Tick lipocalin triggers mammalian IGFBP-3-mediated apoptosis in macrophages and keratinocytes

Ticks secrete several molecules, including lipocalins, in their saliva during blood feeding on a vertebrate host. In this study, we provide novel evidence on the role of <i>Ornithodoros turicata americanus</i> tick lipocalin (Otlip) in modulating cytokine expression and in triggering apoptosis in mammalian macrophages and keratinocytes. The cytokine array analysis revealed significantly increased secretion of insulin-like growth factor-binding protein 3 (IGFBP3) from murine macrophage cell line upon treatment with recombinant Otlip protein (rGST-Otlip) when compared to the secretion noted from cells treated with a control protein (rGST). Similar observation with cytokine protein array analysis was noted when murine macrophages were treated with salivary gland lysates generated from fed <i>O. turicata americanus</i> ticks. In addition, we noted increased expression of IGFBP3 in human keratinocytes cell line upon treatment with rGST-Otlip. The Live/Dead staining and TUNEL microscopic analysis revealed that treatment with rGST-Otlip induced apoptotic cell death in murine macrophage and human keratinocytes cell lines. qRT-PCR analysis showed increased <i>caspase-3</i> and reduced <i>bcl-2</i> (an anti-apoptotic protein) transcripts in both murine macrophages and human keratinocytes cell lines upon treatment with rGST-Otlip. Immunoblotting further showed increased Caspase-3 levels in both cells lines upon treatment with rOtlip. Furthermore, our study noted that siRNA-mediated silencing of <i>igfbp3</i> (carrier protein for IGF) expression inhibited rGST-Otlip-mediated apoptosis in murine macrophages. Taken together, our study not only provides new insights into the role of arthropod salivary molecules in the interactions with mammalian cells but also could lead to the development of strategies to target tick blood feeding.