Luo Jiaoyang, Hu Yichen, Kong Weijun, Yang Meihua
Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, P.R. China.
PLoS One. 2014 Mar 27;9(3):e93516. doi: 10.1371/journal.pone.0093516. eCollection 2014.
Arylnaphthalene lignan lactones have attracted considerable interest because of their anti-tumor and anti-hyperlipidimic activities. However, to our knowledge, few studies have explored the effects of these compounds on human leukemia cell lines. In this study, five arylnaphthalene lignans including 6'-hydroxy justicidin A (HJA), 6'-hydroxy justicidin B (HJB), justicidin B (JB), chinensinaphthol methyl ether (CME) and Taiwanin E methyl ether (TEME) were isolated from Justicia procumbens and their effects on the proliferation and apoptosis of the human leukemia K562 cell line were investigated then used to assess structure-activity relationships. To achieve these aims, cytotoxicity was assayed using the MTT assay, while intracellular SOD activity was detected using the SOD Activity Assay kit. Apoptosis was measured by both the using a cycle TEST PLUS DNA reagent kit as well as the FITC Annexin V apoptosis detection kit in combination with flow cytometry. Activation of caspase-mediated apoptosis was evaluated using a FITC active Caspase-3 apoptosis kit and flow cytometry. The results indicated that HJB, HJA and JB significantly inhibited the growth of K562 cells by decreasing both proliferation and SOD activity and inducing apoptosis. The sequence of anti-proliferative activity induced by the five tested arylnaphthalenes by decreasing strength was HJB > HJA > JB > CME > TEME. HJB, HJA and JB also decreased SOD activity and induced apoptosis in a dose-dependent manner. Activation of caspase-3 further indicated that HJB, HJA and JB induced caspase-dependent intrinsic and/or extrinsic apoptosis pathways. Together, these assays suggest that arylnaphthalene lignans derived from Justicia procumbens induce apoptosis to varying degrees, through a caspase-dependent pathway in human leukemia K562 cells. Furthermore, analysis of structure-activity relationships suggest that hydroxyl substitution at C-1 and C-6' significantly increased the antiproliferative activity of arylnaphthalene lignans while a methoxyl at C-1 significantly decreased the effect.
芳基萘木脂素内酯因其抗肿瘤和抗高血脂活性而备受关注。然而，据我们所知，很少有研究探讨这些化合物对人白血病细胞系的影响。在本研究中，从爵床中分离出五种芳基萘木脂素，包括6'-羟基异爵床素A（HJA）、6'-羟基异爵床素B（HJB）、异爵床素B（JB）、中华萘酚甲醚（CME）和台湾脂素E甲醚（TEME），并研究了它们对人白血病K562细胞系增殖和凋亡的影响，进而评估构效关系。为实现这些目标，使用MTT法检测细胞毒性，同时使用超氧化物歧化酶（SOD）活性检测试剂盒检测细胞内SOD活性。通过使用细胞周期检测PLUS DNA试剂试剂盒以及FITC Annexin V凋亡检测试剂盒结合流式细胞术来检测凋亡。使用FITC活性半胱天冬酶-3凋亡试剂盒和流式细胞术评估半胱天冬酶介导的凋亡激活情况。结果表明，HJB、HJA和JB通过降低增殖和SOD活性并诱导凋亡，显著抑制K562细胞的生长。五种测试芳基萘诱导的抗增殖活性由强到弱的顺序为HJB＞HJA＞JB＞CME＞TEME。HJB、HJA和JB还以剂量依赖性方式降低SOD活性并诱导凋亡。半胱天冬酶-3的激活进一步表明，HJB、HJA和JB诱导了半胱天冬酶依赖性的内源性和/或外源性凋亡途径。总之，这些实验表明，来源于爵床的芳基萘木脂素通过半胱天冬酶依赖性途径在人白血病K562细胞中不同程度地诱导凋亡。此外，构效关系分析表明，C-1和C-6'位的羟基取代显著增加了芳基萘木脂素的抗增殖活性，而C-1位的甲氧基则显著降低了该效应。
