GENETICALLY ENGINEERED PIG MODELS FOR TRANSLATIONAL DIABETES RESEARCH

2013 ◽  
Vol 25 (1) ◽  
pp. 320 ◽  
Author(s):  
Eckhard Wolf
Keyword(s):  
Mouse Models ◽  
Cell Mass ◽  
Physiological Age ◽  
Diabetic Patients ◽  
Diabetes Research ◽  
Rodent Models ◽  
Β Cells ◽  
Β Cell ◽  
Transgenic Pigs ◽  
Genetic Modifications

Animal models play crucial roles for understanding disease mechanisms and for the development and evaluation of therapeutic strategies. In biomedicine, classical rodent models are most widely used for several reasons, including standardization of genetics and environment, cost efficiency, and the possibility to introduce targeted genetic modifications for the generation of tailored disease models. However, due to differences in anatomical and physiological characteristics, rodent models do not always reflect the situation of human patients sufficiently well to be predictive in terms of efficacy and safety of new therapies. In this respect, the pig has been discussed as a missing link between mouse models and human patients. As a monogastric omnivore, the pig shares many anatomical and physiological similarities with humans. Importantly, the techniques for genetic modification of pigs have been refined to a level allowing almost the same spectrum of alterations as in mouse models (Aigner et al. 2010 J. Mol. Med. (Berl.) 88, 653–664). These include inducible transgene expression systems (Klymiuk et al. 2012 FASEB J. 26, 1086–1099) as well as the introduction of targeted genetic modifications (Klymiuk et al. 2012 J. Mol. Med. (Berl.) 90, 597–608). A major focus of our laboratory is the generation, characterisation, and implementation of pig models for translational diabetes research. Transgenic pigs expressing a dominant negative receptor for the incretin hormone glucose-dependent insulinotropic polypeptide (GIP) demonstrated a crucial role of the GIP system for the physiological age-related expansion of pancreatic β-cell mass. Moreover, this animal model shares important characteristics of type 2 diabetes mellitus: impaired incretin effect, reduced glucose tolerance and insulin secretion, and a progressive reduction of β-cell mass (Renner et al. 2010 Diabetes 59, 1228–1238). More recently, we used this model to search for metabolic biomarkers which are associated with progression in the pre-diabetic period and identified specific amino acid and lipid signatures as candidate biomarkers (Renner et al. 2012 Diabetes 61, 2166–2175). Further, we created the first pig model for permanent neonatal diabetes by expression C94Y mutant insulin in the β-cells of transgenic pigs. In addition to their use as biomedical models, pigs may also serve as organ and tissue donors for xenotransplantation. Transplantation of encapsulated porcine pancreatic islets to type 1 diabetic patients with severe unaware hypoglycemia has already entered clinical studies, but encapsulation may shorten the lifespan of the islets. Therefore, in order to overcome the rejection of pig islets by human T-cells, we generated transgenic pigs expressing the optimized CTLA-4Ig variant LEA29Y in the pancreatic β-cells. Islets from LEA29Y transgenic pigs rescued diabetes and were protected against rejection in a humanized mouse model (Klymiuk et al. 2012 Diabetes 61, 1527–1532).

scholarly journals NEUROD1 Is Required for the Early α and β Endocrine Differentiation in the Pancreas

2021 ◽  
Vol 22 (13) ◽  
pp. 6713
Author(s):  
Romana Bohuslavova ◽  
Ondrej Smolik ◽  
Jessica Malfatti ◽  
Zuzana Berkova ◽  
Zaneta Novakova ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Cell Fate ◽  
Cell Mass ◽  
Neonatal Diabetes ◽  
Diabetes Research ◽  
Pancreas Development ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Islet Cell ◽  
Endocrine Differentiation

Diabetes is a metabolic disease that involves the death or dysfunction of the insulin-secreting β cells in the pancreas. Consequently, most diabetes research is aimed at understanding the molecular and cellular bases of pancreatic development, islet formation, β-cell survival, and insulin secretion. Complex interactions of signaling pathways and transcription factor networks regulate the specification, growth, and differentiation of cell types in the developing pancreas. Many of the same regulators continue to modulate gene expression and cell fate of the adult pancreas. The transcription factor NEUROD1 is essential for the maturation of β cells and the expansion of the pancreatic islet cell mass. Mutations of the Neurod1 gene cause diabetes in humans and mice. However, the different aspects of the requirement of NEUROD1 for pancreas development are not fully understood. In this study, we investigated the role of NEUROD1 during the primary and secondary transitions of mouse pancreas development. We determined that the elimination of Neurod1 impairs the expression of key transcription factors for α- and β-cell differentiation, β-cell proliferation, insulin production, and islets of Langerhans formation. These findings demonstrate that the Neurod1 deletion altered the properties of α and β endocrine cells, resulting in severe neonatal diabetes, and thus, NEUROD1 is required for proper activation of the transcriptional network and differentiation of functional α and β cells.

scholarly journals Dual-Reporter β-Cell-Specific Male Transgenic Rats for the Analysis of β-Cell Functional Mass and Enrichment by Flow Cytometry

Endocrinology ◽  
2015 ◽  
Vol 157 (3) ◽  
pp. 1299-1306 ◽  
Author(s):  
Julien Ghislain ◽  
Ghislaine Fontés ◽  
Caroline Tremblay ◽  
Melkam A. Kebede ◽  
Vincent Poitout
Keyword(s):  
Insulin Secretion ◽  
Fluorescent Protein ◽  
Ex Vivo ◽  
Yellow Fluorescent Protein ◽  
Cell Mass ◽  
Renilla Luciferase ◽  
Diabetes Research ◽  
Β Cells ◽  
Transgenic Rats ◽  
Β Cell

Abstract Mouse β-cell-specific reporter lines have played a key role in diabetes research. Although the rat provides several advantages, its use has lagged behind the mouse due to the relative paucity of genetic models. In this report we describe the generation and characterization of transgenic rats expressing a Renilla luciferase (RLuc)-enhanced yellow fluorescent protein (YFP) fusion under control of a 9-kb genomic fragment from the rat ins2 gene (RIP7-RLuc-YFP). Analysis of RLuc luminescence and YFP fluorescence revealed that reporter expression is restricted to β-cells in the adult rat. Physiological characteristics including body weight, fat and lean mass, fasting and fed glucose levels, glucose and insulin tolerance, and β-cell mass were similar between two RIP7-RLuc-YFP lines and wild-type littermates. Glucose-induced insulin secretion in isolated islets was indistinguishable from controls in one of the lines, whereas surprisingly, insulin secretion was defective in the second line. Consequently, subsequent studies were limited to the former line. We asked whether transgene activity was responsive to glucose as shown previously for the ins2 gene. Exposing islets ex vivo to high glucose (16.7 mM) or in vivo infusion of glucose for 24 hours increased luciferase activity in islets, whereas the fraction of YFP-positive β-cells after glucose infusion was unchanged. Finally, we showed that fluorescence-activated cell sorting of YFP-positive islet cells can be used to enrich for β-cells. Overall, this transgenic line will enable for the first time the application of both fluorescence and bioluminescence/luminescence-based approaches for the study of rat β-cells.

scholarly journals Loss of Foxd3 Results in Decreased β-Cell Proliferation and Glucose Intolerance During Pregnancy

Endocrinology ◽  
2011 ◽  
Vol 152 (12) ◽  
pp. 4589-4600 ◽  
Author(s):  
Jennifer L. Plank ◽  
Audrey Y. Frist ◽  
Alison W. LeGrone ◽  
Mark A. Magnuson ◽  
Patricia A. Labosky
Keyword(s):  
Cell Proliferation ◽  
Cell Mass ◽  
Diabetic Patients ◽  
Β Cells ◽  
Β Cell ◽  
Forkhead Box ◽  
Physiological Demands ◽  
Mass Expansion ◽  
Specific Deletion

A complete molecular understanding of β-cell mass expansion will be useful for the improvement of therapies to treat diabetic patients. During normal periods of metabolic challenges, such as pregnancy, β-cells proliferate, or self-renew, to meet the new physiological demands. The transcription factor Forkhead box D3 (Foxd3) is required for maintenance and self-renewal of several diverse progenitor cell lineages, and Foxd3 is expressed in the pancreatic primordium beginning at 10.5 d postcoitum, becoming localized predominantly to β-cells after birth. Here, we show that mice carrying a pancreas-specific deletion of Foxd3 have impaired glucose tolerance, decreased β-cell mass, decreased β-cell proliferation, and decreased β-cell size during pregnancy. In addition, several genes known to regulate proliferation, Foxm1, Skp2, Ezh2, Akt2, and Cdkn1a, are misregulated in islets isolated from these Foxd3 mutant mice. Together, these data place Foxd3 upstream of several pathways critical for β-cell mass expansion in vivo.

scholarly journals The Many Lives of Myc in the Pancreatic β-Cell

2020 ◽  
pp. jbc.REV120.011149
Author(s):  
Carolina Rosselot ◽  
Sharon Baumel-Alterzon ◽  
Yansui Li ◽  
Gabriel Brill ◽  
Luca Lambertini ◽  
...  
Keyword(s):  
Cell Death ◽  
Transcription Factor ◽  
Therapeutic Potential ◽  
Ex Vivo ◽  
Diabetic Patients ◽  
Cell Regeneration ◽  
Diabetes Research ◽  
Β Cells ◽  
Β Cell

Diabetes results from insufficient numbers of functional pancreatic β-cells. Thus, increasing the number of available functional β-cells ex vivo for transplantation, or regenerating them in situ in diabetic patients, is a major focus of diabetes research. The transcription factor, Myc, discovered decades ago, lies at the nexus of most, if not all, known proliferative pathways. Based on this, many studies in the 1990’s and early 2000’s explored the potential of harnessing Myc expression to expand β-cells for diabetes treatment. Nearly all these studies in β-cells used pathophysiological or supraphysiological levels of Myc and reported enhanced β-cell death, de-differentiation or the formation of insulinomas if co-overexpressed with Bcl-xL, an inhibitor of apoptosis. This obviously reduced the enthusiasm for Myc as a therapeutic target for β-cell regeneration. However, recent studies indicate that “gentle” induction of Myc expression enhances β-cell replication without induction of cell death or loss of insulin secretion, suggesting that appropriate levels of Myc could have therapeutic potential for β-cell regeneration. Furthermore, although it has been known for decades that Myc is induced by glucose in β-cells very little is known about how this essential anabolic transcription factor perceives and responds to nutrients and increased insulin demand in vivo. Here we summarize the previous and recent knowledge of Myc in the β-cell, its potential for β-cell regeneration and its physiological importance for neonatal and adaptive β-cell expansion.

scholarly journals Nrf2 Regulates β-cell Mass by Suppressing Cell Death and Promoting Proliferation

2021 ◽  
Author(s):  
Sharon Baumel-Alterzon ◽  
Liora S. Katz ◽  
Gabriel Brill ◽  
Clairete Jean-Pierre ◽  
Yansui Li ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
High Fat Diet ◽  
Ex Vivo ◽  
Cell Mass ◽  
Conditional Knockout ◽  
Diabetes Research ◽  
Loss Of Function ◽  
High Fat ◽  
Β Cells ◽  
Β Cell

SUMMARYFinding therapies that can protect and expand functional β-cell mass is a major goal of diabetes research. Here we generated β-cell-specific conditional knockout and gain-of-function mouse models and used human islet transplant experiments to examine how manipulating Nrf2 levels affects β-cell survival, proliferation and mass. Depletion of Nrf2 in β-cells resulted in decreased glucose-stimulated β-cell proliferation ex vivo and decreased adaptive β-cell proliferation and β-cell mass expansion after a high fat diet in vivo. Nrf2 protects β-cells from apoptosis after a high fat diet. Nrf2 loss-of-function decreases Pdx1 abundance and insulin content. Activating Nrf2 in a β-cell-specific manner increases β-cell proliferation and β-cell mass. Human islets transplanted under the kidney capsule of immunocompromised mice and treated systemically with CDDO-Me, an Nrf2 activator, display increased β-cell proliferation. Thus, Nrf2 regulates β-cell mass and is an exciting therapeutic target for expanding β-cell mass in diabetes.

MiR-221/222 Inhibit Insulin Production of Pancreatic β-Cells in Mice

Endocrinology ◽  
2019 ◽  
Vol 161 (1) ◽  
Author(s):  
Liwen Fan ◽  
Aijing Shan ◽  
Yutong Su ◽  
Yulong Cheng ◽  
He Ji ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Transgenic Mouse ◽  
Mouse Models ◽  
Cell Mass ◽  
Luciferase Reporter ◽  
Transgenic Mouse Models ◽  
Insulin Production ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Β Cells

Abstract Microribonucleic acids (miRNAs) are essential for the regulation of development, proliferation, and functions of pancreatic β-cells. The conserved miR-221/222 cluster is an important regulator in multiple cellular processes. Here we investigated the functional role of miR-221/222 in the regulation of β-cell proliferation and functions in transgenic mouse models. We generated 2 pancreatic β-cell-specific–miR-221/222 transgenic mouse models on a C57BL/6J background. The glucose metabolic phenotypes, β-cell mass, and β-cell functions were analyzed in the mouse models. Adenovirus-mediated overexpression of miR-221/222 was performed on β-cells and mouse insulinoma 6 (MIN6) cells to explore the effect and mechanisms of miR-221/222 on β-cell proliferation and functions. Luciferase reporter assay, histological analysis, and quantitative polymerase chain reaction (PCR) were carried out to study the direct target genes of miR-221/222 in β-cells. The expression of miR-221/222 was significantly upregulated in β-cells from the high-fat diet (HFD)–fed mice and db/db mice. Overexpression of miR-221/222 impaired the insulin production and secretion of β-cells and resulted in glucose intolerance in vivo. The β-cell mass and proliferation were increased by miR-221/222 expression via Cdkn1b and Cdkn1c. MiR-221/222 repressed insulin transcription activity through targeting Nfatc3 and lead to reduction of insulin in β-cells. Our findings demonstrate that miR-221/222 are important regulators of β-cell proliferation and insulin production. The expression of miR-221/222 in β-cells could regulate glucose metabolism in physiological and pathological processes.

scholarly journals Selective β-Cell Loss and α-Cell Expansion in Patients with Type 2 Diabetes Mellitus in Korea

2003 ◽  
Vol 88 (5) ◽  
pp. 2300-2308 ◽  
Author(s):  
Kun Ho Yoon ◽  
Seung Hyun Ko ◽  
Jae Hyoung Cho ◽  
Jung Min Lee ◽  
Yu Bae Ahn ◽  
...  
Keyword(s):  
Cell Mass ◽  
Cell Loss ◽  
Relative Volume ◽  
Diabetic Patients ◽  
Organ Donors ◽  
Β Cells ◽  
Β Cell ◽  
Type 2 Diabetic Patients ◽  
Type 2 Diabetic

In the presence of obesity, β-cell mass needs to be increased to compensate for the accompanying demands and maintain euglycemia. However, in Korea, the majority of type 2 diabetic patients are nonobese. We determined the absolute masses, relative volumes, and ratio of α- and β-cell in the pancreas and islets in normal and diabetic Korean subjects to correlate these findings with the clinical characteristics. Whole pancreases procured from organ donors were divided into 24 parts (control 1, n = 9). Tissue was also obtained by surgical resection after 35 partial pancreatectomies: in 25 diabetic patients, 10 age- and body mass index (BMI)-matched patients of benign or malignant pancreatic tumor without diabetes mellitus (DM) (control 2). Morphometric quantifications were performed. In control 1, the relative volume of β-cells was 2.1 ± 0.9%, and the total β-cell mass was 1.3 ± 0.3 g. The relative volume of β-cells was found to be variable (control 1, 2.1 ± 0.9%; control 2, 1.9 ± 0.7%; DM, 1.4 ± 1.0%; P < 0.05 DM vs. control 1 and 2) and showed good correlation with BMI (control 1, r2 = 0.64; DM, r2 = 0.55; all subjects, r2 = 0.38; P < 0.05). Notably, in type 2 diabetic patients, the ratio of α-cell area to β-cell area in the islet was higher than in control 1 and 2 (0.81 ± 0.4 vs. 0.29 ± 0.2, 0.20 ± 0.1, P < 0.05). Additionally, significant α-cell expansion and a decreased β-cell fraction were predominantly observed in larger islets (islet area, >6415 μm2; P < 0.05) in control 1 and diabetic patients. The relative volume of β-cell was found to be correlated with BMI in diabetic patients and normal organ donors. Moreover, decreased β-cell but increased α-cell proportion in the islets suggests for a selective β-cell loss in the pathogenesis of Korean type 2 diabetes.

scholarly journals Minireview: Directed Differentiation and Encapsulation of Islet β-Cells—Recent Advances and Future Considerations

2015 ◽  
Vol 29 (10) ◽  
pp. 1388-1399 ◽  
Author(s):  
Hubert M. Tse ◽  
Veronika Kozlovskaya ◽  
Eugenia Kharlampieva ◽  
Chad S. Hunter
Keyword(s):  
Cell Mass ◽  
Diabetic Patients ◽  
Directed Differentiation ◽  
Β Cells ◽  
Β Cell ◽  
Cadaveric Donors ◽  
Health Burdens

Abstract Diabetes mellitus has rapidly become a 21st century epidemic with the promise to create vast economic and health burdens, if left unchecked. The 2 major forms of diabetes arise from unique causes, with outcomes being an absolute (type 1) or relative (type 2) loss of functional pancreatic islet β-cell mass. Currently, patients rely on exogenous insulin and/or other pharmacologies that restore glucose homeostasis. Although these therapies have prolonged countless lives over the decades, the striking increases in both type 1 and type 2 diabetic diagnoses worldwide suggest a need for improved treatments. To this end, islet biologists are developing cell-based therapies by which a patient's lost insulin-producing β-cell mass is replenished. Pancreatic or islet transplantation from cadaveric donors into diabetic patients has been successful, yet the functional islet demand far surpasses supply. Thus, the field has been striving toward transplantation of renewable in vitro-derived β-cells that can restore euglycemia. Challenges have been numerous, but progress over the past decade has generated much excitement. In this review we will summarize recent findings that have placed us closer than ever to β-cell replacement therapies. With the promise of cell-based diabetes therapies on the horizon, we will also provide an overview of cellular encapsulation technologies that will deliver critical protection of newly implanted cells.

scholarly journals The RNA-binding protein LARP1 is dispensable for pancreatic β-cell function and mass

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Joao Pedro Werneck-de-Castro ◽  
Flavia Leticia Martins Peçanha ◽  
Diego Henrique Silvestre ◽  
Ernesto Bernal-Mizrachi
Keyword(s):  
Glucose Homeostasis ◽  
Ribosome Biogenesis ◽  
Rna Binding ◽  
Cell Function ◽  
Cell Mass ◽  
Diabetic Patients ◽  
Β Cells ◽  
Β Cell ◽  
Fasting Glycemia ◽  
Mouse Islets

AbstractMechanistic target of rapamycin complex 1 (mTORC1) deficiency or chronic hyperactivation in pancreatic β-cells leads to diabetes. mTORC1 complexes with La-related protein 1 (LARP1) to specifically regulate the expression of 5′ terminal oligopyrimidine tract (5′TOP) mRNAs which encode proteins of the translation machinery and ribosome biogenesis. Here we show that LARP1 is the most expressed LARP in mouse islets and human β-cells, being 2–4-fold more abundant than LARP1B, a member of the family that also interacts with mTORC1. Interestingly, β-cells from diabetic patients have higher LARP1 and LARP1B expression. However, specific deletion of Larp1 gene in β-cells (β-Larp1KO mice) did not impair insulin secretion and glucose metabolism in male and female mice. High fat or high branched-chain amino acid (BCAA) diets did not disturb glucose homeostasis compared to control littermates up to 8 weeks; BCAA diet slightly impaired glucose tolerance in the β-Larp1KO mice at 16 weeks. However, no differences in plasma insulin levels, non-fasting glycemia and β-cell mass were observed in the β-Larp1KO mice. In conclusion, LARP1 is the most abundant LARP in mouse islets and human β-cells, and it is upregulated in diabetic subjects. However, genetically disruption of Larp1 gene did not impact glucose homeostasis in basal and diabetogenic conditions, suggesting no major role for LARP1 in β-cells.

scholarly journals Ontology of the apelinergic system in mouse pancreas during pregnancy and relationship with β-cell mass

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Brenda Strutt ◽  
Sandra Szlapinski ◽  
Thineesha Gnaneswaran ◽  
Sarah Donegan ◽  
Jessica Hill ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Insulin Secretion ◽  
Glucose Transporter ◽  
Cell Mass ◽  
Elevated Serum ◽  
Pancreatic Β Cell ◽  
Β Cells ◽  
Β Cell ◽  
Glucose Transporter 2 ◽  
Glucose Stimulated Insulin Secretion

AbstractThe apelin receptor (Aplnr) and its ligands, Apelin and Apela, contribute to metabolic control. The insulin resistance associated with pregnancy is accommodated by an expansion of pancreatic β-cell mass (BCM) and increased insulin secretion, involving the proliferation of insulin-expressing, glucose transporter 2-low (Ins+Glut2LO) progenitor cells. We examined changes in the apelinergic system during normal mouse pregnancy and in pregnancies complicated by glucose intolerance with reduced BCM. Expression of Aplnr, Apelin and Apela was quantified in Ins+Glut2LO cells isolated from mouse pancreata and found to be significantly higher than in mature β-cells by DNA microarray and qPCR. Apelin was localized to most β-cells by immunohistochemistry although Aplnr was predominantly associated with Ins+Glut2LO cells. Aplnr-staining cells increased three- to four-fold during pregnancy being maximal at gestational days (GD) 9–12 but were significantly reduced in glucose intolerant mice. Apelin-13 increased β-cell proliferation in isolated mouse islets and INS1E cells, but not glucose-stimulated insulin secretion. Glucose intolerant pregnant mice had significantly elevated serum Apelin levels at GD 9 associated with an increased presence of placental IL-6. Placental expression of the apelinergic axis remained unaltered, however. Results show that the apelinergic system is highly expressed in pancreatic β-cell progenitors and may contribute to β-cell proliferation in pregnancy.

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