Novel heat shock protein Hsp70L1 activates dendritic cells and acts as a Th1 polarizing adjuvant

Abstract

IntroductionThe Th1 cellular immune response is crucial for antitumor and antimicrobial immunity, 1,2 and powerful immunomodulatory adjuvants that induce Th1 polarization are an important facet of vaccination strategies. 3,4 While traditional adjuvants, such as aluminum salts and oil emulsions, mainly evoke Th2 responses, characterized by production of antibodies specific for conformational antigenic determinants, 5 new generation adjuvants, including CpG-rich motifs, monophosphoryl lipid A (MPA), and quil a saponin (QS21) promote Th1 polarization. 6 These effects are thought to result from the activation of antigen-presenting cells (APCs), in particular, dendritic cells (DCs). 7,8 In recent years the molecular chaperone heat shock protein (HSP) has been revealed to interact with APCs, and its considerable capacity to induce antigen-specific CD8 ϩ cytotoxic T lymphocyte (CTL) and Th1 responses has attracted much attention. 9,10 Extracellular HSPs can interact with APCs, exhibiting potent adjuvant functions in stimulating the host immune response. 10,11 This interaction triggers a cascade of events, including re-presentation of the chaperoned peptides by the major histocompatibility complex (MHC), secretion of proinflammatory cytokines, and maturation of DCs. [12][13][14] These properties combine to make HSPs a potent adjuvant, eliciting significant immune responses against associated antigens.The Hsp70 subfamily is one of the most important HSP subfamilies. Hsp70 prepared from tumor cells or virus-infected cells are capable of eliciting potent antigen-specific CD8 ϩ CTL responses 10,15 ; these responses are CD4 ϩ T-cell-independent and have been attributed to antigenic peptides bound to the HSP. 16 It also has been shown that Hsp70 complexed with ovalbumin (OVA) antigen-specific epitopes can activate DCs and induce OVAspecific CTL responses. 17 Hsp70 can bind to CD91, CD14, and TLR2/4 receptors on the surface of APCs, activating APCs and facilitating the re-presentation of HSP-associated antigen via a TAP-dependent or TAP-independent route. 13,14,18,19 These findings demonstrate the adjuvant effects of Hsp70 and encourage the potential application of Hsp70 in vaccination.We report here a new member of the Hsp70 subfamily cloned from a human DC cDNA library, Hsp70-like protein 1 (Hsp70L1). Recombinant Hsp70L1 can bind DCs, resulting in DC maturation and activation. While Hsp70L1 shares common receptors (CD91, TLR2, TLR4) on DCs with Hsp70, there are functional differences between Hsp70L1 and Hsp70. Hsp70L1 can induce DCs to secrete IP-10, a cytokine vital for Th1 polarization, but Hsp70 cannot. Moreover, Hsp70L1 is more potent than Hsp70 in the stimulation of IL-12p70, MIP-1␣, MIP-1␤, RANTES, CCR7, and CXCR4 expression by DCs. Hsp70L1 may therefore polarize Th1 responses more effectively than Hsp70. We used OVA as a model antigen to evaluate the adjuvant effects of Hsp70L1 in vivo, finding that Hsp70L1-OVA 257-264 hybrid peptide immunization could strongly Materials and methods Cell linesE.G7-OVA, an OVA-express...

Enhanced T-cell immunity induced by dendritic cells with phagocytosis of heat shock protein 70 gene-transfected tumor cells in early phase of apoptosis

“…In this study, we also demonstrated that apoptotic tumor cells (in late phase of apoptosis) expressing HSP70 stimulate DC maturation after DC phagocytosis of these apoptotic tumor cells, which is consistent with some previous reports. [53][54][55] DCs, with phagocytosis of apoptotic tumor cells (in late phase of apoptosis) expressing HSP70 induced stronger antitumor immunity than DC with phagocytosis of tumor cells in early phase of apoptosis, which is consistent with a recent report from our group. 28 More interestingly, we demonstrated that apoptotic tumor cells expressing HSP70 in early phase of apoptosis more efficiently induce tumor-specific CTL responses and vaccine efficiency than those in late phase of apoptosis in two MCA/HSP and EG7 tumor models.…”

Heat Shock Protein 70, Released from Heat-Stressed Tumor Cells, Initiates Antitumor Immunity by Inducing Tumor Cell Chemokine Production and Activating Dendritic Cells via TLR4 Pathway

“…Twenty-four hours after HT, DC in the spleen and lymph nodes were increased and showed the improved capacity in the induction of tumor Ag-specific CTL, which was confirmed by our FACS assays of DC within spleen and lymph nodes and by our in vitro CTL induction assays. The major involvement of HSP70, but not HSP90 and HSP60, in HT-induced chemoattraction and activation of DC may be due to the differential levels of these HSP released by tumor cells and the different efficiency of these HSP in activating target cells (1,2,5,12,35). When HSC70 was silenced alone, the chemokine production by 3LL cells after HS was much lower than that of HSP70i-silenced 3LL cells, strongly suggesting that the levels of HSP released by tumor cells may significantly affect the relative contribution of each HSP to production of chemokines during HS (data not shown).…”

“…The rHSP proteins were further deprived of LPS contamination as described by using polymycin B agarose (Sigma-Aldrich) (11). The purity (Ͼ92%) and LPS contamination (Ͻ5 pg/ g protein) of rHSP were determined as described previously (12). ELISA kits for measuring chemokines and cytokines were purchased from R&D Systems.…”

TLR4 is essential for dendritic cell activation and anti-tumor T-cell response enhancement by DAMPs released from chemically stressed cancer cells

“…24 Mouse bone marrow-derived DCs (BMDCs) were prepared as previously described. 25 Assays for cytokine and chemokine production Human DCs and BMDCs were cultured for 5 days and adjusted to a final concentration of 5310 5 cells/ml in 24-well plates.…”