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uPARAP/Endo180 is essential for cellular uptake of collagen and promotes fibroblast collagen adhesion.

Engelholm LH, List K, Netzel-Arnett S, Cukierman E, Mitola DJ, Aaronson H, Kjøller L, Larsen JK, Yamada KM, Strickland DK, Holmbeck K, Danø K, Birkedal-Hansen H, Behrendt N, Bugge TH - J. Cell Biol. (2003)

Bottom Line: However, the molecular mechanisms governing this pathway are poorly understood.Here, we show that the urokinase plasminogen activator receptor-associated protein (uPARAP)/Endo180, a novel mesenchymally expressed member of the macrophage mannose receptor family of endocytic receptors, is a key player in this process.These studies identify a central function of uPARAP/Endo180 in cellular collagen interactions.

View Article: PubMed Central - PubMed

Affiliation: Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
The uptake and lysosomal degradation of collagen by fibroblasts constitute a major pathway in the turnover of connective tissue. However, the molecular mechanisms governing this pathway are poorly understood. Here, we show that the urokinase plasminogen activator receptor-associated protein (uPARAP)/Endo180, a novel mesenchymally expressed member of the macrophage mannose receptor family of endocytic receptors, is a key player in this process. Fibroblasts from mice with a targeted deletion in the uPARAP/Endo180 gene displayed a near to complete abrogation of collagen endocytosis. Furthermore, these cells had diminished initial adhesion to a range of different collagens, as well as impaired migration on fibrillar collagen. These studies identify a central function of uPARAP/Endo180 in cellular collagen interactions.

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uPARAP/Endo180 promotes cell adhesion to collagen. Dermal fibroblasts from uPARAP/Endo180−/− (black bars) or littermate uPARAP/Endo180+/+ neonates (gray bars) were allowed to attach for 30 min at 37°C to tissue culture wells coated with collagens type I (A), type II (B), type IV (C), type V (D), total tendon collagen (E), or serum proteins from 10% FCS (F). The number of cells adhering to the different matrices was determined by an MTT assay and expressed as the percentage of adherent uPARAP/Endo180+/+ cells. Error bars indicate SDs of quadruplicate determinations. The data are representative of results obtained with three different sets of uPARAP/Endo180−/− and littermate control fibroblasts. Asterisk in A, P < 0.03; C, P < 0.002; D, P < 0.0004; and E, P < 0.002.
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fig3: uPARAP/Endo180 promotes cell adhesion to collagen. Dermal fibroblasts from uPARAP/Endo180−/− (black bars) or littermate uPARAP/Endo180+/+ neonates (gray bars) were allowed to attach for 30 min at 37°C to tissue culture wells coated with collagens type I (A), type II (B), type IV (C), type V (D), total tendon collagen (E), or serum proteins from 10% FCS (F). The number of cells adhering to the different matrices was determined by an MTT assay and expressed as the percentage of adherent uPARAP/Endo180+/+ cells. Error bars indicate SDs of quadruplicate determinations. The data are representative of results obtained with three different sets of uPARAP/Endo180−/− and littermate control fibroblasts. Asterisk in A, P < 0.03; C, P < 0.002; D, P < 0.0004; and E, P < 0.002.

Mentions: We used a standard cell adhesion assay, in which cells are allowed to attach briefly to immobilized collagens, to investigate if the lack of uPARAP/Endo180 could directly influence cell adhesion. Loss of uPARAP/Endo180 resulted in a 50% reduction in the adhesion of fibroblasts to type V collagen-coated surfaces (Fig. 3 D). This impairment was observed in comparisons of several independent isolates of uPARAP/Endo180−/− fibroblasts with littermate control uPARAP/Endo180+/+ fibroblasts (unpublished data). Interestingly, uPARAP/Endo180−/− cells also demonstrated a reduced ability to adhere to other immobilized collagens, including purified types I and IV collagen and total tendon collagen (Fig. 3, A, C, and E), although the reduction in these cases was less dramatic. In contrast, uPARAP/Endo180 deficiency did not affect the adhesion of fibroblasts to a noncollagen substrate such as fibronectin (Fig. S2 C) or a surface coated with total serum proteins (Fig. 3 F). A time-course study of cell adhesion to collagen V and collagen I showed that the effect of uPARAP/Endo180 deficiency was limited to the initial phase (15–30 min after plating), and wild-type and targeted cells reached the same level of adhesion after 1–2 h (unpublished data).


uPARAP/Endo180 is essential for cellular uptake of collagen and promotes fibroblast collagen adhesion.

Engelholm LH, List K, Netzel-Arnett S, Cukierman E, Mitola DJ, Aaronson H, Kjøller L, Larsen JK, Yamada KM, Strickland DK, Holmbeck K, Danø K, Birkedal-Hansen H, Behrendt N, Bugge TH - J. Cell Biol. (2003)

uPARAP/Endo180 promotes cell adhesion to collagen. Dermal fibroblasts from uPARAP/Endo180−/− (black bars) or littermate uPARAP/Endo180+/+ neonates (gray bars) were allowed to attach for 30 min at 37°C to tissue culture wells coated with collagens type I (A), type II (B), type IV (C), type V (D), total tendon collagen (E), or serum proteins from 10% FCS (F). The number of cells adhering to the different matrices was determined by an MTT assay and expressed as the percentage of adherent uPARAP/Endo180+/+ cells. Error bars indicate SDs of quadruplicate determinations. The data are representative of results obtained with three different sets of uPARAP/Endo180−/− and littermate control fibroblasts. Asterisk in A, P < 0.03; C, P < 0.002; D, P < 0.0004; and E, P < 0.002.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2172772&req=5

fig3: uPARAP/Endo180 promotes cell adhesion to collagen. Dermal fibroblasts from uPARAP/Endo180−/− (black bars) or littermate uPARAP/Endo180+/+ neonates (gray bars) were allowed to attach for 30 min at 37°C to tissue culture wells coated with collagens type I (A), type II (B), type IV (C), type V (D), total tendon collagen (E), or serum proteins from 10% FCS (F). The number of cells adhering to the different matrices was determined by an MTT assay and expressed as the percentage of adherent uPARAP/Endo180+/+ cells. Error bars indicate SDs of quadruplicate determinations. The data are representative of results obtained with three different sets of uPARAP/Endo180−/− and littermate control fibroblasts. Asterisk in A, P < 0.03; C, P < 0.002; D, P < 0.0004; and E, P < 0.002.
Mentions: We used a standard cell adhesion assay, in which cells are allowed to attach briefly to immobilized collagens, to investigate if the lack of uPARAP/Endo180 could directly influence cell adhesion. Loss of uPARAP/Endo180 resulted in a 50% reduction in the adhesion of fibroblasts to type V collagen-coated surfaces (Fig. 3 D). This impairment was observed in comparisons of several independent isolates of uPARAP/Endo180−/− fibroblasts with littermate control uPARAP/Endo180+/+ fibroblasts (unpublished data). Interestingly, uPARAP/Endo180−/− cells also demonstrated a reduced ability to adhere to other immobilized collagens, including purified types I and IV collagen and total tendon collagen (Fig. 3, A, C, and E), although the reduction in these cases was less dramatic. In contrast, uPARAP/Endo180 deficiency did not affect the adhesion of fibroblasts to a noncollagen substrate such as fibronectin (Fig. S2 C) or a surface coated with total serum proteins (Fig. 3 F). A time-course study of cell adhesion to collagen V and collagen I showed that the effect of uPARAP/Endo180 deficiency was limited to the initial phase (15–30 min after plating), and wild-type and targeted cells reached the same level of adhesion after 1–2 h (unpublished data).

Bottom Line: However, the molecular mechanisms governing this pathway are poorly understood.Here, we show that the urokinase plasminogen activator receptor-associated protein (uPARAP)/Endo180, a novel mesenchymally expressed member of the macrophage mannose receptor family of endocytic receptors, is a key player in this process.These studies identify a central function of uPARAP/Endo180 in cellular collagen interactions.

View Article: PubMed Central - PubMed

Affiliation: Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
The uptake and lysosomal degradation of collagen by fibroblasts constitute a major pathway in the turnover of connective tissue. However, the molecular mechanisms governing this pathway are poorly understood. Here, we show that the urokinase plasminogen activator receptor-associated protein (uPARAP)/Endo180, a novel mesenchymally expressed member of the macrophage mannose receptor family of endocytic receptors, is a key player in this process. Fibroblasts from mice with a targeted deletion in the uPARAP/Endo180 gene displayed a near to complete abrogation of collagen endocytosis. Furthermore, these cells had diminished initial adhesion to a range of different collagens, as well as impaired migration on fibrillar collagen. These studies identify a central function of uPARAP/Endo180 in cellular collagen interactions.

Show MeSH
Related in: MedlinePlus