284 Additionally, the supposed danger qualities of HSP are proposed to be mediated through receptor-mediated endo- cytosis (Arnold-Schild et al. 1999). In the latter case TLRs, such as TLR2 and 4, are again implicated as the dominating receptors (Vabulas et al. 2001). There is now a general consensus that the interactions of TLRs with PAMPs evolved to prepare the immune system for tailor-made responses to invading microbes. A diversity of TLRs (and also RIGS, NOD, etc.) enables the immune system to be discriminative at the early level of innate defenses. A microbe that activates TLR4 and TLR5 is likely to be a flagellated gram-negative bacterium, while a microbe that triggers TLR2, 5, and 6 qualifies to be a flagellated gram- positive bacterium. Very aptly, this was called the barcode model of immune recognition (Aderem 2003). However, this concept raises several questions with regard to HSPs. Which advantageous aspect would HSPs serving as DAMPs offer to this system? What would have been the evolutionary advantage of having HSP as a DAMP included in such a recognition strategy? A direct innate response to proteins released by necrotic cells, unidirectionally activating a pro- inflammatory response, would disturb the “bar coding” mediated through TLR and other innate receptors. A feature of TLRs which still remains to be fully appreciated is that they generate not only pro-inflammatory responses but also suppressive responses in innate immune cells (Conroy et al. 2008). TLR2 for example has been shown to be present on Treg cells, and its triggering by an HSP60-derived peptide led to cytokine and contact-dependent suppression (Zanin-Zhorov et al. 2006). Also, TLR2 triggering by Pam3Cys led to Treg induction (Sutmuller et al. 2006). In addition, IL-10-producing T cells were generated in vivo through TLR2 triggering by capsular polysaccharides of commensal bacteria (Round and Mazmanian 2010). TLR4 triggering by LPS was seen to enhance survival and proliferation of Treg cells (Caramalho et al. 2003). This means that these receptors have no danger conveying power on their own. The effect of their triggering is therefore most likely cell-type specific and may depend very much on the context of their triggering, other factors released by other cells in the vicinity, and other receptors triggered simultaneously. In other words, the integration of multiple signals must finally converge within a particular cell to drive the biological outcome. Given the examples of suppressive effects by HSP-derived peptides in this context, it seems even possible that HSPs are dampening inflammatory signals through these innate receptors. The search for dedicated heat shock protein receptors has been intensive over the past years. We are left with a broad array of receptors with rather promiscuous binding charac- teristics. Of the scavenger receptors, heat shock proteins are now known to interact with SRA-1, SREC-1, LOX-1, and FEEL-1 (Calderwood et al. 2007). Interestingly, some of these receptors were found to be immunosuppressive, such W. van Eden et al. as SRA-1 (Wang et al. 2007). As put forth by Calderwood: “Discerning the signaling networks emanating from such an array of receptors will be challenging; however, this is essential if sense is to be made of the heat shock protein signal input” (reported in (Henderson et al. 2010). Whatever the outcome of such an exploration will be, a simply straightforward monotonous pro-inflammatory HSP response will be an unlikely outcome. It is the result of modern cellular and molecular technologies that we tend to study single types of cells in their interactions with single defined molecules through tentative single receptors. However, in reality, the complexity of immune behavior will not be visible by analyzing a linear sequence of consecutive events from cause to effect. We will need a systems biology approach to integrate responses to HSP into a context-dependent unified systemic response. DAMP disqualifying features of HSP A variety of molecules have been nominated to trigger innate receptors in a pro-inflammatory manner. They are often presented as putative DAMPs, despite the fact there may be no discernable patterns associated with their molecular structures. However, this is generally accepted as the name DAMP was coined in keeping with the PAMPs analogy of which the classical examples exhibit obvious molecular patterns. Yet, HSPs are not known to harbor any molecular patterns or to have any sequence or structural similarities between HSP of different families. Possibly with an excep- tion, if we accept the suggestion of hydrophobicity as a damage-associated molecular pattern. In that case, HSP may have hydrophobic binding sites that are exposed in an ATP- dependent manner to allow binding to hydrophobic parts of nascent or unfolded proteins (Seong and Matzinger 2004). Possibly of more relevance to our discussion is the fact that there are no systematic differences between self and microbial HSP. Therefore, the immune system does not seem to possess receptors that may distinguish the self from microbial. As a consequence, if self-HSPs were DAMPs, one would also expect the immunodominant microbial HSPs to exert a continuous pro-inflammatory effect in our immune system: an unlikely and not very attractive situation. DAMPs typically reside intracellularly and are only exposed to the extracellular environment when cells undergo necrosis. However, HSPs are also exposed at the cell surface (Multhoff 2007) and exist as freely circulating HSP (Pockley and Multhoff 2008), and neither of these forms are easily compatible with DAMP-type pro- inflammatory activities of HSP. Some DAMPs are claimed to exert their effects via a receptor associated with pro-inflammatory signaling. This is the case given for HMBG-1 and RAGE. Antibody blocking of RAGE was seen to inhibit the ability of