Odontoblast alkaline phosphatases and Ca2+ transport

Area of Science:

• Biochemistry
• Cell Biology
• Mineralized Tissue Biology

Background:

• Nonspecific alkaline phosphatase (APase) is present in various calcifying tissues like bone and cartilage.
• APase is implicated in the enzymatic degradation of inorganic pyrophosphate (PPi).
• The presence and function of ATP-degrading enzymes in hard tissue formation require further elucidation.

Purpose of the Study:

• To identify and characterize enzymes involved in phosphate compound degradation in calcifying tissues.
• To investigate the localization and potential roles of APase and Ca2+-ATPase in odontoblasts.
• To explore the relationship between APase and Ca2+-ATPase in hard tissue formation.

Main Methods:

• Enzyme assays using p-nitrophenylphosphate (p-NPP) and specific inhibitors.
• Solubilization, separation, and characterization of phosphatases from odontoblasts.
• Subcellular fractionation and electron microscopy (EM) histochemistry with mild fixation techniques.

Main Results:

• Two ATP-degrading enzymes were identified: classical APase and Ca2+-ATPase.
• Both APase and Ca2+-ATPase were primarily localized in the membranes of intercellular vesicles within odontoblasts.
• Enzyme activity was significantly reduced with standard fixation, but preserved with a mild technique.
• No enzyme activity was detected in the cell membranes.

Conclusions:

• Hard tissue-forming cells possess at least two enzymes, APase and Ca2+-ATPase, that degrade phosphate compounds at alkaline pH.
• APase and Ca2+-ATPase exhibit an intimate relationship within these tissues.
• Ca2+-ATPase may play a role in ATP-dependent, intravesicular calcium transport, potentially explaining its function in calcification.