Limits...
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.

Show MeSH

Related in: MedlinePlus

T cell migration mediated by cell uropods is prevented  by blocking with anti–ICAM-3 mAbs or with drugs that impair  uropod formation. T cell migration across HMEC-1 was assayed  as described in Materials and Methods. (A) After treatment of the  first layer of T lymphoblasts with either HP2/19 (uropod) or TP1/ 24 (control) anti–ICAM-3 mAbs, cells were incubated for additional 15 min with the indicated blocking mAbs. (B) Before the  incubation of the first layer of T cells with the EC monolayers,  lymphocytes were treated either with butanedione monoxime or  with colchicine for 30 min at 37°C, and then the drugs were extensively washed. In parallel, drug-treated lymphocytes were allowed to adhere to coverslips coated with 10 μg/ml ICAM-1–Fc  for 30 min at 37°C, fixed and stained for ICAM-3, and the proportion of uropod-bearing cells was calculated as described in  Materials and Methods: untreated control, 11%, uropod, 76%;  butanedione monoxime control, 1%, uropod 4%; colchicine control, 40%, uropod, 81%. A representative experiment out of  three independent ones run in duplicate is shown.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2139764&req=5

Figure 10: T cell migration mediated by cell uropods is prevented by blocking with anti–ICAM-3 mAbs or with drugs that impair uropod formation. T cell migration across HMEC-1 was assayed as described in Materials and Methods. (A) After treatment of the first layer of T lymphoblasts with either HP2/19 (uropod) or TP1/ 24 (control) anti–ICAM-3 mAbs, cells were incubated for additional 15 min with the indicated blocking mAbs. (B) Before the incubation of the first layer of T cells with the EC monolayers, lymphocytes were treated either with butanedione monoxime or with colchicine for 30 min at 37°C, and then the drugs were extensively washed. In parallel, drug-treated lymphocytes were allowed to adhere to coverslips coated with 10 μg/ml ICAM-1–Fc for 30 min at 37°C, fixed and stained for ICAM-3, and the proportion of uropod-bearing cells was calculated as described in Materials and Methods: untreated control, 11%, uropod, 76%; butanedione monoxime control, 1%, uropod 4%; colchicine control, 40%, uropod, 81%. A representative experiment out of three independent ones run in duplicate is shown.

Mentions: The role of uropod in lymphocyte migration was confirmed with blocking experiments. We found that the increase in the migration was virtually abrogated by inhibition of uropod function in the first layer of T cells by using blocking anti–ICAM-3 mAb (Fig. 10 A). In addition, the prevention of uropod formation with the myosin-disrupting drug butanedione monoxime also blocked the increment in transendothelial migration of the second cohort of T cells (Fig. 10 B). This drug has previously been reported to prevent lymphocyte polarization and redistribution of adhesion molecules to the uropod (Campanero et al., 1994; del Pozo et al., 1995).


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)

T cell migration mediated by cell uropods is prevented  by blocking with anti–ICAM-3 mAbs or with drugs that impair  uropod formation. T cell migration across HMEC-1 was assayed  as described in Materials and Methods. (A) After treatment of the  first layer of T lymphoblasts with either HP2/19 (uropod) or TP1/ 24 (control) anti–ICAM-3 mAbs, cells were incubated for additional 15 min with the indicated blocking mAbs. (B) Before the  incubation of the first layer of T cells with the EC monolayers,  lymphocytes were treated either with butanedione monoxime or  with colchicine for 30 min at 37°C, and then the drugs were extensively washed. In parallel, drug-treated lymphocytes were allowed to adhere to coverslips coated with 10 μg/ml ICAM-1–Fc  for 30 min at 37°C, fixed and stained for ICAM-3, and the proportion of uropod-bearing cells was calculated as described in  Materials and Methods: untreated control, 11%, uropod, 76%;  butanedione monoxime control, 1%, uropod 4%; colchicine control, 40%, uropod, 81%. A representative experiment out of  three independent ones run in duplicate is shown.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139764&req=5

Figure 10: T cell migration mediated by cell uropods is prevented by blocking with anti–ICAM-3 mAbs or with drugs that impair uropod formation. T cell migration across HMEC-1 was assayed as described in Materials and Methods. (A) After treatment of the first layer of T lymphoblasts with either HP2/19 (uropod) or TP1/ 24 (control) anti–ICAM-3 mAbs, cells were incubated for additional 15 min with the indicated blocking mAbs. (B) Before the incubation of the first layer of T cells with the EC monolayers, lymphocytes were treated either with butanedione monoxime or with colchicine for 30 min at 37°C, and then the drugs were extensively washed. In parallel, drug-treated lymphocytes were allowed to adhere to coverslips coated with 10 μg/ml ICAM-1–Fc for 30 min at 37°C, fixed and stained for ICAM-3, and the proportion of uropod-bearing cells was calculated as described in Materials and Methods: untreated control, 11%, uropod, 76%; butanedione monoxime control, 1%, uropod 4%; colchicine control, 40%, uropod, 81%. A representative experiment out of three independent ones run in duplicate is shown.
Mentions: The role of uropod in lymphocyte migration was confirmed with blocking experiments. We found that the increase in the migration was virtually abrogated by inhibition of uropod function in the first layer of T cells by using blocking anti–ICAM-3 mAb (Fig. 10 A). In addition, the prevention of uropod formation with the myosin-disrupting drug butanedione monoxime also blocked the increment in transendothelial migration of the second cohort of T cells (Fig. 10 B). This drug has previously been reported to prevent lymphocyte polarization and redistribution of adhesion molecules to the uropod (Campanero et al., 1994; del Pozo et al., 1995).

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