Oral immunosuppression caused by smoking creates a microenvironment to promote the occurrence and development of oral mucosa precancerous lesions. This study aimed to investigate the role of metabolism and macrophage polarization in cigarette-promoting oral leukoplakia. The effects of cigarette smoke extract (CSE) on macrophage polarization and metabolism were studied in vivo and in vitro. The polarity of macrophages was detected by flow cytometric analysis and qPCR. Liquid chromatography-mass spectrometry (LC-MS) was used to perform a metabolomic analysis of Raw cells stimulated with CSE. Immunofluorescence and flow cytometry were used to detect the polarity of macrophages in the condition of glutamine abundance and deficiency. Cell Counting Kit-8 (CCK-8), wound-healing assay, and Annexin V-FITC (fluorescein isothiocyanate)/PI (propidium iodide) double-staining flow cytometry were applied to detect the growth and transferability and apoptosis of Leuk-1 cells in the supernatant of Raw cells which were stimulated with CSE, glutamine abundance and deficiency. Hyperkeratosis and dysplasia of the epithelium were evident in smoking mice. M2 macrophages increased under CSE stimulation in vivo and in vitro. In total, 162 types of metabolites were detected in the CSE group. The metabolites of nicotine, glutamate, arachidic acid, and arginine changed significantly. The significant enrichment pathways were also selected, including nicotine addiction, glutamine and glutamate metabolism, and arginine biosynthesis. The results also showed that the supernatant of Raw cells stimulated by CSE could induce excessive proliferation of Leuk-1 and inhibit apoptosis. Glutamine abundance can facilitate this process. Cigarette smoke promotes oral leukoplakia via regulating glutamine metabolism and macrophage M2 polarization.

吸烟引起的口腔免疫抑制创造了一个微环境，促进了口腔黏膜癌前病变的发生和发展。本研究旨在调查代谢和巨噬细胞极化在促进卷烟的口腔白斑病中的作用。在体内和体外研究了香烟烟雾提取物 (CSE) 对巨噬细胞极化和代谢的影响。通过流式细胞术分析和 qPCR 检测巨噬细胞的极性。液相色谱-质谱 (LC-MS) 用于对 CSE 刺激的原始细胞进行代谢组学分析。采用免疫荧光和流式细胞术检测巨噬细胞在谷氨酰胺丰度和缺乏状态下的极性。Cell Counting Kit-8 (CCK-8)，伤口愈合试验，应用Annexin V-FITC（异硫氰酸荧光素）/PI（碘化丙锭）双染色流式细胞术检测CSE刺激后的Raw细胞上清液中Leuk-1细胞的生长、转移性和凋亡情况、谷氨酰胺丰度和不足。吸烟小鼠的上皮过度角化和发育不良是明显的。M2 巨噬细胞在体内和体外在 CSE 刺激下增加。在 CSE 组中总共检测到 162 种代谢物。尼古丁、谷氨酸、花生酸和精氨酸的代谢物发生显着变化。还选择了重要的富集途径，包括尼古丁成瘾、谷氨酰胺和谷氨酸代谢以及精氨酸生物合成。结果还表明，CSE刺激的Raw细胞上清液可诱导Leuk-1过度增殖并抑制细胞凋亡。谷氨酰胺丰度可以促进这一过程。香烟烟雾通过调节谷氨酰胺代谢和巨噬细胞 M2 极化促进口腔白斑。