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Naringenin Upregulates AMPK-Mediated Autophagy to Rescue Neuronal Cells From β-Amyloid (1–42) Evoked Neurotoxicity

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Abstract

Deposition of an amyloid-β peptide is one of the first events in the pathophysiology of Alzheimer’s disease (AD) and is clinically characterized by Aβ plaques, tau tangles, and behavioral impairments that lead to neuronal death. A substantial number of studies encourage targeting the skewness in the production and degradation of amyloid-β could be among the promising therapies in the disease. Neuronal autophagy has emerged for an essential role in the degradation of such toxic aggregate-prone proteins in various neurodegenerative diseases. We profiled a small library of common dietary compounds and identified those that can enhance autophagy in neuronal cells. Here we noted naringenin in silico exhibits a robust affinity with AMP-activated protein kinase (AMPK) and upregulated AMPK-mediated autophagy signaling in neurons. Naringenin can induce autophagy promoting proteins such as ULK1, Beclin1, ATG5, and ATG7 in Neuro2a cells and primary mouse neurons as well. The knockdown of AMPK by siRNA-AMPK was complemented by naringenin that restored transcript levels of AMPK. Further, naringenin can reduce the levels of Aβ at a nontoxic concentration from neuronal cells. Moreover, it maintained the mitochondrial membrane potential and resisted reactive oxygen species production, which led to the protection against Aβ1–42 evoked neurotoxicity. This highlights the neuroprotective potential of naringenin that can be developed as an anti-amyloidogenic nutraceutical.

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Abbreviations

AD:

Alzheimer’s disease

AMPK:

AMP-activated protein kinase

MAP 2:

Microtubule-associated protein

GFAP :

Glial fibrillary acidic protein

LC3B:

Microtubule-associated protein 1 light chain 3 beta

mTOR :

Mammalian target of rapamycin

Aβ :

Amyloid beta

ULK1:

Unc-51-like kinase

ATG :

Autophagy-related protein

SQSTM1/P62:

Sequestosome 1 protein)

NGN:

Naringenin

Baf:

Bafilomycin A1

TMRE:

Tetramethylrhodamine ethyl ester

MTT :

Thiazolyl blue tetrazolium bromide

LDH :

Lactate dehydrogenase

AICAR:

5-Aminoimidazole-4-carboxamide ribonucleotide

MMP:

Mitochondrial membrane potential

ROS :

Reactive oxygen species

DCFH2-DA:

2', 7'-dichloro-dihydro fluorescein diacetate

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Acknowledgments

We are thankful to UGC (University Grants Commission) for providing financial assistance to Aitizaz Ul Ahsan to carry out this research. We are immensely grateful to Dr. Ajay Kumar for providing his lab during experimental work. We thank Dr. Akeel for his kind help in gifting plasmids. We cannot forget to mention Dr. Abid Hamid for his support and critical suggestions. Thanks are due to Dr. P.R. Sharma for assisting in microscopy and lastly to IIIM, Jammu Kashmir, for keeping facilities available. Dr. Mehak Gupta, Manisha, Mir Faheem Mohammad, Masroor Ahmad, and Adil Qadir, thanks for the help.

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Authors and Affiliations

  1. Cytogenetics Lab, Department of Zoology, Panjab University, Chandigarh, India

    Aitizaz Ul Ahsan, Vijay Lakshmi Sharma & Mani Chopra

  2. Department of Neurology, Washington University School of Medicine, Saint Louis, MO, USA

    Abubakar Wani

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  1. Aitizaz Ul Ahsan
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  2. Vijay Lakshmi Sharma
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  3. Abubakar Wani
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Contributions

A.U.A planned and performed the experiments and analyzed the data and compiled the manuscript. A.W. helped in designing some experiments. V.L.S and M.C. revised and approved the final version of the manuscript.

Corresponding author

Correspondence to Mani Chopra.

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Competing Interests

The authors declare that they have no competing interests.

Ethical Approval

All animal procedures were approved by the Institutional Animals Ethics Committee IAEC (PU/IAEC/S/15/86), Panjab University Chandigarh.

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Highlights

• Naringenin confers neuroprotection against soluble Aβ-evoked neurotoxicity in neuronal cells.

• AMPK-mediated autophagy by naringenin mediates its neuroprotection.

• This study not only establishes the underlying mechanism but also provides the basis for finding the strategy in treating the Aβ pathology.

Electronic Supplementary Material

Supplementary Fig. 1

(a) Screening of various compounds for AMPK induction with metformin (Met) as the standard. (b) Cell viability assay of naringenin at different concentrations in differentiated N2a cells and primary mouse neurons. (c) In-silco evaluation for molecular docking of naringenin with AMPK compared to AICAR. (d) Induction of autophagy by naringenin at different concentrations in N2a cells(PNG 851 kb)

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Ahsan, A.U., Sharma, V.L., Wani, A. et al. Naringenin Upregulates AMPK-Mediated Autophagy to Rescue Neuronal Cells From β-Amyloid (1–42) Evoked Neurotoxicity. Mol Neurobiol 57, 3589–3602 (2020). https://doi.org/10.1007/s12035-020-01969-4

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  • DOI: https://doi.org/10.1007/s12035-020-01969-4

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