Abstract: The human epidermis provides a first line of defense against exogenous pathogens. Resistance to bacterial skin infections, e.g. with Staphylococcus aureus (S. aureus), is based on the function of intact innate immune mechanisms in the epidermis, mainly provided by keratinocytes. They establish the local cytokine and chemokine milieu which is necessary for attracting other cells participating in an immune response. Toll-like receptor (TLR)-2 recognizes components of S. aureus and is known to be expressed on keratinocytes. The aim of this study was to investigate TLR-2-mediated chemokine and cytokine secretion on human primary keratinocytes (HPKs) both on mRNA and on protein level. As there is no selective TLR-2 ligand known so far, we chose Pam3Cys that acts via TLR-2/TLR-1 heterodimers, lipoteichoic acid (LTA) that acts via TLR-2/TLR-6 and peptidoglycan (PGN) which acts via TLR-2 and Nod. Pam3Cys stimulation yielded in an enhanced secretion of CCL20, CCL2, MMP9 and IL-8 in HPK, whereas stimulation with PGN or LTA showed no or solely slight effects. Our findings show evidence for a functional TLR-2/TLR-1 signalling profile in HPKs upon stimulation with Pam3Cys contributing to the defense against bacterial skin infections.