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P-selectin glycoprotein ligand-1 mediates L-selectin-dependent leukocyte rolling in venules.

Sperandio M, Smith ML, Forlow SB, Olson TS, Xia L, McEver RP, Ley K - J. Exp. Med. (2003)

Bottom Line: Immunohistochemistry and flow cytometry failed to show PSGL-1 expression on resting or inflamed endothelium or on platelets.Leukocyte rolling was significantly slower for leukocyte-endothelial than leukocyte-leukocyte interactions.We conclude that leukocyte-expressed PSGL-1 serves as the main L-selectin ligand in inflamed postcapillary venules.

View Article: PubMed Central - PubMed

Affiliation: University of Virginia Health System, Cardiovascular Research Center, MR5 Bldg, Rm. 1013, P.O. Box 801 394, Charlottesville, VA 22908-1394, USA.

ABSTRACT
Leukocyte rolling in postcapillary venules of inflamed tissues is reduced in L-selectin-deficient mice and mice treated with L-selectin blocking antibodies, but the glycoprotein ligand for L-selectin in inflamed venules is unknown. Here, we show that L-selectin-dependent rolling after P-selectin blockade is completely absent in P-selectin glycoprotein ligand-1 (PSGL-1)-/- mice or wild-type mice treated with a PSGL-1 blocking monoclonal antibody. Immunohistochemistry and flow cytometry failed to show PSGL-1 expression on resting or inflamed endothelium or on platelets. To investigate whether leukocyte-expressed PSGL-1 is mediating L-selectin-dependent rolling, we reconstituted lethally irradiated wild-type mice with PSGL-1-/- bone marrow cells. These chimeric mice showed no L-selectin-dependent rolling, suggesting that leukocyte-expressed PSGL-1 mediates L-selectin-dependent rolling. Frame-to-frame video analysis of L-selectin-dependent rolling in wild-type mice showed that the majority of observed L-selectin-dependent leukocyte rolling was between free flowing leukocytes and already adherent leukocytes or possibly leukocyte fragments, followed by E-selectin-dependent leukocyte rolling along the endothelium. Leukocyte rolling was significantly slower for leukocyte-endothelial than leukocyte-leukocyte interactions. We conclude that leukocyte-expressed PSGL-1 serves as the main L-selectin ligand in inflamed postcapillary venules. L-selectin binding to PSGL-1 initiates tethering events that enable L-selectin-independent leukocyte-endothelial interactions. These findings provide a molecular mechanism for the inflammatory defects seen in L-selectin-deficient mice.

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Leukocyte rolling velocity distribution in untreated cremaster muscle venules of (A) wild-type mice treated with P-selectin blocking mAb RB40.34 (anti-P), (B) PSGL-1−/− mice treated with anti-P, and (C) wild-type mice treated with anti-P and anti-L (L-selectin blocking F(ab′)2 of mAb MEL-14) >60 min after exteriorization. Rolling was completely blocked by adding L- and E-selectin mAb in panel A, or E-selectin mAb in panels B and C. Absolute cell numbers are shown for leukocyte velocity groups with bin sizes of 10 μm/s.
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fig3: Leukocyte rolling velocity distribution in untreated cremaster muscle venules of (A) wild-type mice treated with P-selectin blocking mAb RB40.34 (anti-P), (B) PSGL-1−/− mice treated with anti-P, and (C) wild-type mice treated with anti-P and anti-L (L-selectin blocking F(ab′)2 of mAb MEL-14) >60 min after exteriorization. Rolling was completely blocked by adding L- and E-selectin mAb in panel A, or E-selectin mAb in panels B and C. Absolute cell numbers are shown for leukocyte velocity groups with bin sizes of 10 μm/s.

Mentions: Leukocyte rolling velocities were measured more than 1 h after surgery and after injection of P-selectin blocking mAb RB40.34 (Fig. 3 A). This rolling was L-selectin dependent, because it was completely abolished by injecting F(ab)′2 fragments of MEL-14. In wild-type mice, leukocyte rolling velocities (n = 127) distributed broadly with an average rolling velocity of 64 ± 4 μm/s (Fig. 3 A). This is consistent with an earlier report on L-selectin–dependent rolling velocities (33). In PSGL-1–deficient mice, L-selectin–dependent rolling was abolished so that leukocyte rolling velocities of only 11 leukocytes could be measured, which averaged 40 ± 13 μm/s (Fig. 3 B). This rolling was blocked by injection of mAb 9A9 to E-selectin. Analysis of E-selectin–dependent leukocyte rolling velocities in wild-type mice treated with blocking antibodies against P-selectin (RB40.34) and against L-selectin (F(ab′)2 MEL-14) revealed a similar rolling velocity in the few rolling cells left (23 ± 5 μm/s, n = 15; Fig. 3 C).


P-selectin glycoprotein ligand-1 mediates L-selectin-dependent leukocyte rolling in venules.

Sperandio M, Smith ML, Forlow SB, Olson TS, Xia L, McEver RP, Ley K - J. Exp. Med. (2003)

Leukocyte rolling velocity distribution in untreated cremaster muscle venules of (A) wild-type mice treated with P-selectin blocking mAb RB40.34 (anti-P), (B) PSGL-1−/− mice treated with anti-P, and (C) wild-type mice treated with anti-P and anti-L (L-selectin blocking F(ab′)2 of mAb MEL-14) >60 min after exteriorization. Rolling was completely blocked by adding L- and E-selectin mAb in panel A, or E-selectin mAb in panels B and C. Absolute cell numbers are shown for leukocyte velocity groups with bin sizes of 10 μm/s.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2193782&req=5

fig3: Leukocyte rolling velocity distribution in untreated cremaster muscle venules of (A) wild-type mice treated with P-selectin blocking mAb RB40.34 (anti-P), (B) PSGL-1−/− mice treated with anti-P, and (C) wild-type mice treated with anti-P and anti-L (L-selectin blocking F(ab′)2 of mAb MEL-14) >60 min after exteriorization. Rolling was completely blocked by adding L- and E-selectin mAb in panel A, or E-selectin mAb in panels B and C. Absolute cell numbers are shown for leukocyte velocity groups with bin sizes of 10 μm/s.
Mentions: Leukocyte rolling velocities were measured more than 1 h after surgery and after injection of P-selectin blocking mAb RB40.34 (Fig. 3 A). This rolling was L-selectin dependent, because it was completely abolished by injecting F(ab)′2 fragments of MEL-14. In wild-type mice, leukocyte rolling velocities (n = 127) distributed broadly with an average rolling velocity of 64 ± 4 μm/s (Fig. 3 A). This is consistent with an earlier report on L-selectin–dependent rolling velocities (33). In PSGL-1–deficient mice, L-selectin–dependent rolling was abolished so that leukocyte rolling velocities of only 11 leukocytes could be measured, which averaged 40 ± 13 μm/s (Fig. 3 B). This rolling was blocked by injection of mAb 9A9 to E-selectin. Analysis of E-selectin–dependent leukocyte rolling velocities in wild-type mice treated with blocking antibodies against P-selectin (RB40.34) and against L-selectin (F(ab′)2 MEL-14) revealed a similar rolling velocity in the few rolling cells left (23 ± 5 μm/s, n = 15; Fig. 3 C).

Bottom Line: Immunohistochemistry and flow cytometry failed to show PSGL-1 expression on resting or inflamed endothelium or on platelets.Leukocyte rolling was significantly slower for leukocyte-endothelial than leukocyte-leukocyte interactions.We conclude that leukocyte-expressed PSGL-1 serves as the main L-selectin ligand in inflamed postcapillary venules.

View Article: PubMed Central - PubMed

Affiliation: University of Virginia Health System, Cardiovascular Research Center, MR5 Bldg, Rm. 1013, P.O. Box 801 394, Charlottesville, VA 22908-1394, USA.

ABSTRACT
Leukocyte rolling in postcapillary venules of inflamed tissues is reduced in L-selectin-deficient mice and mice treated with L-selectin blocking antibodies, but the glycoprotein ligand for L-selectin in inflamed venules is unknown. Here, we show that L-selectin-dependent rolling after P-selectin blockade is completely absent in P-selectin glycoprotein ligand-1 (PSGL-1)-/- mice or wild-type mice treated with a PSGL-1 blocking monoclonal antibody. Immunohistochemistry and flow cytometry failed to show PSGL-1 expression on resting or inflamed endothelium or on platelets. To investigate whether leukocyte-expressed PSGL-1 is mediating L-selectin-dependent rolling, we reconstituted lethally irradiated wild-type mice with PSGL-1-/- bone marrow cells. These chimeric mice showed no L-selectin-dependent rolling, suggesting that leukocyte-expressed PSGL-1 mediates L-selectin-dependent rolling. Frame-to-frame video analysis of L-selectin-dependent rolling in wild-type mice showed that the majority of observed L-selectin-dependent leukocyte rolling was between free flowing leukocytes and already adherent leukocytes or possibly leukocyte fragments, followed by E-selectin-dependent leukocyte rolling along the endothelium. Leukocyte rolling was significantly slower for leukocyte-endothelial than leukocyte-leukocyte interactions. We conclude that leukocyte-expressed PSGL-1 serves as the main L-selectin ligand in inflamed postcapillary venules. L-selectin binding to PSGL-1 initiates tethering events that enable L-selectin-independent leukocyte-endothelial interactions. These findings provide a molecular mechanism for the inflammatory defects seen in L-selectin-deficient mice.

Show MeSH
Related in: MedlinePlus