Comparison of the Response of Human Embryonic Stem Cells and Their Differentiated Progenies to Oxidative Stress

Objective: To investigate effects of oxidative stress on human embryonic stem cells (hESC) and their spontaneously differentiated fibroblastic progenies (at passage 5).

Background Data: In ischemic disease models, high levels of free radicals and reactive oxygen species are critical factors in decreasing survivability and engraftment of transplanted/transfused cells. Hence, it is imperative to characterize response of hESC and their differentiated progenies to oxidative stress.

Methods: Oxidative stress was induced either by (i) varying durations (0 to 40 min) of photodynamic treatment (diode laser, 664 nm, 30 mW) in the presence of 10 microM methylene blue as a photosensitizer, or by (ii) exposure to varying concentrations of hydrogen peroxide (0 to 50 microM) for a fixed duration of 40 min. Additionally, the effects of heat shock and mild oxidative stress preconditioning on oxidative stress response was also investigated.

Results: Consistently higher survivability (MTT assay) of hESC was observed compared to their differentiated fibroblastic progenies, upon exposure to equivalent levels of oxidative stress. Further experiments demonstrated that heat-shock pretreatment (42 degrees C for 90 min) did not enhance the resistance of either hESC or their differentiated progenies to oxidative stress (photodynamic treatment), but in fact had a slightly detrimental effect on their survivability upon subsequent exposure to oxidative stress. Similarly, preconditioning of both undifferentiated hESC and their differentiated progenies with low levels of oxidative stress also did not enhance cellular survivability upon subsequent exposure to much higher levels of oxidative stress induced by photodynamic treatment.

Conclusions: Undifferentiated hESC are intrinsically more resistant to oxidative stress compared to their spontaneously differentiated fibroblastic progenies.