Immunofluorescent localization of structural collagen types in endochondral fracture repair

Abstract

A nonimmobilized rat tibial fracture model of endochondral osseous repair was examined for the unique localizations of specific collagen genetic types. At various stages of the healing process, the demineralized callus was reacted with immunofluorescent antibodies directed against the type specific forms of matrix collagen. Type III collagen rapidly appeared (day 8-10) and remained in the primitive mesenchymal callus until remodeled. It was particularly prominent in the highly vasoformative regions and the pericallus encapsulation but not present in preexisting cortical and neoformed lamellar bone. The type II collagen, a marker of cartilage, was uniquely located only in areas of chondroid differentiation and calcification. Type II collagen was absent from all bone and was not identified beneath the repairing intact periosteum. The differentiating chondrocytes synthesized type II collagen on an underlayer of type III collagen already within the mesenchymal matrix. From these studies of genetically unique collagen markers, it appears that only in areas of motion or anoxia does an intermediate of chondroid tissue appear. The utilization of specific type II and type III collagen immunofluorescent antibodies has facilitated the understanding of the fracture repair process and has acted as an indicator for unique matrix components.

Major fibrillar collagens and fibronectin in an experimental nonunion: An immunohistochemical study

“…The collagens produced in the matrix of a healing fracture can be quantitated by biochemical methods (Penttinen 1972, Paavolainen et al 1989). However, these methods cannot localize the proteins, and therefore the distribution of specific collagens during fracture healing was studied by immunohistochemistry (Lane et al 1986). The early phases of fracture healing in this experimental model appeared to follow closely a normal repair pattern, as described in previously published immunohistochemical studies using a rat tibial fracture model (Lane et al 1986, Ashhurst 1990.…”

“…However, these methods cannot localize the proteins, and therefore the distribution of specific collagens during fracture healing was studied by immunohistochemistry (Lane et al 1986). The early phases of fracture healing in this experimental model appeared to follow closely a normal repair pattern, as described in previously published immunohistochemical studies using a rat tibial fracture model (Lane et al 1986, Ashhurst 1990. During the first week of the healing process, loose connective tissue is found at the fracture site, containing mostly collagen 111 and also collagen I, which is expressed throughout the normal fracture healing process.…”

“…During the first week of the healing process, loose connective tissue is found at the fracture site, containing mostly collagen 111 and also collagen I, which is expressed throughout the normal fracture healing process. New periosteum is developing along the periphery of loose connective tissue and contains collagens I, 111 and V. Collagen V is mainly found in the same tissues as collagens I and 111 in close contact with blood vessels and is thought to be located in the fibrils of collagens I and 111, controlling the diameter of the fibers (Lane et al 1986, Ashhurst 1990, Liu et al 1995.…”

“…It is normally the last major collagen synthesized in the endochondral fracture healing cascade and found in the matrix of the interfragmentary zone (Lane et al 1986, Ashhurst 1990. Collagen I1 was formed on the fibrous matrix, and type I11 and type V collagens remained.…”

Alteration of fracture stability influences chondrogenesis, osteogenesis and immigration of macrophages

“…Fracture healing is a complex phenomenon involving the coordinated participation of immigration, differentiation and proliferation of inflammatory cells, angioblasts, fibroblasts, chondroblasts and osteoblasts which synthesize and release bioactive substances of extracellular matrix components, e.g., collagen I, 11, 111, V or IX, growth factors [Transforming Growth Factor-p1 (TGF-Pl), Platelet-derived Growth Factor (PDGF) or Fibroblast Growth Factor (FGF)] and intercellular mediators (Interleukin-1, Interleukin-6, Granulocytemacrophage Colony Stimulating Factor or Macrophage-colony Stimulating Factor) [2,3,10,20,28,31]. Cell proliferation during the healing of fractured rat femora, *Corresponding author.…”