Although the epidermis is importantly affected by steroid hormones, little is known about the effects of dehydroepiandrosterone (DHEA) on human keratinocytes, in spite of its abundance in human serum. Here, we demonstrate for the first time a protective role of DHEA against apoptosis in keratinocytes, using non-cancerous immortalized human HaCaT cells. We show that DHEA transmits its signal via specific G protein-coupled, membrane binding sites and inhibits apoptosis, through prevention of mitochondrial disruption and altered balance of Bcl-2 proteins. DHEA conjugated to the membrane impermeable molecule BSA, as well as DHEA-S, the most abundant form of DHEA in human serum exhibit similar anti-apoptotic effect. Our data provide new insights in the treatment of the epidermis with steroid hormones in apoptosis-related conditions.