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The Effect of Stress-Induced Senescence on Aging Human Cord Blood-Derived Endothelial Cells.
Cheung TM, Ganatra MP, Fu JJ, Truskey GA
Abstract:
PURPOSE:
We sought to determine the effect of stress-induced senescence on the permeability to albumin of aging endothelial progenitor cells.
METHODS:
Human umbilical cord blood derived endothelial cells (hCB-ECs) and human aortic endothelial cells (HAECs) were treated with 200 μM H2O2 and permeability to FITC-bovine serum albumin was measured. Some samples were subsequently treated with 100μM 8-pCPT-2'-O-Me-cAMP, a cAMP analog that activates the Epac1-Rap1 pathway. Cell proliferation was measured with the EdU assay. Phase contrast, and immunofluorescence images were taken to observe morphological changes in cells after exposure to H2O2.
RESULTS:
hCB-ECs exposed to H2O2 exhibited a significant increase in permeability, but their response differed from the HAECs. Low passage hCB-ECs had a permeability increase of about 82% (p<0.01) compared to aged cells which had a permeability increase of about 37% (p<0.05). This increase in permeability was reduced by treating the cells with 100 μM 8-pCPT-2'-O-Me-cAMP. The younger cells exhibited a significant decrease in proliferation after being subjected to various concentrations of H2O2 whereas the aged cells exhibited a more gradual decrease in the percent of cells in S-phase. These changes also correlated with changes in cell morphology and junction staining. When placed back in the original media, the morphology and permeability of the hCB-ECs returned to the control condition, while the HAECs did not.
CONCLUSIONS:
The permeability of low and high passage hCB-ECs and HAECs initially increases in response to oxidative stress. hCB-ECs, but not HAECs, were able to recover from the stress 24 hours later. Early passage hCB-ECs were more susceptible to exogenous H2O2 than late passage hCB-ECs. The increase in permeability of hCB-ECs to H2O2 also correlated with decreased cell proliferation and changes in cell junctions.
We sought to determine the effect of stress-induced senescence on the permeability to albumin of aging endothelial progenitor cells.
METHODS:
Human umbilical cord blood derived endothelial cells (hCB-ECs) and human aortic endothelial cells (HAECs) were treated with 200 μM H2O2 and permeability to FITC-bovine serum albumin was measured. Some samples were subsequently treated with 100μM 8-pCPT-2'-O-Me-cAMP, a cAMP analog that activates the Epac1-Rap1 pathway. Cell proliferation was measured with the EdU assay. Phase contrast, and immunofluorescence images were taken to observe morphological changes in cells after exposure to H2O2.
RESULTS:
hCB-ECs exposed to H2O2 exhibited a significant increase in permeability, but their response differed from the HAECs. Low passage hCB-ECs had a permeability increase of about 82% (p<0.01) compared to aged cells which had a permeability increase of about 37% (p<0.05). This increase in permeability was reduced by treating the cells with 100 μM 8-pCPT-2'-O-Me-cAMP. The younger cells exhibited a significant decrease in proliferation after being subjected to various concentrations of H2O2 whereas the aged cells exhibited a more gradual decrease in the percent of cells in S-phase. These changes also correlated with changes in cell morphology and junction staining. When placed back in the original media, the morphology and permeability of the hCB-ECs returned to the control condition, while the HAECs did not.
CONCLUSIONS:
The permeability of low and high passage hCB-ECs and HAECs initially increases in response to oxidative stress. hCB-ECs, but not HAECs, were able to recover from the stress 24 hours later. Early passage hCB-ECs were more susceptible to exogenous H2O2 than late passage hCB-ECs. The increase in permeability of hCB-ECs to H2O2 also correlated with decreased cell proliferation and changes in cell junctions.
