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ICAMs redistributed by chemokines to cellular uropods as a mechanism for recruitment of T lymphocytes.

del Pozo MA, Cabañas C, Montoya MC, Ager A, Sánchez-Mateos P, Sánchez-Madrid F - J. Cell Biol. (1997)

Bottom Line: Quantitative analysis revealed that the induction of uropods results in a 5-10-fold increase in cell recruitment.Additional studies showed that the cell recruitment mediated by uropods was abrogated with antibodies to ICAM-1, -3, and LFA-1, whereas mAb to CD43, CD44, CD45, and L-selectin did not have a significant effect, thus indicating that the interaction of LFA-1 with ICAM-1 and -3 appears to be responsible for this process.An enhancement of T cell migration was observed under conditions of uropod formation, and this increase was prevented by incubation with either blocking anti-ICAM-3 mAbs or drugs that impair uropod formation.

View Article: PubMed Central - PubMed

Affiliation: Servicio de Immunología, Hospital de la Princesa, Universidad Autónoma de Madrid, Spain.

ABSTRACT
The recruitment of leukocytes from the bloodstream is a key step in the inflammatory reaction, and chemokines are among the main regulators of this process. During lymphocyte-endothelial interaction, chemokines induce the polarization of T lymphocytes, with the formation of a cytoplasmic projection (uropod) and redistribution of several adhesion molecules (ICAM-1,-3, CD43, CD44) to this structure. Although it has been reported that these cytokines regulate the adhesive state of integrins in leukocytes, their precise mechanisms of chemoattraction remain to be elucidated. We have herein studied the functional role of the lymphocyte uropod. Confocal microscopy studies clearly showed that cell uropods project away from the cell bodies of adhered lymphocytes and that polarized T cells contact other T cells through the uropod structure. Time-lapse videomicroscopy studies revealed that uropod-bearing T cells were able, through this cellular projection, to contact, capture, and transport additional bystander T cells. Quantitative analysis revealed that the induction of uropods results in a 5-10-fold increase in cell recruitment. Uropod-mediated cell recruitment seems to have physiological relevance, since it was promoted by both CD45R0+ peripheral blood memory T cells as well as by in vivo activated lymphocytes. Additional studies showed that the cell recruitment mediated by uropods was abrogated with antibodies to ICAM-1, -3, and LFA-1, whereas mAb to CD43, CD44, CD45, and L-selectin did not have a significant effect, thus indicating that the interaction of LFA-1 with ICAM-1 and -3 appears to be responsible for this process. To determine whether the increment in cell recruitment mediated by uropod may affect the transendothelial migration of T cells, we carried out chemotaxis assays through confluent monolayers of endothelial cells specialized in lymphocyte extravasation. An enhancement of T cell migration was observed under conditions of uropod formation, and this increase was prevented by incubation with either blocking anti-ICAM-3 mAbs or drugs that impair uropod formation. These data indicate that the cell interactions mediated by cell uropods represent a cooperative mechanism in lymphocyte recruitment, which may act as an amplification system in the inflammatory response.

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Uropod induction increases migration across HEC.  (a) Interaction through a uropod-like structure between migrated  T cells and bound lymphocytes. D shows the usual morphology of  high endothelial vein cells. A–C show the same field photographed in a different plane of focus, from the level of HEC (A)  to the level of surface bound lymphocytes (C). E shows a different field. White arrows point to cell–cell contacts through a uropodlike structure. (b) T lymphoblasts preincubated with the indicated  mAb were added to confluent cultures of HEC in multichamber  polystyrene slides and incubated for different periods of time. After washing, adherent lymphocytes were identified as either “surface bound” or “transmigrated” by counterstaining with toluidine  blue and high power light microscopy; the migration index was  calculated as stated in Materials and Methods. A representative  experiment out of three independent ones run in triplicate is  shown. (c) In parallel, some specimens were stained for ICAM-3  after fixation as described in Materials and Methods. Same fields  were photographed under bright field (A) and epifluorescent (B)  conditions. Arrows point to ICAM-3+ uropods.
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Figure 9: Uropod induction increases migration across HEC. (a) Interaction through a uropod-like structure between migrated T cells and bound lymphocytes. D shows the usual morphology of high endothelial vein cells. A–C show the same field photographed in a different plane of focus, from the level of HEC (A) to the level of surface bound lymphocytes (C). E shows a different field. White arrows point to cell–cell contacts through a uropodlike structure. (b) T lymphoblasts preincubated with the indicated mAb were added to confluent cultures of HEC in multichamber polystyrene slides and incubated for different periods of time. After washing, adherent lymphocytes were identified as either “surface bound” or “transmigrated” by counterstaining with toluidine blue and high power light microscopy; the migration index was calculated as stated in Materials and Methods. A representative experiment out of three independent ones run in triplicate is shown. (c) In parallel, some specimens were stained for ICAM-3 after fixation as described in Materials and Methods. Same fields were photographed under bright field (A) and epifluorescent (B) conditions. Arrows point to ICAM-3+ uropods.

Mentions: When the role of uropod in lymphocyte migration was studied by the migration assay described by Ager and Mistry (1988), using HEC, similar results were found. In this assay, T cells were coincubated with HEC, and after different periods of time, the number of lymphocytes bound to the apical surface or underneath the HEC monolayer were counted. We have also found that the induction of the uropod caused an important increment in the migration index in short periods of time, and that this effect vanished at longer times (Fig. 9 b). Interestingly, we observed that in these assays the interaction between migrated lymphocytes and bound T cells was also mediated through ICAM-3–bearing uropod structures (Fig. 9, a and c), indicating that this interaction takes place during transendothelial migration.


ICAMs redistributed by chemokines to cellular uropods as a mechanism for recruitment of T lymphocytes.

del Pozo MA, Cabañas C, Montoya MC, Ager A, Sánchez-Mateos P, Sánchez-Madrid F - J. Cell Biol. (1997)

Uropod induction increases migration across HEC.  (a) Interaction through a uropod-like structure between migrated  T cells and bound lymphocytes. D shows the usual morphology of  high endothelial vein cells. A–C show the same field photographed in a different plane of focus, from the level of HEC (A)  to the level of surface bound lymphocytes (C). E shows a different field. White arrows point to cell–cell contacts through a uropodlike structure. (b) T lymphoblasts preincubated with the indicated  mAb were added to confluent cultures of HEC in multichamber  polystyrene slides and incubated for different periods of time. After washing, adherent lymphocytes were identified as either “surface bound” or “transmigrated” by counterstaining with toluidine  blue and high power light microscopy; the migration index was  calculated as stated in Materials and Methods. A representative  experiment out of three independent ones run in triplicate is  shown. (c) In parallel, some specimens were stained for ICAM-3  after fixation as described in Materials and Methods. Same fields  were photographed under bright field (A) and epifluorescent (B)  conditions. Arrows point to ICAM-3+ uropods.
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Related In: Results  -  Collection

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Figure 9: Uropod induction increases migration across HEC. (a) Interaction through a uropod-like structure between migrated T cells and bound lymphocytes. D shows the usual morphology of high endothelial vein cells. A–C show the same field photographed in a different plane of focus, from the level of HEC (A) to the level of surface bound lymphocytes (C). E shows a different field. White arrows point to cell–cell contacts through a uropodlike structure. (b) T lymphoblasts preincubated with the indicated mAb were added to confluent cultures of HEC in multichamber polystyrene slides and incubated for different periods of time. After washing, adherent lymphocytes were identified as either “surface bound” or “transmigrated” by counterstaining with toluidine blue and high power light microscopy; the migration index was calculated as stated in Materials and Methods. A representative experiment out of three independent ones run in triplicate is shown. (c) In parallel, some specimens were stained for ICAM-3 after fixation as described in Materials and Methods. Same fields were photographed under bright field (A) and epifluorescent (B) conditions. Arrows point to ICAM-3+ uropods.
Mentions: When the role of uropod in lymphocyte migration was studied by the migration assay described by Ager and Mistry (1988), using HEC, similar results were found. In this assay, T cells were coincubated with HEC, and after different periods of time, the number of lymphocytes bound to the apical surface or underneath the HEC monolayer were counted. We have also found that the induction of the uropod caused an important increment in the migration index in short periods of time, and that this effect vanished at longer times (Fig. 9 b). Interestingly, we observed that in these assays the interaction between migrated lymphocytes and bound T cells was also mediated through ICAM-3–bearing uropod structures (Fig. 9, a and c), indicating that this interaction takes place during transendothelial migration.

Bottom Line: Quantitative analysis revealed that the induction of uropods results in a 5-10-fold increase in cell recruitment.Additional studies showed that the cell recruitment mediated by uropods was abrogated with antibodies to ICAM-1, -3, and LFA-1, whereas mAb to CD43, CD44, CD45, and L-selectin did not have a significant effect, thus indicating that the interaction of LFA-1 with ICAM-1 and -3 appears to be responsible for this process.An enhancement of T cell migration was observed under conditions of uropod formation, and this increase was prevented by incubation with either blocking anti-ICAM-3 mAbs or drugs that impair uropod formation.

View Article: PubMed Central - PubMed

Affiliation: Servicio de Immunología, Hospital de la Princesa, Universidad Autónoma de Madrid, Spain.

ABSTRACT
The recruitment of leukocytes from the bloodstream is a key step in the inflammatory reaction, and chemokines are among the main regulators of this process. During lymphocyte-endothelial interaction, chemokines induce the polarization of T lymphocytes, with the formation of a cytoplasmic projection (uropod) and redistribution of several adhesion molecules (ICAM-1,-3, CD43, CD44) to this structure. Although it has been reported that these cytokines regulate the adhesive state of integrins in leukocytes, their precise mechanisms of chemoattraction remain to be elucidated. We have herein studied the functional role of the lymphocyte uropod. Confocal microscopy studies clearly showed that cell uropods project away from the cell bodies of adhered lymphocytes and that polarized T cells contact other T cells through the uropod structure. Time-lapse videomicroscopy studies revealed that uropod-bearing T cells were able, through this cellular projection, to contact, capture, and transport additional bystander T cells. Quantitative analysis revealed that the induction of uropods results in a 5-10-fold increase in cell recruitment. Uropod-mediated cell recruitment seems to have physiological relevance, since it was promoted by both CD45R0+ peripheral blood memory T cells as well as by in vivo activated lymphocytes. Additional studies showed that the cell recruitment mediated by uropods was abrogated with antibodies to ICAM-1, -3, and LFA-1, whereas mAb to CD43, CD44, CD45, and L-selectin did not have a significant effect, thus indicating that the interaction of LFA-1 with ICAM-1 and -3 appears to be responsible for this process. To determine whether the increment in cell recruitment mediated by uropod may affect the transendothelial migration of T cells, we carried out chemotaxis assays through confluent monolayers of endothelial cells specialized in lymphocyte extravasation. An enhancement of T cell migration was observed under conditions of uropod formation, and this increase was prevented by incubation with either blocking anti-ICAM-3 mAbs or drugs that impair uropod formation. These data indicate that the cell interactions mediated by cell uropods represent a cooperative mechanism in lymphocyte recruitment, which may act as an amplification system in the inflammatory response.

Show MeSH
Related in: MedlinePlus