Utilization of commercial collagens for preparing well-differentiated human beta cells for confocal microscopy

Brianna R. Brennecke; USeong Yang; Siming Liu; Fatma S. Ilerisoy; Beyza N. Ilerisoy; Aditya Joglekar; Lucy B. Kim; Spencer J. Peachee; Syreine L. Richtsmeier; Samuel B. Stephens; Edward A. Sander; Stefan Strack; Thomas O. Moninger; James A. Ankrum; Yumi Imai

doi:10.3389/fendo.2023.1187216

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Utilization of commercial collagens for preparing well-differentiated human beta cells for confocal microscopy

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Utilization of commercial collagens for preparing well-differentiated human beta cells for confocal microscopy

Brianna R. Brennecke, USeong Yang, Siming Liu, Fatma S. Ilerisoy, Beyza N. Ilerisoy, Aditya Joglekar, Lucy B. Kim, Spencer J. Peachee, Syreine L. Richtsmeier, Samuel B. Stephens, …

Frontiers in endocrinology (Lausanne), Vol.14, 1187216

05/01/2023

DOI: 10.3389/fendo.2023.1187216

PMCID: PMC10248405

PMID: 37305047

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Abstract

IntroductionWith technical advances, confocal and super-resolution microscopy have become powerful tools to dissect cellular pathophysiology. Cell attachment to glass surfaces compatible with advanced imaging is critical prerequisite but remains a considerable challenge for human beta cells. Recently, Phelps et al. reported that human beta cells plated on type IV collagen (Col IV) and cultured in neuronal medium preserve beta cell characteristics.MethodsWe examined human islet cells plated on two commercial sources of Col IV (C6745 and C5533) and type V collagen (Col V) for differences in cell morphology by confocal microscopy and secretory function by glucose-stimulated insulin secretion (GSIS). Collagens were authenticated by mass spectrometry and fluorescent collagen-binding adhesion protein CNA35. ResultsAll three preparations allowed attachment of beta cells with high nuclear localization of NKX6.1, indicating a well-differentiated status. All collagen preparations supported robust GSIS. However, the morphology of islet cells differed between the 3 preparations. C5533 showed preferable features as an imaging platform with the greatest cell spread and limited stacking of cells followed by Col V and C6745. A significant difference in attachment behavior of C6745 was attributed to the low collagen contents of this preparation indicating importance of authentication of coating material. Human islet cells plated on C5533 showed dynamic changes in mitochondria and lipid droplets (LDs) in response to an uncoupling agent 2-[2-[4-(trifluoromethoxy)phenyl]hydrazinylidene]-propanedinitrile (FCCP) or high glucose + oleic acid. DiscussionAn authenticated preparation of Col IV provides a simple platform to apply advanced imaging for studies of human islet cell function and morphology.

CNA35

lipid droplets

mitochondria

type IV collagen

type V collagen

Details

Title: Subtitle: Utilization of commercial collagens for preparing well-differentiated human beta cells for confocal microscopy
Creators: Brianna R. Brennecke - Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, IA, United States
USeong Yang - Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, IA, United States
Siming Liu - University of Iowa
Fatma S. Ilerisoy - Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, IA, United States
Beyza N. Ilerisoy - Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, IA, United States
Aditya Joglekar - Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, IA, United States
Lucy B. Kim - Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, IA, United States
Spencer J. Peachee - Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, IA, United States
Syreine L. Richtsmeier - Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, IA, United States
Samuel B. Stephens - University of Iowa
Edward A. Sander - University of Iowa
Stefan Strack - University of Iowa
Thomas O. Moninger - University of Iowa
James A. Ankrum - University of Iowa
Yumi Imai - University of Iowa
Resource Type: Journal article
Publication Details: Frontiers in endocrinology (Lausanne), Vol.14, 1187216
DOI: 10.3389/fendo.2023.1187216
PMID: 37305047
PMCID: PMC10248405
NLM abbreviation: Front Endocrinol (Lausanne)
eISSN: 1664-2392
Publisher: Frontiers Media S.A
Grant note: DOI: 10.13039/100000002, name: National Institutes of Health, award: R01-DK090490, U24DK098085, S10 RR025439-01; DOI: 10.13039/100000005, name: U.S. Department of Defense, award: W81XWH-20-1-0200; DOI: 10.13039/100000738, name: U.S. Department of Veterans Affairs, award: I01 BX005107
Language: English
Date published: 05/01/2023
Academic Unit: Roy J. Carver Department of Biomedical Engineering; Pathology; Iowa Neuroscience Institute; Orthopedics and Rehabilitation; Craniofacial Anomalies Research Center; Fraternal Order of Eagles Diabetes Research Center; Neuroscience and Pharmacology; Chemical and Biochemical Engineering; Endocrinology and Metabolism; Internal Medicine
Record Identifier: 9984421702602771

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