Buzhong Yiqitang Regulates Endoplasmic Reticulum Stress via Nrf2/ROS/PERK/CHOP Signaling Pathway to Attenuate Cisplatin Resistance in NSCLC
10.13422/j.cnki.syfjx.20242025
- VernacularTitle:补中益气汤通过Nrf2/ROS/PERK/CHOP信号通路调控内质网应激改善NSCLC顺铂耐药的分子机制
- Author:
He LI
1
;
Yuetong LIU
1
;
Jingyi HUANG
1
;
Qirui MU
1
;
Chunying LIU
1
;
Yuan GAO
1
Author Information
1. College of Integrated Traditional Chinese and Western Medicine， Liaoning University of Traditional Chinese Medicine， Shenyang 110847， China
- Publication Type:Journal Article
- Keywords:
Buzhong Yiqitang;
non-small cell lung cancer;
chemotherapy resistance;
endoplasmic reticulum stress;
reactive oxygen species
- From:
Chinese Journal of Experimental Traditional Medical Formulae
2025;31(10):79-89
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo explore the molecular mechanism of Buzhong Yiqitang in attenuating cisplatin resistance of non-small cell lung cancer （NSCLC） cells （A549/DDP） by regulating endoplasmic reticulum stress （ERS） via the nuclear factor E2-related factor 2 （Nrf2）/reactive oxygen species （ROS）/double-stranded RNA-activated protein kinase R （PKR）-like endoplasmic reticulum kinase （PERK）/CCAAT enhancer-binding protein homologous protein （CHOP） signaling pathway. MethodsSprague Dawley （SD） rats were used to prepare blank serum and Buzhong Yiqitang-containing serum samples， and A549/DDP cells were sub-cultured and randomly allocated into 9 groups： Blank （10% blank rat serum medium）， model （10% blank rat serum medium+20 mg·L-1 cisplatin）， traditional Chinese medicine（TCM） （10% Buzhong Yiqitang-containing rat serum medium+20 mg·L-1 cisplatin）， TBHQ （10% blank rat serum medium+5 μmol·L-1 TBHQ+20 mg·L-1 cisplatin）， TBHQ+TCM （10% Buzhong Yiqitang-containing rat serum medium+5 μmol·L-1 TBHQ+20 mg·L-1 cisplatin）， NAC （10% blank rat serum medium+600 μmol·L-1 NAC+20 mg·L-1 cisplatin）， NAC+TCM （10% Buzhong Yiqitang-containing rat serum medium+600 μmol·L-1 NAC+20 mg·L-1 cisplatin）， Salubrinal （10% blank rat serum medium+20 μmol·L-1 Salubrinal+20 mg·L-1 cisplatin）， and Salubrinal+TCM （10% Buzhong Yiqitang-containing rat serum medium+20 μmol·L-1 Salubrinal+20 mg·L-1 cisplatin）. The median inhibitory concentration （IC50） of cisplatin in each group was determined by the cell counting kit-8 （CCK-8） method. The ROS level was detected by flow cytometry. The ultrastructure of endoplasmic reticulum （ER） was observed by transmission electron microscopy. Western blot was employed to determine the protein levels of Nrf2， phosphorylated （p）-Nrf2， CHOP， PERK， p-PERK， eukaryotic translation initiation factor 2α （eIF2α）， p-eIF2α， and activated transcription factor 4 （ATF4）. The fluorescence intensity of CHOP and p-PERK was detected by confocal fluorescence localization. ResultsCompared with the model group， the TCM group showed a decrease in IC50 of cisplatin in A549/DDP cells （P<0.01）. TBHQ， NAC， and Salubrinal groups showed increases in IC50 of cisplatin in A549/DDP cells （P<0.01）. TCM combined with TBHQ， NAC， and Salubrinal reduced the IC50 （P<0.01）. Compared with that in the blank group， the ROS level in the model group elevated （P<0.01）. Compared with that in the model group， the ROS level in the TCM group elevated （P<0.01）. The ROS levels in the TBHQ， NAC， and Salubrinal groups were lower than that in the model group （P<0.01）. The combinations of TBHQ， NAC， and Salubrinal with TCM raised the ROS levels of A549/DDP cells （P<0.01）. A flat saclike and neatly arranged ER structure was observed in the blank group， and slight expansion of ER was observed in the model group. Significant expansion of ER was observed in the TCM group. The expansion of ER was not obvious in the TBHQ， NAC， and Salubrinal groups but significant after TBHQ， NAC， and Salubrinal were combined with TCM. CHOP and p-PERK showed higher fluorescence intensity in the model group than in the blank group （P<0.01）， and the fluorescence signal intensity in the TCM group was higher than that in the model group （P<0.01）. The fluorescence intensity in the TBHQ， NAC， and Salubrinal groups was lower than that in the model group （P<0.01）. The fluorescence intensity in the groups of TBHQ， NAC， and Salubrinal combined with TCM was higher than that in corresponding groups without TCM （P<0.01）. Compared with those in the blank group， the expression levels of Nrf2 and p-Nrf2 were up-regulated in the model group （P<0.05）. Compared with the model group， the TCM group showed down-regulated expression levels of Nrf2 and p-Nrf2 （P<0.05）. TBHQ， NAC， and Salubrinal increased the expression levels of Nrf2 and p-Nrf2 （P<0.05）， while their combinations with TCM decreased the expression levels of Nrf2 and p-Nrf2 compared with the corresponding groups without TCM （P<0.01）. There were no significant differences in the expression levels of PERK and eIF2α between groups. The expression levels of CHOP， p-PERK/PERK， p-eIF2α/eIF2α， and ATF4 were up-regulated in the model group compared with those in the blank group （P<0.05）. Compared with the model group， the TCM group showed up-regulated expression levels of CHOP， p-PERK/PERK， p-eIF2α/eIF2α， and ATF4 （P<0.05）， while the TBHQ， NAC， and Salubrinal groups showed down-regulated expression levels （P<0.05）. The groups of TBHQ， NAC， and Salubrinal combined with TCM had higher expression levels than the groups without TCM （P<0.01）. ConclusionBuzhong Yiqitang regulates ERS via the Nrf2/ROS/PERK/CHOP signaling pathway to attenuate cisplatin resistance in NSCLC.
