6.3. Accumulation and distribution of type I procollagen and pN-collagen in sarcoidosis and fibrosing alveolitis as detected in immunohistochemical evaluation of open lung biopsy specimens (IV)

Small propeptide molecules disappear during the processing of histological specimens. Intracellularly, aminoterminal propeptides are attached to the procollagen molecules, and the antibodies to procollagen peptides obviously react with these. In the extracellular matrix, however, the aminoterminal domains still exist in pN-collagens as part of collagen fibres (Karttunen et al 1989, Risteli & Risteli 1990) and can be detected with antibodies to aminoterminal propeptides. The immunoreactivity of PINP detects intracellularly type I procollagen and extracellularly N-terminal extension of type I pN-collagen molecules in the collagen molecule, whereas PIIINP detects type III procollagen and type III pN-collagen correspondingly.

A notable portion of the previous data concerning the histological findings on collagens in lungs in interstitial disease are old and only pertain to mature collagens. No previous histochemical data are available on immunoreactivity for PIIINP and PINP in normal human lung. In the present study, type I pN-collagen was localised mainly in the ­middle of the interstitium of the alveolar wall, while type III pN-collagen was observed near the alveolar epithelium cells beneath the basement membrane. This suggests that type I pN-collagen may have a connection with the other extracellular matrix proteins of the interstitium, whereas type III pN-collagen may be associated with BM proteins, such as collagen IV and laminin. No sign of intracellular procollagens suggesting active production of collagens was detected, which is in line with the previous findings on the absence of immunoreactivity for PICP in normal human lung (McDonald et al. 1986, Roman et al. 1995). It is known that collagen I is localised in alveolar septa, whereas collagen III is localised in capillary walls and as aggregates of collagen in normal lung (Madri & Furthmayr 1980). Some collagen I has also been detected in alveolar walls, and especially, in pleural, peribronchial and perivascular connective tissue of normal lung (Bateman et al. 1981).

In sarcoidosis, the present study shows that procollagen type I was expressed intracellularly within spindle-shaped cells. It is not known if some of the fibroblasts of sarcoid granulomas could eventually be myofibroblasts. In some cases, type I procollagen was occasionally observed in multinucleated giant cells of sarcoid granulomas, suggesting that macrophages or cells derived from monocytes may produce type I procollagen in sarcoidosis. Taken together, the present findings on elevated PINP and PIIINP in ELF in sarcoidosis (I) suggest that alveolar macrophages may be able to synthesise type I and III collagens. The expression of type III pN-collagen was more prominent around lymphocytes than that of type pN-collagen around the granuloma. Type III procollagen was not observed intracellularly in sarcoidosis. Both collagens I and III have been found previously in sarcoid granulomas (Bateman et al. 1981, Peyrol et al. 1986). In the early stages they are distributed only peripherally but in the mature stages they can be found both around and within granulomas (Peyrol et al. 1986). Previously, immunoreactivity for PICP has been shown to concentrate in cells in the granulomas, where it was distributed within multinucleated giant cells, macrophages and surrounding fibroblasts and in fibroblasts scattered throughout the collagen fibrils between the granulomas. (Roman et al. 1995).

In the present study, the most prominent immunohistochemical accumulation of both procollagens was localised in the areas of alveolar epithelial damage, newly formed fibrosis and epithelial cell regeneration in fibrosing alveolitis. In the temporal process of fibrosing alveolitis, however, the expressions of type I pN-collagen and type III pN-collagen were very different. Type I pN-collagen was expressed mostly in the areas of damaged or deficient alveolar epithelium, while type III pN-collagen accumulated mainly at the newly formed fibrosis underneath the regenerative alveolar or bronchiolar-type epithelium. It is possible that the presence of type I pN-collagen may be important during epithelial damage, while type III pN-collagen operates mainly in the local healing process. Type III pN-collagen immunoreactivity was usually more abundant than the immunoreactivity of type I pN-collagen. The immunoreactivity for type III pN-collagen was strongest underneath the BM of regenerating metaplastic alveolar or bronchiolar type epithelium. Type III pN-collagen immunoreactivity was also present in the areas of denser fibrosis, and not only in the loose, newly formed fibromyxoid lesions. This suggests that immunoreactivity for type III pN-collagen rather than that for type I pN-collagen is present in areas of old fibrosis. Old fibrosis in fibrotic lung diseases has been shown to mainly compose of type I collagen (Bateman et al. 1981). It has to be emphasised that the type I procollagens and type I and III pN-collagens in the present study reflect mainly the active synthesis of collagen types I and III rather than the total amount of mature type I and III collagen. Collagen III is known to be located at subepithelial sites or in loosely arranged fibrils between densely packed type I collagen fibres in alveolar walls (Bateman et al. 1983, Takiya et al. 1983). Interstitial and alveolar fibroblasts stained intensely with PICP in active fibrosis, but not in clinically quiescent disease (McDonald et al. 1986). Not all of the patients with UIP had collagen-synthesising fibroblasts, and PICP-positive fibroblasts showed highly variable staining with PICP. Stained fibroblasts made up sub­epithelial clusters, so called fibroblast foci, near the air-tissue interface, and the cells were large and intensively stained. (Kuhn et al. 1989, Kuhn & McDonald 1991). Immunoreactivity of PINP has been detected in myofibroblasts in fibroblast foci in fibrosing alveolitis (Bensadoun et al. 1996). In the present study, immunoreactivity for type I procollagen was typically observed as intracellular spots within the clusters of fibroblasts and myofibroblasts in the fibroblast foci. These results are also in agreement with the previous findings of immunoreactivity of PICP (Kuhn et al. 1989).