Table2_Associations of Porphyromonas gingivalis Infection and Low Beclin1 Expression With Clinicopathological Parameters and Survival of Esophageal Squamous Cell Carcinoma Patients.XLSX

Purpose: The present study focused on exploring the associations of Porphyromonas gingivalis (P. gingivalis) infection and low Beclin1 expression with clinicopathological parameters and survival of esophageal squamous cell carcinoma (ESCC) patients, so as to illustrate its clinical significance and prognostic value.
Methods: Immunohistochemistry (IHC) was used to detect P. gingivalis infection status and Beclin1 expression in 370 ESCC patients. The chi-square test was adopted to illustrate the relationship between categorical variables, and Cohen’s kappa coefficient was used for correlation analysis. Kaplan-Meier survival curves with the log-rank test were used to analyse the correlation of P. gingivalis infection and low Beclin1 expression with survival time. The effects of P. gingivalis infection and Beclin1 downregulation on the proliferation, migration and antiapoptotic abilities of ESCC cells in vitro were detected by Cell Counting Kit-8, wound healing and flow cytometry assays. For P. gingivalis infection of ESCC cells, cell culture medium was replaced with antibiotic-free medium when the density of ESCC cells was 70–80%, cells were inoculated with P. gingivalis at a multiplicity of infection (MOI) of 10.
Result:P. gingivalis infection was negatively correlated with Beclin1 expression in ESCC tissues, and P. gingivalis infection and low Beclin1 expression were associated with differentiation status, tumor invasion depth, lymph node metastasis, clinical stage and prognosis in ESCC patients. In vitro experiments confirmed that P. gingivalis infection and Beclin1 downregulation potentiate the proliferation, migration and antiapoptotic abilities of ESCC cells (KYSE150 and KYSE30). Our results provide evidence that P. gingivalis infection and low Beclin1 expression were associated with the development and progression of ESCC.
Conclusion: Long-term smoking and alcohol consumption causes poor oral and esophageal microenvironments and ESCC patients with these features were more susceptible to P. gingivalis infection and persistent colonization, and exhibited lower Beclin1 expression, worse prognosis and more advanced clinicopathological features. Our findings indicate that effectively eliminating P. gingivalis colonization and restoring Beclin1 expression in ESCC patients may contribute to preventation and targeted treatment, and yield new insights into the aetiological research on ESCC.

研究目的：本研究旨在探讨牙龈卟啉单胞菌（Porphyromonas gingivalis, P. gingivalis）感染与Beclin1低表达分别及联合与食管鳞状细胞癌（esophageal squamous cell carcinoma, ESCC）患者临床病理参数及生存状况的关联，以期阐明二者的临床意义与预后价值。

研究方法：采用免疫组化（Immunohistochemistry, IHC）检测370例食管鳞状细胞癌患者组织中的牙龈卟啉单胞菌感染状态与Beclin1表达水平。采用卡方检验分析分类变量间的关联，以科恩κ系数（Cohen’s kappa coefficient）进行相关性分析；采用Kaplan-Meier生存曲线联合log-rank检验分析牙龈卟啉单胞菌感染、Beclin1低表达与患者生存时间的相关性。通过细胞计数试剂盒-8（Cell Counting Kit-8）、划痕愈合实验及流式细胞术分别检测牙龈卟啉单胞菌感染与Beclin1下调对食管鳞状细胞癌细胞体外增殖、迁移及抗凋亡能力的影响。针对食管鳞状细胞癌细胞的牙龈卟啉单胞菌感染实验：当食管鳞状细胞癌细胞密度达到70%~80%时，更换无抗生素培养基，以感染复数（multiplicity of infection, MOI）为10的比例接种牙龈卟啉单胞菌。

研究结果：牙龈卟啉单胞菌感染与食管鳞状细胞癌组织中Beclin1表达呈负相关；牙龈卟啉单胞菌感染及Beclin1低表达均与食管鳞状细胞癌患者的分化状态、肿瘤浸润深度、淋巴结转移、临床分期及预后显著相关。体外实验证实，牙龈卟啉单胞菌感染与Beclin1下调可增强食管鳞状细胞癌细胞（KYSE150与KYSE30）的增殖、迁移及抗凋亡能力。本研究结果提示，牙龈卟啉单胞菌感染与Beclin1低表达与食管鳞状细胞癌的发生发展密切相关。

研究结论：长期吸烟与过量饮酒可导致口腔及食管微环境恶化，合并该特征的食管鳞状细胞癌患者更易发生牙龈卟啉单胞菌感染及持续定植，且Beclin1表达水平更低、预后更差、临床病理分期更晚。本研究结果表明，针对食管鳞状细胞癌患者有效清除牙龈卟啉单胞菌定植、恢复Beclin1表达，可为该病的预防与靶向治疗提供新思路，并为食管鳞状细胞癌的病因学研究提供新视角。