayers derived from postnatal rat pancreatic islets

2004 ◽  
Vol 182 (1) ◽  
pp. 113-122 ◽  
Author(s):  
R Wang ◽  
J Li ◽  
N Yashpal
Keyword(s):  
Stem Cell ◽  
Progenitor Cells ◽  
Matrix Protein ◽  
Ex Vivo ◽  
Stem Cell Marker ◽  
Haematopoietic Stem Cell ◽  
Extracellular Matrix Protein ◽  
Type I ◽  
Cytokeratin 20 ◽  
Epithelial Monolayers

The limitation of available islets for transplantation is a major obstacle for the treatment of diabetes through islet therapy. However, islet monolayers expanded ex vivo may provide a source of progenitor cells and a model to help understand islet development from precursor cell types. The existence of progenitor cells within the islets is highly likely, yet, to date, no fully defined or characterized postnatal stem cell has been isolated, expanded or marked. Our study evaluates the expression of progenitor markers, including the haematopoietic stem cell marker c-Kit, in epithelial monolayers derived from postnatal rat islets through immunofluorescence and RT-PCR, and the ability of precursor-rich monolayers to reform islet-like structures. Islets formed confluent monolayers when cultured on a type I collagen gel which lacked endocrine phenotypes but were positive for cytokeratin 20 and contained an increased proportion of proliferating c-Kit-expressing cells, with the proportion reaching a maximum of 45+/-6% at 8 weeks of culture. Evaluation of transcription factors at the mRNA level revealed constant PDX-1, ngn3 and Pax4 expression, while undifferentiated cell markers, such as Oct4 and alpha-fetoprotein, were also detected frequently after 4 weeks of culture. Changing the extracellular matrix protein to laminin-rich Matrigel, the monolayers re-formed islet-like clusters that secreted insulin in a glucose-responsive fashion. Our data show that islets can be expanded ex vivo to form epithelial monolayers with rich undifferentiating cell populations that are characterized by cells expressing the progenitor markers. These monolayers are capable of extensive proliferation and retain plasticity to form new islet cells, and c-Kit-expressing cells may play an important role in new islet cluster formation.

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