Background

Lactucin, a naturally occurring active sesquiterpene lactone, is abundantly found in chicory and romaine lettuce. A recent study reported that lactucin could induce apoptosis in leukemia cells. However, its cytotoxicity and potential molecular mechanisms underlying cancer cell death remain unclear.

Objective

Therefore, in this study, we aimed to investigate the direct effect and underlying mechanism of action of lactucin on renal cancer cells.

Methods

MTT assay and flow cytometry were performed to evaluate the rate of cell proliferation and apoptosis, respectively. Western blotting, reverse transcription polymerase chain reaction, and protein stability analyses were performed to analyze the effect of lactucin on the expression of apoptosis-related proteins such as B-cell lymphoma 2 (BCL-2) and CFLAR (CASP8 and FADD like apoptosis regulator) long isoform (CFLAR_L) in Caki-1 human renal cancer cells. In addition, reactive oxygen species (ROS) generation was evaluated using flow cytometry.

Results

Lactucin treatment induced apoptosis in Caki-1 cells in a dose-dependent manner via activation of the caspase pathway. It downregulated BCL-2 and CFLAR_L expression levels by suppressing BCL-2 transcription and CFLAR_L protein stability, respectively. Pretreatment with N-acetyl-1-cysteine, a ROS scavenger, attenuated the lactucin-induced apoptosis and restored the BCL-2 and CFLAR_L expression to basal levels. Lactucin-facilitated BCL-2 downregulation was regulated at the transcriptional level through the inactivation of the NF-κB pathway.

Conclusions

Our study is the first to demonstrate that lactucin-induced apoptosis is mediated by ROS production, which in turn activates the caspase-dependent apoptotic pathway by inhibiting BCL-2 and CFLAR_L expression in Caki-1 cells.

中文翻译：

---

Lactucin 通过活性氧介导的 BCL-2 和 CFLARL 下调 Caki-1 细胞诱导细胞凋亡

背景

Lactucin 是一种天然存在的活性倍半萜内酯，大量存在于菊苣和长叶莴苣中。最近的一项研究报告说，lactucin 可以诱导白血病细胞凋亡。然而，其细胞毒性和癌细胞死亡的潜在分子机制仍不清楚。

客观的

因此，在本研究中，我们旨在探讨乳素对肾癌细胞的直接作用和潜在作用机制。

方法

MTT法和流式细胞术分别评估细胞增殖和凋亡率。采用Western blotting、逆转录聚合酶链反应和蛋白稳定性分析，分析了lactucin对凋亡相关蛋白如B细胞淋巴瘤2（BCL-2）和CFLAR（CASP8和FADD样凋亡调节因子）表达的影响。 ) Caki -1 人肾癌细胞中的长亚型 (CFLAR _{L )。}此外，使用流式细胞仪评估活性氧 (ROS) 的产生。

结果

Lactucin 处理通过激活半胱天冬酶途径以剂量依赖性方式诱导 Caki-1 细胞凋亡。它分别通过抑制BCL-2转录和 CFLAR _L蛋白稳定性来下调 BCL-2 和 CFLAR _{L表达水平。}用 ROS 清除剂N-乙酰基-1-半胱氨酸进行预处理，减弱了 lactucin 诱导的细胞凋亡，并将 BCL-2 和 CFLAR _L表达恢复到基础水平。Lactucin 促进的 BCL-2 下调通过 NF-κB 通路的失活在转录水平上受到调节。

结论

我们的研究首次证明了 lactucin 诱导的细胞凋亡是由 ROS 产生介导的，ROS 反过来通过抑制 Caki -1 细胞中BCL-2 和 CFLAR _{L的表达来激活 caspase 依赖性凋亡途径。}