Cytochemical Demonstration and Metabolic Significance of Reduced Diphospho-Pyridinenucleotide and Triphospho-Pyridinenucleotide Reductases in Human Gingiva

¹ Institute of Stomatological Research, Brookline, Massachusetts

Diphospho-Pyridinenucleotide nitro blue tetrazolium reductase (DPNH₂R) and Triphospho-Pyridinenucleotide nitro blue tetrazolium reductase (TPNH₂R) were demonstrated histochemically in human-gingiva biopsy samples. DPNH₂R exhibited considerably higher activity than TPNH₂R in the epithelium and the connective tissue. Positive reaction, as evidenced by intense purple and blue-purple formazan, was observed for each enzyme in the cytoplasm of all epithelial cells except those of parakeratin and keratin. However, the parakeratin layer consistently exhibited endogenous reducing activity characterized by pink-violet to violet colors. Cyanide intensification of formazan deposition was observed in all tissue sections. Keratin remained unreactive in all cases. Positive purple and blue-purple formazan DPNH₂R and TPNH₂R reactions were observed in blood vessels, nerves, and cells of the connective tissue. Characteristic contrasting patterns of distribution of the enzyme activities were found. DPNH₂R was most active in the basal epithelium, decreasing progressively to and through the stratum granulosum. TPNH₂R was most active in the stratum granulosum and decreased progressively to and through the stratum spinosum, becoming more reactive in the stratum germinativum. Desmosomes, tonofibrils, intercellular bridges, and nucleoli of the epithelial cells exhibited positive purple formazan-staining reactions for both enzymes, suggesting a broader metabolic role for these organelles than previously indicated. Many TPNH₂R specific blue-staining granules (probably keratohyalin) were observed in the stratum granulosum sparsely extending into the parakeratin. These granules were present in lesser amounts with the DPNH₂R disclosure. An oxidative metabolic pathway has been proposed for the formation of the keratin-precursor substance and keratin, whose reductive phase primarily is dependent upon TPN-mediated metabolism (e.g., the pentose shunt) and secondarily upon DPN-mediated metabolism (e.g., the Krebs cycle), and whose oxidative phase is dependent on a cyanide-sensitive oxidase, probably cytochrome oxidase, in the stratum granulosum, and an unidentified cyanide-sensitive oxidative system in the parakeratin.