Objective: To explore the mechanism that mediates the inhibitory effects of β-hydroxybutyrate (BHB) on lung adenocarcinoma cells.
Methods: A549 and LLC cell lines treated with 5 or 10 mmol/L BHB were examined for changes in cell viability, proliferation, migration, and invasion using CCK-8 assay, EdU staining, scratch assay, and Transwell assay. The differential expression of GPR109A in lung adenocarcinoma and normal lung tissue was analyzed using GEPIA database. GPR109A expressions in BHB-treated lung adenocarcinoma cells were determined using RT-PCR and Western blotting. The changes in IC50 of BHB were examined in A549 and LLC cells with GPR109A knockdown. The effect of BHB administered via gavage for 21 days on tumor growth was evaluated in nude mouse and Balb/c mouse models bearing xenografts derived A549 and LLC cells with or without GPR109A knockdown.
Results: Treatment with BHB concentration-dependently repressed the viability, proliferation, migration and invasion of A549 and LLC cells. GPR109A expression was significantly decreased in lung adenocarcinoma tissues and A549 and LLC cell lines (P<0.05). Loss of function experiments showed that the inhibitory effects of BHB on A549 and LLC cells were partly mediated by GPR109A, and in the tumor-bearing mouse models, BHB significantly inhibited tumor growth partly by regulating GPR109A expression (P<0.05).
Conclusion: BHB can repress the malignant behaviors of A549 and LLC cells and inhibit tumor growth in mice, and these effects are mediated partly by regulating GPR109A expression.
目的: 探究β-羟基丁酸（BHB）对肺腺癌细胞（A549细胞、LLC细胞）的作用及其调控机制。
方法: 分别用0 mmol/L（PBS）、5 mmol/L、10 mmol/L的BHB处理人肺腺癌A549细胞和LLC细胞，用CCK-8、EdU染色、细胞划痕和transwell实验检测细胞的活性、增殖、迁移和侵袭。GEPIA在线分析GPR109A在正常和肺腺癌样本之间的表达差异。RT-PCR和Western blot检测不同剂量BHB处理对GPR109A表达的影响。敲低GPR109A后观察A549细胞和LLC细胞对BHB的反应，探究GPR109A是否参与介导BHB的抑癌作用。在裸鼠和Balb/c小鼠右腋皮下分别接种A549细胞、LLC细胞及相应系的GPR109A敲低细胞，给予BHB或等体积无菌水灌胃21 d，观察肿瘤生长情况。
结果: BHB以浓度依赖方式抑制A549细胞和LLC细胞的活性、增殖、迁移和侵袭（P＜0.05）。GEPIA在线分析发现GPR109A在肺腺癌病人中表达下调，且体外验证发现GPR109A的mRNA表达量和蛋白表达量随BHB浓度升高而上升（P＜0.05）。BHB对A549细胞和LLC细胞的抑制作用是由GPR109A部分介导，体内实验结果进一步显示BHB可通过介导GPR109A的表达来抑制裸鼠和balb/c小鼠体内肿瘤的生长（P＜0.05）。
结论: BHB可部分介导GPR109A的表达来抑制肺腺癌细胞的恶性行为，并抑制小鼠体内的肿瘤生长。
Keywords: A549 cells; G protein coupled receptor; GPR109A; lung adenocarcinoma; β-hydroxybutyrate.