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P-selectin glycoprotein ligand-1 (PSGL-1) on T helper 1 but not on T helper 2 cells binds to P-selectin and supports migration into inflamed skin.

Borges E, Tietz W, Steegmaier M, Moll T, Hallmann R, Hamann A, Vestweber D - J. Exp. Med. (1997)

Bottom Line: PSGL-1 on Th2 cells, although expressed at similar levels as on Th1 cells, did not support binding to P-selectin.Thus, the P-selectin-binding form of PSGL-1 distinguishes Th1 cells from Th2 cells.Furthermore, PSGL-1 is relevant for the entry of Th1 cells into inflamed areas of the skin.

View Article: PubMed Central - PubMed

Affiliation: Institut für Zellbiologie, ZMBE, Universität Münster, Germany.

ABSTRACT
We have shown recently that mouse Th1 cells but not Th2 cells are selectively recruited into inflamed sites of a delayed-type hypersensitivity (DTH) reaction of the skin. This migration was blocked by monoclonal antibodies (mAb) against P- and E-selectin. Here we show that Th1 cells bind to P-selectin via the P-selectin glycoprotein ligand-1 (PSGL-1). This is the only glycoprotein ligand that was detectable by affinity isolation with a P-selectin-Ig fusion protein. Binding of Th1 cells to P-selectin, as analyzed by flow cytometry and in cell adhesion assays, was completely blocked by antibodies against PSGL-1. The same antibodies blocked partially the migration of Th1 cells into cutaneous DTH reactions. This blocking activity, in combination with that of a mAb against E-selectin, was additive. PSGL-1 on Th2 cells, although expressed at similar levels as on Th1 cells, did not support binding to P-selectin. Thus, the P-selectin-binding form of PSGL-1 distinguishes Th1 cells from Th2 cells. Furthermore, PSGL-1 is relevant for the entry of Th1 cells into inflamed areas of the skin. This is the first demonstration for the importance of PSGL-1 for mouse leukocyte recruitment in vivo.

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Partial inhibition of Th1 cell immigration into inflamed skin  by antibodies against PSGL-1. Radiolabeled Th1 cells were injected together with PBS (no Ab) or the same buffer containing 100 μg of nonimmune rabbit IgG Fab fragments (Co Fab), 100 μg of affinity-purified  anti–PSGL-1 Fab fragments (PSGL-1), 200 μg of mAb RB40 against  mouse P-selectin (P-Sel), 200 μg of mAb UZ4 against mouse E-selectin  (E-Sel). Immigration of cells into the noninflamed control skin region of  the same mice is depicted as solid bars. For each determination, four mice  were analyzed. Experiments shown by the left graph were performed  with a different preparation of Th1 cells than the experiments depicted by  the right graph. Numbers on the left refer to the percentage of injected  cells that were found in the analyzed skin area of 2.5 cm2.
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Figure 4: Partial inhibition of Th1 cell immigration into inflamed skin by antibodies against PSGL-1. Radiolabeled Th1 cells were injected together with PBS (no Ab) or the same buffer containing 100 μg of nonimmune rabbit IgG Fab fragments (Co Fab), 100 μg of affinity-purified anti–PSGL-1 Fab fragments (PSGL-1), 200 μg of mAb RB40 against mouse P-selectin (P-Sel), 200 μg of mAb UZ4 against mouse E-selectin (E-Sel). Immigration of cells into the noninflamed control skin region of the same mice is depicted as solid bars. For each determination, four mice were analyzed. Experiments shown by the left graph were performed with a different preparation of Th1 cells than the experiments depicted by the right graph. Numbers on the left refer to the percentage of injected cells that were found in the analyzed skin area of 2.5 cm2.

Mentions: To determine whether PSGL-1 would be involved in the P-selectin–mediated step during migration of Th1 cells into inflamed sites of the skin, we examined the effect of affinitypurified antibodies against PSGL-1 on Th1 cell homing in a DNFB-elicited cutaneous DTH reaction. 51Cr-labeled cells were preincubated with the antibodies for 15 min and injected into mice intravenously. Organs were taken 1 h later and analyzed for the presence of infiltrated lymphocytes by measuring radioactivity. Affinity-purified, complete rabbit antibodies against PSGL-1 blocked T cell migration into inflamed skin by 43% but also caused some trapping of these cells in the lung (not shown). Therefore, Fab fragments were generated. As shown in Fig. 4, anti–PSGL-1 Fab fragments reduced the entry of Th1 cells into the inflamed skin area by 26% (± 5.2%), while no effect was seen with Fab fragments from preimmune serum. Migration of Th1 cells into noninflamed control skin of the same mice was low. No effect of the Fab fragments on Th1 cell accumulation in other organs was observed (not shown). The anti–P-selectin antibody RB40.34 and the anti–E-selectin mAb UZ4 blocked Th1 cell entry into inflamed skin by 42% (± 9.8%) or 48% (± 14%), respectively, when injected together with the cells. The effect of anti–PSGL-1 Fab fragments and the anti–E-selectin mAb were additive, resulting in 83% (± 5.2%) reduction of Th1 cell entry into inflamed skin. This is in agreement with the additive inhibitory effect of 92% that we observed when the anti–P- and anti–E-selectin mAbs were simultaneously injected (19).


P-selectin glycoprotein ligand-1 (PSGL-1) on T helper 1 but not on T helper 2 cells binds to P-selectin and supports migration into inflamed skin.

Borges E, Tietz W, Steegmaier M, Moll T, Hallmann R, Hamann A, Vestweber D - J. Exp. Med. (1997)

Partial inhibition of Th1 cell immigration into inflamed skin  by antibodies against PSGL-1. Radiolabeled Th1 cells were injected together with PBS (no Ab) or the same buffer containing 100 μg of nonimmune rabbit IgG Fab fragments (Co Fab), 100 μg of affinity-purified  anti–PSGL-1 Fab fragments (PSGL-1), 200 μg of mAb RB40 against  mouse P-selectin (P-Sel), 200 μg of mAb UZ4 against mouse E-selectin  (E-Sel). Immigration of cells into the noninflamed control skin region of  the same mice is depicted as solid bars. For each determination, four mice  were analyzed. Experiments shown by the left graph were performed  with a different preparation of Th1 cells than the experiments depicted by  the right graph. Numbers on the left refer to the percentage of injected  cells that were found in the analyzed skin area of 2.5 cm2.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2196023&req=5

Figure 4: Partial inhibition of Th1 cell immigration into inflamed skin by antibodies against PSGL-1. Radiolabeled Th1 cells were injected together with PBS (no Ab) or the same buffer containing 100 μg of nonimmune rabbit IgG Fab fragments (Co Fab), 100 μg of affinity-purified anti–PSGL-1 Fab fragments (PSGL-1), 200 μg of mAb RB40 against mouse P-selectin (P-Sel), 200 μg of mAb UZ4 against mouse E-selectin (E-Sel). Immigration of cells into the noninflamed control skin region of the same mice is depicted as solid bars. For each determination, four mice were analyzed. Experiments shown by the left graph were performed with a different preparation of Th1 cells than the experiments depicted by the right graph. Numbers on the left refer to the percentage of injected cells that were found in the analyzed skin area of 2.5 cm2.
Mentions: To determine whether PSGL-1 would be involved in the P-selectin–mediated step during migration of Th1 cells into inflamed sites of the skin, we examined the effect of affinitypurified antibodies against PSGL-1 on Th1 cell homing in a DNFB-elicited cutaneous DTH reaction. 51Cr-labeled cells were preincubated with the antibodies for 15 min and injected into mice intravenously. Organs were taken 1 h later and analyzed for the presence of infiltrated lymphocytes by measuring radioactivity. Affinity-purified, complete rabbit antibodies against PSGL-1 blocked T cell migration into inflamed skin by 43% but also caused some trapping of these cells in the lung (not shown). Therefore, Fab fragments were generated. As shown in Fig. 4, anti–PSGL-1 Fab fragments reduced the entry of Th1 cells into the inflamed skin area by 26% (± 5.2%), while no effect was seen with Fab fragments from preimmune serum. Migration of Th1 cells into noninflamed control skin of the same mice was low. No effect of the Fab fragments on Th1 cell accumulation in other organs was observed (not shown). The anti–P-selectin antibody RB40.34 and the anti–E-selectin mAb UZ4 blocked Th1 cell entry into inflamed skin by 42% (± 9.8%) or 48% (± 14%), respectively, when injected together with the cells. The effect of anti–PSGL-1 Fab fragments and the anti–E-selectin mAb were additive, resulting in 83% (± 5.2%) reduction of Th1 cell entry into inflamed skin. This is in agreement with the additive inhibitory effect of 92% that we observed when the anti–P- and anti–E-selectin mAbs were simultaneously injected (19).

Bottom Line: PSGL-1 on Th2 cells, although expressed at similar levels as on Th1 cells, did not support binding to P-selectin.Thus, the P-selectin-binding form of PSGL-1 distinguishes Th1 cells from Th2 cells.Furthermore, PSGL-1 is relevant for the entry of Th1 cells into inflamed areas of the skin.

View Article: PubMed Central - PubMed

Affiliation: Institut für Zellbiologie, ZMBE, Universität Münster, Germany.

ABSTRACT
We have shown recently that mouse Th1 cells but not Th2 cells are selectively recruited into inflamed sites of a delayed-type hypersensitivity (DTH) reaction of the skin. This migration was blocked by monoclonal antibodies (mAb) against P- and E-selectin. Here we show that Th1 cells bind to P-selectin via the P-selectin glycoprotein ligand-1 (PSGL-1). This is the only glycoprotein ligand that was detectable by affinity isolation with a P-selectin-Ig fusion protein. Binding of Th1 cells to P-selectin, as analyzed by flow cytometry and in cell adhesion assays, was completely blocked by antibodies against PSGL-1. The same antibodies blocked partially the migration of Th1 cells into cutaneous DTH reactions. This blocking activity, in combination with that of a mAb against E-selectin, was additive. PSGL-1 on Th2 cells, although expressed at similar levels as on Th1 cells, did not support binding to P-selectin. Thus, the P-selectin-binding form of PSGL-1 distinguishes Th1 cells from Th2 cells. Furthermore, PSGL-1 is relevant for the entry of Th1 cells into inflamed areas of the skin. This is the first demonstration for the importance of PSGL-1 for mouse leukocyte recruitment in vivo.

Show MeSH
Related in: MedlinePlus