State Key Laboratory of Oral & Maxillofacial reconstruction and Regeneration, National Clinical Research Center for Oral Diseases, Shaanxi Key Laboratory of Stomatology, Department of Operative Dentistry and Endodontics, School of Stomatology, Air Force Medical University, 145 West Chang-le Road, Xi'an, China.
Odontology. 2024 Jan;112(1):125-137. doi: 10.1007/s10266-023-00838-5. Epub 2023 Jul 26.
Perfect intercellular junctions are key for odontoblast barrier function. However, whether Partitioning defective-3 (Par3) is expressed in odontoblasts and its potential effects on odontoblast junctions are unknown. Herein, we investigated the effect of Par3 on cellular junctions and the biological behavior of odontoblast-lineage cells (OLCs). Whole-transcriptome sequencing was used to analyze the effects of Par3 on OLCs and the underlying molecular mechanism. Par3 was detected under physiological and inflammatory conditions in OLCs. To investigate the regulatory effect of Par3 on junctions between mouse OLCs, the effects of Par3 downregulation on the proliferation, migration, cycle and apoptosis of OLCs were detected by 5-ethyl-2'-deoxyuridine (EdU) and Transwell assays and flow cytometry. Western blotting and alizarin red S and alkaline phosphatase (ALP) staining were used to observe the effect of Par3 downregulation on OLC mineralization. Whole-transcriptome sequencing was used to investigate the biological role of Par3 in OLCs and potential molecular mechanisms. Par3 was located along the odontoblast layer in the rat pulp tissue and in the cytoplasm of OLCs. Par3 expression was downregulated under inflammatory conditions. The OLC junctions were discontinuous, and total Zona occluden-1 (ZO-1) expression and expression of ZO-1 at the membrane in OLCs were reduced after Par3 silencing (P < 0.05). Expression of a junction-related protein (ZO-1) was downregulated after the downregulation of Par3 (P < 0.05), and ZO-1 moved from the cell membrane to the cytoplasm. OLC proliferation and migration were enhanced, but apoptosis and mineralization were inhibited in shPar3-transfected cells (P < 0.05). Sequencing identified 2996 differentially expressed genes (DEGs), which were mainly enriched in the response to stimuli and binding. Downregulation of Par3 could overactivate the PI3k-AKT pathway by promoting AKT phosphorylation (P < 0.05). Downregulation of Par3 may disrupt junctions between OLCs by affecting ZO-1 expression and distribution and promote OLC proliferation and migration but inhibit OLC mineralization. Par3 may interact with 14-3-3 proteins for PI3K-AKT pathway activation to affect OLC junctions and function.
理想的细胞间连接对于成牙本质细胞屏障功能至关重要。然而，Par3 是否在成牙本质细胞中表达及其对成牙本质细胞连接的潜在影响尚不清楚。本研究旨在探讨 Par3 对成牙本质细胞系（ODCs）细胞连接和生物学行为的影响。通过全转录组测序分析 Par3 对 ODCs 的影响及其潜在的分子机制。在生理和炎症条件下检测 ODCs 中 Par3 的表达。为了研究 Par3 对小鼠 ODC 之间连接的调节作用，通过 5-乙基-2'-脱氧尿苷（EdU）和 Transwell 测定以及流式细胞术检测 Par3 下调对 ODC 增殖、迁移、周期和凋亡的影响。Western blot 和茜素红 S 和碱性磷酸酶（ALP）染色用于观察 Par3 下调对 ODC 矿化的影响。通过全转录组测序研究 Par3 在 ODCs 中的生物学作用及其潜在的分子机制。Par3 定位于大鼠牙髓组织中成牙本质细胞层和 ODC 细胞质中。在炎症条件下，Par3 表达下调。Par3 沉默后 OLC 连接不连续，总 ZO-1（紧密连接相关蛋白）表达和 OLC 细胞膜上 ZO-1 表达减少（P < 0.05）。Par3 下调后，连接相关蛋白（ZO-1）的表达下调（P < 0.05），ZO-1 从细胞膜转移到细胞质。shPar3 转染细胞中 OLC 增殖和迁移增强，但凋亡和矿化受到抑制（P < 0.05）。测序鉴定出 2996 个差异表达基因（DEGs），主要富集在对刺激和结合的反应中。Par3 下调通过促进 AKT 磷酸化过度激活 PI3k-AKT 通路（P < 0.05）。Par3 下调可能通过影响 ZO-1 表达和分布破坏 OLC 之间的连接，促进 OLC 增殖和迁移，但抑制 OLC 矿化。Par3 可能与 14-3-3 蛋白相互作用激活 PI3K-AKT 通路，影响 OLC 连接和功能。
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