Withanolide D induces apoptosis in leukemia by targeting the activation of neutral sphingomyelinase-ceramide cascade mediated by synergistic activation of c-Jun N-terminal kinase and p38 mitogen-activated protein kinase.
Withanolide D induces apoptosis in leukemia by targeting the activation of neutral sphingomyelinase-ceramide cascade mediated by synergistic activation of c-Jun N-terminal kinase and p38 mitogen-activated protein kinase.
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DOI：
10.1186/1476-4598-9-239
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发表时间：
2010-09-13
期刊：
影响因子：
37.3
通讯作者：
Mandal C
中科院分区：
文献类型：
--
作者：
Mondal S;Mandal C;Sangwan R;Chandra S;Mandal C
神经酰胺是一种重要的第二信使，具有多种细胞和生物学效应。它是一种特异性和有效的细胞凋亡诱导剂和细胞生长抑制剂。在白血病中，化学抗性通常是由于神经酰胺代谢失调而产生的。在白血病的组合治疗策略中，很少有组分具有增加神经酰胺产生的能力。因此，通过生理学和药理学调节剂对神经酰胺产生的操纵将在白血病化疗中产生累加效应。我们发现从催眠睡茄中提取的纯中药化合物WithanidD（C4β-C5β，C6β-epoxy-1-oxo-，20 β，dihydroxy-20 S，22 R-witha-2，24-dienetransferase; WithaD）能以剂量和时间依赖的方式有效诱导骨髓细胞（K562）和淋巴细胞（MOLT-4）凋亡，对正常淋巴细胞和对照增殖细胞无毒性。WithaD可能增加这些细胞中神经酰胺的产生。在神经酰胺下游，WithaD作用于MKK组蛋白，显著增加JNK和p38 MAPK磷酸化。p38 MAPK和JNK的药理学抑制证明了它们在WithaD诱导的细胞死亡中的协同作用。分析神经酰胺产生的原因，我们发现中性鞘磷脂酶的激活，并显示中性鞘磷脂酶2（N-SMase 2）是WithaD诱导细胞凋亡的关键介质。通过siRNA和N-SMase抑制剂（GW 4869）敲低N-SMase 2显著减少WithaD诱导的神经酰胺产生和MKK 4和MKK 3/6的磷酸化，而MKK 7的磷酸化在白血病细胞中受到适度调节。此外，无论是通过沉默N-SMase 2和/或GW 4869阻断保护这些细胞从WithaD介导的死亡和抑制凋亡，而伏马菌素B1，神经酰胺合成酶的抑制剂，没有任何影响。此外，WithaD有效地诱导了患者新鲜分离的淋巴母细胞的凋亡，并且通过JNK和p38 MAPK激活有效的细胞杀伤活性。我们的研究结果表明，WithaD通过激活N-SMase 2，调节JNK和p38 MAPK的磷酸化，以及诱导白血病患者的髓样和淋巴样细胞以及原代细胞沿着的细胞凋亡，从而增强神经酰胺的积累。两者合计，这种纯草药化合物（WithaD）可以考虑作为一个潜在的替代工具，与传统的化疗治疗相结合，从而加速传统药物开发的过程。
Ceramide is an important second messenger that has diverse cellular and biological effect. It is a specific and potent inducer of apoptosis and suppressor of cell growth. In leukemia, chemoresistance generally developed due to deregulated ceramide metabolism. In combinatorial treatment strategies of leukemia, few components have the capability to increases ceramide production. Manipulation in ceramide production by physiological and pharmacological modulators therefore will give additive effect in leukemia chemotherapy. Here, we show that Withanolide D (C4β-C5β,C6β-epoxy-1-oxo-,20β, dihydroxy-20S,22R-witha-2,24-dienolide; WithaD), a pure herbal compound isolated from Withania somnifera could effectively induces apoptosis in a dose and time dependant manner both in myeloid (K562) and lymphoid (MOLT-4) cells being nontoxic to normal lymphocytes and control proliferative cells. WithaD potentially augment ceramide production in these cells. Downstream of ceramide, WithaD acted on MKK group of proteins and significantly increased JNK and p38MAPK phosphorylation. Pharmacological inhibition of p38MAPK and JNK proves their cooperative action on WithaD-induced cell death. Dissecting the cause of ceramide production, we found activation of neutral sphingomyelinase and showed neutral-sphingomyelinase 2 (N-SMase 2) is a critical mediator of WithaD-induced apoptosis. Knockdown of N-SMase 2 by siRNA and inhibitor of N-SMase (GW4869) significantly reduced WithaD-induced ceramide generation and phosphorylation of MKK4 and MKK3/6, whereas phosphorylation of MKK7 was moderately regulated in leukemic cells. Also, both by silencing of N-SMase 2 and/or blocking by GW4869 protects these cells from WithaD-mediated death and suppressed apoptosis, whereas Fumonisin B1, an inhibitor of ceramide synthase, did not have any effect. Additionally, WithaD effectively induced apoptosis in freshly isolated lymphoblasts from patients and the potent cell killing activity was through JNK and p38MAPK activation. Our results demonstrate that WithaD enhance the ceramide accumulation by activating N-SMase 2, modulate phosphorylation of the JNK and p38MAPK and induced apoptosis in both myeloid and lymphoid cells along with primary cells derived from leukemia patients. Taken together, this pure herbal compound (WithaD) may consider as a potential alternative tool with additive effects in conjunction with traditional chemotherapeutic treatment, thereby accelerate the process of conventional drug development.
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DOI：
10.1073/pnas.251194298
发表时间：
2001-11-20
影响因子：
11.1
作者：
Bennett, BL;Sasaki, DT;Anderson, DW
通讯作者：
Anderson, DW
影响因子：
11.4
作者：
Jaffrezou, JP;Levade, T;Laurent, G
通讯作者：
Laurent, G
DOI：
10.1016/j.bbagrm.2009.08.006
发表时间：
2009-11-01
影响因子：
4.7
作者：
Ito, Hiromi;Murakami, Masashi;Murate, Takashi
通讯作者：
Murate, Takashi
影响因子：
4.1
作者：
Fleming, Y;Armstrong, CG;Cohen, P
通讯作者：
Cohen, P
影响因子：
6.5
作者：
Ghosh, S;Bandyopadhyay, S;Mandal, C
通讯作者：
Mandal, C