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Chronic hyperglycemia impairs mitochondrial unfolded protein response and precipitates proteotoxicity in experimental diabetic neuropathy: focus on LonP1 mediated mitochondrial regulation

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Abstract

Background

Disturbed mitochondrial homeostasis has been identified to contribute to the pathogenesis of diabetic neuropathy (DN). However, the role of Mitochondrial Lon peptidase 1 (Lonp1) and Heat shock proteins (HSP’s) in DN remains elusive. Here we studied the role of these proteins in experimental DN.

Methods

Rats were injected with STZ (55 mg/kg, ip) to induce diabetes. After confirmation of diabetes, animals were maintained for 8 weeks to develop neuropathy. Resveratrol was administered at two dose levels 10 and 20 mg/kg for last 2 weeks. Neuronal PC12 cells was challenged with 30 mM of β-d glucose to evaluate the molecular changes.

Results

Diabetic rats showed reduced expression of various mitochondrial proteases in dorsal root ganglions (DRG). This effect may increase proteotoxicity and diminish electron transport chain (ETC) activity as evident by increased protein oxidation and reduced ETC complexes activities under diabetic condition. In particular, we focused on our efforts to characterize the expression pattern of Lonp1 which was found to be significantly (p < 0.01 vs. control group) under expressed in DRG of diabetic rats. We used Resveratrol to characterize the importance of Lonp1 in regulation of mitochondrial function. High glucose (HG) (30 mM) exposed PC12 cells suggested that Resveratrol treatment attenuated the HG induced mitochondrial damage via induction of mitochondrial proteases. Moreover, siRNA directed against Lonp1 has impaired the activity of Resveratrol in attenuating the HG induced mitochondrial dysfunction.

Conclusion

These results would signify the importance of modulating mitochondrial proteases for the therapeutic management of DN

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Acknowledgements

Authors would like to acknowledge the financial support provided by the Department of Pharmaceuticals, Ministry of Chemicals and Fertilizers, Government of India for carrying out this work. The abstract of this paper was presented on the NEURODIAB conference 2017, (https://www.neurodiab2017.com/AbstractBook.aspx) and awarded as Best poster and Young Investigators Award in NEURODIAB 2017.

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Authors and Affiliations

  1. Department of Pharmacology and Toxicology, National Institute of Pharmaceutical Education and Research (NIPER)-Hyderabad, Bala Nagar, Hyderabad, Telangana, 500037, India

    Anil Kumar Kalvala, Bhoomika Sherkhane, Chayanika Gundu, Vijay Arruri, Rahul Kumar & Ashutosh Kumar

  2. Keenan Research Centre for Biomedical Science and Li Ka Shing Knowledge Institute, St. Michael’s Hospital, Toronto, ON, Canada

    Veera Ganesh Yerra

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Contributions

Mr. AKK performed studies, evaluated data, prepared figures and wrote manuscript; Dr. VGY, performed studies, evaluated data and wrote manuscript; Mrs. BS, performed studies, evaluated data and reviewed manuscript; Mrs. CG, performed studies, evaluated data and reviewed manuscript, Mr. VA, performed studies, evaluated data and reviewed manuscript, Dr. RK, performed studies, evaluated data and reviewed manuscript; Dr. AK, performed studies, evaluated data and wrote manuscript.

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Correspondence to Ashutosh Kumar.

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43440_2020_147_MOESM1_ESM.tif

Supplementary Figure 1: Effect of resveratrol and Silencing of Lonp1 gene on mitochondrial superoxides and mitochondrial membrane potential (Δψm) in high glucose exposed PC12 cells (A) Dot plot monitored by FACS analysis represents Δψm depolarization of JC-1 mitochondrial potential marker staining, (B) Graphic representation of mean values for JC-1 ratio (% cells with trimeric JC-1/% cells with monomeric or depolarized mitochondria) and (C) Bar graph represents the Mitochondrial ROS levels measured by Mitosox staining in PC12 cells of various groups measured by FACS analysis. Results are expressed as mean ± SEM (n=3). NC: PC12 cells under normal conditions, HG: PC12 cells exposed to high glucose (30 mM) condition, HG+RSV: PC12 cells coincubated with HG stimulus and RSV (20μM), HG+RSV+LONP1-T: PC12 cells exposed to glucose and RSV and Lonp1 siRNA (0.5μM), LONP1-T: PC12 cells transfected with Lonp1 siRNA (0.5 μM) and siRNA negative control represents scramble control. ***p<0.001, **p<0.01 vs. HG and ###p<0.001 vs. NC (TIF 83885 kb)

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Kalvala, A.K., Yerra, V.G., Sherkhane, B. et al. Chronic hyperglycemia impairs mitochondrial unfolded protein response and precipitates proteotoxicity in experimental diabetic neuropathy: focus on LonP1 mediated mitochondrial regulation. Pharmacol. Rep 72, 1627–1644 (2020). https://doi.org/10.1007/s43440-020-00147-6

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