6-Methylsulfinylhexyl isothiocyanate activates carbonic anhydrase-dependent HCO3 -/H+/Na+/Ca2+ transport via SLC4As-NHE-NCX-PMCA axis in odontoblasts.

Ca2+-permeable transient receptor potential channels, which play an important role in the developmental/reactionary dentinogenesis by odontoblasts, are sensitive to wasabi sulfinyls. We investigated the effects of the wasabi sulfinyls 6-methylsulfinylhexyl isothiocyanate (6-MSITC) and its eight derivatives on ion transport mechanisms which promote mineralisation by odontoblasts. 6-MSITC significantly increased the mineralisation efficiency in cultured odontoblasts, and we also observed a significant increase in medium pH. Inhibitors of carbonic anhydrase (CA) and plasma membrane Ca2+-ATPase (PMCA) significantly reduced 6-MSITC-induced mineralisation. Odontoblasts expressed the HCO3 --transporting solute carrier family 4 (SLC4A) members SLC4A1, SLC4A2, SLC4A3, SLC4A4, SLC4A8 and SLC4A9, as well as CA I and CA II. 6-MSITC enhanced reactionary dentinogenesis beneath the cavities prepared on rat mandibular first molars. We recorded 6-MSITC-induced outward currents, which were suppressed by inhibitors of CA, Na+-H+ exchanger (NHE), and Na+-Ca2 + exchanger (NCX). These results indicated that 6-MSITC has a strong ability to form reactionary dentin by activating or upregulating intracellular CA and electrically neutral HCO3 -/H+ transport via SLC4As/NHE. Exchanging Na+ with H+ using NHE resulted in the reversal of the transmembrane Na+ gradient. This activated the Ca2+ influx mode of NCX, and the subsequent accumulation of intracellular Ca2+ was then extruded by PMCA activity to produce reactionary dentin. Thus, 6-MSITC activates CA-mediated SLC4As-NHE-NCX-PMCA coupling and is useful in dentin regenerative medicine. KEY POINTS: Ca2+ signalling in odontoblasts plays an important role not only in developmental/reactionary dentinogenesis, but also in the generation of dentinal (tooth) pain. The wasabi sulfinyls including 6-methylsulfinylhexyl isothiocyanate (6-MSITC) promote mineralisation by odontoblasts and significantly increase pH in medium with cultured odontoblasts. We showed that 6-MSITC has a strong ability to form reactionary dentin through the upregulation and activation of intracellular carbonic anhydrase (CA) and electrically neutral HCO3 -/H+ transport by members of the HCO3 --transporting solute carrier family 4 (SLC4As) and Na+-H+ exchanger (NHE). Na+ accumulation by NHE activity resulted in the reversal of the transmembrane Na+ gradient. This activated the Ca2+ influx mode of the Na+-Ca2 + exchanger (NCX), resulting in the accumulation of intracellular Ca2+. It was then extruded using plasma membrane Ca2+-ATPase (PMCA) to produce reactionary dentin. 6-MSITC activates CA-mediated SLC4As-NHE-NCX-PMCA coupling and is useful in dentin regenerative medicine.