Suppression of Epithelial-to-Mesenchymal Transitioning Enhances Ex Vivo Reprogramming of Human Exocrine Pancreatic Tissue Toward Functional Insulin-Producing β-Like Cells

Lima, Maria João; Muir, Kenneth; Docherty, Hilary M.; Drummond, Robert J.; McGowan, Neil; Forbes, Shareen; Heremans, Yves; Houbracken, Isabelle; Ross, James A.; Forbes, Stuart J.; Ravassard, Philippe; Heimberg, Harry; Casey, John; Docherty, Kevin

doi:10.2337/db12-1256

Cited by 58 publications

(69 citation statements)

References 49 publications

(53 reference statements)

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“…More recently, the above results were confirmed in vitro using first the AR42J acinar cell line (113,114), and subsequently primary human pancreatic exocrine cells cultured in conditions that prevented epithelial-tomesenchymal transition (114). While not as ready as hematopoietic stem cells for clinical prime time, reprogramming of human exocrine tissues offers a most promising alternative to the use of stem cells, especially considering that the exocrine compartment of the pancreas makes up to 95% of the pancreas and is now routinely discarded after every clinical islet preparation.…”

Section: Cell Reprogrammingsupporting

confidence: 63%

Cell Replacement Strategies Aimed at Reconstitution of the β-Cell Compartment in Type 1 Diabetes

et al. 2014

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Emerging technologies in regenerative medicine have the potential to restore the b-cell compartment in diabetic patients, thereby overcoming the inadequacies of current treatment strategies and organ supply. Novel approaches include: 1) Encapsulation technology that protects islet transplants from host immune surveillance; 2) stem cell therapies and cellular reprogramming, which seek to regenerate the depleted b-cell compartment; and 3) whole-organ bioengineering, which capitalizes on the innate properties of the pancreas extracellular matrix to drive cellular repopulation. Collaborative efforts across these subfields of regenerative medicine seek to ultimately produce a bioengineered pancreas capable of restoring endocrine function in patients with insulin-dependent diabetes.Regenerative medicine promises to contribute to the advancement of b-cell replacement strategies through the development and implementation of microencapsulation technology, the production of insulin-producing cells either by regeneration from endogenous cells or reprogramming from nonendocrine adult cell sources, and, more recently, the exploitation of bioengineered microenvironments (1). Considering the shortage of available pancreata, the search for alternative cell sources has recently been categorized within one of the following three "Rs" of pancreatic b-cell replenishment: replacement (from stem cells or xenoislets), regeneration (from endogenous progenitor cells), and reprogramming (from nonendocrine adult cell sources) (2).The purpose of this article is to comprehensively and critically review the main regenerative medicine2based strategies that are currently being developed to treat type 1 diabetes. The first part of the article will illustrate and discuss islet encapsulation technology, which has been extensively investigated over the past 40 years (3) and represents one of the most promising regenerative medicine2based approaches to achieve immunoisolation of transplantable organs. The second part will focus on cell bioengineering, where different types of progenitor and differentiated cell sources are manipulated to ultimately yield insulin-producing cells. The last part of the article will summarize the most recent advancements in the study of the pancreatic extracellular matrix (ECM) as a platform for endocrine pancreas bioengineering. ISLET ENCAPSULATION TECHNOLOGYEncapsulation technology has been used to protect islets from the host immune system. This strategy holds the promise to allow implantation of islets without immunosuppression not only across genetically different individuals from the same species, but also across species barriers. MicroencapsulationThe first approach to immunoisolate cells consists of the microencapsulation of one to three islets per semipermeable immunoprotective capsule. The spherical configuration of these microcapsules results in a higher surface-tovolume ratio and a higher diffusion rate (4). Furthermore, microcapsules can be injected in large numbers, are durable, and are difficult to disrup...

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Section: Cell Reprogrammingsupporting

confidence: 63%

Cell Replacement Strategies Aimed at Reconstitution of the β-Cell Compartment in Type 1 Diabetes

et al. 2014

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“…Reprogramming variability is also observed in exocrine cell populations. In fact, the incapacity for b-cell differentiation of contaminating stromal cells arising from unselected exocrine cultures was recently described using transcription factor-based protocols (Lima et al, 2013). Hence, we were able to derive insulin + cells from HDDCs but not from freshly plated DCs or from stromal cells.…”

Section: Discussionmentioning

confidence: 83%

β-Cell Differentiation of Human Pancreatic Duct–Derived Cells AfterIn VitroExpansion

Corritore

Dugnani

Pasquale

et al. 2014

Cellular Reprogramming

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β-Cell replacement therapy is a promising field of research that is currently evaluating new sources of cells for clinical use. Pancreatic epithelial cells are potent candidates for β-cell engineering, but their large-scale expansion has not been evidenced yet. Here we describe the efficient expansion and β-cell differentiation of purified human pancreatic duct cells (DCs). When cultured in endothelial growth-promoting media, purified CA19-9(+) cells proliferated extensively and achieved up to 22 population doublings over nine passages. While proliferating, human pancreatic duct-derived cells (HDDCs) downregulated most DC markers, but they retained low CK19 and SOX9 gene expression. HDDCs acquired mesenchymal features but differed from fibroblasts or pancreatic stromal cells. Coexpression of duct and mesenchymal markers suggested that HDDCs were derived from DCs via a partial epithelial-to-mesenchymal transition (EMT). This was supported by the blockade of HDDC appearance in CA19-9(+) cell cultures after incubation with the EMT inhibitor A83-01. After a differentiation protocol mimicking pancreatic development, HDDC populations contained about 2% of immature insulin-producing cells and showed glucose-unresponsive insulin secretion. Downregulation of the mesenchymal phenotype improved β-cell gene expression profile of differentiated HDDCs without affecting insulin protein expression and secretion. We show that pancreatic ducts represent a new source for engineering large amounts of β-like-cells with potential for treating diabetes.

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“…A similar observation was also reported in the transformation of mesenchymal cells. Glucagonsecreting alpha-like cells were generated by transfecting monolayer mesenchymal cells derived from pancreatic tissue exocrine with adenoviruses containing Pdx1, Ngn3, MafA, and Pax4 genes (Lima, 2013). This study also suggested that the formation of epithelial-like clusters of exocrine-derived mesenchymal stromal cells promotes the generation of IPCs.…”

Section: Discussionmentioning

confidence: 99%

Improved insulin-secreting properties of pancreatic islet mesenchymalstem cells by constitutive expression of Pax4 and MafA

Açıksarı¹,

Duruksu²,

Karaöz³

2017

Turk J Biol

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For long-term treatment of diabetes type 1, transplantation of insulin-producing beta cells may be a promising method, but the limited number of islets for transplantation requires the development of different approaches. In this study, we aimed to generate betalike insulin-producing cells. For this purpose, MafA, Pax4, and Ngn3 genes were transferred into pancreatic islet-derived mesenchymal stem cells, and the effect of their ectopic expressions on differentiation efficiency was examined. Stemness properties of pancreatic islet stem cells were characterized. The 3 genes were transfected by electroporation and expressed constitutively. The transfected cells were further stimulated to differentiate by using chemical induction. Pax4 expression had significant effects on differentiation into insulin-producing cells. Although it caused morphological alterations in cells, similar to epithelial cells, the insulin secretion levels remained lower than those of the cell line cotransfected with MafA and Pax4. Cotransfection of the 3 transcription factors did not further improve the beta-like cell generation. MafA and Pax4 ectopic expression resulted in improved differentiation efficiency into insulin-secreting cells. However, support of this differentiation process using additional chemical induction may suffice to overcome control by endogenous regulatory pathways.

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Suppression of Epithelial-to-Mesenchymal Transitioning Enhances Ex Vivo Reprogramming of Human Exocrine Pancreatic Tissue Toward Functional Insulin-Producing β-Like Cells

Cited by 58 publications

References 49 publications

Cell Replacement Strategies Aimed at Reconstitution of the β-Cell Compartment in Type 1 Diabetes

Cell Replacement Strategies Aimed at Reconstitution of the β-Cell Compartment in Type 1 Diabetes

β-Cell Differentiation of Human Pancreatic Duct–Derived Cells AfterIn VitroExpansion

Improved insulin-secreting properties of pancreatic islet mesenchymalstem cells by constitutive expression of Pax4 and MafA

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