Radiofrequency radiations constitute a new form of environmental pollution. Among them, millimeter waves (MMW) will be widely used in the near future for high speed communication systems. This study aimed therefore to evaluate the biocompatibility of MMW at 60 GHz. For this purpose, we used a whole gene expression approach to assess the effect of acute 60 GHz exposure on primary cultures of human keratinocytes. Controls were performed to dissociate the electromagnetic from the thermal effect of MMW. Microarray data were validated by RT-PCR, in order to ensure the reproducibility of the results. MMW exposure at 20 mW/cm2, corresponding to the maximum incident power density authorized for public use (local exposure averaged over 1 cm2), led to an increase of temperature and to a strong modification of keratinocyte gene expression (665 genes differentially expressed). Nevertheless, when temperature is artificially maintained constant, no modification in gene expression was observed after MMW exposure. However, a heat shock control did not mimic exactly the MMW effect, suggesting a slight but specific electromagnetic effect under hyperthermia conditions (34 genes differentially expressed). By RT-PCR, we analyzed the time course of the transcriptomic response and 7 genes have been validated as differentially expressed: ADAMTS6, NOG, IL7R, FADD, JUNB, SNAI2 and HIST1H1A. Our data evidenced a specific electromagnetic effect of MMW, which is associated to the cellular response to hyperthermia. This study raises the question of co-exposures associating radiofrequencies and other environmental sources of cellular stress.