Adenine, guanine, and inosine nucleotides of chick growth cartilage: Relationship between energy status and the mineralization process

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Abstract
The major aim of this investigation was to measure the nucleotide content of the developing chick epiphysis and to relate changes in nucleotide levels to chondrocyte maturation and the development of mineralization. Using a cryostat, sections of cartilage were isolated from the proximal head of the tibial growth cartilage, care being taken to preserve the metabolic integrity of the tissue. Sections were identified microscopically, pooled, and the nucleotide and nucleoside content of each sample determined by HPLC. Procedures used for the study were shown to minimize degradation of nucleotides. Their effectiveness was assessed through an evaluation of the rapid freezing technique and by examination of the effects of apatite on the recovery of endogenous and added nucleotides. Analysis of nucleotide levels in the growth cartilage indicated that chondrocytes undergo a profound change in energy metabolism during development and maturation. Thus, in the premineralized resting and proliferative zones, ATP and, to a lesser extent, GTP values were high, suggesting that the chondrocytes obtained metabolic energy through both glycolytic and mitochondrial oxidative processes. In the hypertrophic zone and in calcified cartilage, there was a profound decrease in the ATP concentration and a corresponding fall in the energy charge and the ATP/ADP ratios. The nucleotide levels in this zone indicated that there was increased reliance on nonoxidative metabolism. Measurement of nucleoside levels in premineralized cartilage suggested that there was little resynthesis of nucleotides through the salvage pathway. These observed changes in nucleotide values are consistent with earlier observations concerning chondrocyte redox and the low pO2 tension of the hypertrophic zone. The decrease in the level of ATP at the site of cartilage calcification lends support to the notion that nucleotides may serve as a source of phosphate ions for the initiation of mineralization.
Funding Information
  • National Institutes of Health (AR-34411, DE-02623, 06533)