scholarly journals Microporous scaffolds support assembly and differentiation of pancreatic progenitors into β-cell clusters

2019 ◽  
Vol 96 ◽  
pp. 111-122 ◽  
Author(s):  
Richard L. Youngblood ◽  
Joshua P. Sampson ◽  
Kimberly R. Lebioda ◽  
Lonnie D. Shea
Keyword(s):  
Β Cell ◽  
Cell Clusters ◽  
Pancreatic Progenitors ◽  
Microporous Scaffolds

scholarly journals Microporous Scaffolds Drive Assembly and Maturation of Progenitors into β-Cell Clusters

2019 ◽  
Author(s):  
Richard L. Youngblood ◽  
Joshua P. Sampson ◽  
Kimberly R. Lebioda ◽  
Graham Spicer ◽  
Lonnie D. Shea
Keyword(s):  
Β Cell ◽  
Cell Clusters ◽  
Microporous Scaffolds

scholarly journals Cooperative function of Pdx1 and Oc1 in multipotent pancreatic progenitors impacts postnatal islet maturation and adaptability

2018 ◽  
Vol 314 (4) ◽  
pp. E308-E321 ◽  
Author(s):  
Peter A. Kropp ◽  
Jennifer C. Dunn ◽  
Bethany A. Carboneau ◽  
Doris A. Stoffers ◽  
Maureen Gannon
Keyword(s):  
Expression Patterns ◽  
Cell Maturation ◽  
Placental Lactogen ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Progenitors ◽  
Glucose Stimulated Insulin Secretion ◽  
Double Heterozygosity ◽  
And Function ◽  
The Impact

The transcription factors pancreatic and duodenal homeobox 1 (Pdx1) and onecut1 (Oc1) are coexpressed in multipotent pancreatic progenitors (MPCs), but their expression patterns diverge in hormone-expressing cells, with Oc1 expression being extinguished in the endocrine lineage and Pdx1 being maintained at high levels in β-cells. We previously demonstrated that cooperative function of these two factors in MPCs is necessary for proper specification and differentiation of pancreatic endocrine cells. In those studies, we observed a persistent decrease in expression of the β-cell maturity factor MafA. We therefore hypothesized that Pdx1 and Oc1 cooperativity in MPCs impacts postnatal β-cell maturation and function. Here our model of Pdx1-Oc1 double heterozygosity was used to investigate the impact of haploinsufficiency for both of these factors on postnatal β-cell maturation, function, and adaptability. Examining mice at postnatal day (P) 14, we observed alterations in pancreatic insulin content in both Pdx1 heterozygotes and double heterozygotes. Gene expression analysis at this age revealed significantly decreased expression of many genes important for glucose-stimulated insulin secretion (e.g., Glut2, Pcsk1/2, Abcc8) exclusively in double heterozygotes. Analysis of P14 islets revealed an increase in the number of mixed islets in double heterozygotes. We predicted that double-heterozygous β-cells would have an impaired ability to respond to stress. Indeed, we observed that β-cell proliferation fails to increase in double heterozygotes in response to either high-fat diet or placental lactogen. We thus report here the importance of cooperation between regulatory factors early in development for postnatal islet maturation and adaptability.

scholarly journals Microporous scaffolds drive assembly and maturation of progenitors into β-cell clusters

2019 ◽  
Author(s):  
Richard L. Youngblood ◽  
Joshua P. Sampson ◽  
Kimberly R. Lebioda ◽  
Graham Spicer ◽  
Lonnie D. Shea
Keyword(s):  
Basement Membrane ◽  
Suspension Cultures ◽  
Cell Interactions ◽  
Cell Maturation ◽  
Β Cells ◽  
Β Cell ◽  
Cell Clusters ◽  
Matrix Deposition ◽  
Cell Cell

AbstractHuman pluripotent stem cells (hPSCs) represent a promising cell source for the development of β-cells for use in therapies for type 1 diabetes. Current culture approaches provide the signals to drive differentiation towards β-cells, with the cells spontaneously assembling into clusters. Herein, we adapted the current culture systems to cells seeded on microporous biomaterials, with the hypothesis that the pores can guide the assembly into β-cell clusters of defined size that can enhance maturation. The microporous scaffold culture allows hPSC-derived pancreatic progenitors to form clusters at a consistent size as cells undergo differentiation to immature β-cells. By modulating the scaffold pore sizes, we observed 250-425 µm pore size scaffolds significantly enhance insulin expression and key β-cell maturation markers compared to suspension cultures. Furthermore, when compared to suspension cultures, the scaffold culture showed increased insulin secretion in response to glucose stimulus indicating the development of functional β-cells. In addition, scaffolds facilitated cell-cell interactions enabled by the scaffold design and cell-mediated matrix deposition of extracellular matrix (ECM) proteins associated with the basement membrane of islet cells. We further investigated the influence of ECM on cell development by incorporating an ECM matrix on the scaffold prior to cell seeding; however, their presence did not further enhance maturation. These results suggest the microporous scaffold culture facilitates 3D cluster formation, supports cell-cell interactions, and provides a matrix similar to a basement membrane to drive in vitro hPSC-derived β-cell maturation and demonstrates the feasibility of these scaffolds as a biomanufacturing platform.

scholarly journals Pancreatic β-cell replacement: advances in protocols used for differentiation of pancreatic progenitors to β-like cells

2015 ◽  
Author(s):  
Muhammad Waseem Ghani ◽  
Li Ye ◽  
Zhao Yi ◽  
Hammad Ghani ◽  
Muhammad Waseem Birmani ◽  
...  
Keyword(s):  
Pancreatic Β Cell ◽  
Β Cell ◽  
Cell Replacement ◽  
Pancreatic Progenitors

scholarly journals Retinoblastoma tumor suppressor protein in pancreatic progenitors controls α- and β-cell fate

2013 ◽  
Vol 110 (36) ◽  
pp. 14723-14728 ◽  
Author(s):  
Erica P. Cai ◽  
Xiaohong Wu ◽  
Stephanie A. Schroer ◽  
Andrew J. Elia ◽  
M. Cristina Nostro ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Cell Fate ◽  
Tumor Suppressor Protein ◽  
Β Cell ◽  
Retinoblastoma Tumor Suppressor Protein ◽  
Pancreatic Progenitors ◽  
Retinoblastoma Tumor Suppressor ◽  
Retinoblastoma Tumor

scholarly journals Increase in β-Cell Mass in Transplanted Porcine Neonatal Pancreatic Cell Clusters Is Due to Proliferation of β-Cells and Differentiation of Duct Cells*

Endocrinology ◽  
2001 ◽  
Vol 142 (5) ◽  
pp. 2115-2122 ◽  
Author(s):  
N. Trivedi ◽  
J. Hollister-Lock ◽  
M. D. Lopez-Avalos ◽  
J. J. O’Neil ◽  
M. Keegan ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cell Mass ◽  
Insulin Content ◽  
Double Immunostaining ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Cell ◽  
Cell Clusters ◽  
Duct Cells ◽  
Neonatal Pig

Abstract A 20-fold increase in β-cell mass has been found after transplantation of porcine neonatal pancreatic cell clusters (NPCCs). Here the mechanisms leading to this increased β-cell mass were studied. NPCCs (4000 islet equivalents) generated after 8 days culture of digested neonatal pig pancreas were transplanted beneath the renal capsule of streptozotocin (STZ) diabetic and normoglycemic nude mice. Grafts were removed at 10 days, 6 weeks, and 20 weeks after transplantation for immunostaining and insulin content. Proliferation of β-cells and duct cells was assessed morphometrically using double immunostaining for Ki-67 with insulin or cytokeratin 7 (CK7). Graft maturation was assessed with double immunostaining of CK7 and insulin. Apoptosis was determined using propidium iodide staining. β-cell proliferation in NPCCs was higher after 8 days of culture compared with that found in neonatal pig pancreas. After transplantation, β-cell proliferation remained high at 10 days, decreased somewhat at 6 weeks, and was much lower 20 weeks after transplantation. Diabetic recipients not cured at 6 weeks after transplantation had significantly higherβ -cell proliferation compared with those cured and to normoglycemic recipients. The size of individual β-cells, as determined by cross-sectional area, increased as the grafts matured. Graft insulin content was 20-fold increased at 20 weeks after transplantation compared with 8 days cultured NPCCs. The proliferation index of duct cells was significantly higher in neonatal pig pancreas than in 8 days cultured NPCCs and in 10-day-old grafts. The incidence of apoptosis in duct cells appeared to be low. About 20% of duct cells 10 days post transplantation showed costaining for CK7 and insulin, a marker of protodifferentiation. In conclusion, the increase in β-cell mass after transplantation of NPCCs is due to both proliferation of differentiated β-cells and differentiation of duct cells intoβ -cells.

Accelerated Functional Maturation of Isolated Neonatal Porcine Cell Clusters: In Vitro and In Vivo Results in NOD Mice

2005 ◽  
Vol 14 (5) ◽  
pp. 249-261 ◽  
Author(s):  
Giovanni Luca ◽  
Claudio Nastruzzi ◽  
Mario Calvitti ◽  
Ennio Becchetti ◽  
Tiziano Baroni ◽  
...  
Keyword(s):  
Sertoli Cells ◽  
Time Lag ◽  
Cell Mass ◽  
Nod Mice ◽  
Β Cell ◽  
Cell Clusters ◽  
Porcine Islets ◽  
Neonatal Porcine Islets

Neonatal porcine cell clusters (NPCCs) might replace human for transplant in patients with type 1 diabetes mellitus (T1DM). However, these islets are not immediately functional, due to their incomplete maturation/differentiation. We then have addressed: 1) to assess whether in vitro coculture of islets with homologous Sertoli cells (SC) would shorten NPCCs' functional time lag, by accelerating the β-cell biological maturation/differentiation; 2) to evaluate metabolic outcome of the SC preincubated, and microencapsulated NPCCs, upon graft into spontaneously diabetic NOD mice. The islets, isolated from <3 day piglets, were examined in terms of morphology/viability/function and final yield. SC effects on the islet maturation pathways, both in vitro and in vivo, upon microencapsulation in alginate/poly-L-ornithine, and intraperitoneal graft into spontaneously diabetic NOD mice were determined. Double fluorescence immunolabeling showed increase in β-cell mass for SC+ neonatal porcine islets versus islets alone. In vitro insulin release in response to glucose, as well as mRNA insulin expression, were significantly higher for SC+ neonatal porcine islets compared with control, thereby confirming SC-induced increase in viable and functional β-cell mass. Graft of microencapsulated SC+ neonatal porcine islets versus encapsulated islets alone resulted in significantly longer remission of hyperglycemia in NOD mice. We have preliminarily shown that the in vitro NPCCs' maturation time lag can dramatically be curtailed by coincubating these islets with SC. Graft of microencapsulated neonatal porcine islets, precultured in Sertoli cells, has been proven successful in correcting hyperglycemia in stringent animal model of spontaneous diabetes.

scholarly journals TET1 dioxygenase is required for FOXA2-associated chromatin remodeling in pancreatic beta-cell differentiation

2020 ◽  
Author(s):  
Xinwei Wu ◽  
Jianfang Li ◽  
Minjung Lee ◽  
Qingping Lan ◽  
Jia Li ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Pancreatic Beta Cell ◽  
Embryonic Stem ◽  
Β Cell ◽  
Whole Genome Analysis ◽  
Knock Out ◽  
Pancreatic Progenitors ◽  
Pioneer Transcription Factor ◽  
Unique Cell

AbstractExisting knowledge of the role of epigenetic modifiers in pancreas development has exponentially increased. However, the function of TET dioxygenases in pancreatic endocrine specification remains obscure. We set out to tackle this issue using a human embryonic stem cell (hESC) differentiation system, in which TET1/TET2/TET3 triple knock-out cells displayed severe defects in pancreatic β-cell specification. Integrative whole-genome analysis identified unique cell-type-specific hypermethylated regions (hyper-DMRs) displaying reduced chromatin activity and remarkable enrichment for the binding of FOXA2, a pioneer transcription factor essential for pancreatic endoderm specification. Intriguingly, hundreds of hyper-DMRs recently identified in type-2 diabetes pathogenesis overlapped with the hyper-DMRs we found in TET-deficient cells. Furthermore, transduction of TET1 in TET-deficient cells effectively rescued β-cell differentiation and reversed hypermethylation and suppression of the β-cell determinant PAX4. Genome-wide mapping of TET1 showed that TET1 co-localized at a subset of FOXA2 targets featuring high levels of active chromatin modifications in pancreatic progenitors. Taking these findings together with the defective generation of functional β-cells upon TET1-inactivation, our study not only unveils an essential role of TET1-dependent epigenetic regulation in establishing β-cell identity but also provides a new mechanistic clue regarding the complex crosstalk between TET dioxygenases and pioneer transcription factors in lineage specification.

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