Introduction: The process of angiogenesis is important in the wound healing cascade. Endothelial cells play a central role in this process via its proliferation and concerted efforts. Topical antimicrobials (TA), which are frequently employed to treat wounds, have been shown to be cytotoxic to endothelial cells. While hyperbaric oxygen (HBO) at the appropiate atomospheric pressure and duration has been shown to cause proliferation of cultured endothelial cells, its effects with TA are unknown. This study was performed to evaluate hyperbaric oxygen on the cytotoxic effects of TA on endothelial cells. Methods: Human umbilical endothelial cells (HUEC) were treated with Silver Sulfadiazine (0.01%), Chlorhexidine Gluconate (0.001%), and Povidone Iodine (0.001%). These were then exposed to a HBO at 2.4 ATA for 15, 60, and 120 minutes. Using a commercial quick cell proliferation assay kit (BioVision) and an automated microplate reader (Bio-Tek Instruments, Inc.) at 450-nm, cell proliferation and survival were determined. Results: The topical antimicrobials were cytotoxic to the endothelial cells. A 120-minute HBO treatment was most optimal in cell proliferation (119%, p<0.01) and was beneficial in increasing cell survival when topical antimicrobials were added. Silver Sulfadiazine (0.01%) with a 120 minute HBO treatment had the highest proliferation of endothelial cells compared to the other antimicrobials tested (117%. p<0.01). Conclusion: HBO causes endothelial cell proliferation. TAs are detrimental to endothelial cells. However, HBO has a reversal effects on the cytotoxicity of topical antimicrobials on endothelial cells.