Morimoto Satoshi, Tanaka Yumi, Sasaki Kaori, Tanaka Hiroyuki, Fukamizu Tomohide, Shoyama Yoshinari, Shoyama Yukihiro, Taura Futoshi
Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka 812-8582, Japan.
J Biol Chem. 2007 Jul 13;282(28):20739-51. doi: 10.1074/jbc.M700133200. Epub 2007 May 17.
Cannabinoids are secondary metabolites stored in capitate-sessile glands on leaves of Cannabis sativa. We discovered that cell death is induced in the leaf tissues exposed to cannabinoid resin secreted from the glands, and identified cannabichromenic acid (CBCA) and Delta(1)-tetrahydrocannabinolic acid (THCA) as unique cell death mediators from the resin. These cannabinoids effectively induced cell death in the leaf cells or suspension-cultured cells of C. sativa, whereas pretreatment with the mitochondrial permeability transition (MPT) inhibitor cyclosporin A suppressed this cell death response. Examinations using isolated mitochondria demonstrated that CBCA and THCA mediate opening of MPT pores without requiring Ca(2+) and other cytosolic factors, resulting in high amplitude mitochondrial swelling, release of mitochondrial proteins (cytochrome c and nuclease), and irreversible loss of mitochondrial membrane potential. Therefore, CBCA and THCA are considered to cause serious damage to mitochondria through MPT. The mitochondrial damage was also confirmed by a marked decrease of ATP level in cannabinoid-treated suspension cells. These features are in good accord with those of necrotic cell death, whereas DNA degradation was also observed in cannabinoid-mediated cell death. However, the DNA degradation was catalyzed by nuclease(s) released from mitochondria during MPT, indicating that this reaction was not induced via a caspase-dependent apoptotic pathway. Furthermore, the inhibition of the DNA degradation only slightly blocked the cell death induced by cannabinoids. Based on these results, we conclude that CBCA and THCA have the ability to induce necrotic cell death via mitochondrial dysfunction in the leaf cells of C. sativa.
大麻素是储存于大麻叶片上的头状无柄腺中的次生代谢产物。我们发现，暴露于腺分泌的大麻素树脂的叶片组织中会诱导细胞死亡，并从该树脂中鉴定出大麻色原酸（CBCA）和Δ⁽¹⁾-四氢大麻酚酸（THCA）作为独特的细胞死亡介质。这些大麻素能有效诱导大麻叶片细胞或悬浮培养细胞的细胞死亡，而用线粒体通透性转换（MPT）抑制剂环孢菌素A预处理可抑制这种细胞死亡反应。使用分离的线粒体进行的检测表明，CBCA和THCA介导MPT孔的开放，无需Ca²⁺和其他胞质因子，导致线粒体高幅度肿胀、线粒体蛋白（细胞色素c和核酸酶）释放以及线粒体膜电位的不可逆丧失。因此，CBCA和THCA被认为通过MPT对线粒体造成严重损害。大麻素处理的悬浮细胞中ATP水平显著降低也证实了线粒体损伤。这些特征与坏死性细胞死亡的特征高度一致，而在大麻素介导的细胞死亡中也观察到了DNA降解。然而，DNA降解是由MPT期间从线粒体释放的核酸酶催化的，表明该反应不是通过半胱天冬酶依赖性凋亡途径诱导的。此外，DNA降解的抑制仅略微阻断了大麻素诱导的细胞死亡。基于这些结果，我们得出结论，CBCA和THCA有能力通过大麻叶片细胞中的线粒体功能障碍诱导坏死性细胞死亡。
