Nakagawa Hiroshi, Hasumi Keiji, Takami Masamichi, Aida-Hyugaji Sachiko, Woo Je-Tae, Nagai Kazuo, Ishikawa Toshihisa, Wachi Masaaki
Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8501, Japan.
Biochem Pharmacol. 2007 Jan 1;73(1):34-43. doi: 10.1016/j.bcp.2006.09.002. Epub 2006 Sep 12.
(-)-Epigallocatechin gallate (EGCG) induces cell death of osteoclasts in an Fe(2+)- and H(2)O(2)-dependent manner. In the present study, we further explore the cytotoxic mechanism of EGCG using four EGCG analogues. Molecules methylated at position 4' in the B ring (EGCG-4'-O-Me) or at position 4'' in the D-ring (EGCG-4''-O-Me) showed markedly decreased cytotoxicity to osteoclasts, indicating that hydroxyl groups at these two positions of EGCG are crucial for inducing cell death of osteoclasts. EGCG-4'-O-Me also showed the lowest Fe(3+)-reducing activity among five EGCGs. The Fe(3+)-reducing activity of EGCG was enhanced under conditions whereby protonated EGCG levels were increased, indicating that the protonated status of EGCG was involved in the Fe(3+)-reducing activity. The hydroxyl group at position 4'' in the D-ring was shown by quantum chemical calculation to be preferentially deprotonated among all of the hydroxyl groups in EGCGs. It was also shown that the highest occupied molecular orbital (HOMO) was localized to the B-ring of EGCGs, except for EGCG-4'-O-Me. We report here that the HOMO on the B-ring plays crucial roles in both the Fe(3+)-reducing activity of EGCG and the cytotoxicity of EGCG to osteoclasts, while deprotonation of the hydroxyl group at position 4'' in the D-ring plays a supplementary role.
（-）-表没食子儿茶素没食子酸酯（EGCG）以铁离子（Fe(2+)）和过氧化氢（H(2)O(2)）依赖的方式诱导破骨细胞死亡。在本研究中，我们使用四种EGCG类似物进一步探究EGCG的细胞毒性机制。在B环4'位甲基化的分子（EGCG-4'-O-Me）或在D环4''位甲基化的分子（EGCG-4''-O-Me）对破骨细胞的细胞毒性显著降低，这表明EGCG这两个位置的羟基对于诱导破骨细胞死亡至关重要。EGCG-4'-O-Me在五种EGCG中还表现出最低的铁离子（Fe(3+)）还原活性。在质子化EGCG水平增加的条件下，EGCG的铁离子（Fe(3+)）还原活性增强，这表明EGCG的质子化状态参与了铁离子（Fe(3+)）还原活性。通过量子化学计算表明，在所有EGCG的羟基中，D环4''位的羟基优先去质子化。还表明，除了EGCG-4'-O-Me外，最高占据分子轨道（HOMO）定位于EGCG的B环。我们在此报告，B环上的HOMO在EGCG的铁离子（Fe(3+)）还原活性和EGCG对破骨细胞的细胞毒性中均起关键作用，而D环4''位羟基的去质子化起辅助作用。
