Mechanism Of Phagocytosis Of Spores Or Microparticles By Non-professional Phagocytes

Endocytosis is a cellular process in which substances are brought into the cell.Endocytosis pathways include phagocytosis,macropinocytosis,clathrin-mediated endocytosis,and caveolinmediated endocytosis.Phagocytosis is the process by which cells bind and internalize particles large than 0.5 μm in diameter.Phagocytosis plays a central role in host defense against infection and inflammation.Phagocytes are divided into professional and non-professional phagocytes.Professional phagocytes include macrophage,dendritic cells,neutrophils and monocytes.Apart from professional phagocytes,various cells,such as epithelial cells,fibroblasts,are capable of phagocytosis;these cells are called non-professional phagocytes.Non-professional phagocytes exhibit low efficiency of phagocytosis compared to professional phagocytes.However,it has been shown that non-professional phagocytes are functionally redundant with professional phagocytes for clearance of apoptotic cells.Phagocytic process is studied very well in professional phagocytes.However,the mechanism in non-professional phagocytes is poorly understood.The yeast Saccharomyces cerevisiae forms spores in response to environmental stress.We have studied to use spores as microparticles.During this study,we performed an experiment to incubate spores with mammalian cells.We found that non-professional phagocytes could internalize spores.This finding led us to use spores as a model for studying the mechanism of phagocytosis in nonprofessional phagocytes.The main results are followings:(1)We found that various non-professional phagocytes,such as HEK293 T,HEK293,He La,HUC and mouse primary Alveolar epithelial type II cells(ATII)could internalize spores.Further studies show internalization of spores in HEK293 T cells was inhibited by inhibitors of Spleen tyrosine kinase(SYK)and Phosphatidylinositol-3 kinase(PI3K),that revealed non-professional phagocytes internalize spores by phagocytosis pathway.(2)Phagocytosis in non-professional phagocytes is induced by polynucleotides.We found that RNA attached to the spore wall,and that RNA fragments serve as ligands to induce phagocytosis in non-professional phagocytes.Microparticles decorated with polynucleotides were phagocytosed by non-professional phagocytes.Thus,polynucleotides serve as ligands to induce phagocytosis in nonprofessional phagocytes.Nucleotide sequence is not critical for induction of phagocytosis.(3)Receptor for advanced glycation end-products(RAGE)is the receptor to induce phagocytosis in non-professional phagocytes.RAGE is a member of immunoglobulin receptor superfamily.We found that overexpression of RAGE could significantly increase the ability of cells to internalize spores in non-professional phagocytes.By contrast,phagocytosis of spores was significantly decreased by knockout of RAGE.(4)Various molecules are internalized via RAGE-mediated phagocytosis.RAGE is a multiligand receptor;its ligands include polynucleotides and high mobility group box 1(HMGB1).Apart from polynucleotides,microparticles decorated with HMGB1 or DNA/HMGB1 complex were phagocytosed by non-professional phagocytes in a manner dependent on RAGE.Additionally,we found that histone is a novel ligand for RAGE.Microparticles decorated with histone or DNA/histone complex were phagocytosed by non-professional phagocytes.Our results show that various microparticle decorated with RAGE ligands are internalized by non-professional phagocytes via RAGE-mediated phagocytosis process.Finally,we examined whether RAGE is required for HEK293 T cells to internalize apoptotic cells,which are well-known targets for phagocytosis by nonprofessional phagocytes in vivo.Our results demonstrate that phagocytosis of apoptotic cells was significantly decreased by knockout of RAGE.These results show that efferocytosis by nonprofessional phagocytes is also mediated by a RAGE-dependent pathway.