Table 1.

Effect of Hypoxia on Proliferation and Differentiation Properties of ESC, hMSC, and ADSC

Cell Type	Species	Oxygen Tension	Cultivation Conditions	Expansion (compared to 20% O2)	Differentiation (compared to 20% O2)	Reference
ESC	Human	1%, 3%, 5%	Monolayer culture of ES cells and embryoid body formation	3 and 5% : ○ proliferation rate of ES colonies 1% : Slightly ↓ proliferation of ES colonies	Better maintenance of undifferentiated state ↑ EB formation, ↑ SSEA-4, ↑Oct-4, ↓SSEA-1	Ezashi et al.50
		2%	Monolayer culture of ES cells	↑ (Increase in clone recovery)	Smaller and less granular cells ○ SSEA4, SSEA1, TRA-1-60, or TRA-1-81 ↓ chromosomal aberrations	Forsyth et al.60
	Mouse	1%	EB formation	No changes in first 5 days	↑ mRNA levels of Aldolase A and VEGF	Gassman et al.52
		5%, 20%, 40%	Monolayer culture of ES cells and EB formation	Suppressed proliferation of ES cells at 5% in comparison to 20 and 40% O2	5% and 20%: ↓ specific AP activity in ES cells 40%: ○ AP activity in ES cells, ↑ Oct-4 expression in comparison to 5 and 20% (ES cells as well as EBs)	Kurosawa et al.51
Adipose-derived MSC	Human	5%	Expansion in monolayer or/and alginate beads in chondrogenic medium: up to 14 days	○ proliferation in expansion medium ↓ proliferation in chondrogenic cultures	Chondrogenesis: ↑ GAG synthesis at certain time points; ○ collagen type II ↑ protein content, ↑ total collagen synthesis	Wang et al.74
		2%	Monolayer; osteogenic and adipogenic differentiation	N/A	Adipogenesis: No intracellular lipid droplets Osteogenesis: No mineralization	Lee and Kemp75
		5%	Cell aggregate culture in chondrogenic medium, 14 days.	↓ cell proliferation	Chondrogenesis: ↑ GAG content, ↑collagen type II, ↑IX, ↑XI, ↑ SOX6, ↑ SOX5, ↑ SOX9, ↑ HIF2α not HIF1α	Khan et al.76
		5%	Elastin-like polypeptide scaffold; 7 days DMEM/F-12 supplemented with ascorbate 2-phosphate	N/A	Chondrogenesis: ↓ expression of Collagen type II, ↑ expression of SOX9, ↓ expression of Collagen X	Betre et al.77
	Mouse	2%	Micromass culture in chondrogenic medium; up to 12 days	↑ cell proliferation	Chondrogenesis: ↓ collagen type II, ↓ GAG content	Malladi et al.78
			Monolayer culture in osteogenic medium; up to 3 weeks		Osteogenesis: ↓ AP activity, ↓ extent of mineralization
			Monolayer in basal medium Micromass culture in chondrogenic medium in normoxia; up to 6 days	↑ cell proliferation	Chondrogenesis: ↑ PG nodule formation, Micromasses rapidly condensed	Xu et al.79
			Monolayer; Osteogenic differentiation in normoxia; up to 3 weeks		Osteogenesis: ↓ AP activity, ↓ extent of mineralization
BM-derived MSC	Human	2%	3D PET Scaffolds (4 weeks) Osteogenic and adipogenic differentiation at 20 % O2	Slower proliferation during initial period but maintained expansion phase longer	↑ Oct-4, ↑ Rex-1 expression, ↑ CFU-F Osteogenesis: ↑ Mineralization (von Kossa) ↑ AP activity (max. levels) ○ Osteonectin Adipogenesis: ↑ LPL expression	Grayson et al.64
			Monolayer; Osteogenic and adipogenic differentiation at 20 % O2	↑ (30 times greater expansion over 6 passages)	↑ Oct-4, ↑ HIF-2α, ↑ connexin-34 expression Osteogenesis: ○ mineralization (von Kossa), ○ AP activity Adipogenesis: ○ lipid vacuole formation	Grayson et al.61
		3%	Long term monolayer culturesfollowed by osteogenic, adipogenic differentiation.	↑ proliferative lifespan over 100 days. Delayed onset of senescent phenotype (↓ AGE)	○ expression of hypoxia related genes: HIF-1α, PH-4, HIF1AN, VHL, Hyou1, HIG1, HIG2. Osteogenesis: ↓ calcium, ↓ AP mRNA, ↓ IBSP mRNA. Adipogenesis: ↓ lipid-droplet containing cells, ↓ FABP4, ↓ LPL Osteogenic and adipogenic potential regained when cells induced in normoxia.	Fehrer et al.67
		1.5%	Monolayer cultures exposed to hypoxia for 24 h, followed by osteogenic (14 days), adipogenic (21 days) and chondrogenic (21 day-pellets) differentiation in normoxia	↑ rate of cell division and proliferation	↑HIF1α, ↑BNIP3, ↑VEGFA, ↑AK3L1/L2, ↑STC1, ↑EPHA3, ↓EDG1expression Osteogenesis: ○ mineralization, ○ RUNX2 expression Adipogenesis: ○ number of lipid droplets, ○ (PPAR)-α Chondrogenesis: ↑ pellet size, ↑SOX9 expression	Martin-Rendon et al.65
		4%	Monolayer cultures exposed to hypoxia for 48 h, followed by osteogenic differentiation (up to 28 days)	Cell survival not affected when lasted up to 72 h. Cell death (35%) occurred when cultured for 120 h.	↑ bFGF mRNA but ↓bFGF protein secretion, ↑VEGF mRNA, ↑VEGF protein secretion, ○ IL-6, MCP-1, TIMP-1, TIMP-2 Osteogenesis: ○ AP expression, ○ BMP-2, ○ BSP, slightly ↓ expression of cbfa-1/Runx2 and Osteocalcin (day 0 and 14); no changes at day 28, ↑ Osteopontin, ↓ Collagen type I expression	Potier et al.25
	Mouse	8%	Monolayer cultures in expansion medium (IMDM) and adipogenic differentiation	↑ cell proliferation (2.8 fold increase in cell number after 8 days)	↑ in number of AP positive, flattened cells, ↑HIF-1α, ↑ H-2Dd, ↑ CD44, ○ IA-d and CD13, ↓Oct4, ○ VEGF expression Adipogenesis: ↑ in number of lipid droplets (5.6 fold)	Ren et al.66
	Rat	5%	Monolayer culture, osteogenic differentiation; in parallel ceramic scaffolds implanted in vivo (up to 6 weeks)	↑ number of colonies	Osteogenesis in vitro: ↑ AP activity, ↑ calcium content, ↑ mineralization (von Kossa) Osteogenesis in vivo: ↑ bone formation	Lennon et al.63

AGE, advanced glycation endproducts; SSEA, stage-specific embryonic antigen; ES, embryonic stem; EB, embryoid body; VEGF, vascular endothelial growth factor; AP, alkaline phosphatase; GAG, glycosaminoglycans; PG, proteoglycan; HIF, hypoxia-inducible factor; BSP, bone sialoprotein; PPAR, peroxisome proliferator-activated receptor; BMP, bone morphogenetic protein; TIMP, tissue inhibitor of metalloproteinase; MCP, monocyte chemoattractant protein; IL, interleukin; bFGF, basic fibroblast growth factor; EDG, endothelial differentiation gene; STC, stanniocalcin; LPL, lipoprotein lipase; FABP4, fatty acid–binding protein; HIG, hypoxia-inducible gene; Hyou, hypoxia upregulated.

↑, increase;

↓, decrease;

○, unchanged (or change not quantified).