**4.2.2 Accumulation of chemokines**

In addition to pro-inflammatory cytokines, chemo-attractive cytokines are also produced by different cell types involved in the inflammatory response at the bone-implant interface including: polymorphonuclear leukocytes, monocytes, macrophages, osteoblasts, fibroblasts and synovial cells. Monocyte chemoattractant protein-1 (MCP-1) is one of the most abundantly released chemokines and is an immediate early stress-responsive factor (Tuan et al. 2008). Macrophage inhibitory protein-1 (MIP-1 or CCL2) and interleukin 8 (IL-8) are also involved in these events.

*MCP-1* is a C-C chemokine ( chemokine subfamily) and its gene location in humans is 17q11.2. MCP-1 is primarily secreted by monocytes, macrophages and dendritic cells. With regards to bone, MCP-1 is also expressed by osteoblasts and osteoclasts. MCP-1 can bind to several receptors (CCR2, CCR4, CCR5) but binds preferentially to the CCR2. CCR2 has two isoforms: CCR2A and CCR2B; the expression of the different isoforms is restricted to specific cell types. Mononuclear cells and vascular smooth muscle cells express CCR2A whereas CCR2B is mainly expressed by monocytes and activated NK cells (Deshmane et al. 2009). MCP-1 regulates the migration and infiltration of monocytes, dendritic cells, memory T-lymphocytes and NK cells. Production of MCP-1 is under the direct control of the transcription factor NFB.

*MIP-1* belongs to the same subfamily of chemokines as MCP-1 (the C-C chemokine family), and is secreted primarily by activated macrophages and T-lymphocytes. Other cells may also secrete MIP-1. MIP-1 has two main isoforms: MIP-1 (CCL3) and MIP-1 (CCL4). In mice, MIP-1 and MIP-1 are encoded by one gene each, however in humans, MIP-1 is encoded by at least three genes and MIP-1 by two, all located in chromosome 17 (Menten et al. 2002). Inducers of MIP-1 release are IL-1, IL-7, LPS and specific adhesion molecules (e.g. intercellular adhesion molecule-1, ICAM-1). High levels of MIP-1 have been observed when macrophages from retrieved periprosthetic tissues have been challenged with titanium alloy and polymethylmethacrylate particles (Nakashima et al. 1999). The release of MIP-1 increases the systemic recruitment of macrophages and has a paracrine effect enhancing IL-1 and IL-6 release (Cook 1996; Menten et al. 2002). Although MCP-1 has a specific receptor (CCR2), MIP-1 acts through several receptors (CCR1, CCR5), (Proudfoot 2002).

*Interleukin-8* (IL-8) is a member of the CXC chemokine family, and also has the designation CXCL8. It is an early stress responsive chemokine and the main cellular sources are: macrophages, endothelial cells, epithelial cells, and cells of the mesenchymal lineage such as mesenchymal stem cells and osteoblasts. IL-8 is a potent chemokine that induces the chemotaxis of neutrophils and monocytes/macrophages, and increases the mobility of osteoclasts (Baggiolini and Clark-Lewis 1992). Il-8 is also an angiogenic factor. IL-8 does not have a specific receptor but acts through the coupled G protein receptors CXCR1 and CXCR2, which are receptors for other chemokines (e.g. GCP-2, NAP-2, Gro- and others), (Proudfoot 2002). When osteoblasts are challenged with phagocytosable titanium particles, the IL-8 peak in gene expression occurs within 1-2 hours after particle exposure (Fritz et al. 2002). The biological mechanism of IL-8 expression involves first a protein tyrosine phosphorylation after contact with Ti particles and then activation of the pro-inflammatory NF-B through two subunits, p50 (NF-B1) and p65 (RelA) to bind the IL-8 gene promoter.
