Type VI collagen is phagocytosed by fibroblasts and digested in the lysosomal apparatus: involvement of collagenase, serine proteinases and lysosomal enzymes.
Type VI collagen is present in most connective tissues, where it is considered to play a crucial role in the attachment of cells to the extracellular matrix and/or in the three-dimensional organization of the collagen meshwork. Although some information is available on its formation, the mechanisms involved in its degradation are not understood. Here, we present evidence for lysosomal digestion of type VI collagen by fibroblasts of periosteal explants. In the lysosomal apparatus of these cells, broad-banded filamentous aggregates characterized by 100-nm periodicity were found, which proved to consist of type VI collagen as indicated by their stainability with anti-type VI collagen antibodies. By interfering with synthesis (ascorbate or alpha, alpha-dipyridyl), intracellular translocation of collagen-containing vesicles (colchicine) as well as phagocytosis (cytochalasin B), it was shown that the intracellular broad-banded type VI collagen represented phagocytosed material. In the presence of acidotropic agents (NH4Cl and methylamine) the amount of intracellular type VI collagen increased significantly (5- to 10-fold), suggesting that a rise of pH in the endosomal/lysosomal apparatus causes inhibition of its degradation. By using a variety of proteinase inhibitors, it was found that inhibition of collagenase (when used in combination with NH4Cl), or inhibition of cysteine proteinases (both with and without NH4Cl), resulted in an increased amount of intracellular type VI collagen, whereas inhibition of serine proteinases significantly lowered the level of intracellular type VI collagen. The data presented are the first to indicate a pathway by which type VI collagen degradation may occur: fibroblasts phagocytose type VI collagen and subsequently digest this collagen in their lysosomal apparatus. Degradation depends on the activity of several enzymes, among them collagenase and serine proteinases, probably exerting their activity in the extracellular space just before the actual internalization. After uptake, digestion involves pH-sensitive lysosomal enzymes, including those belonging to the class of cysteine proteinases.
Everts
V
V
Laboratory of Cell Biology and Histology, Academic Medical Center, Amsterdam, The Netherlands.
Korper
W
W
Niehof
A
A
Jansen
I
I
Beertsen
W
W
D000818:Animals; D003094:Collagen / Q000737:chemistry / Q000378:metabolism*; D017364:Collagenases / Q000378:metabolism*; D005347:Fibroblasts / Q000502:physiology* / Q000648:ultrastructure; D006863:Hydrogen-Ion Concentration; D007150:Immunohistochemistry; D008247:Lysosomes / Q000201:enzymology*; D010521:Periosteum / Q000166:cytology / Q000648:ultrastructure; D010587:Phagocytosis; D011480:Protease Inhibitors / Q000494:pharmacology; D011817:Rabbits; D012697:Serine Endopeptidases / Q000378:metabolism*
D011480:Protease Inhibitors; D003094:Collagen; D012697:Serine Endopeptidases; D017364:Collagenases