title: Characterization of pancreatic ductal cells in human islet preparations.
abstract: Substantial amounts of nonendocrine cells are implanted as part of human islet grafts, and a possible influence of nonendocrine cells on clinical islet transplantation outcome has been postulated. There are currently no product release criteria specific for nonendocrine cells due to lack of available methods. The aims of this study were to develop a method for the evaluation of pancreatic ductal cells (PDCs) for clinical islet transplantation and to characterize them regarding phenotype, viability, and function. We assessed 161 human islet preparations using laser scanning cytometry (LSC/iCys) for phenotypic analysis of nonendocrine cells and flow cytometry (FACS) for PDC viability. PDC and beta-cells obtained from different density fractions during the islet cell purification were compared in terms of viability. Furthermore, we examined PDC ability to produce proinflammatory cytokines/chemokines, vascular endothelial growth factor (VEGF) and tissue factor (TF) relevant to islet graft outcome. Phenotypic analysis by LSC/iCys indicated that single staining for CK19 or CA19-9 was not enough for identifying PDCs, and that double staining for amylase and CK19 or CA19-9 allowed for quantitative evaluation of acinar cells and PDC content in human islet preparation. PDC showed a significantly higher viability than beta-cells (PDC vs beta-cell: 75.5+/-13.9 and 62.7+/-18.7%; P<0.0001). Although beta-cell viability was independent of its density, that of PDCs was higher as the density from which they were recovered increased. There was no correlation between PDCs and beta-cell viability (R(2)=0.0078). PDCs sorted from high-density fractions produced significantly higher amounts of proinflammatory mediators and VEGF, but not TF. We conclude that PDCs isolated from different fractions had different viability and functions. The precise characterization and assessment of these cells in addition to beta-cells in human islet cell products may be of assistance in understanding their contribution to islet engraftment and in developing strategies to enhance islet graft function.
authors.affiliation: Cell Transplant Center, Diabetes Research Institute, University of Miami Leonard M. Miller School of Medicine, Miami, FL 33136, USA. hichii@med.miami.edu
authors.forename: Hirohito
authors.initials: H
authors.lastname: Ichii
authors.forename: Atsushi
authors.initials: A
authors.lastname: Miki
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Yamamoto
authors.forename: Ruth D
authors.initials: RD
authors.lastname: Molano
authors.forename: Scott
authors.initials: S
authors.lastname: Barker
authors.forename: Atsuyoshi
authors.initials: A
authors.lastname: Mita
authors.forename: Rayner
authors.initials: R
authors.lastname: Rodriguez-Diaz
authors.forename: Dagmar
authors.initials: D
authors.lastname: Klein
authors.forename: Ricardo
authors.initials: R
authors.lastname: Pastori
authors.forename: Rodolfo
authors.initials: R
authors.lastname: Alejandro
authors.forename: Luca
authors.initials: L
authors.lastname: Inverardi
authors.forename: Antonello
authors.initials: A
authors.lastname: Pileggi
authors.forename: Camillo
authors.initials: C
authors.lastname: Ricordi
mesh_terms: D000818:Animals; D018395:CA-19-9 Antigen; D003921:Diabetes Mellitus, Experimental; D006801:Humans; D050417:Insulin-Secreting Cells / Q000145:classification / Q000166:cytology*; D007515:Islets of Langerhans / Q000166:cytology; D016381:Islets of Langerhans Transplantation; D053539:Keratin-19; D047569:Laser Scanning Cytometry; D051379:Mice; D008819:Mice, Nude; D010183:Pancreatic Ducts / Q000166:cytology*; D010641:Phenotype
chemical_list: D018395:CA-19-9 Antigen; D053539:Keratin-19
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