Potency assays for human adipose-derived stem cells as a medicinal product toward wound healing

Table 3 In vitro biological potency assays testing the effects of hASCs in the proliferation phase of wound healing

MoA	Target	Assay	References
Granulation tissue formation by fibroblasts
Proliferation	Dehydrogenases activity	CCK8	[46, 52,53,54]
	Cell number	Manual cell counting	[55,56,57]
	NAD(P)H-dependent dehydrogenase enzyme activity	MTT	[32, 45, 58, 59]
Viability
Migration	Invasion	Scratch assay	[32, 38, 46, 53, 55, 56, 58, 60,61,62,63]
	Migration	Transwell assay	[38, 45, 46, 53]
ECM production	Collagen I	qPCR	[38, 39, 52, 55,56,57, 62, 63]
		WB	[52, 55, 57, 62]
		IF	[57]
	Collagen III	qPCR	[38, 52, 56, 57, 62]
		WB	[52, 57, 64]
		IF	[57]
	Elastin	qPCR	[38, 63]
		WB	[52]
	Fibronectin	qPCR	[62]
		WB	[52]
ECM modulation	MMP1	qPCR	[57, 62, 65]
		WB	[55]
	MMP2	qPCR	[52, 57, 65]
	TIMP1	ELISA	[65]
		qPCR	[57]
Paracrine milieu	VEGF	qPCR	[52, 63]
	TGF-β	qPCR	[52]
Proliferation		ELISA	[65]
Vascularization by endothelial cells
Proliferation	NAD(P)H-dependent dehydrogenase enzyme activity	MTT	[61, 66]
	Dehydrogenases activity	CCK8	[46, 48, 52, 67]
Migration	Invasion	Transwell assay	[46, 48, 52, 67,68,69]
	Migration	Scratch assay	[46, 48, 52, 61, 66, 67, 70]
Angiogenesis	VEGFA	WB	[52]
		qPCR	[52, 70, 71]
	Sprouting	Aortic ring assay	[68, 72, 73]
	3D organization	Spheroid-based angiogenesis assay	[61, 74]
	2D organization	Tube formation assay	[46, 48, 52, 59, 66,67,68,69, 74,75,76]
Re-epithelialization by keratinocytes
Proliferation	Dehydrogenases activity	CCK8	[52, 53, 77, 78]
	NAD(P)H-dependent dehydrogenase enzyme activity	MTS	[47, 79]
		MTT	[39, 45, 59]
Migration	MMP2	WB	[78]
		qPCR	[52]
	MMP9	WB	[78]
		qPCR	[52]
	Chemotaxis assay		[47, 80]
	Scratch assay		[47, 48, 53, 58, 61, 77,78,79,80,81]
	Transwell assay		[45, 52, 53, 77, 78]

MoA, mode of action; CCK8, cell counting kit-8; NAD(P)H, reduced nicotinamide adenine dinucleotide phosphate; MMT, 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide; qPCR, quantitative polymerase chain reaction; WB, western blotting; IF, immunofluorescence microscopy; MMP-1/2/9, matrix metalloproteinase-1/2/9; TIMP-1, TIMP metallopeptidase inhibitor 1; VEGFA, vascular endothelial growth factor A; TGF-β, transforming growth factor beta; 3/2 D, three/two dimensional, MTS, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium

ISSN: 1757-6512