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ARS Home » Northeast Area » Beltsville, Maryland (BARC) » Beltsville Agricultural Research Center » Animal Biosciences & Biotechnology Laboratory » Research » Publications at this Location » Publication #333615

Research Project: DEVELOPING GENETIC BIOTECHNOLOGIES FOR INCREASED FOOD ANIMAL PRODUCTION, INCLUDING NOVEL ANTIMICROBIALS FOR IMPROVED HEALTH & PRODUCT SAFETY

Location: Animal Biosciences & Biotechnology Laboratory

Title: Derivation and characterization of a pig embryonic stem cell-derived exocrine pancreatic cell line

Author
item Talbot, Neil
item Garrett, Wesley
item Shannon, Amy
item Phillips, Caitlin

Submitted to: Pancreas
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/6/2017
Publication Date: N/A
Citation: N/A

Interpretive Summary: The establishment and initial characterization of a pig embryonic stem cell-derived pancreatic cell line, PICM-31, is described. The PICM-31 cell line was started from the in vitro culture of the inner cells of an early pig embryo. The cells were small and grew relatively slowly. They frequently self-organized into radially symmetrical groupings of cells, i.e., wheel-like and similar to the acinar structures inside the pancreas. Electron microscopic examination confirmed that the cells were organized into a functional acinar-like unit and that they contained secretory granules as expected of the cells of the pancreas that secrete digestive enzymes. The PICM-31 cells were shown to be expressing the genes for digestive enzymes, and the secreted enzymes themselves were also identified by mass spectroscopy. The PICM-31 cells also expressed 'marker' genes that indicated the cells may be 'progenitor' pancreas cells, i.e., that the PICM-31 cells might be able to change into the “endocrine” cells of the pancreas given the right conditions. Endocrine pancreas cells include the beta-cells that are responsible for making and secreting insulin. Lack of insulin secretion is a key problem in some forms of diabetes, so if the PICM-31 cells could be induced to become beta-cells, the cell line might be a source of insulin for diabetic people. The PICM-31 cell line can be used to study and develop useful genetic metabolic changes in pigs, and they are the first demonstration of normal pig pancreas cells that can be grown in culture.

Technical Abstract: The establishment and initial characterization of a pig embryonic stem cell-derived pancreatic cell line, PICM-31, and a colony-cloned derivative cell line, PICM-31A, is described. The cell lines were propagated for several months at split ratios of 1:3 or 1:5 at each passage on STO feeder cells after establishment. The cells were relatively small, grew slowly (72-96 h doubling time), and at approximately one week post-passage, they frequently self-organized into radially symmetrical groupings of cells, i.e., acinar-like arrangements. Electron microscopic examination showed the acinar-like structures to be organized units of cells joined by tight-junction-like cell-to-cell unions at their apical aspect. The cells' apical surface had microvilli projecting into the acinar-like space and, most notably, the cells contained prominent, dark secretory granules as well as granules with a dense core surrounded by an unstained “halo”. The PICM-31 and -31A cells were positive by RT-PCR analysis for the expression of exocrine pancreas secretory products (digestive enzymes) and transthyreitin, a protein produced by pancreatic alpha-cells. Also, a number of transcription factors typical of pancreatic progenitor cells were expressed by the cells. Cellular proteomic analysis confirmed the production of digestive enzymes by the PICM-31 and -31A cells and identified over 2658 pig proteins that were semi-quantitatively ranked for relative abundance by spectral count analysis. The secreted pancreatic enzyme, pig carboxypeptidase A1 (CPA1), was the second most abundant protein in the cells as specified by spectral count ranking. The cell lines provide in vitro models of fetal pig pancreatic exocrine cells, and perhaps pancreatic progenitor cells, and they are the first demonstration of continuous cultures of normal pig pancreatic cells as cell lines.