Khajah Maitham A, Khushaish Sarah, Luqmani Yunus A
Faculty of Pharmacy, Kuwait University, Kuwait City, Kuwait.
Front Pharmacol. 2021 Oct 20;12:747001. doi: 10.3389/fphar.2021.747001. eCollection 2021.
Lactate dehydrogenase (LDH) plays an important role in cancer pathogenesis and enhanced expression/activity of this enzyme has been correlated with poor prognosis. In this study we determined the expression profile of LDH-A and B in normal as well as in endocrine-resistant and -responsive breast cancer cells and the effect of their knockdown on LDH activity, lactate production, proliferation and cell motility. Knockdown experiments were performed using siRNA and shRNA. The expression profile of LDH and signaling molecules was determined using PCR and western blotting. Intracellular LDH activity and extracellular lactate levels were measured by a biochemical assay. Cell motility was determined using wound healing, while proliferation was determined using MTT assay. LDH-A was expressed in all of the tested cell lines, while LDH-B was specifically expressed only in normal and endocrine-resistant breast cancer cells. This was correlated with significantly enhanced LDH activity and lactate production in endocrine resistant breast cancer cells when compared to normal or endocrine responsive cancer cells. LDH-A or -B knockdown significantly reduced LDH activity and lactate production, which led to reduced cell motility. Exogenous lactate supplementation enhanced cell motility co-incident with enhanced phosphorylation of ERK1/2 and reduced E-cadherin expression. Also, LDH-A or -B knockdown reduced ERK 1/2 phosphorylation. Enhanced cell motility in endocrine resistant breast cancer cells is at least in part mediated by enhanced extracellular lactate levels, and LDH inhibition might be a promising therapeutic target to inhibit cancer cell motility.
乳酸脱氢酶（LDH）在癌症发病机制中起重要作用，该酶表达/活性增强与预后不良相关。在本研究中，我们测定了正常以及内分泌抵抗和内分泌反应性乳腺癌细胞中LDH-A和LDH-B的表达谱，以及敲低它们对LDH活性、乳酸生成、增殖和细胞运动的影响。使用小干扰RNA（siRNA）和短发夹RNA（shRNA）进行敲低实验。使用聚合酶链反应（PCR）和蛋白质免疫印迹法测定LDH和信号分子的表达谱。通过生化分析测定细胞内LDH活性和细胞外乳酸水平。使用伤口愈合实验测定细胞运动，而使用MTT法测定增殖。LDH-A在所有测试细胞系中均有表达，而LDH-B仅在正常和内分泌抵抗性乳腺癌细胞中特异性表达。与正常或内分泌反应性癌细胞相比，这与内分泌抵抗性乳腺癌细胞中显著增强的LDH活性和乳酸生成相关。敲低LDH-A或LDH-B可显著降低LDH活性和乳酸生成，从而导致细胞运动减少。外源性补充乳酸可增强细胞运动，同时伴有细胞外信号调节激酶1/2（ERK1/2）磷酸化增强和E-钙黏蛋白表达降低。此外，敲低LDH-A或LDH-B可降低ERK 1/2磷酸化。内分泌抵抗性乳腺癌细胞中增强的细胞运动至少部分是由细胞外乳酸水平升高介导的，抑制LDH可能是抑制癌细胞运动的一个有前景的治疗靶点。
