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Biomed Eng Online. 2018 Jul 31;17(1):102. doi: 10.1186/s12938-018-0535-z.

Wound healing of human embryonic stem cell-derived retinal pigment epithelial cells is affected by maturation stage.

Author information

1
Faculty of Biomedical Sciences and Engineering, BioMediTech, Tampere University of Technology, Arvo Ylpön Katu 34, Tampere, Finland.
2
Faculty of Medical and Life Sciences, BioMediTech, University of Tampere, Arvo Ylpön Katu 34, Tampere, Finland.
3
Department of Ophthalmology, Institute of Clinical Medicine, University of Eastern Finland, Kuopio, Finland.
4
Department of Ophthalmology, Kuopio University Hospital, Kuopio, Finland.
5
Faculty of Medical and Life Sciences, BioMediTech, University of Tampere, Arvo Ylpön Katu 34, Tampere, Finland. kati.juuti-uusitalo@staff.uta.fi.

Abstract

BACKGROUND:

Wound healing of retinal pigment epithelium (RPE) is a complex process that may take place in common age-related macular degeneration eye disease. The purpose of this study was to evaluate whether wounding and wound healing has an effect on Ca2+ dynamics in human embryonic stem cell (hESC)-RPEs cultured different periods of time.

METHODS:

The 9-day-cultured or 28-day-cultured hESC-RPEs from two different cell lines were wounded and the dynamics of spontaneous and mechanically induced intracellular Ca2+ activity was measured with live-cell Ca2+ imaging either immediately or 7 days after wounding. The healing time and speed were analyzed with time-lapse bright field microscopy. The Ca2+ activity and healing speed were analysed with image analysis. In addition the extracellular matrix deposition was assessed with confocal microscopy.

RESULTS:

The Ca2+ dynamics in hESC-RPE monolayers differed depending on the culture time: 9-day-cultured cells had higher number of cells with spontaneous Ca2+ activity close to freshly wounded edge compared to control areas, whereas in 28-day-cultured cells there was no difference in wounded and control areas. The 28-day-cultured, wounded and 7-day-healed hESC-RPEs produced wide-spreading intercellular Ca2+ waves upon mechanical stimulation, while in controls propagation was restricted. Most importantly, both wave spreading and spontaneous Ca2+ activity of cells within the healed area, as well as the cell morphology of 28-day-cultured, wounded and thereafter 7-day-healed areas resembled the 9-day-cultured hESC-RPEs.

CONCLUSIONS:

This acquired knowledge about Ca2+ dynamics of wounded hESC-RPE monolayers is important for understanding the dynamics of RPE wound healing, and could offer a reliable functionality test for RPE cells. The data presented in here suggests that assessment of Ca2+ dynamics analysed with image analysis could be used as a reliable non-invasive functionality test for RPE cells.

KEYWORDS:

Ca2+ waves; Cell maturation; Image analysis; Mechanical stimulation; Mechanically induced intercellular Ca2+ waves; RPE; Spontaneous [Ca2+]i increases; Wound healing; hESC-RPE

PMID:
30064430
PMCID:
PMC6069779
DOI:
10.1186/s12938-018-0535-z
[Indexed for MEDLINE]
Free PMC Article

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