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The adhesion molecule L1 regulates transendothelial migration and trafficking of dendritic cells.

Maddaluno L, Verbrugge SE, Martinoli C, Matteoli G, Chiavelli A, Zeng Y, Williams ED, Rescigno M, Cavallaro U - J. Exp. Med. (2009)

Bottom Line: In agreement with these findings, L1 was expressed in cutaneous DCs that migrated to draining lymph nodes, and its ablation reduced DC trafficking in vivo.Within the skin, L1 was found in Langerhans cells but not in dermal DCs, and L1 deficiency impaired Langerhans cell migration.Our results implicate L1 in the regulation of DC trafficking and shed light on novel mechanisms underlying transendothelial migration of DCs.

View Article: PubMed Central - PubMed

Affiliation: The FIRC Institute of Molecular Oncology, 20139 Milan, Italy.

ABSTRACT
The adhesion molecule L1, which is extensively characterized in the nervous system, is also expressed in dendritic cells (DCs), but its function there has remained elusive. To address this issue, we ablated L1 expression in DCs of conditional knockout mice. L1-deficient DCs were impaired in adhesion to and transmigration through monolayers of either lymphatic or blood vessel endothelial cells, implicating L1 in transendothelial migration of DCs. In agreement with these findings, L1 was expressed in cutaneous DCs that migrated to draining lymph nodes, and its ablation reduced DC trafficking in vivo. Within the skin, L1 was found in Langerhans cells but not in dermal DCs, and L1 deficiency impaired Langerhans cell migration. Under inflammatory conditions, L1 also became expressed in vascular endothelium and enhanced transmigration of DCs, likely through L1 homophilic interactions. Our results implicate L1 in the regulation of DC trafficking and shed light on novel mechanisms underlying transendothelial migration of DCs. These observations might offer novel therapeutic perspectives for the treatment of certain immunological disorders.

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L1 regulates the adhesion of DCs to endothelium. (A and B)CFSE-labeled bone marrow–derived DCs fromL1floxed andTie2-Cre;L1floxed mice were seededon TNF-α–stimulated MELC (A) or SV-LEC (B)monolayers and allowed to adhere for the indicated time lengths. Afterwashing and fixation, cell adhesion was measured as described inMaterials and methods. Data represent the means ± SD of asingle representative experiment performed in triplicate. The experimentwas independently repeated five times, each time using DCs fromdifferent mice. The insets in B show the morphology of DCs seeded onSV-LEC monolayers. Bar, 10 µm. *, P < 0.05;**, P < 0.005 (relative toL1floxed DCs). (C) Mouse inguinal lymphnode cells were enriched for CD11c+ cells and thenFACS sorted into CD11c+/L1+ andCD11c+/L1− DCs (top,postsorting cell populations), which were then labeled with CFSE (green)and PKH26 (red), respectively, before adhesion assays onTNF-α–stimulated SV-LEC monolayers (bottom left,example of DC adhesion; bar, 30 µm). Data in the bottom rightrepresent the means ± SD from three independent experiments,each performed with lymph nodes from three mice. *, P <0.05 (relative to L1-positive DCs).
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fig2: L1 regulates the adhesion of DCs to endothelium. (A and B)CFSE-labeled bone marrow–derived DCs fromL1floxed andTie2-Cre;L1floxed mice were seededon TNF-α–stimulated MELC (A) or SV-LEC (B)monolayers and allowed to adhere for the indicated time lengths. Afterwashing and fixation, cell adhesion was measured as described inMaterials and methods. Data represent the means ± SD of asingle representative experiment performed in triplicate. The experimentwas independently repeated five times, each time using DCs fromdifferent mice. The insets in B show the morphology of DCs seeded onSV-LEC monolayers. Bar, 10 µm. *, P < 0.05;**, P < 0.005 (relative toL1floxed DCs). (C) Mouse inguinal lymphnode cells were enriched for CD11c+ cells and thenFACS sorted into CD11c+/L1+ andCD11c+/L1− DCs (top,postsorting cell populations), which were then labeled with CFSE (green)and PKH26 (red), respectively, before adhesion assays onTNF-α–stimulated SV-LEC monolayers (bottom left,example of DC adhesion; bar, 30 µm). Data in the bottom rightrepresent the means ± SD from three independent experiments,each performed with lymph nodes from three mice. *, P <0.05 (relative to L1-positive DCs).

Mentions: Next, we investigated whether L1 is involved in the interaction of DCs with thelymphatic vessel endothelium, a key process in DC trafficking to lymphoid organs(12). To this goal, DCs derivedfrom L1floxed orTie2-Cre;L1floxed bone marrows were subjected toadhesion assays on monolayers of lymphatic ECs (LECs). Two mouse LEC lines wereused, MELCs (13) and SV-LECs (14). In both cases,Tie2-Cre;L1floxed DCs exhibited a loweradhesion capacity to lymphatic endothelium as compared with DCs from controlL1floxed mice (Fig. 2, A and B). Furthermore, L1-positive DCs spread andextended cellular protrusions upon adhesion to LECs, whereasTie2-Cre;L1floxed DCs retained a roundmorphology (Fig. 2 B, inset). Thestronger adhesion of L1-expressing bone marrow–derived DCs was not theresult of an L1-dependent regulation of β2 integrins because nodifference in β2 expression was observed betweenL1floxed andTie2-Cre;L1floxed DCs and the two cellpopulations adhered to purified ICAM-2 (a major β2 ligand) with similarefficiency (unpublished data). The role of L1 in the interaction of DCs with thelymphatic endothelium was also assessed using DCs freshly isolated from lymphnodes. In this case, L1+ and L1− DCswere separated by FACS sorting and labeled with different dyes before adhesionassays on SV-LEC monolayers. As shown in Fig. 2C, lymph node–derived L1+ DCs adheredtwice more efficiently than L1− cells to the lymphaticendothelium, confirming the results obtained with bone marrow–derivedDCs. L1− DCs isolated from the lymph nodes ofTie2-Cre;L1floxed mice showed an adhesionrate to lymphatic endothelium comparable to that of L1− DCsfrom L1floxed mice (unpublished data). These resultssupported the notion that L1 is required for DC–LEC interaction.


The adhesion molecule L1 regulates transendothelial migration and trafficking of dendritic cells.

Maddaluno L, Verbrugge SE, Martinoli C, Matteoli G, Chiavelli A, Zeng Y, Williams ED, Rescigno M, Cavallaro U - J. Exp. Med. (2009)

L1 regulates the adhesion of DCs to endothelium. (A and B)CFSE-labeled bone marrow–derived DCs fromL1floxed andTie2-Cre;L1floxed mice were seededon TNF-α–stimulated MELC (A) or SV-LEC (B)monolayers and allowed to adhere for the indicated time lengths. Afterwashing and fixation, cell adhesion was measured as described inMaterials and methods. Data represent the means ± SD of asingle representative experiment performed in triplicate. The experimentwas independently repeated five times, each time using DCs fromdifferent mice. The insets in B show the morphology of DCs seeded onSV-LEC monolayers. Bar, 10 µm. *, P < 0.05;**, P < 0.005 (relative toL1floxed DCs). (C) Mouse inguinal lymphnode cells were enriched for CD11c+ cells and thenFACS sorted into CD11c+/L1+ andCD11c+/L1− DCs (top,postsorting cell populations), which were then labeled with CFSE (green)and PKH26 (red), respectively, before adhesion assays onTNF-α–stimulated SV-LEC monolayers (bottom left,example of DC adhesion; bar, 30 µm). Data in the bottom rightrepresent the means ± SD from three independent experiments,each performed with lymph nodes from three mice. *, P <0.05 (relative to L1-positive DCs).
© Copyright Policy - openaccess
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2664975&req=5

fig2: L1 regulates the adhesion of DCs to endothelium. (A and B)CFSE-labeled bone marrow–derived DCs fromL1floxed andTie2-Cre;L1floxed mice were seededon TNF-α–stimulated MELC (A) or SV-LEC (B)monolayers and allowed to adhere for the indicated time lengths. Afterwashing and fixation, cell adhesion was measured as described inMaterials and methods. Data represent the means ± SD of asingle representative experiment performed in triplicate. The experimentwas independently repeated five times, each time using DCs fromdifferent mice. The insets in B show the morphology of DCs seeded onSV-LEC monolayers. Bar, 10 µm. *, P < 0.05;**, P < 0.005 (relative toL1floxed DCs). (C) Mouse inguinal lymphnode cells were enriched for CD11c+ cells and thenFACS sorted into CD11c+/L1+ andCD11c+/L1− DCs (top,postsorting cell populations), which were then labeled with CFSE (green)and PKH26 (red), respectively, before adhesion assays onTNF-α–stimulated SV-LEC monolayers (bottom left,example of DC adhesion; bar, 30 µm). Data in the bottom rightrepresent the means ± SD from three independent experiments,each performed with lymph nodes from three mice. *, P <0.05 (relative to L1-positive DCs).
Mentions: Next, we investigated whether L1 is involved in the interaction of DCs with thelymphatic vessel endothelium, a key process in DC trafficking to lymphoid organs(12). To this goal, DCs derivedfrom L1floxed orTie2-Cre;L1floxed bone marrows were subjected toadhesion assays on monolayers of lymphatic ECs (LECs). Two mouse LEC lines wereused, MELCs (13) and SV-LECs (14). In both cases,Tie2-Cre;L1floxed DCs exhibited a loweradhesion capacity to lymphatic endothelium as compared with DCs from controlL1floxed mice (Fig. 2, A and B). Furthermore, L1-positive DCs spread andextended cellular protrusions upon adhesion to LECs, whereasTie2-Cre;L1floxed DCs retained a roundmorphology (Fig. 2 B, inset). Thestronger adhesion of L1-expressing bone marrow–derived DCs was not theresult of an L1-dependent regulation of β2 integrins because nodifference in β2 expression was observed betweenL1floxed andTie2-Cre;L1floxed DCs and the two cellpopulations adhered to purified ICAM-2 (a major β2 ligand) with similarefficiency (unpublished data). The role of L1 in the interaction of DCs with thelymphatic endothelium was also assessed using DCs freshly isolated from lymphnodes. In this case, L1+ and L1− DCswere separated by FACS sorting and labeled with different dyes before adhesionassays on SV-LEC monolayers. As shown in Fig. 2C, lymph node–derived L1+ DCs adheredtwice more efficiently than L1− cells to the lymphaticendothelium, confirming the results obtained with bone marrow–derivedDCs. L1− DCs isolated from the lymph nodes ofTie2-Cre;L1floxed mice showed an adhesionrate to lymphatic endothelium comparable to that of L1− DCsfrom L1floxed mice (unpublished data). These resultssupported the notion that L1 is required for DC–LEC interaction.

Bottom Line: In agreement with these findings, L1 was expressed in cutaneous DCs that migrated to draining lymph nodes, and its ablation reduced DC trafficking in vivo.Within the skin, L1 was found in Langerhans cells but not in dermal DCs, and L1 deficiency impaired Langerhans cell migration.Our results implicate L1 in the regulation of DC trafficking and shed light on novel mechanisms underlying transendothelial migration of DCs.

View Article: PubMed Central - PubMed

Affiliation: The FIRC Institute of Molecular Oncology, 20139 Milan, Italy.

ABSTRACT
The adhesion molecule L1, which is extensively characterized in the nervous system, is also expressed in dendritic cells (DCs), but its function there has remained elusive. To address this issue, we ablated L1 expression in DCs of conditional knockout mice. L1-deficient DCs were impaired in adhesion to and transmigration through monolayers of either lymphatic or blood vessel endothelial cells, implicating L1 in transendothelial migration of DCs. In agreement with these findings, L1 was expressed in cutaneous DCs that migrated to draining lymph nodes, and its ablation reduced DC trafficking in vivo. Within the skin, L1 was found in Langerhans cells but not in dermal DCs, and L1 deficiency impaired Langerhans cell migration. Under inflammatory conditions, L1 also became expressed in vascular endothelium and enhanced transmigration of DCs, likely through L1 homophilic interactions. Our results implicate L1 in the regulation of DC trafficking and shed light on novel mechanisms underlying transendothelial migration of DCs. These observations might offer novel therapeutic perspectives for the treatment of certain immunological disorders.

Show MeSH
Related in: MedlinePlus