Department of Restorative Dentistry, School of Dentistry, University of São Paulo, São Paulo, Brazil.
Department of Biological Sciences, Dental School of Bauru, University of São Paulo, Bauru, Brazil.
J Endod. 2020 Jun;46(6):810-817. doi: 10.1016/j.joen.2020.03.015. Epub 2020 Apr 21.
Stem cells of apical papilla (SCAP) may be affected by inflammatory mediators released by activation with lipopolysaccharide (LPS) from infected pulpal cavities of necrotic immature teeth. Therefore, this study aimed to investigate the presence of a local renin-angiotensin system (RAS) and the role of angiotensin II (Ang II) on the modulation of SCAP in vitro.
Primary cultures of SCAP were incubated with LPS (0.1-10 μg/mL) for cell viability and quantification of the chemokine CCL2. Components of RAS were searched by gene expression of angiotensinogen (AGTN), angiotensin converting enzyme (ACE), renin, angiotensin receptor 1 (AT1) and 2 (AT2), and Mas receptor. Ang II was investigated in SCAP supernatants. Immunofluorescence was used to detect AGTN and AT1. Next, cells were treated with Ang II for viability/proliferation assessment, quantification of CCL2 and interleukin 6, and mineralization assay. Data were evaluated by analysis of variance using Tukey post hoc comparisons or the Student t test. P values <.05 were considered to be significant.
LPS increased CCL2 production at 1 and 10 μg/mL. The gene expression of AGTN, renin, ACE, and AT1 was detected, but only ACE was increased by LPS. Ang II peptide was found in SCAP supernatants but unaltered by LPS. Both AGTN and AT1 proteins were detected by immunostaining. Ang II significantly induced SCAP proliferation, increased CCL2 production, down-regulated IL-6 release, and reduced the SCAP mineralization rate.
A local RAS was found at the apical papilla, and Ang II was able to modulate SCAP function in vitro.
来自感染性坏死未成熟牙髓腔的脂多糖（LPS）激活后释放的炎症介质可能会影响根尖乳头干细胞（SCAP）。因此，本研究旨在探讨局部肾素-血管紧张素系统（RAS）的存在，以及血管紧张素 II（Ang II）在体外调节 SCAP 中的作用。
将 SCAP 的原代培养物与 LPS（0.1-10μg/ml）孵育，以评估细胞活力和趋化因子 CCL2 的定量。通过血管紧张素原（AGTN）、血管紧张素转换酶（ACE）、肾素、血管紧张素受体 1（AT1）和 2（AT2）和 Mas 受体的基因表达来搜索 RAS 的成分。研究了 SCAP 上清液中的 Ang II。通过免疫荧光检测 AGTN 和 AT1。然后，用 Ang II 处理细胞，以评估细胞活力/增殖、CCL2 和白细胞介素 6 的定量以及矿化测定。使用方差分析和 Tukey 事后比较或学生 t 检验评估数据。P 值<.05 被认为具有统计学意义。
LPS 在 1 和 10μg/ml 时增加了 CCL2 的产生。检测到 AGTN、肾素、ACE 和 AT1 的基因表达，但只有 LPS 增加了 ACE。在 SCAP 上清液中发现了 Ang II 肽，但 LPS 未改变其含量。免疫染色检测到 AGTN 和 AT1 蛋白。Ang II 显著诱导 SCAP 增殖，增加 CCL2 的产生，下调 IL-6 释放，并降低 SCAP 矿化率。
在根尖乳头中发现了局部 RAS，Ang II 能够在体外调节 SCAP 的功能。
