2. Academic Validation
  3. Mechanistic Insights into Salidroside's Mitochondrial Protection via AMPK/Sirt1/HIF-1α Pathway in Hypoxic HT22 Cells

Mechanistic Insights into Salidroside's Mitochondrial Protection via AMPK/Sirt1/HIF-1α Pathway in Hypoxic HT22 Cells

  • J Vis Exp. 2025 Apr 25:(218). doi: 10.3791/66923.
Xiaobo Wang 1,  Yating Zhang 2,  Ya Hou 2,  Hong Jiang 2,  Tingting Kuang 2,  Rui Li 3,  Xianli Meng 4
Affiliations

Affiliations

  • 1 School of Pharmacy/School of Modern Chinese Medicine Industry, Chengdu University of Traditional Chinese Medicine; Innovative Institute of Chinese Medicine and Pharmacy/Academy for Interdiscipline, Chengdu University of Traditional Chinese Medicine; Research Service Office, Traditional Chinese Medicine Hospital of Meishan; Vitadrwang@cdutcm.edu.cn.
  • 2 School of Pharmacy/School of Modern Chinese Medicine Industry, Chengdu University of Traditional Chinese Medicine.
  • 3 Research Service Office, Traditional Chinese Medicine Hospital of Meishan; ruilee@cdutcm.edu.cn.
  • 4 School of Pharmacy/School of Modern Chinese Medicine Industry, Chengdu University of Traditional Chinese Medicine; Innovative Institute of Chinese Medicine and Pharmacy/Academy for Interdiscipline, Chengdu University of Traditional Chinese Medicine; Research Service Office, Traditional Chinese Medicine Hospital of Meishan; xlm999@cdutcm.edu.cn.
PMID: 40354296 DOI: 10.3791/66923
Abstract

Salidroside (Sal), an active ingredient of Rhodiola crenulata (Hook. f. et Thoms.) H. Ohba has been found to exert mitochondrial protective effects by improving metabolism and enhancing the energy supply of brain cells under hypoxic conditions. However, its mechanism of action has not been fully clarified. In the present study, high-performance liquid chromatography was first employed to analyze the effects of Sal on nucleotide (ATP, ADP, and AMP) levels. The cellular thermal shift assay (CETSA), a widely used molecular interaction method for validating and quantifying drug target engagement in cells and tissues across different species, was then chosen to confirm the affinity of Sal for AMPK/SIRT1/HIF-1α pathway-related proteins. The results revealed that Sal increased ATP and ADP levels in hypoxic HT22 cells while reducing AMP levels. Moreover, Sal exhibited stable binding to AMPKα, p-AMPKα, SIRT1, and HIF-1α proteins. In conclusion, Sal may exert mitochondrial protective effects by modulating the AMPK/SIRT1/HIF-1α pathway to regulate nucleotide content. This study provides a methodological reference for nucleotide content analysis in cell samples and contributes to the identification and discovery of targets for compounds derived from traditional Chinese medicine.

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