Department of Endodontics, Affiliated Stomatology Hospital of Guangzhou Medical University, Guangdong Engineering Research Center of Oral Restoration and Reconstruction, Guangzhou Key Laboratory of Basic and Applied Research of Oral Regenerative Medicine, Guangzhou, 510182, China.
Department of Endodontics, Stomatological Hospital, Southern Medical University, Guangzhou, China.
BMC Oral Health. 2022 Jul 8;22(1):279. doi: 10.1186/s12903-022-02295-y.
Enterococcus faecalis is a dominant pathogen in the root canals of teeth with persistent apical periodontitis (PAP), and osteoblast apoptosis contributes to imbalanced bone remodelling in PAP. Here, we investigated the effect of E. faecalis OG1RF on apoptosis in primary human calvarial osteoblasts. Specifically, the expression of apoptosis-related genes and the role of anti-apoptotic and pro-apoptotic members of the BCL-2 family were examined.
Primary human calvarial osteoblasts were incubated with E. faecalis OG1RF at multiplicities of infection corresponding to infection time points. Flow cytometry, terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay, caspase-3/-8/-9 activity assay, polymerase chain reaction (PCR) array, and quantitative real-time PCR were used to assess osteoblast apoptosis.
E. faecalis infection increased the number of early- and late-phase apoptotic cells and TUNEL-positive cells, decreased the mitochondrial membrane potential (ΔΨm), and activated the caspase-3/-8/-9 pathway. Moreover, of all 84 apoptosis-related genes in the PCR array, the expression of 16 genes was upregulated and that of four genes was downregulated in the infected osteoblasts. Notably, the mRNA expression of anti-apoptotic BCL2 was downregulated, whereas that of the pro-apoptotic BCL2L11, HRK, BIK, BMF, NOXA, and BECN1 and anti-apoptotic BCL2A1 was upregulated.
E. faecalis OG1RF infection triggered apoptosis in human calvarial osteoblasts, and BCL-2 family members acted as regulators of osteoblast apoptosis. Therefore, BCL-2 family members may act as potential therapeutic targets for persistent apical periodontitis.
粪肠球菌是持续性根尖周炎（PAP）牙齿根管中的主要病原体，成骨细胞凋亡导致 PAP 中骨重塑失衡。本研究旨在探讨 OG1RF 粪肠球菌对原代人颅骨成骨细胞凋亡的影响。具体而言，检测了凋亡相关基因的表达，并研究了 BCL-2 家族的抗凋亡和促凋亡成员的作用。
采用感染复数（MOI）分别对应感染时间点的方式，将原代人颅骨成骨细胞与 OG1RF 粪肠球菌共孵育。采用流式细胞术、末端脱氧核苷酸转移酶 dUTP 缺口末端标记（TUNEL）检测、caspase-3/-8/-9 活性检测、聚合酶链反应（PCR）阵列和实时定量 PCR 检测评估成骨细胞凋亡。
粪肠球菌感染增加了早期和晚期凋亡细胞及 TUNEL 阳性细胞的数量，降低了线粒体膜电位（ΔΨm），并激活了 caspase-3/-8/-9 途径。此外，在 PCR 阵列中 84 个凋亡相关基因中，16 个基因的表达上调，4 个基因的表达下调。值得注意的是，抗凋亡 BCL2 的 mRNA 表达下调，而促凋亡 BCL2L11、HRK、BIK、BMF、NOXA 和 BECN1 以及抗凋亡 BCL2A1 的 mRNA 表达上调。
OG1RF 粪肠球菌感染可触发人颅骨成骨细胞凋亡，BCL-2 家族成员作为成骨细胞凋亡的调节因子发挥作用。因此，BCL-2 家族成员可能是持续性根尖周炎的潜在治疗靶点。
