Dong Siqi, Zheng Lei, Jiang Tao
Department of Andrology, The Second Hospital of Dalian Medical University, Dalian116000, China.
J Oncol. 2023 Jan 21;2023:2830306. doi: 10.1155/2023/2830306. eCollection 2023.
Ferroptosis and autophagy have an important role in the occurrence and development of cancer, and lactate in cells and microenvironment is one of the influencing factors of ferroptosis and autophagy. The lactate/proton monocarboxylate transporter 4 (MCT4), which is expressed in the cell membrane, regulates the transport of intracellular lactic acid and lactate. The knockout of MCT4 can affect intracellular and extracellular lactic acid levels, thereby affecting the growth, proliferation, and metastasis of tumor cells via regulation of the oxidative stress in cells. However, whether MCT4 affects ferroptosis and autophagy in bladder cancer cells remains unclear.
Colony formation assay and bladder cancer xenograft animal model were used to assess the effect of MCT4 on the growth in 5637 cells. Reactive oxygen species (ROS) assay, lipid ROS assay, lipid peroxidation assay (MDA), and transmission electron microscopy were performed to assess the level of lipid peroxidation in 5637 cells. RNA-sequence, RT-PCR, and Western Blot were used to analyze the mechanism of MCT4 of ferroptosis and autophagy. AdPlus-mCherry-GFP-LC3B reporter system was used to detect the effect of MCT4 on autophagy in 5637 cells, and the effect of knockdown of MCT4 on apoptosis was analyzed by flow cytometry.
The mRNA level of MCT4 was significantly upregulated in patients with bladder cancer, which was associated with a poor prognosis. and studies demonstrated that knockdown of MCT4 could inhibit the proliferation of bladder cancer cells. Furthermore, knockdown of MCT4 led to the significant increase of ROS and MDA levels in 5637 cells and ferroptosis in 5637 cells induced by ferroptosis inducers including RSL3 (APExBIO) and erastin (APExBIO) via inhibition of AMPK-related proteins. Moreover, knockdown of MCT4 inhibited autophagy in 5637 cells, while siMCT4 promoted inhibition of autophagy by CQ (an autophagy inhibitor), which increased the level of apoptosis.
This study confirmed that knockdown of MCT4 could affect oxidative stress and induce ferroptosis and inhibition of autophagy, thus suggesting that MCT4 may be a potential target for the treatment of bladder cancer.
铁死亡和自噬在癌症的发生发展中起重要作用，细胞及微环境中的乳酸是影响铁死亡和自噬的因素之一。细胞膜上表达的乳酸/质子单羧酸转运蛋白4（MCT4）调节细胞内乳酸和乳酸根的转运。敲除MCT4可影响细胞内和细胞外乳酸水平，从而通过调节细胞内氧化应激影响肿瘤细胞的生长、增殖和转移。然而，MCT4是否影响膀胱癌细胞中的铁死亡和自噬仍不清楚。
采用集落形成试验和膀胱癌异种移植动物模型评估MCT4对5637细胞生长的影响。进行活性氧（ROS）检测、脂质ROS检测、脂质过氧化检测（MDA）和透射电子显微镜观察以评估5637细胞中的脂质过氧化水平。采用RNA测序、逆转录-聚合酶链反应（RT-PCR）和蛋白质免疫印迹法分析MCT4影响铁死亡和自噬的机制。利用AdPlus-mCherry-GFP-LC3B报告系统检测MCT4对5637细胞自噬的影响，并通过流式细胞术分析敲低MCT4对细胞凋亡的影响。
膀胱癌患者中MCT4的mRNA水平显著上调，这与预后不良相关。 和 研究表明，敲低MCT4可抑制膀胱癌细胞的增殖。此外，敲低MCT4导致5637细胞中ROS和MDA水平显著升高，并通过抑制AMPK相关蛋白使包括RSL3（APExBIO）和厄洛替尼（APExBIO）在内的铁死亡诱导剂诱导5637细胞发生铁死亡。此外，敲低MCT4抑制5637细胞中的自噬，而siMCT4增强了自噬抑制剂CQ对自噬的抑制作用，从而增加了细胞凋亡水平。
本研究证实敲低MCT4可影响氧化应激并诱导铁死亡和抑制自噬，因此表明MCT4可能是治疗膀胱癌的潜在靶点。
