Bovine spleen cathepsin B1 and collagenolytic cathepsin. A comparative study of the properties of the two enzymes in the degradation of native collagen

Etherington, David J.

doi:10.1042/bj1530199

Cited by 96 publications

(24 citation statements)

References 35 publications

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“…The reason for this is un clear, but may reflect the presence of other proteases being released by the neurons that are not inhibited by HS-LFA, yet are in volved in the degradation of collagen. A number of proteinases in addition to col lagénase are capable of degrading type I col lagen [Burleigh et al, 1974;Etherington, 1976;Starkey, 1977],…”

Section: Resultsmentioning

confidence: 99%

Neurite Penetration into Collagen Gels Requires Ca<sup>2+</sup>-Dependent Metalloproteinase Activity

Pittman¹,

Williams²

1989

Dev Neurosci

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Cultured sympathetic neurons from neonatal rats release a Ca²⁺-dependent metalloproteinase capable of degrading native and denatured type I collagen. A large fraction of the metalloproteinase activity is released from growth cones of growing neurites, which suggests that this proteinase may be involved in neurite penetration into collagen-rich extracellular matrices during development. In the present study, we investigated the possibility that neurite penetration into 3-dimensional collagen gels requires the Ca²⁺-dependent metalloproteinase. Sympathetic neurons were plated on top of collagen gels and the neuronal metalloproteinase activity was inhibited with the collagenase inhibitor HSCH₂CH[CH₂CH(CH₃)₂]-CO-Phe-Ala-NH₂ (HS-LFA). HS-LFA inhibited the Ca²⁺-dependent metalloproteinase activity, penetration of neurites into collagen gels and release of ³H from ³H-collagen gels in a dose-dependent manner over very similar concentration ranges (EC₅₀ = 150–200 nM). Highly significant correlations existed between neurite penetration into collagen gels and metalloproteinase activity (r = 0.99), and metalloproteinase activity and release of ³H from ³H-collagen gels (r = 0.99). Although HS-LFA inhibited the growth of neurites within collagen gels, it had no effect on neurite outgrowth on collagen films. Thus, the neuronal Ca²⁺-dependent metalloproteinase appears to be required for neurite penetration into and growth within 3-dimensional collagen matrices, but not for growth along a 2-dimensional collagen substrate.

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Section: Resultsmentioning

confidence: 99%

Neurite Penetration into Collagen Gels Requires Ca<sup>2+</sup>-Dependent Metalloproteinase Activity

Pittman¹,

Williams²

1989

Dev Neurosci

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“…6A). Since cathepsin B degrades collagen by an initial cleavage of telopeptides to release fibrils (27,28), our results suggest that the A-7 antibody (21) recognizes an epitope exposed during the initial unfolding of collagen by cathepsin B. Accordingly, we used the A-7 antibody and flow cytometry to examine during phagocytosis the temporal degradation of native collagen coated on the beads.…”

Section: Fig 2 Phagosome-associated Proteinsmentioning

confidence: 99%

“…Phagosome Acidification Is Required for Collagen Degradation but Not Phagosomal Maturation-Since cathepsin B requires an acidic phagosomal compartment (28,29) to effect collagen degradation (27), we examined the pH of the beadassociated compartment. To calibrate the intracellular response of FITC-streptavidin-biotinylated collagen beads to changes in pH, we used nigericin-KCl (30), which enabled us to equilibrate the pH in the cell with that of the medium (Fig.…”

Section: Figmentioning

confidence: 99%

A Novel Model System for Characterization of Phagosomal Maturation, Acidification, and Intracellular Collagen Degradation in Fibroblasts

Arora

Manolson²,

Downey³

et al. 2000

Journal of Biological Chemistry

104

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Intracellular collagen degradation by fibroblasts is an important but poorly understood pathway for the physiological remodeling of mature connective tissues. The objective of this study was to determine whether gingival fibroblasts that express endogenous ␣ 2 ␤ 1 integrin, the collagen receptor, would exhibit the cellular machinery required for phagosomal maturation and collagen degradation. There was a time-dependent increase of collagen bead internalization and a time-dependent decrease of bead-associated ␣ 2 ␤ 1 integrin after initial bead binding. ␤-Actin and gelsolin associated transiently with beads (0 -30 min) followed by LAMP-2 (60 -240 min) and cathepsin B (30 -240 min). Cytochalasin D prevented phagosome formation and also prevented the sequential fusion of early endosomes with lysosomes. Collagen bead-associated pH was progressively reduced from 7.25 to 5.4, which was contemporaneous with progressive increases in degradation of bead-associated collagen (30 -120 min). Concanamycin blocked acidification of phagolysosomes and collagen degradation but not phagosome maturation. Phagosomal acidification was partly dependent on elevated intracellular calcium. These studies demonstrate that the cellular machinery required for intracellular collagen degradation in fibroblasts closely resembles the vacuolar system in macrophages.Phagocytosis is central to the uptake and degradation of microorganisms as well as damaged or senescent cells and is therefore an essential process in host defense, tissue remodeling, and inflammation. Although "professional" phagocytic cells such as macrophages and neutrophils have been studied in considerable depth (1), several types of nonphagocytic cells such as epithelial cells and fibroblasts can internalize particles and matrix proteins in vivo and in culture. Collagen fibril phagocytosis is thought to be an important pathway for physiological degradation of extracellular matrix in mature connective tissues (2). Uterine, wounded dermal, and periodontal connective tissues exhibit rapid physiological turnover of matrix proteins, processes that are mediated by the intracellular degradation pathway (3). While in inflamed periodontal sites, extracellular matrix metalloproteinases are believed to be responsible for the bulk degradation of connective tissues (4), in normal turnover, fibroblasts are thought to use solely the intracellular vacuolar system for focal proteolysis of collagen (3,5). However, at present, the vacuolar system that mediates intracellular collagen degradation is incompletely characterized.In professional phagocytes, the phagocytic process is initiated by binding of particles to receptors on the plasma membrane, an event that subsequently generates a phagocytic signal (6). In fibroblasts, the initial internalization of intracellular collagen degradation is a specific process that is mediated by the adhesive interactions between ligand and collagen receptors (i.e. ␣ 2 ␤ 1 integrins; Ref. 7). However, little is known about the regulation of the downstream events and...

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“…Cathepsin B emerged in the breakthrough peak. Cathepsin L, absorbed on the ionexchange resin, was eluted with 0.15 M phosphate buffer (pH 7.50) according to [11 ]. Cathepsin L was isolated by a single gel filtration on Sephadex G-75 (superfine).…”

Section: Preparation Of Human Liver Cathepsin Lmentioning

confidence: 99%

Inhibition of human liver cathepsin L by α‐thiol proteinase inhibitor

Pagano¹,

Engler²

1982

FEBS Letters

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Bovine spleen cathepsin B1 and collagenolytic cathepsin. A comparative study of the properties of the two enzymes in the degradation of native collagen

Cited by 96 publications

References 35 publications

Neurite Penetration into Collagen Gels Requires Ca<sup>2+</sup>-Dependent Metalloproteinase Activity

Neurite Penetration into Collagen Gels Requires Ca<sup>2+</sup>-Dependent Metalloproteinase Activity

A Novel Model System for Characterization of Phagosomal Maturation, Acidification, and Intracellular Collagen Degradation in Fibroblasts

Inhibition of human liver cathepsin L by α‐thiol proteinase inhibitor

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