A pressure ulcer (PU) is a localized injury to the skin and/or to underlying tissues, typically over a weight-bearing bony prominence. PUs often develop in ischemic tissues. Other than being relevant to the etiology of PUs, ischemic factors such as glucose levels, acidity and temperature could potentially affect healing processes as well, particularly, the rate of damage repair. Using an in vitro cell culture model, the goal of the present study was to determine the influence of ischemic factors: low temperature (35 °C), low glucose (1 g/L) and acidic pH (6.7) on the migration rate of NIH3T3 fibroblasts, 3T3L1 preadipocytes and C2C12 myoblasts, which could all be affected by PUs. Cell migration into a local damage site, produced by crushing cells under a micro-indentor, was monitored over ~16 h under controlled temperature and pH conditions. We found that in the NIH3T3 cultures, acidosis significantly hindered the migration rate as well as delayed the times for onset and end of mass cell migration. The effects of temperature and glucose however were not significant. Additionally, under control conditions (temperature 37 °C, glucose 4.5 g/L, pH 7.6), migration rates and times differed significantly across the different cell types. The present findings motivate further studies related to the effects of pH levels on migration performances, particularly in PU where bacterial contamination-associated with an acidic environment-is involved.