Wound Healing and Collagen Formation

Abstract

The uptake, intracellular transport, and secretion of protein by guinea pig wound fibroblasts was studied by electron microscope radioautography using L-proline-S,4-H a as a tracer. Experiments were performed to determine the curve of concentration of free amino acid in the blood after intraperitoneal administration of the labeled proline. Radioautographs were quantitatively analyzed and the concentration of isotope, in grains per unit area, was determined for the following cellular and extracellular compartments: ergastoplasm, Golgi complex, peripheral cytoplasmic structures, and collagen. The concentration of label, expressed as number of grains per unit area of each subcellular system, reveals the period during which each cellular compartment is maximally labeled, and presents a clearer picture of the passage of the label through each of these compartments. The data demonstrate appearance of the label at maximum concentration in the ergastoplasm 15 minutes after injection, and this compartment remains maximally labeled for 2 hours. In the Golgi complex, concentration is not maximal until 60 minutes after injection of isotope, and appears to decrease before or at about the same rate as that of the ergastoplasm. The present experiment is consistent with previous light microscope radioautographic studies, and no storage phase was found in the fibroblasts. The findings are not simply consistent with a direct precursor-product relationship between the contents of the ergastoplasm and those of the Golgi complex. Morphologic observations of regions in the fibroblast interpretable as possible sites of communication between the ergastoplasm and the extracellular space, together with the kinetic studies, permit the suggestion of an alternate pathway of passage of at least some of the synthesized protein directly from the ergastoplasmic cisternae to the cell exterior.

Location of procollagen in chick corneal and tendon fibroblasts with ferritin-conjugated antibodies.

“…Its importance has been the subject of debate in the case of procollagen, since there is not always an obvious system of condensed secretory granules present, and since there have been indications from autoradiography that, in fibroblasts at least, transport through the Goigi apparatus would have to be rapid to account for the relatively small proportion of [SH]prolinelabeled product found in this organelle (27,28). …”

“…It is possible that they represent the compartment in which procollagen is transported to the cell surface (11,12,24,26). It is not possible either with this technique or with autoradiography to rule out direct transport of some procollagen from the endoplasmic reticulum to the exterior via small vesicles (27). The antibody-labeled small vesicles observed in well-fragmented Golgi zones, however, may have been transporting procollagen to the Golgi zone.…”

“…The intracellular transport of the secretory protein, procollagen, and the role of the Golgi apparatus in this function have been the subject of much discussion and study (3,5,11,12,24,(26)(27)(28)34). Procollagen is unusual insofar as it is secreted by several distinct cell types.…”

“…The question of whether passage of procollagen through the Golgi apparatus is obligatory in connective tissue cells was first approached by electron microscope autoradiography using [SH]proline; the large Golgi vacuoles of chondrocytes were readily labeled (25), and location in the Golgi zone of fibroblasts was also observed (27). However, quantitative electron microscope autoradiographic studies gave conflicting results as to the amount of proline-rich protein passing from endoplasmic reticulum into the Golgi vacuoles of THE JOURNAL OF CELL BIOLOGY • VOLUME 65, 1975 • pages 75-87several cell types (12,27,28,34) and suggested that two intracellular pathways might exist for collagen transport (27,28).…”

“…However, quantitative electron microscope autoradiographic studies gave conflicting results as to the amount of proline-rich protein passing from endoplasmic reticulum into the Golgi vacuoles of THE JOURNAL OF CELL BIOLOGY • VOLUME 65, 1975 • pages 75-87several cell types (12,27,28,34) and suggested that two intracellular pathways might exist for collagen transport (27,28).…”