CD300 receptors regulate efferocytosis, importance in inflammation, cancer and autoimmunity

稳态需要凋亡细胞（AC）在成为促炎坏死细胞之前进行免疫沉默清除。CD300f是一种磷脂酰丝氨酸受体，已知可正向调节巨噬细胞的efferocytosis， CD300f基因缺陷的小鼠易患狼疮样疾病。我们发现CD300f在巨噬细胞和树突状细胞（DC）上均表达，并对它们的efferocytosis进行差异调节。CD300f作为激活受体增强巨噬细胞的efferocytosis，但抑制DC对AC的吞噬。虽然目前尚不清楚CD300f如何在巨噬细胞和DC中差异调节efferocytosis，但值得注意的是，CD300f在其细胞质尾部有五个基于酪氨酸的基序：两个基于免疫受体酪氨酸的基序（ITIM），一个基于免疫受体酪氨酸的开关基序（ITSM），一个pi3k结合基序和一个生长因子受体结合蛋白2 （Grb2）结合基序。AC结合CD300f导致PI3K结合基序的磷酸化，从而促进PI3K的募集和激活，从而启动AC吞噬所需的信号转导事件；其他四个酪氨酸基序中的每一个都启动抑制信号，用于抑制efferocytosis。AC是否被吞噬取决于吞噬细胞内可用的酪氨酸激酶和/或下游信号传导能力。在巨噬细胞中，这些能力的平衡有利于AC吞噬，而DC则相反。

Homeostasis requires the immunologically silent clearance of apoptotic cells (AC) before they become pro-inflammatory necrotic cells. CD300f is a phosphatidylserine receptor known to positively regulate efferocytosis by macrophages, and CD300f gene-deficient mice are predisposed to develop a lupus-like disease. We showed that CD300f is expressed on both macrophages and dendritic cells (DC), and differentially regulates their efferocytosis. CD300f functions as an activating receptor to enhance macrophage efferocytosis, but inhibits the AC engulfment by DC. Although it is not clear at this point how CD300f differentially regulates efferocytosis in macrophages and DC, of note is the fact that CD300f has five tyrosine-based motifs in its cytoplasmic tail: two immunoreceptor tyrosine-based motifs (ITIM), an immunoreceptor tyrosine-based switch motif (ITSM), a PI3K-binding motif and a growth factor receptor-bound protein 2 (Grb2)-binding motif. AC binding to CD300f leads to phosphorylation of the PI3K-binding motif thereby promoting PI3K recruitment and activation, which initiates the signaling events required for AC engulfment; each of the other four tyrosine-based motifs initiate inhibitory signals that serve to suppress efferocytosis. Whether AC are engulfed or not depends on the available tyrosine kinases and/or downstream signaling capacity within the phagocytic cell. In macrophages the balance of these capabilities favors AC engulfment, while the opposite is true for DC. Our data show that CD300f-deficient macrophages are less efficient in clearing AC and that the engulfment of AC by DC is enhanced. Efferocytosis by DC is highly relevant to T-cell priming, since DC are efficient at processing and presentation of engulfed antigens. CD8+ DC engulf AC and migrate to the T-cell zone of the spleen for cross-presentation of AC-associated antigens. In correlation with the increased phagocytic capability of CD300f-deficient DC, more DC engulfed AC in the spleen of CD300f-deficient mice, compared with WT mice. The increased availability of AC, combined with enhanced efferocytic potential of CD300f-deficient DC, likely leads to elevated presentation and cross-presentation of AC-derived self-antigens to T cells, resulting in the observed sustained presence of activated T-cell populations and an expanded population of memory T cells in aged mice. This creates more potential to trigger self-reactive B cells to secrete auto-antibodies, which would explain the increased levels of ANA that we observe in aged mice. The increase in the memory T-cell compartment and auto-antibody levels did not lead to overt signs of autoimmune disease development in CD300f-deficient mice. In elderly people, it has also been observed that there are higher levels of auto-antibodies without clinical relevance. Such individuals also have higher levels of memory and effector T cells, yet are in general hypo-responsive and referred to as immunosenescent. Other than the higher frequency of memory and activated T cells in aged CD300f-deficient mice and the presence of elevated ANA, the T-cell proliferation in response to TCR stimulation, the activation status of macrophages or DC based on co-stimulatory molecule expression, and the levels of inflammatory or anti-inflammatory cytokines were similar between CD300f-deficient and WT mice, suggesting an immunosenescent status in aged mice. Thus, our data indicate that the autoimmune-risk status generated by CD300f deficiency occurs in the context of immunosenescence, which explains the absence of overt disease pathologies. Although CD300f-deficient mice do not develop overt disease, they clearly are more prone than WT mice to develop overt autoimmune symptoms if stressed. CD300f-deficient mice developed a more severe autoimmune inflammation than the WT mice, when the mice were stressed with excess AC in the pristane-induced autoimmune disease model, which fits the factthat repeated exposure to AC leads to exacerbated disease in autoimmune-prone mice. Moreover, our previous study showed that combined CD300f/FcgRIIB deficient mice are predisposed to develop more severe splenomegaly and have a higher fatality rate than in Fcgr2b deficient mice. We performed an in-depth analysis to identify cellular and molecular abnormalities behind this predisposition. In the double-deficient mice, germinal center B cells and plasma cells were expanded and inflammatory cytokines were increased. An enhanced accumulation of AC was found in spleen tissues of double-deficient mice. These data indicate that the autoimmune-risk status due to CD300f deficiency advances to a fully activated autoimmune response once the immune activation threshold is decreased by combining CD300f deficiency with FcgRIIB deficiency, or the immune system is over-loaded with excess AC. Receptor CD300b is implicated in regulating the immune response to bacterial infection by an unknown mechanism. We identified CD300b as a lipopolysaccharide (LPS)-binding receptor and determined the mechanism underlying CD300b augmentation of septic shock. In vivo depletion and adoptive transfer studies identified CD300b-expressing macrophages as the key cell type augmenting sepsis. We showed that CD300b, and its adaptor DAP12, associated with Toll-like receptor 4 (TLR4) upon LPS binding, thereby enhancing TLR4-adaptor MyD88- and TRIF-dependent signaling that resulted in an elevated pro-inflammatory cytokine storm. LPS engagement of the CD300b-TLR4 complex led to the recruitment and activation of spleen tyrosine kinase (Syk) and phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K). This resulted in an inhibition of the ERK1/2 protein kinase- and NFkappaB transcription factor-mediated signaling pathways, which subsequently led to a reduced interleukin-10 (IL-10) production. Collectively, our data describe a mechanism of TLR4 signaling regulated by CD300b in myeloid cells in response to LPS.