This study examined the effects of red light generated from a light emitting diode (LED) upon proliferation and mitochondrial stress in human gingival fibroblasts (hGFs). Cells were exposed to LED-generated red light at a clinically relevant intensity and distance with a 610-630 nm wavelength for various times (0-48 min). At different exposure times, cells were processed for the analysis of succinate dehydrogenase (SDH) activity, proliferation, mitochondrial membrane potential (MMP) and cytotoxicity. Cell cycle progression was also investigated by flow cytometry after staining with propidium iodide. Red light exposure was found to inhibit SDH activity and DNA synthesis in hGFs in a time-dependent manner. Light exposure also reduced the MMP levels in these cells and this was closely associated with a G_O/G₁ arrest. In contrast, exposure of hGFs to red light for 48 min led to a dramatic loss of MMP with an attendant increase in cytotoxicity. These findings demonstrate that LED-generated red light may cause mitochondrial stress and growth inhibition in hGFs during tooth whitening therapy, depending on the length of the exposure.