Wang Yanchun, Dong Yilong, Zhang Wenbo, Wang Yanmei, Jao Yang, Liu Jianjun, Zhang Mingzhu, He Hongbing
School and Hospital of Stomatology, Kunming Medical University, Kunming, Yunnan, People's Republic of China.
School of Medicine, Yunnan University, Kunming, Yunnan, People's Republic of China.
Apoptosis. 2023 Aug;28(7-8):1012-1023. doi: 10.1007/s10495-023-01839-z. Epub 2023 Apr 4.
Epithelial disruption is the initiation of most infectious disease. Regulation of epithelium apoptosis may play a key role in balance the survival competition between resident bacteria and host cells. The role of the mTOR/p70S6K pathway in preventing apoptosis of human gingival epithelial cells (hGECs) infected with Porphyromonas gingivalis (Pg) was investigated in order to further understand the survival strategy of the epithelial cells in during Pg infecting. hGECs was challenged with Pg for 4, 12, and 24 h. Additionally, hGECs was pretreated with LY294002 (PI3K signaling inhibitor) or Compound C (AMPK inhibitor) for 12 h and exposed them to Pg for 24 h. Subsequently, apoptosis was detected using flow cytometry, and expression and activity of Bcl-2, Bad, Bax, PI3K, AKT, AMPK, mTOR, and p70S6K proteins were analyzed using western blotting. Pg-infecting did not increase apoptosis of hGECs; but the expression ratio of Bad to Bcl-2 was increased after infecting. In contrast, Bad phosphorylation was promoted, accompanied by a significant reduction of mTOR/p70S6K and PI3K/AKT signaling, along with the upregulation of AMPK signaling. Morrover, the PI3K inhibitor LY294002 promoted Pg-mediated reduction of mTOR/p70S6K expression, and the increase of AMPK signaling and Bad phosphorylation rate, eventually decreasing apoptosis. While Compound C inhibited Pg-mediated activation of AMPK and downregulation of mTOR/p70S6K signaling, significantly reduced the Bad phosphorylation rate, thereby increasing apoptosis. Thus, hGECs prevent apoptosis via an inherent cellular-homeostasis, pro-survival mechanism during Pg infection, the AMPK/mTOR/p70S6K pathway helps prevent apoptosis in hGECs infected with Pg by regulating Bad phosphorylation.
上皮细胞破坏是大多数传染病的起始环节。上皮细胞凋亡的调控可能在平衡常驻细菌与宿主细胞之间的生存竞争中起关键作用。为了进一步了解牙龈卟啉单胞菌（Pg）感染期间上皮细胞的生存策略，研究了mTOR/p70S6K信号通路在预防感染Pg的人牙龈上皮细胞（hGECs）凋亡中的作用。用Pg刺激hGECs 4、12和24小时。此外，将hGECs用LY294002（PI3K信号抑制剂）或Compound C（AMPK抑制剂）预处理12小时，然后使其暴露于Pg中24小时。随后，使用流式细胞术检测凋亡情况，并通过蛋白质印迹法分析Bcl-2、Bad、Bax、PI3K、AKT、AMPK、mTOR和p70S6K蛋白的表达及活性。Pg感染并未增加hGECs的凋亡；但感染后Bad与Bcl-2的表达比值增加。相反，Bad磷酸化得到促进，同时mTOR/p70S6K和PI3K/AKT信号显著降低，伴随AMPK信号上调。此外，PI3K抑制剂LY294002促进了Pg介导的mTOR/p70S6K表达降低、AMPK信号增加以及Bad磷酸化率升高，最终导致凋亡减少。而Compound C抑制了Pg介导的AMPK激活以及mTOR/p70S6K信号下调，显著降低了Bad磷酸化率，从而增加了凋亡。因此，hGECs在Pg感染期间通过一种内在的细胞稳态、促生存机制来预防凋亡，AMPK/mTOR/p70S6K信号通路通过调节Bad磷酸化来帮助预防感染Pg的hGECs凋亡。
