Apoptotic cell thrombospondin-1 and heparin-binding domain lead to dendritic-cell phagocytic and tolerizing states

Abstract

Apoptotic cells were shown to induce dendritic cell immune tolerance. We applied a proteomic approach to identify molecules that are secreted from apoptotic monocytes, and thus may mediate engulfment and immune suppression. Supernatants of monocytes undergoing apoptosis were collected and compared using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and differentially expressed proteins were identified using tandem mass spectrometry. Thrombospondin-1 (TSP-1) and its cleaved 26-kDa heparin-binding domain (HBD) were identified. We show that TSP-1 is expressed upon induction of monocyte apoptosis in a caspase-dependent pattern and the HBD is cleaved by chymotrypsinlike serine protease. We further show that CD29, CD36, CD47, CD51, and CD91 simultaneously participate in engulfment induction and generation of an immature dendritic cell (iDC) tolerogenic and phagocytic state. We conclude that apoptotic cell TSP-1, and notably its HBD, creates a signalosome in iDCs to improve engulfment and to tolerate engulfed material prior to the interaction with apoptotic cells. IntroductionIn recent years, it has become apparent that upon induction of apoptosis, apoptotic cells play an active role in their own engulfment by signaling professional phagocytes and/or antigenpresenting cells, without triggering an inflammatory or autoimmune response. [1][2][3][4][5] This process seems to play an important role in homeostasis, resolution of inflammation, and peripheral tolerance induction. 4,[6][7][8] Apoptotic cells have been shown to signal the innate immune system in a variety of ways. "Eat me" signals on apoptotic cells serve as markers for phagocytes to specifically recognize these cells and subsequently ingest them. Such signals can appear on apoptotic cell membranes. Direct signals include alteration in cell surface phospholipid composition, 9 changes in cell surface glycoprotein expression, distinct alterations in cell surface charge, 10,11 or expression of specific molecules. 12 Alternatively, certain serum or phagocyte-derived proteins can opsonize an apoptotic cell surface and signal phagocytes to engulf the opsonized cells. 4,[13][14][15][16][17] Viable cells actively express "do not eat me" signals by restriction of phosphatydilserine to the inner leaflet of their membrane, or "stay away" signals using CD31 expression. 18 Recently, attention has been given not only to apoptotic cell membrane changes and phagocyte receptors, but also to the release of a membrane-derived phospholipid, lysophosphatidylcholine, which acts as a "find me" signal that is important for phagocytic cell recruitment. 19 Most of these mechanisms suggest efficient identification and clearance of cells undergoing apoptosis, with noninflammatory and nonautoimmune consequences.We decided to further explore whether apoptotic cells can actively express and secrete molecules that have a physiological significance for their own engulfment and for the environmental immune suppression. We examined whether apoptosis-induced immune suppress...

Exogenous heat shock protein 27 uniquely blocks differentiation of monocytes to dendritic cells

“…High concentrations of HSP-27 (1000 ng/mL) might induce such elevated PD-L1 levels, thereby triggering increased apoptosis in MO?iDC differentiation cultures. Uptake of apoptotic cells inhibits MO?iDC differentiation and HSP-27-mediated increased apoptosis could be a mechanism of HSP-27 inhibitory activity [40,41]. rhHSP-27 induced some apoptosis in MO?iDC differentiation culture at the concentration of 1000 ng/mL (Fig.…”

From regulation of dying cell engulfment to development of anti-cancer therapy

“…Recently, thrombospondin-1, a calcium-binding protein released from the engulfing cell, was also identified as a protein that is synthesized de novo when apoptosis is induced in monocytes, and that tolerizes immature DCs. 48 Necrotic cells release massive amount of high mobility group box 1 (HMGB-1) protein, which incites inflammatory responses of macrophages through pathways mediated by Toll-like receptor 2 (TLR2) and TLR4. 8,52 Whether HMGB-1 protein is specifically released by necrotic cells but not by apoptotic or secondary necrotic cells remains controversial.…”

Altered dendritic cells with tolerizing phenotype in patients with systemic lupus erythematosus

“…Apoptotic cells were labeled with DiI as described elsewhere [5,6,18] and added to iDC at different ratios for 2 h on day 6 of culture, followed by iDC staining with DC-SIGN FITC. Interaction was assessed with the FACScan TM .…”

“…Stuart et al [4] demonstrated in mice that DC that ingest apoptotic cells have a diminished ability to produce IL-12 in response to external stimuli, a property that corresponds to a failure to upregulate CD86. Earlier, we were able to show the generation of tolerizing, monocyte-derived DC that express low CD86 and MHC class II, and upregulate CCR7, as a result of interaction with iC3b-opsonized apoptotic cells [5], or via interaction with thrombospondin-1 produced by apoptotic cell [6]. In the present work, we are interested in studying the generation of tolerizing DC in a systemic autoimmune disease characterized by dysregulation of peripheral tolerance.…”