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Alpha 6 integrins are required for Langerhans cell migration from the epidermis.

Price AA, Cumberbatch M, Kimber I, Ager A - J. Exp. Med. (1997)

Bottom Line: RGD-containing peptides were also without effect on LC migration from skin explants.In contrast, alpha 4 integrins, or other integrin-dependent interactions with fibronectin that are mediated by the RGD recognition sequence, did not influence LC migration from the epidermis.In addition, alpha 4 integrins did not affect the accumulation of LCs as DCs in draining lymph nodes.

View Article: PubMed Central - PubMed

Affiliation: Division of Cellular Immunology, National Institute for Medical Research, London, United Kingdom.

ABSTRACT
Topical exposure of mice to chemical allergens results in the migration of epidermal Langerhans cells (LCs) from the skin and their accumulation as immunostimulatory dendritic cells (DCs) in draining lymph nodes. Epidermal cell-derived cytokines have been implicated in the maturation and migration of LCs, but the adhesion molecules that regulate LC migration have not been studied. We hypothesized that integrin-mediated interactions with extracellular matrix components of the skin and lymph node may regulate LC/DC migration. We found that alpha 6 integrins and alpha 4 integrins were differentially expressed by epidermal LCs and lymph node DCs. A majority of LCs (70%) expressed the alpha 6 integrin subunit, whereas DCs did not express alpha 6 integrins. In contrast, the alpha 4 integrin subunit was expressed at high levels on DCs but at much lower levels on LCs. The anti-alpha 6 integrin antibody, GoH3, which blocks binding to laminin, completely prevented the spontaneous migration of LCs from skin explants in vitro and the rapid migration of LCs from mouse ear skin induced after intradermal administration of TNF-alpha in vivo. GoH3 also reduced the accumulation of DCs in draining lymph nodes by a maximum of 70% after topical administration of the chemical allergen oxazolone. LCs remaining in the epidermis in the presence of GoH3 adopted a rounded morphology, rather than the interdigitating appearance typical of LCs in naive skin, suggesting that the cells had detached from neighboring keratinocytes and withdrawn cellular processes in preparation for migration, but were unable to leave the epidermis. The anti-alpha 4 integrin antibody PS/2, which blocks binding to fibronectin, had no effect on LC migration from the epidermis either in vitro or in vivo, or on the accumulation of DCs in draining lymph nodes after oxazolone application. RGD-containing peptides were also without effect on LC migration from skin explants. These results identify an important role for alpha 6 integrins in the migration of LC from the epidermis to the draining lymph node by regulating access across the epidermal basement membrane. In contrast, alpha 4 integrins, or other integrin-dependent interactions with fibronectin that are mediated by the RGD recognition sequence, did not influence LC migration from the epidermis. In addition, alpha 4 integrins did not affect the accumulation of LCs as DCs in draining lymph nodes.

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The effect of anti-α6  and anti-α4 integrin antibodies  on the migration of epidermal  LCs in vitro. Skin explants were  derived from the ears of naive  mice and incubated on medium  containing either (A) anti-α6  (GoH3) or (B) anti-α4 (PS/2) integrin antibodies at 10, 50, or 100  μg/ml (solid bars). Controls included explants cultured on medium containing either no antibody (open bars) or isotype-matched control antibody at 100  μg/ml (hatched bars). Epidermal  sheets were prepared from fresh  ear skin (0 h) and explants after  72 h of incubation and the number of LCs/mm2 determined by  immunohistochemistry. The results are expressed as means ± SD  (n = 18). At all concentrations  tested, in cultures containing  anti-α6 antibodies, the frequency  of LCs did not differ significantly  from that found in fresh explants.  These same values were significantly (P <0.0001) higher than  those found in fresh explants cultured for 72 h in the absence of  antibody, or with an isotype-matched control antibody (A). Similar treatment of explant cultures with  anti-α4 antibody failed to result, at any concentration, in a significant increase in LC numbers compared with controls (B).
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Figure 2: The effect of anti-α6 and anti-α4 integrin antibodies on the migration of epidermal LCs in vitro. Skin explants were derived from the ears of naive mice and incubated on medium containing either (A) anti-α6 (GoH3) or (B) anti-α4 (PS/2) integrin antibodies at 10, 50, or 100 μg/ml (solid bars). Controls included explants cultured on medium containing either no antibody (open bars) or isotype-matched control antibody at 100 μg/ml (hatched bars). Epidermal sheets were prepared from fresh ear skin (0 h) and explants after 72 h of incubation and the number of LCs/mm2 determined by immunohistochemistry. The results are expressed as means ± SD (n = 18). At all concentrations tested, in cultures containing anti-α6 antibodies, the frequency of LCs did not differ significantly from that found in fresh explants. These same values were significantly (P <0.0001) higher than those found in fresh explants cultured for 72 h in the absence of antibody, or with an isotype-matched control antibody (A). Similar treatment of explant cultures with anti-α4 antibody failed to result, at any concentration, in a significant increase in LC numbers compared with controls (B).

Mentions: The differential expression of α6 and β4 integrin subunits by LCs and lymphoid DCs suggested that the α6β4 integrin may regulate the early stages of LC migration from the epidermis. Conversely, the higher level of expression of the α4 integrin subunit by DCs in comparison with LCs raises the possibility that α4 integrins may be involved in DC localization within the lymph node. To study the roles of these integrins in regulating the initial stages of LC migration from the epidermis, blocking antibodies to α6 and α4 integrin subunits were included in the culture medium of skin explants and their effects on LC migration were determined (Fig. 2). Affinity-purified anti-α6 (GoH3) and anti-α4 (PS/2) integrin antibodies were added to the culture medium at 10, 50, and 100 μg/ml. Control explants were incubated on culture medium containing isotype-matched control antibody at 100 μg/ml, or on culture medium alone. Explants were incubated for 72 h, the epidermis was removed, and the number of LCs/mm2 was determined. The number of LCs in fresh epidermis ranged from 1,000 to 1,200 LCs/mm2. After 72 h of incubation in culture medium, this number decreased by ∼70% to 300–350 LCs/mm2. Inclusion of 100 μg/ml MAC 193 (rat IgG2a) or HRPN (rat IgG2b) had no effect on the number of LCs remaining in the epidermis after 72 h and these antibodies were therefore used as isotype-matched controls.


Alpha 6 integrins are required for Langerhans cell migration from the epidermis.

Price AA, Cumberbatch M, Kimber I, Ager A - J. Exp. Med. (1997)

The effect of anti-α6  and anti-α4 integrin antibodies  on the migration of epidermal  LCs in vitro. Skin explants were  derived from the ears of naive  mice and incubated on medium  containing either (A) anti-α6  (GoH3) or (B) anti-α4 (PS/2) integrin antibodies at 10, 50, or 100  μg/ml (solid bars). Controls included explants cultured on medium containing either no antibody (open bars) or isotype-matched control antibody at 100  μg/ml (hatched bars). Epidermal  sheets were prepared from fresh  ear skin (0 h) and explants after  72 h of incubation and the number of LCs/mm2 determined by  immunohistochemistry. The results are expressed as means ± SD  (n = 18). At all concentrations  tested, in cultures containing  anti-α6 antibodies, the frequency  of LCs did not differ significantly  from that found in fresh explants.  These same values were significantly (P <0.0001) higher than  those found in fresh explants cultured for 72 h in the absence of  antibody, or with an isotype-matched control antibody (A). Similar treatment of explant cultures with  anti-α4 antibody failed to result, at any concentration, in a significant increase in LC numbers compared with controls (B).
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Related In: Results  -  Collection

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Figure 2: The effect of anti-α6 and anti-α4 integrin antibodies on the migration of epidermal LCs in vitro. Skin explants were derived from the ears of naive mice and incubated on medium containing either (A) anti-α6 (GoH3) or (B) anti-α4 (PS/2) integrin antibodies at 10, 50, or 100 μg/ml (solid bars). Controls included explants cultured on medium containing either no antibody (open bars) or isotype-matched control antibody at 100 μg/ml (hatched bars). Epidermal sheets were prepared from fresh ear skin (0 h) and explants after 72 h of incubation and the number of LCs/mm2 determined by immunohistochemistry. The results are expressed as means ± SD (n = 18). At all concentrations tested, in cultures containing anti-α6 antibodies, the frequency of LCs did not differ significantly from that found in fresh explants. These same values were significantly (P <0.0001) higher than those found in fresh explants cultured for 72 h in the absence of antibody, or with an isotype-matched control antibody (A). Similar treatment of explant cultures with anti-α4 antibody failed to result, at any concentration, in a significant increase in LC numbers compared with controls (B).
Mentions: The differential expression of α6 and β4 integrin subunits by LCs and lymphoid DCs suggested that the α6β4 integrin may regulate the early stages of LC migration from the epidermis. Conversely, the higher level of expression of the α4 integrin subunit by DCs in comparison with LCs raises the possibility that α4 integrins may be involved in DC localization within the lymph node. To study the roles of these integrins in regulating the initial stages of LC migration from the epidermis, blocking antibodies to α6 and α4 integrin subunits were included in the culture medium of skin explants and their effects on LC migration were determined (Fig. 2). Affinity-purified anti-α6 (GoH3) and anti-α4 (PS/2) integrin antibodies were added to the culture medium at 10, 50, and 100 μg/ml. Control explants were incubated on culture medium containing isotype-matched control antibody at 100 μg/ml, or on culture medium alone. Explants were incubated for 72 h, the epidermis was removed, and the number of LCs/mm2 was determined. The number of LCs in fresh epidermis ranged from 1,000 to 1,200 LCs/mm2. After 72 h of incubation in culture medium, this number decreased by ∼70% to 300–350 LCs/mm2. Inclusion of 100 μg/ml MAC 193 (rat IgG2a) or HRPN (rat IgG2b) had no effect on the number of LCs remaining in the epidermis after 72 h and these antibodies were therefore used as isotype-matched controls.

Bottom Line: RGD-containing peptides were also without effect on LC migration from skin explants.In contrast, alpha 4 integrins, or other integrin-dependent interactions with fibronectin that are mediated by the RGD recognition sequence, did not influence LC migration from the epidermis.In addition, alpha 4 integrins did not affect the accumulation of LCs as DCs in draining lymph nodes.

View Article: PubMed Central - PubMed

Affiliation: Division of Cellular Immunology, National Institute for Medical Research, London, United Kingdom.

ABSTRACT
Topical exposure of mice to chemical allergens results in the migration of epidermal Langerhans cells (LCs) from the skin and their accumulation as immunostimulatory dendritic cells (DCs) in draining lymph nodes. Epidermal cell-derived cytokines have been implicated in the maturation and migration of LCs, but the adhesion molecules that regulate LC migration have not been studied. We hypothesized that integrin-mediated interactions with extracellular matrix components of the skin and lymph node may regulate LC/DC migration. We found that alpha 6 integrins and alpha 4 integrins were differentially expressed by epidermal LCs and lymph node DCs. A majority of LCs (70%) expressed the alpha 6 integrin subunit, whereas DCs did not express alpha 6 integrins. In contrast, the alpha 4 integrin subunit was expressed at high levels on DCs but at much lower levels on LCs. The anti-alpha 6 integrin antibody, GoH3, which blocks binding to laminin, completely prevented the spontaneous migration of LCs from skin explants in vitro and the rapid migration of LCs from mouse ear skin induced after intradermal administration of TNF-alpha in vivo. GoH3 also reduced the accumulation of DCs in draining lymph nodes by a maximum of 70% after topical administration of the chemical allergen oxazolone. LCs remaining in the epidermis in the presence of GoH3 adopted a rounded morphology, rather than the interdigitating appearance typical of LCs in naive skin, suggesting that the cells had detached from neighboring keratinocytes and withdrawn cellular processes in preparation for migration, but were unable to leave the epidermis. The anti-alpha 4 integrin antibody PS/2, which blocks binding to fibronectin, had no effect on LC migration from the epidermis either in vitro or in vivo, or on the accumulation of DCs in draining lymph nodes after oxazolone application. RGD-containing peptides were also without effect on LC migration from skin explants. These results identify an important role for alpha 6 integrins in the migration of LC from the epidermis to the draining lymph node by regulating access across the epidermal basement membrane. In contrast, alpha 4 integrins, or other integrin-dependent interactions with fibronectin that are mediated by the RGD recognition sequence, did not influence LC migration from the epidermis. In addition, alpha 4 integrins did not affect the accumulation of LCs as DCs in draining lymph nodes.

Show MeSH
Related in: MedlinePlus