Traditionally, heat shock proteins have long been regarded as intracellular molecules, with functions restricted to the intracellular compartment. However, recent observations demonstrating their regulated extracellular release led to a paradigm shift towards studying its immune functions. This study investigates the role of the extracellular heat shock protein 70 (eHsp70) in intestinal immune homeostasis. We propose Hsp70 as an endogenous toll-like receptor(TLR) ligand that selectively drives differentiation of regulatory IL-10 immune responses in the gut. Intriguingly, the N-terminus of Hsp70 shares homology with LcrV, a virulence factor for Yersinia pestis, the causative agent for bubonic plague and a potential threat for bioterrorism. LcrV effectively and lethally shuts down the host immune response via induction of the potent anti-inflammatory cytokine IL-10. Given the amino acid alignment of the two seemingly unrelated proteins, Hsp70 and LcrV, we sought for their functional homology. Challenging previous reports suggesting Hsp70 as a danger signal, treatment of dendritic cells(DCs) with Hsp70 resulted in attenuation of TNF-alpha induced by LPS and flagellin. Similar to LcrV, immune suppression by Hsp70 was due to its strong induction of IL-10. This was dependent on TLR 6, which we recently reported to selectively promote an anti-inflammatory response through induction of tolerogenic DCs subsequently leading to IL-10 producing regulatory T cells(Tr1). Thus, Hsp70 demonstrates immune modulatory properties via its effect on innate immunity, revealing a novel function of an ancient and ubiquitous stress protein.
While Hsp70 is an inducible protein whose basal expression is low, its expression is strikingly robust and constituitive from the mouse cecum to colon, an area richly populated by microflora. Accordingly, we chose colitis as a model to further investigate the physiological function of Hsp70 in immune modulation. Using the DSS colitis model, we found that Hsp70-deficient mice had worse disease compared to wildtype littermates and that Hsp70 overexpression in the gut epithelium showed improved recovery and reduced inflammatory cytokines. Thus, Hsp70 may be an endogenous intestinal factor driving differentiation of tolerogenic DC and Tr1 cells that are critical in the prevention of uncontrolled inflammation.