Program

PO4-3-53

Zinc promotes differentiation and proliferation of MC3T3-E1 cells through activation of Ca2+-activatide K+ channels

[Speaker] Takashi Yoshida:1
[Co-author] Mengkai Guang:1,2,3, Kaori Takahashi:1, Takashi Nakamura:1, Ping Gong:2, Minoru Wakamori:1
1:Division of Molecular Pharmacology and Cell Biophysics, Department of Oral Biology, Graduate School of Dentistry, Tohoku University, Japan, 2:Department of Oral Implantology, State Key Laboratory of Oral Diseases, West China College of Stomatology, Sichuan University, China, 3:China-Japan Friendship Hospital, China

Zinc, as the divalent cation (Zn2+) and the second most abundant trace metal in the human body, plays pivotal roles in the proliferation and differentiation of osteoblasts. It is well known that the Ca2+-activated K+ (KCa) channels regulate membrane potential and calcium signaling in the non-excitable cells. Although KCa channels are expressed in osteoblast-like cells including MC3T3-E1, their functions in osteoblasts have not been fully clarified. The molecular mechanisms of Zn2+ induced proliferation and differentiation of osteoblasts also remain unclear. In this research, the mechanisms of proliferation and differentiation through KCa channel activation by Zn2+ in MC3T3-E1 cells are investigated.
To clarify the physiological roles of Zn2+ in osteoblasts, we applied whole-cell mode of the patch-clamp technique as well as calcium and zinc imaging in MC3T3-E1 cells. One millimolar of Zn2+ hyperpolarized membrane potential under current-clamp mode and induced outward K+ currents and blockade of non-selective cation channel currents under voltage-clamp mode. Internal and external Ca2+ was necessary for the Zn2+-induced K+ currents. The K+ currents were inhibited by intermediate-conductance Ca2+-activated K+ (IK) channel blockers and a large-conductance Ca2+-activated K+ (BK) channel blocker, but not by a small-conductance Ca2+-activated K+ (SK) channel blocker. The phospholipase C (PLC) inhibitor and the non-hydrolysable GDP analog (GDPγS) blocked the increase in intracellular Ca2+ and Zn2+-induced K+ currents, respectively. TPEN (10 μM), a Zn2+-chelator, in the pipette solution abolished BK currents, but not IK currents, indicating that intracellular Zn2+ passed through TRP channels and/or Zrt-, Irt-related proteins (ZIPs) activates BK channel directly, but not IK and SK channels. Zn2+ promoted cell proliferation, and enhanced osteoprotegerin (OPG) mRNA expression. Interestingly, these biological effects of Zn2+ were diminished by TRAM-34, an IK inhibitor.
These results suggest that Zn2+ can activate IK and BK channels through two different pathways in MC3T3-E1 cells. External administration of Zn2+ increases intracellular free-Ca2+ and free-Zn2+ concentration, and hyperpolarizes MC3T3-E1 cells synergistically. Finally, Zn2+ promotes their proliferation and differentiation of MC3T3-E1 cells.
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