Human skin was labeled with purified antibodies against type I and m collagens and against their extension aminopropeptides by using the ferritin technique. Both aminopropeptides were visualized mainly along thin collagenous fibrils (diameter, 20-40 nm) and rarely in nonfibrillar regions of the skin. The labeling showed a periodicity of 60-65 nm resembling the D (67 nm) stagger of collagen molecules. Blocking of antibodies with aminopropeptides and treatment of tissues with procollagen NH2-terminal protease abolished labeling. Antibodies against type I collagen uniformly labeled -80% of the fibrils (diameter, 20-80 nm), while reaction with antibodies against type m collagen was restricted to thin fibrils. It is currently thought that the aminopropeptides of procollagen molecules are cleaved after they are released from the cell and before fibril formation. Our data indicate that aminopropeptides are removed at the fibrillar level and that fibril growth can be regulated by extracellular procollagen processing. Type I and III collagens are major components of the dermis and are organized into fibrils varying considerably in diameter. Distinct growth of fibrils is observed during development and may be disturbed in certain skin diseases (1). The mechanism of controlling this process is unknown. It has also not been established whether type I and III collagens are present in the same fibrils. Both collagens can form D-staggered fibrils in vitro showing identical cross-striations when examined in the electron microscope (2). Because specific antibodies against various collagens and procollagens are available (3, 4), it is feasible to identify distinct types of collagenous proteins at the ultrastructural level in situ, as was recently done for type III collagen (5).Interstitial collagens are synthesized in the form of procollagens that possess additional extension amino-and carboxypropeptides. Each precursor-specific peptide is removed by specific proteases (procollagen NH2-terminal and COOH-terminal proteases) presumably after release of procollagens from the cell (6). Cell and organ culture studies indicated different kinetics in the processing of type I and III procollagens (7-9). After cleavage, the aminopropeptides persist for some time in the extracellular space, as shown by immunofluorescence staining with antibodies against these peptides (3, 4).A functional role for extension aminopropeptides in the control of fibril growth was first suggested in studies of dermatosparactic animals, which, due to a defective NH2-terminal protease, accumulate an intermediate form of type I procollagen (pN-collagen) in the skin and other organs (10, 11). The collagenous fibrils appeared thin and hieroglyphic (12, 13) and could be stained with antibodies against the aminopropeptide by using a ferritin label (13). Small amounts of type I and III pN-collagens could also be extracted from the skin of growing animals (14-18). It was suggested (3, 14, 19) that these aminopropeptides are structural elements ofimmature or th...
Fibroblasts derived from a skin biopsy of a patient with scleroderma in the sclerotic stage were shown to have a higher rate of DNA synthesis, and to synthesize more collagen than fibroblasts from a healthy control. The addition of procollagen peptides to the culture medium of scleroderma fibroblasts almost normalized the collagen synthesis. This observation indicates that the mechanism for the regulation of collagen synthesis by feed back inhibition of prollagen peptides is functioning in this disease. It is suggested that the level of biologically active procollagen peptides is lowered.
Scleroderma skin and the subcutaneous tissue was studied by indirect immunofluorescence with specific antibodies against interstitial collagens and procollagens, against fibronectin and against the basement membrane proteins Type IV collagen and laminin. Staining for Type I procollagen and fibronectin was distinctly increased in the lower dermis and subcutaneous tissue. When compared with normal skin the data suggests that fibrosis may start around capillaries and in close proximity to adipose cells. Additional changes in the distribution to Type IV collagen and laminin were found in some patients and probably reflect the alterations in small blood vessels.
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