Patients with diabetes mellitus experience impaired wound healing, often resulting in chronic foot ulcers. Healing can be accelerated by application of growth factors like platelet-derived growth factor (PDGF). We investigated the mitogenic responses, measured by 3[H]thymidine incorporation, of fibroblasts cultured from diabetic ulcers, non-diabetic ulcers, and non-lesional diabetic and age-matched controls, to recombinant human PDGF-AB, epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and insulin-like growth factor (IGF-I). We determined the optimal concentration of these factors and investigated which single factor, or combination of factors, added simultaneously or sequentially, induced the highest mitogenic response. For single growth factor additions, in all fibroblast populations significant differences in mitogenic response to different growth factors were observed, with PDGF-AB consistently inducing the highest response and IGF-I the lowest (p < 0.043). IGF-I produced only a 1.7-fold stimulation over control in diabetic ulcer fibroblasts, versus 2.95-fold for chronic ulcer, 3.2-fold for non-lesional (p = 0.007) and 5-fold for age-matched fibroblasts (p = 0.007). The highest mitogenic response induced by EGF was significantly less for chronic ulcer fibroblasts compared with age-matched and nonlesional controls (p < 0.03), chronic ulcer fibroblasts also needed significantly more EGF to reach this optimal stimulus (p < 0.02 versus age-matched and non-lesional controls). The simultaneous addition of FGF-IGF-I, PDGF-IGF-I and FGF-PDGF to diabetic ulcer fibroblasts always produced a higher stimulatory response than sequential additions (p < or = 0.05). Also the addition of bFGF, PDGF-AB and EGF prior to IGF-I induced a higher 3[H]thymidine uptake in all fibroblasts compared to the combination of each in reverse order. Significant differences were observed when comparing the combinations of growth factors with the highest stimulatory responses (PDGF-IGF-I, FGF-PDGF and EGF-PDGF added simultaneously) to a double dose of PDGF, with the highest mean rank for the combination PDGF-IGF-I (p = 0.018). In conclusion, combinations such as PDGF-AB and IGF-I may be more useful than PDGF-AB alone for application in chronic diabetic wounds.