Celastrol attenuates 6-hydroxydopamine-induced neurotoxicity by regulating the miR-146a/PI3K/Akt/mTOR signaling pathways in differentiated rat pheochromocytoma cells

J Affect Disord. 2022 Nov 1:316:233-242. doi: 10.1016/j.jad.2022.08.026. Epub 2022 Aug 15.

Abstract

Background: Parkinson's disease (PD) is a neurological disorder. Recently, celastrol (Cel) has been reported to have neuroprotective properties. We investigated the protective effects of Cel on PD in a cell model with 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in PC12 cells and further addressed the underlying protective mechanisms of Cel.

Methods: PC12 cells were treated with 6-OHDA, and Cel was added to the medium at various concentrations. The CCK-8 assay, Hoechst/PI staining, and flow cytometry analysis were performed to detect cellular viability and apoptosis. Mitochondrial membrane potential (MMP) was examined by JC-1 staining. ROS level was quantified by ROS staining. The effects of Cel on the expression of miR-146a and PI3K/Akt/mTOR pathway were then clarified using real-time PCR and Western blotting. Moreover, a miR-146a mimic was synthesized and transfected into PC12 cells to further determine the mechanisms of Cel's neuronal protection against 6-OHDA-induced neurotoxicity.

Results: Cel greatly improved cell viability and lessened apoptosis. Flow cytometry showed that Cel especially inhibited early apoptosis. Cel also obviously restored the MMP and decreased ROS level destroyed by 6-OHDA. Moreover, 6-OHDA increased the expression of miR-146a and decreased pAkt/mTOR protein levels, whereas Cel reversed these changes. In particular, miR-146a targeted and inhibited the expression of PI3K, an upstream molecule of Akt/mTOR. Transfection of 6-OHDA-treated neurons with miR-146a mimic notably attenuated Cel's protective effects.

Limitations: There were no animal experiments in our study.

Conclusions: Cel exerts neuroprotective activity against 6-OHDA-caused neurotoxicity by regulating miR-146a/PI3K/Akt/mTOR pathway, which provides a potential application of Cel for treating neurodegenerative diseases.

Keywords: 6-OHDA; Celastrol; Neurotoxicity; PI3K/Akt/mTOR; miR-146a.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adrenal Gland Neoplasms*
  • Animals
  • Apoptosis
  • MicroRNAs* / genetics
  • MicroRNAs* / metabolism
  • Neuroprotective Agents* / pharmacology
  • Oxidopamine / toxicity
  • Parkinson Disease*
  • Pentacyclic Triterpenes
  • Pheochromocytoma*
  • Phosphatidylinositol 3-Kinases / metabolism
  • Proto-Oncogene Proteins c-akt / metabolism
  • Rats
  • Reactive Oxygen Species
  • Signal Transduction / physiology
  • TOR Serine-Threonine Kinases / metabolism

Substances

  • MicroRNAs
  • Neuroprotective Agents
  • Pentacyclic Triterpenes
  • Reactive Oxygen Species
  • Oxidopamine
  • mTOR protein, rat
  • Proto-Oncogene Proteins c-akt
  • TOR Serine-Threonine Kinases
  • celastrol