Type IIA procollagen in development of the human intervertebral disc: Regulated expression of the NH2-propeptide by enzymic processing reveals a unique developmental pathway

Type II collagen can be synthesized in two forms generated by alternative s plicing of the precursor mRNA Type IIA procollagen, which contains a cystei ne-rich domain in the NH2-propeptide (exon 2), is produced by precartilage and noncartilage epithelial and mesenchymal cells, and type IIB procollagen , without the cysteine-rich domain, is characteristic of chondrocytes. Mice lacking type II collagen fail to develop intervertebral discs. We have pre viously shown that the human intervertebral disc and notochord synthesize p rimarily the type IIA form of procollagen. Therefore, we investigated the d istribution of type IIA procollagen during early disc development in humans . By processes of radioactive in situ hybridization and fluorescence immuno histochemistry, we localized mRNA and protein of type IIA procollagen, type I collagen, and type III collagen in fetal intervertebral disc specimens r anging from day 42 (embryonic stage 17) to day 101 (week 14.5) of gestation , Antibodies to the three distinct domains of type IIA procollagen: the NH2 -propeptide, the fibrillar domain, and the COOH-propeptide were used. The e arliest stage of developing intervertebral disc (42 days, stage 17) was cha racterized by diffuse synthesis of types I and III collagens in the dense z one (intervertebral area) and synthesis of type IIA procollagen by the chon drocyte progenitor cells surrounding the disc. The notochord cells synthesi zed and deposited into the notochordal sheath all three fibrillar collagens . By 54 days (stage 22), the developing disc was clearly divided into three regions: 1.) the outer annulus, characterized by synthesis and deposition of types I and III collagens; 2.) the inner annulus, characterized by synth esis and deposition of type IIA collagen containing the NH2-propeptide but devoid of the COOH-propeptide (pN-procollagen); and 3.) the notochord, the cells of which synthesized and deposited of all three fibrillar collagens. In later stages of fetal development (72-101 days), a change in type IIA pr ocollagen processing was observed in the cells of the inner annulus: even t hough these cells continued to synthesize type IIA procollagen, they deposi ted into the extracellular matrix (ECM) only the processed fibrillar domain , with the NH2-propeptide removed. This finding indicates that there is a d evelopmentally regulated change in the processing of type IIA procollagen N H,propeptide in the cells of the inner annulus. This mechanism is in contra st to previously shown developmental regulation of the cysteine-rich domain of the NH2-propeptide by alternative splicing of the precursor mRNA Althou gh the cells of the inner annulus have been identified as chondrocytes, bas ed on their shape and synthesis of characteristic ECM components, they appe ar to represent a distinct developmental pathway characterized by their syn thesis and differential processing of type IIA procollagen. This developmen tal pattern may prove important for disc regeneration. (C) 2001 Wiley-Liss, Inc.