5-氨基酮戊酸光动力疗法促进内质网应激在 HR-HPV 感染的 HeLa 细胞治疗中的机制。
The mechanism of 5-aminolevulinic acid photodynamic therapy in promoting endoplasmic reticulum stress in the treatment of HR-HPV-infected HeLa cells.
机构信息
Department of Dermatology and Venereology, Nanfang Hospital, Southern Medical University, Guangzhou, China.
Molecular Diagnosis and Treatment Center for Infectious Diseases, Dermatology Hospital, Southern Medical University, Guangzhou, China.
出版信息
Photodermatol Photoimmunol Photomed. 2021 Jul;37(4):348-359. doi: 10.1111/phpp.12663. Epub 2021 Feb 8.
BACKGROUND
5-aminoketovaleric acid, as a precursor of the strong photosensitizer protoporphyrin IX (PpIX), mainly enters the mitochondria after entering the cell, and the formed PpIX is also mainly localized in the mitochondria. So at present the research on the mechanism of 5-aminoketovalerate photodynamic therapy (ALA-PDT) mainly focuses on its impact on mitochondria. There are few reports on whether ALA-PAT can affect the endoplasmic reticulum and trigger endoplasmic reticulum stress (ERS).
AIMS/OBJECTIVES: Here we investigated the effects of ALA-PDT on endoplasmic reticulum and its underlying mechanisms in high-risk human papillomavirus (HR-HPV) infection.
MATERIALS AND METHODS
The human cervical cancer cell line HeLa (containing whole genome of HR-HPV18) was treated with ALAPDT, and cell viability, ROS production, the level of Ca2+ in the cytoplasm and apoptosis were evaluated by CCK8, immunofluorescence and flow cytometry, respectively. The protein expression of the markers of ERS and autophagy and CamKKβ-AMPK pathway was examined by western blot.
RESULTS
The results showed that ALA-PDT inhibited cell viability of HeLa cells in vitro; ALA-PDT induced autophagy in HeLa cells ; ALA-PDT induced autophagy via the Ca2+-CamKKβ-AMPK pathway, which could be suppressed by the inhibition of ERS;ALA-PDT induced ERS-specific apoptosis via the activation of caspase 12.
CONCLUSIONS
Our study demonstrated that ALA-PDT could exert a killing effect by inducing HeLa cell apoptosis, including endoplasmic reticulum-specific apoptosis. Meanwhile, ERS via the Ca2+ -CamKKβ-AMPK pathway promoted the occurrence of autophagy in HeLa cells. Inhibition of autophagy could increase the apoptosis rate of HeLa cells after ALA-PDT, suggesting that autophagy may be one of the mechanisms of PDT resistance; The Ca2+-CamKKβ-AMPK pathway and autophagy may be targets to improve the killing effect of ALA-PDT in treating HR-HPV infection.
背景
5-氨基酮戊酸作为强光敏剂原卟啉 IX（PpIX）的前体，进入细胞后主要进入线粒体，形成的 PpIX 也主要定位于线粒体。因此，目前 5-氨基酮戊酸光动力疗法（ALA-PDT）的机制研究主要集中在其对线粒体的影响上。关于 ALA-PAT 是否能影响内质网并引发内质网应激（ERS）的报道较少。
目的/目标：本研究旨在探讨 ALA-PDT 对高危型人乳头瘤病毒（HR-HPV）感染内质网的影响及其机制。
材料和方法
用 ALA-PDT 处理人宫颈癌细胞系 HeLa（含 HR-HPV18 全基因组），通过 CCK8、免疫荧光和流式细胞术分别评估细胞活力、ROS 产生、细胞质 Ca2+水平和细胞凋亡。通过 Western blot 检测 ERS 和自噬及 CamKKβ-AMPK 通路标志物的蛋白表达。
结果
结果表明，ALA-PDT 抑制 HeLa 细胞的体外细胞活力；ALA-PDT 诱导 HeLa 细胞自噬；ALA-PDT 通过 Ca2+-CamKKβ-AMPK 通路诱导自噬，该通路可被 ERS 抑制；ALA-PDT 通过激活 caspase 12 诱导内质网特异性凋亡。
结论
本研究表明，ALA-PDT 通过诱导 HeLa 细胞凋亡（包括内质网特异性凋亡）发挥杀伤作用。同时，通过 Ca2+-CamKKβ-AMPK 通路的 ERS 促进 HeLa 细胞自噬的发生。抑制自噬可增加 ALA-PDT 后 HeLa 细胞的凋亡率，提示自噬可能是 PDT 耐药的机制之一；Ca2+-CamKKβ-AMPK 通路和自噬可能是提高 ALA-PDT 治疗 HR-HPV 感染杀伤效果的靶点。
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