Rapid clearance of adoptively transferred Cd47-null (Cd47−/−) cells in congeneic WT mice suggests a critical self-recognition mechanism, in which CD47 is the ubiquitous marker of self, and its interaction with macrophage signal regulatory protein α (SIRPα) triggers inhibitory signaling through SIRPα cytoplasmic immunoreceptor tyrosine-based inhibition motifs and tyrosine phosphatase SHP-1/2. However, instead of displaying self-destruction phenotypes, Cd47−/− mice manifest no, or only mild, macrophage phagocytosis toward self-cells except under the nonobese diabetic background. Studying our recently established Sirpα-KO (Sirpα−/−) mice, as well as Cd47−/− mice, we reveal additional activation and inhibitory mechanisms besides the CD47-SIRPα axis dominantly controlling macrophage behavior. Sirpα−/− mice and Cd47−/− mice, although being normally healthy, develop severe anemia and splenomegaly under chronic colitis, peritonitis, cytokine treatments, and CFA-/LPS-induced inflammation, owing to splenic macrophages phagocytizing self-red blood cells. Ex vivo phagocytosis assays confirmed general inactivity of macrophages from Sirpα−/− or Cd47−/− mice toward healthy self-cells, whereas they aggressively attack toward bacteria, zymosan, apoptotic, and immune complex-bound cells; however, treating these macrophages with IL-17, LPS, IL-6, IL-1β, and TNFα, but not IFNγ, dramatically initiates potent phagocytosis toward self-cells, for which only the Cd47-Sirpα interaction restrains. Even for macrophages from WT mice, phagocytosis toward Cd47−/− cells does not occur without phagocytic activation. Mechanistic studies suggest a PKC-Syk–mediated signaling pathway, to which IL-10 conversely inhibits, is required for activating macrophage self-targeting, followed by phagocytosis independent of calreticulin. Moreover, we identified spleen red pulp to be one specific tissue that provides stimuli constantly activating macrophage phagocytosis albeit lacking in Cd47−/− or Sirpα−/− mice.

CD47, a ‘self’ recognition marker expressed on tissue cells, interacts with immunoreceptor SIRPα expressed on the surface of macrophages to initiate inhibitory signaling that prevents macrophage phagocytosis of healthy host cells. Previous studies have suggested that cells may lose the surface CD47 during aging or apoptosis to enable phagocytic clearance. In the present study, we demonstrate that the level of cell surface CD47 is not decreased but the distribution pattern of CD47 is altered during apoptosis. On non-apoptotic cells, CD47 molecules are clustered in lipid rafts forming ‘punctates’ on the surface, whereas on apoptotic cells, CD47 molecules are diffused on the cell surface following the disassembly of lipid rafts. We show that clustering of CD47 in lipid rafts provides a high binding avidity for cell surface CD47 to ligate macrophage SIRPα, which also presents as clusters, and elicit SIRPα-mediated inhibitory signaling that prevents phagocytosis. In contrast, dispersed CD47 on the apoptotic cell surface is associated a significant reduction of the binding avidity to SIRPα and failure to trigger SIRPα signal transduction. Disruption of lipid rafts with methyl-β-cyclodextrin (MβCD) disrupted CD47 cluster formation on the cell surfaces, leading to decrease of the binding avidity to SIRPα and a concomitant increase of cells being engulfed by macrophages. Taken together, our study reveals that CD47 normally is clustered in lipid rafts on non-apoptotic cells but is diffused in the plasma membrane when apoptosis occurs, and this transformation of CD47 greatly reduces the strength of CD47-SIRPα engagement, resulting in the phagocytosis of apoptotic cells.

Chronic diseases are often associated with altered inflammatory response, leading to increased host vulnerability to new inflammatory challenges. Employing streptozotocin (STZ)-induced diabetes as a model, we further investigate mechanisms leading to enhanced neutrophil (polymorphonuclear leukocytes, PMN) response under hyperglycemia and compare to that under chronic colitis. We show that, different from colitis under which PMN response is significantly potentiated, the existence of a proinflammatory state associated with broad increases in macrophages in various organs plays a dominant role in promoting PMN inflammatory response in diabetic mice. Studies of PMN infiltration during zymosan-induced peritonitis reveal that hyperglycemia enhances PMN recruitment not through inducing a high level of IL-17, which is the case in colitis, but through increasing F4/80+ macrophages in the peritoneal cavity, resulting in elevations of IL-6, IL-1β, TNF-α, and CXCL1 production. Insulin reversal of hyperglycemia, but not the neutralization of IL-17, reduces peritoneal macrophage numbers and ameliorates PMN infiltration during peritonitis. Significantly increased macrophages are also observed in the liver, kidneys, and intestines under hyperglycemia, and are attributable to exacerbated nephropathy and colitis when inflammatory conditions are induced by doxorubicin and dextran sulfate sodium (DSS), respectively. Furthermore, analyses of monocyte production and macrophage proliferation in tissues suggest significant monocytosis of inflammatory F4/80+Gr-1+ monocytes from the spleen and macrophage proliferation in situ synergistically contribute to the increased macrophage population under hyperglycemia. In conclusion, our results demonstrate that STZ-induced hyperglycemic mice develop a systemic proinflammatory state mediated by broad infiltration of macrophages.

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Study Design: A retrospective cohort of prospective data. Objective: Determine the frequency of various symptoms in a surgical cohort of cervical myelopathy (CM). Summary of Background Data: CM can be difficult to diagnose as there is no sine qua non “myelopathic symptom.” Despite extensive literature, the likelihood or frequency of symptoms at presentation remains unclear. Materials and Methods: A total of 484 patients treated at a single academic center were reviewed. Preoperative symptoms included: axial neck pain; upper extremity (UE) pain; UE sensory or motor deficit; lower extremity (LE) sensory or motor deficit; and sphincter dysfunction. It was noted whether a symptom was the chief complaint (CC) and/or one of a list of overall symptoms (OS) reported by the patient. Magnetic resonance imaging was assessed for the maximal cord compression level and T2 hyperintensity. Results: The most common CC was UE sensory deficit (46.5%), whereas the most common OS were UE and LE motor deficits (82.6% and 81.2%). Neck pain was significantly less common (32.6% CC, 55.4% OS), and sphincter dysfunction was rare (0.6% CC, 16.5% OS). UE pain as a CC was significantly higher when the maximal compression involved a more distal level. The presence of T2 hyperintensity was negatively associated with neck pain but positively associated with sensory and motor deficits of LE. Conclusions: The most common CC in CM related to UE sensation, whereas the most common OS related to upper and lower motor function. UE pain was more common with more distal cord compression. Those with T2 hyperintensity had worse myelopathy and were less likely to have neck pain, but more likely to have LE symptoms. To our knowledge, this study is the largest to quantify the frequency of myelopathic symptom presentation in a surgical population. These findings provide valuable insight into the symptomatic presentation of CM in clinical practice and can be used to better inform diagnosis and treatment in this complex patient population. Level of Evidence: Level II—retrospective study.