Table 2.
Ca2+-permeable ion channels regulation by hypoxia.
| Ion Channel | Cell Type | Methodology | Hypoxia Technique and Treatment Time | Effect of Hypoxia on Channel’s Activity/Expression | Effect of Hypoxia on Ca2+ Signals | Cellular Function | Ref. |
|---|---|---|---|---|---|---|---|
| Piezo1 | Mouse and human sickle red blood cells (RBCs) | Cell-attached and nystatin-permeabilized patch clamp Calcium imaging |
Deoxygenation obtained by exposure with a superfusate gassed 30 min prior to the experiment with 100% N2 | Deoxygenation activates a Ca2+- and cation-permeable conductance in a reversible manner, and this current is sensitive to inhibition by GsMTx-4; 1 mM | Increased Ca2+ influx | Not assessed | [94] |
| Pulmonary arterial endothelial cells (PASMCs) of patients with pulmonary arterial hypertension (PAH) | Calcium imaging EdU and cell counting proliferation assay Western Blot |
/ | Piezo1 expression and activity are increased in idiopathic pulmonary arterial hypertension and pulmonary arterial smooth muscle cells | Increased Ca2+ influx and increased intracellular Ca2+ release | Increased PAH-PASMCs’ proliferation | [95] | |
| Pulmonary artery smooth muscle cells of mice and rats’ models with experimental chronic hypoxia-induced pulmonary hypertension (PH) Human pulmonary artery endothelial cells (hPAECs) |
Western Blot Calcium imaging |
Hypoxia induced by incubation in 3% O2 for 4 h–12 h or in 10% O2 for a total of 6 weeks | Piezo1 is significantly upregulated in the lung tissue of PH rats and in chronic hypoxia-induced PH models. Piezo1 protein is transiently upregulated also in hPAECs after 6 h exposition to hypoxic conditions. Hypo-osmotic conditions upregulate Piezo1 protein levels in hPAECs |
Hypo-osmotic upregulation of Piezo1 promotes Ca2+ influx, promoting Akt and Erk signalling pathways activation, with downstream upregulation of Notch ligand | GsMTx4-mediated Piezo1 blockade partially reduces the chronic hypoxia-induced PH in mice with chronic hypoxia-induced pulmonary hypertension | [96] | |
| TRPM2 | TRPM2 WT and knockout (KO) neonatal hypoxic-ischemic (HI) brain injury mouse model | Western Blot | Hypoxia damage was induced in ischemic mice models by incubating the pups in a hypoxic chamber for 2 h | TRPM2 is acutely overexpressed 24 h after hypoxia-ischemic injury in brain tissue samples from mouse pups | Not assessed | Brain damage and inflammation are reduced in TRPM2 KO mice 7 days following hypoxic-ischemic brain injury. TRPM2 inhibits cell survival pathways after HI injury |
[97] |
| Primary cultures of rat cortical neurons subjected to oxidative stress | Calcium imaging Trypan Blue exclusion assay |
Oxidative stress induced by 1 mM or 50 µM H2O2 treatment | Not assessed | H2O2 induces TRPM2-mediated intracellular calcium rise | SiTRPM2 prevents H2O2-mediated neuronal cell death | [98] | |
| TRPM2-overexpressing HEK293 cells | Whole-cell Patch Clamp | Hypoxia induced by cell incubation with gas mixture containing 5% O2 for 30 and 60 min | TRPM2 activation is induced by 30- and 60-min exposure to hypoxic conditions | Not assessed | Hypoxia treatment enhances cell death, probably via TRPM2-mediated Ca2+ influx | [99] | |
| ARPE-19 retinal pigment epithelial cells | Patch Clamp Calcium imaging Propidium iodide cell death assay |
Hypoxia induced by CoCl2 (200 μM) for 24 h | Hypoxia induces activation of TRPM2 currents and upregulates TRPM2 protein levels | Hypoxia induces TRPM2-mediated intracellular calcium rise | Hypoxia causes mitochondrial oxidative cell cytotoxicity and cell death via TRPM2-mediated Ca2+ signals | [100] | |
| Primary IGR39 melanoma cells TRPM2-overexpressing HEK293 cells |
Patch Clamp Calcium imaging |
Treatment with chloramine-T (Chl-T) oxidant agent | Amount of 0.5 mM Chl-T activates TRPM2 in IGR39 and in TRPM2-expressing HEK293 cells | Chl-T treatments induce a significant increase in cytosolic Ca2+ levels | Chl-T-induced TRPM2 activation and increased Ca2+ influx activate BK and KCa3.1 potassium channels | [101] | |
| PC3 prostate cancer cells | Calcium imaging MTT and TUNEL assay |
Treatment with 0.5 to 4 mM H2O2 for 6 h | H2O2 induces TRPM2 activation | H2O2 treatment leads to TRPM2-mediated intracellular Ca2+ increase in a concentration-dependent manner | H2O2 induces TRPM2-Ca2+-CaMKII cascade that promotes ROS production, mitochondrial fragmentation, and inhibition of autophagy, inducing cell death | [102] | |
| TRPM2-L and TRPM2-S-expressing SH-SY5Y neuroblastoma cells | Calcium imaging | Treatment with 250 μM H2O2 for 20 min | Not assessed | H2O2 treatment leads to TRPM2-L-mediated intracellular Ca2+ increase and a decrease in TRPM2-S | TRPM2-L-expressing cells show higher HIF-1/2α levels with respect to TRPM2 short isoform and promote tumour growth in vivo | [103] | |
| Human breast cancer cells | Calcium imaging qPCR |
Co-culture with neutrophils or H2O2 treatment | Neutrophil-derived H2O2 induces decrease in TRPM2 expression in H2O2-selected tumour cells | Not assessed | TRPM2 activation by neutrophil-derived H2O2 and following Ca2+ entry promotes cancer cells’ death | [104] | |
| TRPM6 | Hepatic ischemia-reperfusion rat model | qPCR | Ischemia was obtained by 60 min clamping the left hepatic artery and the portal vein | TRPM6 expression is increased in liver tissue from ischemia-reperfusion rat model | Not assessed | Not assessed | [105] |
| TRPM7 | TRPM7-overexpressing HEK293T cells Cortical neurons |
Ca2+ imaging Patch clamp PI cell death assay |
Hypoxia induced by anaerobic chamber containing ˂0.2% O2 atmosphere for 1, 1.5 and 2 h. | Hypoxia induces TRPM7 channel activation | Hypoxia increases Ca2+ entry | Hypoxia-activated TRPM7 mediated-Ca2+ entry determines cell death in cortical neurons | [106] |
| Hepatic ischemia-reperfusion rat model | qPCR | Ischemia was obtained by 60 min clamping the left hepatic artery and the portal vein | TRPM7 expression is increased in liver tissue from ischemia-reperfusion rat model | Not assessed | Not assessed | [105] | |
| TRPV1 | HEK293T cells overexpressing rat TRPV1 | Patch Clamp Calcium imaging |
Hypoxic solution obtained by bubbling with 100% N2 gas for at least 20 min before the perfusion (PO2, 3%) | Acute hypoxia weakly increases TRPV1 activity, but negatively affects capsaicin induced TRPV1 currents | Hypoxia leads to a slight increase in cytosolic Ca2+ levels | Not assessed | [107] |
| Rat DRG neurons hTRPV1/rTRPV1-expressing HEK293 cells |
Whole-cell patch-clamp | Overnight (18–20 h) exposition to hypoxia (4% O2) | Overnight exposure to hypoxic/high glucose conditions increases TRPV1 mean peak current densities in both cell lines, without affecting its expression | Not assessed | Not assessed | [108] | |
| Rat pulmonary artery smooth muscle cells (PASMCs) | Calcium imaging qPCR Western Blot Wound Healing assay BrdU proliferation assay |
24–48 h long exposition to hypoxia (1% and 10% O2) | Hypoxic conditions do not affect TRPV1 expression, but they increase TRPV1 activity | No assessed | Hypoxia-mediated TRPV1 activation enhances PASMCs migratory abilities and proliferation | [109] | |
| Human pulmonary artery smooth muscle cells (PASMCs) | Calcium imaging qPCR Western Blot Cell count proliferation assay |
72 h long exposition to hypoxia (3% O2) | Chronic hypoxia upregulates both TRPV1 gene and protein levels | Chronic hypoxia increases cytosolic Ca2+ levels | The proliferation of PASMCs is increased under hypoxia | [110] | |
| TRPV2 | HepG2 and Huh-7 human hepatoma cell lines | RT-PCR Western Blot Flow cytometry |
50, 100, 200, and 400 Μm H2O2 treatment for 24 h | H2O2 upregulates the expression of TRPV2 at mRNA and protein levels | Not assessed | Overexpression of TRPV2 promotes H2O2-induced cell death | [111] |
| TRPV3 | Rat myocardial cells | MTT and Edu staining assay Western Blot Caspase-3 and LDH activity assay |
12 h long exposition to hypoxia (1% O2) | TRPV3 is overexpressed in myocardial cells induced by ischemia/hypoxia | Not assessed | TRPV3 silencing protects cardiomyocytes from hypoxia-induced cell death and decreases the secretion of proinflammatory cytokines | [112] |
| Primary rat pulmonary artery smooth muscle cells (PASMCs) | Western Blot Flow cytometry MTT assay |
24 h long exposition to hypoxia (3% O2) | TRPV3 protein expression is enhanced in PASMCs from hypoxic rats | Not assessed | TRPV3 mediates hypoxia-induced PASMCs’ proliferation via PI3K/AKT signalling | [113] | |
| TRPV3-overexpressing HEK293 | Patch Clamp | 12 h long exposition to hypoxia (1% O2) | Pre-incubation in hypoxic conditions potentiates TRPV3 currents in response to 2-APB treatment | Not assessed | Not assessed | [114] | |
| TRPV4 | Rat cardiomyocytes | Western Blot qPCR Calcium imaging |
6 h long exposition to hypoxia (95% N2) in a controlled hypoxic chamber | TRPV4 gene and protein expression levels are increased after 6 h exposure to hypoxia | Hypoxia increases TRPV4-mediated Ca2+ influx responses to 300 nM GSK | Hypoxia-mediated activation of TRPV4 induces cytosolic Ca2+ overload in cardiomyocytes, leading to ROS production and oxidative injury in vitro and in vivo | [115] |
| Adult rat hippocampal astrocytes | Patch Clamp qPCR Western Blot Calcium imaging |
Hypoxia/ischemia (H/I) is induced by occlusion of the common carotids in combination with hypoxic conditions (from 1 h up to 7 days, 6% O2) | TRPV4 mRNA and protein expression are significantly increased 1 h after H/I. H/I also activates TRPV4 channel | H/I enhances the response of 4aPDD, inducing TRPV4-mediated Ca2+ oscillations | Not assessed | [116] | |
| TRPA1 | Several breast and lung cancer cell lines | Calcium imaging Cell viability and apoptosis assay via PI and Annexin IV staining |
Treatment with 10 µM H2O2 for 15 min for calcium measurements, 1, 20, and 100 µM for 72–96 h-long exposition for cell viability and cell death assays | H2O2 treatment activates TRPA1 channel | H2O2 treatment increases TRPA1-mediated calcium entry | TRPA1-mediated calcium entry promotes cell survival by upregulating anti-apoptotic pathways and promoting oxidative stress resistance | [117] |
| Oligodendrocytes | Calcium imaging | Ischemia inducing solution | Not assessed | Ischemia-induced intracellular acidosis promotes Ca2+ entry via TRPA1 | Ischemia-induced intracellular acidosis and consequent Ca2+ entry via TRPA1 mediate myelin damage | [118] | |
| TRPC1 | U-87 MG glioma cells | qPCR, western blot | Hypoxia induced by exposition to 1% O2 | Not assessed | Not assessed | TRPC1 participates in hypoxia-induced VEGF gene and protein expression | [119] |
| MDA-MB-468 breast cancer cells | qPCR, calcium imaging | Hypoxia induced by exposition to 1% O2 for 24 h | Hypoxia upregulates TRPC1 via HIF1α | siTRPC1 reduces non-stimulated Ca2+ entry and increases Store-Operated Ca2+ entry in hypoxic conditions | TRPC1 overexpression promotes Snail EMT marker upregulation and decrease in claudin-4 epithelial marker in hypoxic conditions. TRPC1 regulates HIF-1α protein levels via Akt-dependent pathway and promotes hypoxia-induced STAT3 and EGFR phosphorylation. TRPC1 also regulates hypoxia-induced LC3BII levels via effects on EGFR. | [120] | |
| TRPC5 | MCF-7/WT and adriamycin-treated (MCF-7/ADM) human breast cancer cells | Western Blot, immunofluorescence, | Not assessed | Not assessed | Not assessed | TRPC5 promotes HIF-1alpha translocation to the nucleus and HIF-1alpha-mediated VEGF expression, boosting tumour angiogenesis | [121] |
| SW620 colon cancer cells | Western blot, transwell invasion, and migration assay, MTT proliferation assay | Not assessed | Not assessed | Not assessed | TRPC5 activates HIF-1alpha-Twist signalling to induce EMT, supporting colon cancer cells’ migration, invasion, and proliferation | [122] | |
| TRPC6 | Murine pancreatic stellate cells (mPSCs) | Time-lapse single-cell random migration assay Bead-based cytokine assay qPCR Western Blot Ca2+ signals quantification by Mn2+ quench technique |
24 h incubation in hypoxic conditions (1% O2, 5%CO2, and 94% N2) or chemically induced hypoxia by pretreatment with 0.5 mmol/l DMOG | Hypoxic conditions enhance TRPC6 expression and activate the channel | Hypoxia stimulates Ca2+ influx mediated by TRPC6 channels | Hypoxia-induced TRPC6 activation enhances mPSCs migration via secretion of pro-migratory factors | [123] |
| lx-2 human hepatic stellate cells (HSCs) | Calcium imaging qPCR Western Blot |
Hypoxia induced by 100 μmol/L CoCl2 treatment | Hypoxic conditions enhance TRPC6 expression and activate the channel | Hypoxia stimulates Ca2+ influx mediated by TRPC6 channels | Hypoxia-induced TRPC6 activation and consequent calcium entry promote the synthesis of ECM proteins, which facilitate the fibrotic activation of HSCs | [124] | |
| Huh7 and HepG2 hepatocellular carcinoma cells (HCCs) | Confocal Calcium imaging Western Blot |
Hypoxia induced by cell incubation in a low oxygen atmosphere with 1% O2, 5%CO2, and 94% N2 for 6 h | Hypoxic conditions activate the channel | Hypoxia promotes calcium influx | Hypoxia-induced TRPC6-mediated calcium entry promotes HCCs drug resistance via STAT3 pathway | [125] | |
| U373MG and HMEC-1 glioblastoma cell lines | qPCR Western Blot Calcium imaging Proliferation assay Matrigel invasion assay Endothelial cell tube formation assay |
Hypoxia induced by 100 μmol/L CoCl2 treatment | Hypoxia enhances TRPC6 expression via Notch pathway | Hypoxia stimulates Ca2+ influx mediated by TRPC6 channels | Hypoxia-induced TRPC6-mediated calcium entry promotes HCCs proliferation, colony formation, and invasion via NFAT pathway | [126] | |
| ORAI1/STIM1 | Primary Aortic Smooth Muscle Cells and HEK293 cells transfected with ORAI1 and STIM1 | Patch Clamp Calcium imaging |
Hypoxia was induced with 3 methods: (1) sodium dithionite (Na2S2O4) treatment to 1 mM final concentration, pH adjustment to pH 7.4, and bubbling with 100% N2. (2) cell culture media with 30 min-long bubbling with 100% N2. (3) cell culture media with 30 min-long bubbling with 3% O2 | Intracellular acidification induced by hypoxia in HEK293 cells leads to inhibition of SOCE by disrupting the electrostatic ORAI1/STIM1 binding and closing ORAI1 channel. | Hypoxia-induced intracellular acidification reduces SOCE in Primary Aortic Smooth Muscle Cells and HEK293 cells transfected with ORAI1 and STIM1 | Not assessed | [92] |
| A549 non-small cell lung cancer cells | Western Blot qPCR BrdU cell proliferation assay Calcium imaging Scrape-wound migration assay Matrigel transwell invasion assay |
Hypoxia induced by Nicotine treatment for 48 h | Nicotine treatment-induced hypoxia determines ORAI1 overexpression at gene and protein levels | Nicotine treatment-induced hypoxia increases intracellular basal calcium levels and SOCE | Nicotine treatment-induced hypoxia increases A549 cells’ proliferation and migration | [127] | |
| MDA-MB 231 and BT549 breast cancer cell lines and Human Microvascular Endothelial Cell line-1 (HMEC-1) | Western Blot qPCR Calcium imaging Migration assay (Wound healing and transwell migration assay) Matrigel transwell invasion assay Tube formation assay in vitro |
Hypoxia induced by cell incubation in low oxygen atmosphere | Hypoxia promotes ORAI1 gene and protein upregulation via activation of Notch1 signalling | Hypoxia increases thapsigargin-induced SOCE, with consequent rise in cytosolic calcium entry | Hypoxia-induced ORAI1 overexpression and consequent increase in SOCE promote NFAT4 activation and enhance neuroblastoma cells’ migration, invasion, and angiogenesis | [128] | |
| HCT-116 and SW480 human colon cancer cells and Human Microvascular Endothelial Cell line-1 (HMEC-1) | Western Blot qPCR Calcium imaging Transwell migration assay Matrigel transwell invasion assay Tube formation assay in vitro Cell attachment and detachment assays |
Hypoxia induced by 100 μmol/L CoCl2 treatment | Hypoxia promotes ORAI1 gene and protein upregulation via activation of Notch1 signalling | Hypoxia increases thapsigargin-induced SOCE | Hypoxia-induced ORAI1 overexpression and consequent increase in SOCE promote NFATc3 activation and enhance neuroblastoma cells’ migration, invasion, and angiogenesis | [129] |