Investigation of the inhibitory potential of caffeic acid phenethyl ester on prion replication, amplification, and fibril formation in vitro
10.3760/cma.j.cn112150-20240325-00249
- VernacularTitle:咖啡酸苯乙酯对朊病毒体外复制、扩增和纤维形成的抑制作用研究
- Author:
Zhiyue CHAO
1
;
Xiaoxi JIA
;
Jiafeng ZENG
;
Yuezhang WU
;
Kang XIAO
;
Liping GAO
;
Qi SHI
;
Xiaoping DONG
;
Cao CHEN
Author Information
1. 传染病溯源预警与智能决策全国重点实验室 国家卫生健康委医学病毒和病毒病重点实验室 中国疾病预防控制中心病毒病预防控制所，北京 102206
- Keywords:
Caffeic acid phenethyl ester;
Prion;
Prion protein
- From:
Chinese Journal of Preventive Medicine
2024;58(7):1011-1019
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effects and possible mechanisms of caffeic acid phenethyl ester (CAPE) on the replication, amplification, and fibre formation of prions (PrP Sc). Methods:The CCK8 assay was used to detect the cell viability of the prion-infected cell model SMB-S15 after CAPE treatment for 3 days and 7 days and the maximum safe concentration of CAPE for SMB-S15 was obtained. The cells were treated with a concentration within a safe range, and the content of PrP Sc in the cells before and after CAPE treatment was analyzed by western blot. Protein misfolding cycle amplification (PMCA) and western blot were used to assess changes in PrP Sc level in amplification products following CAPE treatment. Real-time-quaking induced conversion assay (RT-QuIC) technology was employed to explore the changes in fibril formation before and after CAPE treatment. The binding affinity between CAPE and murine recombinant full-length prion protein was determined using a molecular interaction assay. Results:CCK8 cell viability assay results demonstrated that treatment with 1 μmol/L CAPE for 3 and 7 days did not exhibit statistically significant differences in cell viability compared to the control group (all P<0.05). However, when the concentration of CAPE exceeded 1 μmol/L, a significant reduction in cell viability was observed in cells treated with CAPE for 3 and 7 days, compared to the control group (all P<0.05). Thus, 1 μmol/L was determined as the maximum safe concentration of CAPE treatment for SMB-S15 cells. The western blot results revealed that treatment with CAPE for both 3 and 7 days led to a detectable reduction in the levels of PrP Sc in SMB-S15 cells (all P<0.05). The products of PMCA experiments were assessed using western blot. The findings revealed a significant decrease in the levels of PrP Sc (relative grey value) in the PMCA amplification products of adapted-strains SMB-S15, 139A, and ME7 following treatment with CAPE, as compared to the control group (all P<0.05). The RT-QuIC experimental results demonstrated a reduction in fibril formation (as indicated by ThT peak values) in CAPE-treated mouse-adapted strains 139A, ME7, and SMB-S15, as well as in SMB-S15 cells infected with prions. Furthermore, CAPE exhibited varying degrees of inhibition towards different seed fibrils formation, with statistically significant differences observed (all P<0.05). Notably, CAPE exhibited a more pronounced inhibitory effect on ME7 seed fibrils. Molecular interaction analyses demonstrated significant binding between CAPE and murine recombinant prion protein, and the association constant was (2.92±0.41)×10 -6 mol/L. Conclusions:CAPE inhibits PrP Sc replication, amplification, and fibril formation in vitro possibly due to specific interactions with the prion protein at the molecular level.
