Table 2.
Effect of hypoxia on MSC viability, proliferation, and clonogenicity.
| Treatment conditions | Types of stem cells | The effect compared to normoxia (methods of analysis) | Ref. | |
|---|---|---|---|---|
| O2 concentration | Time/passage | |||
| 1% | 2 d | hBM-MSC | Proliferation (DNA Quant-iT Picrogreen assay), clonogenicity (Giemsa staining), and metabolic activity (Vybrant assay) increased; HIF-1α downregulated (qRT-PCR), the proapoptotic genes: BAX, BCL-2, and CASP-3 downregulated (qRT-PCR) | [57] |
| 1% | 2 d | rBM-MSC | The proliferation decreased (Trypan Blue staining, cell count) | [80] |
| 1% | 7 d | hBM-MSC | Proliferation significantly reduced (MTS proliferation assay) | [58] |
| 1% | 7 d | hBM-MSC | HIF-1α upregulated three-folds (qRT-PCR) | [58] |
| 1% | 9 d | hAD-MSC | Proliferation increased 1.7-folds (Trypan Blue staining, cell count) | [62] |
| 1% | 10 d | hBM-MSC | Proliferation (DNA Quant-iT Picrogreen assay) decreased, and metabolic activity increased (Vybrant assay), HIF-1α downregulated (qRT-PCR), the proapoptotic genes BCL-2 and CASP-3 downregulated, BAX upregulated (qRT-PCR) | [57] |
| 1% | 14 d | mBM-MSC | Viability (MTT viability assay) and proliferation (BrdU cell proliferation assay) increased, the main metabolic regulators like Hk2 upregulated (sqRT-PCR), shift to anaerobic glycolysis, the Slc16a3 (MCT-4) gene upregulated under prolonged hypoxia (qRT-PCT), the MCT-4 level increased under prolonged hypoxia (WB) | [68] |
| 1% | 14 d | rBM-MSC | Clonogenicity increased (crystal violet staining) | [80] |
| 1% | 21 d | hAD-MSC | Cell aging reduced, telomeres longer 1.5-folds (qPCR) | [62] |
| 1% | 21 d | hBM-MSC | A slowdown of cell cycle progression, accumulation in G1 phase under prolonged hypoxia (flow cytometry) | [58] |
| 1-3% | 16 h | hBM-MSC | Viability and proliferation (flow cytometry) maintained, Akt signaling pathway activated (WB) | [81] |
| 1.5% | 1 d | hBM-MSC hUCB-MSC |
Proliferation increased (Trypan Blue staining, cell count) and the cell cycle faster progression (flow cytometry), HIF-1α increased (WB) | [82] |
| 1.5% | 3 d | hUC-MSC | Proliferation decreased (Trypan Blue staining, cell count), LDHA, GLUT-1, and PDK-1 upregulated (RT-PCR), glutamate production decreased (HPLC), glucose consumption significantly increased (YSI 2700 analyzer) | [69] |
| 2% | 2 d | hBM-MSC | Proliferation (DNA Quant-iT Picrogreen assay), clonogenicity (Giemsa staining), and viability (flow cytometry) increased | [57] |
| 2% | 2 d | hWJ-MSC | Expression of the genes HIF1-α, HIF-2α, Notch2, and JAGGED1 increased (RT-PCR) | [64] |
| 2% | 7 d | hBM-MSC | A high growth rate maintained even after confluency–multilayer formation (cell count, growth curve), HIF-2α upregulated (RT-PCR) | [38] |
| 2% | 7 d | hBM-MSC | Clonogenicity increased (crystal violet staining) | [63] |
| 2% | 12 d | hBM-MSC | Higher proliferation rate (Trypan Blue staining, cell count), the number of actively dividing cells significantly increased (PKH26 Red Fluorescent Cell Linker kit), the cellular division started earlier in the cell cycle (PKH26 staining, flow cytometry) | [63] |
| 2% | 20 d | hBM-MSC | Clonogenicity (colony count from microscopic images) and doubling time (cell count and growth curve) maintained, cellular senescence reduced (β-galactosidase staining, histochemistry) | [71] |
| 2% | Passages 2-7 | hBM-MSC | Higher cell number in each passage from 2 to 7 (Trypan Blue staining, cell count) | [38] |
| 2% | 10 passages | hWJ-MSC | Faster growth rates and higher total cell number yielded (cell area count, image analysis), normal karyotype maintained (Giemsa staining) | [64] |
| 2% | 64 d | hBM-MSC | Homogenous morphology of rapidly self-renewing cells maintained up to 52 d (microscopy analysis) | [83] |
| 2.5% | 3 d | hUC-MSC | Proliferation increased (cell counting under a microscope), HIF-1α increased (WB), PDK-1, GLUT-1, and LDHA upregulated (RT-PCR), glutamate production diminished (HPLC), glucose consumption significantly increased (YSI 2700 SELECT analyzer) | [69] |
| 2.5% ∗ | >3 d∗ | hUCB-MSC | Cell viability (at 24 h and 2 d) increased (Trypan Blue staining, cell count, and MTT); proliferation (at 3 d) increased (Trypan Blue staining, cell count), CFU-F number in vitro significantly enhanced (Giemsa staining) | [60] |
| 2.5% ∗ | >3 d∗ | hUCB-MSCs | Cell metabolic activity (MTT), CFU-F number (Giemsa staining), and proliferation (at 2 and 3 d) (Trypan Blue staining, cell count) increased, doubling time reduced (at 2 and 3 d) (Trypan Blue staining, cell count), cell death inhibited (at 2 and 3 d) (microscope analysis) | [84] |
| 3% | ~100 d Passage 1 |
hBM-MSC | Proliferative lifespan with additional 10 PD improved (flow cytometry), transcription of hypoxia-related the genes encoding VHL, HIF-1, PH-4, HYOU1, HIF1AN, HIG, and HIG unaltered (qPCR) | [33] |
| 3% | Over 25 passages | hBM-MSC | Cell growth improved (Trypan Blue staining, cell count), population doublings increased (Trypan Blue staining, cell count), oxidative stress reactions (DHE, flow cytometry) and nuclear alterations such as damage of DNA, telomere shortening, and chromosomal abnormalities (DAPI, Q-FISH, Breast Aneusomy Multicolor Probe kit) limited, glycolysis increased (OCR/ECAR, F96 Flux analyzer) | [32] |
| 5% | 2 d | hBM-MSC | Proliferation rate lowered (DNA Quant-iT Picrogreen assay), clonogenicity (Giemsa staining), and metabolic activity elevated (Vybrant assay) | [57] |
| 5% | 3 d | hUC-MSC | Proliferation increased (Trypan Blue staining, cell count), LDHA, PDK-1, and GLUT-1 upregulation (RT-PCR) | [69] |
| 5% | 4 d | rBM-MSC | Proliferation rate increased (flow cytometry) | [85] |
| 5% | 4 d | hBM-MSC | Clonogenicity (crystal violet staining), proliferation (EDU Proliferation kit), and metabolic activity (Alamar Blue staining) increased | [65] |
| 5% | 14 d | hBM-MSC | Clonogenicity decreased at primary cells and the passage 1 but increased at the passages 2 and 3 (crystal violet staining) | [66] |
| 5% | 20 d | hBM-MSC | Colony formation significantly reduced (colony count from microscopic images), doubling time maintained (Trypan Blue staining, cell count, growth curve), cellular senescence reduced (β-galactosidase staining, histochemistry, blue stained cell count) | [71] |
| 5% | Passage 1-10 | hBM-MSC | The number of population doublings increased (Trypan Blue staining, cell count), cellular senescence reduced (β-galactosidase staining, histochemistry, blue stained cell count) | [72] |
∗Hypoxic preconditioning in 2.5% O2 for 15 minutes, then reoxygenation at 21% O2 for 30 minutes, and again hypoxia preconditioning at 2.5% O2 for 3 days; d: day/days; h: human; m: mouse; r: rat; PD: population doublings; MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; BrdU: 5-bromo tetrazolium inner salt-20-deoxyuridine; MTS: tetrazolium inner salt; WB: Western Blotting.