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Phosphatidylserine (PS) induces PS receptor-mediated macropinocytosis and promotes clearance of apoptotic cells.

Hoffmann PR, deCathelineau AM, Ogden CA, Leverrier Y, Bratton DL, Daleke DL, Ridley AJ, Fadok VA, Henson PM - J. Cell Biol. (2001)

Bottom Line: Further, recognition of PS was found to be dependent on the presence of the PS receptor (PSR).Both PS and anti-PSR antibodies stimulated membrane ruffling, vesicle formation, and "bystander" uptake of cells bound to the surface of the phagocyte.We propose that the phagocytosis of apoptotic cells requires two events: tethering followed by PS-stimulated, PSR-mediated macropinocytosis.

View Article: PubMed Central - PubMed

Affiliation: Program in Cell Biology, Department of Pediatrics, National Jewish Medical and Research Center, Denver, CO 80206, USA.

ABSTRACT
Efficient phagocytosis of apoptotic cells is important for normal tissue development, homeostasis, and the resolution of inflammation. Although many receptors have been implicated in the clearance of apoptotic cells, the roles of these receptors in the engulfment process have not been well defined. We developed a novel system to distinguish between receptors involved in tethering of apoptotic cells versus those inducing their uptake. Our results suggest that regardless of the receptors engaged on the phagocyte, ingestion does not occur in the absence of phosphatidylserine (PS). Further, recognition of PS was found to be dependent on the presence of the PS receptor (PSR). Both PS and anti-PSR antibodies stimulated membrane ruffling, vesicle formation, and "bystander" uptake of cells bound to the surface of the phagocyte. We propose that the phagocytosis of apoptotic cells requires two events: tethering followed by PS-stimulated, PSR-mediated macropinocytosis.

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Anti-PSR antibodies and PS stimulate macropinocytosis. (A) HMDM were stimulated with M-CSF (as a positive control), antibodies to the PSR or CD36, PS, or PC. Vesicle formation was visualized using LY (green). Nuclei were stained with DAPI (blue), and cells were stained for actin with rhomadine–phalloidin (red). Macrophages stimulated with either anti-PSR antibodies or PS formed vesicles similar to those formed after stimulation with M-CSF, a known inducer of macropinocytosis. Representative images are shown. (B) Accumulation of Cascade blue was quantified by fluorimetry. MEM were untreated or stimulated with M-CSF, anti-PSR antibody, or an IgM isotype control antibody, in the presence of dye. At the designated time point, cells were washed, lysed, and fluorescence of the lysate was measured. Similar results were obtained on four separate occasions. Bar, 5 μm.
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fig7: Anti-PSR antibodies and PS stimulate macropinocytosis. (A) HMDM were stimulated with M-CSF (as a positive control), antibodies to the PSR or CD36, PS, or PC. Vesicle formation was visualized using LY (green). Nuclei were stained with DAPI (blue), and cells were stained for actin with rhomadine–phalloidin (red). Macrophages stimulated with either anti-PSR antibodies or PS formed vesicles similar to those formed after stimulation with M-CSF, a known inducer of macropinocytosis. Representative images are shown. (B) Accumulation of Cascade blue was quantified by fluorimetry. MEM were untreated or stimulated with M-CSF, anti-PSR antibody, or an IgM isotype control antibody, in the presence of dye. At the designated time point, cells were washed, lysed, and fluorescence of the lysate was measured. Similar results were obtained on four separate occasions. Bar, 5 μm.

Mentions: Even though membrane ruffling is closely associated with the process of macropinocytosis, stimulation of membrane ruffling does not always result in vesicle formation (Li et al., 1997; Bretscher and Aguado-Velasco, 1998). HMDM were stimulated with either the anti-PSR antibody or PS-containing liposomes and examined for uptake of a water-soluble, membrane-impermeable fluorescent dye, Lucifer yellow (LY). Whereas untreated macrophages showed little or no uptake of LY, treatment of HMDM with M-CSF, which has been shown to induce macropinocytosis in macrophages (Racoosin and Swanson, 1989, 1992), resulted in uptake of LY into large macropinocytotic vesicles (Fig. 7 A). Large vesicles containing the dye were also detectable in PS liposome–treated cells. However, little uptake of dye was seen when cells were stimulated with liposomes containing only PC. We also attempted to induce macropinocytosis with an anti-CD36 IgM antibody and saw little or no LY uptake. As observed for M-CSF and PS, anti-PSR antibody also stimulated the uptake of LY into large vesicles.


Phosphatidylserine (PS) induces PS receptor-mediated macropinocytosis and promotes clearance of apoptotic cells.

Hoffmann PR, deCathelineau AM, Ogden CA, Leverrier Y, Bratton DL, Daleke DL, Ridley AJ, Fadok VA, Henson PM - J. Cell Biol. (2001)

Anti-PSR antibodies and PS stimulate macropinocytosis. (A) HMDM were stimulated with M-CSF (as a positive control), antibodies to the PSR or CD36, PS, or PC. Vesicle formation was visualized using LY (green). Nuclei were stained with DAPI (blue), and cells were stained for actin with rhomadine–phalloidin (red). Macrophages stimulated with either anti-PSR antibodies or PS formed vesicles similar to those formed after stimulation with M-CSF, a known inducer of macropinocytosis. Representative images are shown. (B) Accumulation of Cascade blue was quantified by fluorimetry. MEM were untreated or stimulated with M-CSF, anti-PSR antibody, or an IgM isotype control antibody, in the presence of dye. At the designated time point, cells were washed, lysed, and fluorescence of the lysate was measured. Similar results were obtained on four separate occasions. Bar, 5 μm.
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Related In: Results  -  Collection

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fig7: Anti-PSR antibodies and PS stimulate macropinocytosis. (A) HMDM were stimulated with M-CSF (as a positive control), antibodies to the PSR or CD36, PS, or PC. Vesicle formation was visualized using LY (green). Nuclei were stained with DAPI (blue), and cells were stained for actin with rhomadine–phalloidin (red). Macrophages stimulated with either anti-PSR antibodies or PS formed vesicles similar to those formed after stimulation with M-CSF, a known inducer of macropinocytosis. Representative images are shown. (B) Accumulation of Cascade blue was quantified by fluorimetry. MEM were untreated or stimulated with M-CSF, anti-PSR antibody, or an IgM isotype control antibody, in the presence of dye. At the designated time point, cells were washed, lysed, and fluorescence of the lysate was measured. Similar results were obtained on four separate occasions. Bar, 5 μm.
Mentions: Even though membrane ruffling is closely associated with the process of macropinocytosis, stimulation of membrane ruffling does not always result in vesicle formation (Li et al., 1997; Bretscher and Aguado-Velasco, 1998). HMDM were stimulated with either the anti-PSR antibody or PS-containing liposomes and examined for uptake of a water-soluble, membrane-impermeable fluorescent dye, Lucifer yellow (LY). Whereas untreated macrophages showed little or no uptake of LY, treatment of HMDM with M-CSF, which has been shown to induce macropinocytosis in macrophages (Racoosin and Swanson, 1989, 1992), resulted in uptake of LY into large macropinocytotic vesicles (Fig. 7 A). Large vesicles containing the dye were also detectable in PS liposome–treated cells. However, little uptake of dye was seen when cells were stimulated with liposomes containing only PC. We also attempted to induce macropinocytosis with an anti-CD36 IgM antibody and saw little or no LY uptake. As observed for M-CSF and PS, anti-PSR antibody also stimulated the uptake of LY into large vesicles.

Bottom Line: Further, recognition of PS was found to be dependent on the presence of the PS receptor (PSR).Both PS and anti-PSR antibodies stimulated membrane ruffling, vesicle formation, and "bystander" uptake of cells bound to the surface of the phagocyte.We propose that the phagocytosis of apoptotic cells requires two events: tethering followed by PS-stimulated, PSR-mediated macropinocytosis.

View Article: PubMed Central - PubMed

Affiliation: Program in Cell Biology, Department of Pediatrics, National Jewish Medical and Research Center, Denver, CO 80206, USA.

ABSTRACT
Efficient phagocytosis of apoptotic cells is important for normal tissue development, homeostasis, and the resolution of inflammation. Although many receptors have been implicated in the clearance of apoptotic cells, the roles of these receptors in the engulfment process have not been well defined. We developed a novel system to distinguish between receptors involved in tethering of apoptotic cells versus those inducing their uptake. Our results suggest that regardless of the receptors engaged on the phagocyte, ingestion does not occur in the absence of phosphatidylserine (PS). Further, recognition of PS was found to be dependent on the presence of the PS receptor (PSR). Both PS and anti-PSR antibodies stimulated membrane ruffling, vesicle formation, and "bystander" uptake of cells bound to the surface of the phagocyte. We propose that the phagocytosis of apoptotic cells requires two events: tethering followed by PS-stimulated, PSR-mediated macropinocytosis.

Show MeSH
Related in: MedlinePlus