Modulation of cellular immunity by antibodies against calreticulin

Abstract

Although caltreticulin (CRT) is mainly a residential ER protein, it is also expressed on the membrane surface of various types of cells exhibiting multiple functions. We report here that intraperitoneal administration of a soluble recombinant CRT fragment (rCRT/39-272) led to a substantial decrease in delayed type hypersensitivity (DTH) responses in BALB/c mice and EAE in C57BL/6 mice. In the recall response against keyhole limpet hemocyanin (KLH) in vitro, draining lymph node cells from the rCRT/39-272-treated mice produced less IFN-γ but more IL-4 as compared with the cells from the control group. The immunomodulating effect of intraperitoneally administered rCRT/39-272 was attributed to anti-CRT Abs thereby induced, because, in passive transfer experiments, the CRTspecific antiserum could suppress DTH in BALB/c mice. B-cell-deficient μMT mice were not susceptible to rCRT/39-272-mediated DTH suppression. Furthermore, CRT appears on the surface of murine T cells soon after activation and remains detectable (at relatively low level) by flow cytometry for approximately 5 days in vitro. Anti-CRT Abs were able to inhibit AKT phosphorylation, proliferation, and cytokine production by activated murine T cells. We propose that cell surface CRT could play a role in the function of effector T cells and may be considered a target for immunological manipulation.Keywords: Anti-CRT antibodies r Calreticulin r Immunoregulation Supporting Information available online IntroductionCalreticulin (CRT) is a multifunctional glycoprotein of 46 kDa, and is most abundant in the ER of the cell [1][2][3][4][5][6]. It folds into three domains: (i) a lectin-like N domain, (ii) a proline-rich P domain, and (iii) a Ca 2+ -binding C-domain. CRT has well-recognized physiological roles in intracellular Ca 2+ storage and signaling and also as a molecular chaperone [1,5,6]. It has been detected in soluble form in the sera of patients with rheumatoid arthritis (RA) and systemic lupus erythematosus [7,8]. We have shown that a recombinant CRT fragment 39-272 containing its partial NCorrespondence: Prof. Xiao-Ming Gao e-mail: xmgao@suda.edu.cn and P-domains (CRT/39-272) exhibits potent immunostimulatory activity and strong adjuvanticity [7], implying soluble CRT may be involved in immunopathological and inflammatory reactions leading to the development of RA and/or systemic lupus erythematosus. Moreover, CRT can be exposed at the membrane surface of various types of cells and exhibits multiple cellular functions as a surface molecule [6]. For example, cell-surface CRT (csCRT) mediates thrombospondin (TSP) triggered disassembly of focal contacts, exhibits antithrombotic effects, and inhibits melanoma cell spreading and angiogenesis [5,9,10]. Ligation of neuronal csCRT by complement C1q triggers increased levels of cellular * These authors contributed equally to this work. Eur. J. Immunol. 2012. 42: 2419-2430 reactive oxygen species [11]. There is also ample evidence showing that surface exposure of CRT enhances the immunogenicity of tumor cells and ...

Recombinant murine calreticulin fragment 39-272 expands CD1dhiCD5+ IL-10-secreting B cells that modulate experimental autoimmune encephalomyelitis in C57BL/6 mice

“…Some of the animals (4 from each group) were sacrificed on day 9 for splenocytes and the rest (6 mice per group) were scored daily for EAE severity up to 28 days. In line with our previous report (Qiu et al, 2012), i.p. administration of rCRT/39-272, but not rEGFP or PBS, mildly reduced EAE severity (Fig.…”

“…We have recently demonstrated that i.p. injection of rCRT/39-272, a potent stimulator of B cells and macrophages (Hong et al, 2010), could modulate DTH as well as EAE in mice (Qiu et al, 2012). In subsequent experiments addressing the question if EAE modulation by rCRT/39-272 was related to Th2/Th1 balance shift, groups of naïve C57BL/6 mice were s.c. immunized, on day 0, with MOG 35-55 in CFA (100 g/mouse), followed by i.p.…”

“…We have recently shown that CRT-specific IgG Abs can modulate DTH in mice, likely through binding to CRT on the surface of activated T cells and interfering Th2/Th1differentiation (Qiu et al, 2012). Passively transferred CRT antisera suppressed DTH in mice (Qiu et al, 2012), but did not reduce EAE severity in mice (Supplementary Fig.…”

“…administered rCRT/39-272 was attributed to CRT-Abs that were able to affect Th2/Th1 balance through binding to CRT on the surface of activated T cells (Qiu et al, 2012). However, passive transfer of such Abs did not show clear inhibition of EAE in subsequent animal experiments ( Supplementary Fig.…”

“…Additionally, intraperitoneal (i.p.) administration of rCRT/39-272 effectively suppressed delayed-type hypersensitivity (DTH) in mice, which was attributable to production of anti-CRT Abs following rCRT/39-272 injection (Qiu et al, 2012). The present study focuses on rCRT/39-272-induced activation/expansion of CD1d hi CD5 + B10 cells capable of modulating EAE in C57BL/6 mice.…”