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.

The effects of foliar-applied ZnO nanoparticles (ZnO NPs) and ZnSO on the winter wheat ( Triticum aestivum L.) grain yield and grain quality were studied under field conditions, with the distribution and speciation of Zn within the grain examined using synchrotron-based X-ray fluorescence microscopy and X-ray absorption spectroscopy. Although neither of the two Zn compounds improved the grain yield or quality, both increased the grain Zn concentration (average increments were 5 and 10 mg/kg for ZnSO and ZnO NP treatments, respectively). Across all treatments, this Zn was mainly located within the aleurone layer and crease of the grain, although the application of ZnO NPs also slightly increased Zn within the endosperm. This Zn within the grain was found to be present as Zn phosphate, regardless of the form in which Zn was applied. These results indicate that the foliar application of ZnO NPs appears to be a promising approach for Zn biofortification, as required to improve human health.

Background Signal-regulatory protein α (SIRPα) is an essential signaling molecule that modulates leukocyte inflammatory responses. However, the regulation of selective SIRPα synthesis and its dynamic changes in leukocytes under inflammatory stimulation remain incompletely understood. Objective We sought to identify the microRNAs (miRNAs) that posttranscriptionally regulate SIRPα synthesis and their roles in modulating macrophage inflammatory responses. Methods SIRPα was induced in SIRPα-negative promyelocytic cells by retinoic acid or phorbol 12-myristate 13-acetate, and the differential expression of miRNAs was assessed by means of microarray and quantitative RT-PCR assays. The roles of identified miRNAs in controlling SIRPα synthesis in leukocytes and leukocyte inflammatory responses were determined. Results We identified SIRPα as a common target gene of miR-17, miR-20a, and miR-106a. During SIRPα induction, levels of these 3 miRNAs were all reduced, and their downregulation by retinoic acid or phorbol 12-myristate 13-acetate occurred through suppression of the c-Myc signaling pathway. All miR-17, miR-20a, and miR-106a specifically bound to the same seed sequence within the SIRPα 3′ untranslated region and correlated inversely with SIRPα protein levels in various cells. In macrophages upregulation of miR-17, miR-20a, and miR-106a by LPS served as the mechanism underlying LPS-induced SIRPα reduction and macrophage activation. Both in vitro and in vivo assays demonstrate that miR-17, miR-20a, and miR-106a regulate macrophage infiltration, phagocytosis, and proinflammatory cytokine secretion through targeting SIRPα. Conclusion These findings demonstrate for the first time that miR-17, miR-20a, and miR-106a regulate SIRPα synthesis and SIRPα-mediated macrophage inflammatory responses in a redundant fashion, providing a novel pathway in which a panel of miRNAs can modulate immune polarization through regulation of macrophage activation.