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Spermine Protects Cardiomyocytes from High Glucose-Induced Energy Disturbance by Targeting the CaSR-gp78-Ubiquitin Proteasome System

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Abstract

Purpose

To determine the mediation of spermine on energy metabolism disorder and diabetic cardiomyopathy (DCM) development as well as the underlying mechanisms.

Methods

An in vitro model of DCM was established by incubating primary cultured neonatal rat cardiomyocytes with high glucose (HG). Spermine content was assessed by RP-HPLC. The protein levels were detected by western blot. Mitochondrial functions were analyzed using the respiratory chain complex assay kit and immunofluorescence staining.

Results

The endogenous content of spermine was decreased in the HG group, and the protein levels of ornithine decarboxylase, respiratory chain complex (I–V), mitochondrial fusion-related protein (Mfn1, Mfn2), Cx43, N-cadherin, CaSR, and β-catenin (in cytomembrane) were also down-regulated by HG. In contrast, the protein levels of spermine-N1-acetyltransferase, gp78, Fis1, Drp1, and β-catenin were up-regulated by HG. Meanwhile, we observed that HG increased ubiquitination levels of Mfn1, Mfn2, and Cx43, decreased membrane potential (ΔΨm), and the opening of mitochondrial permeability transport pore (mPTP) followed by intracellular ATP leakage. The supplement of spermine or siRNA-mediated knockdown of gp78 significantly alleviated the detrimental effects of HG, while downregulation of CaSR aggravated the development of DCM. We further confirmed that the lower level of spermine by HG activates the gp78-ubiquitin-proteasome pathway via downregulation of CaSR protein level, which in turn damages mitochondrial gap junction intercellular communication and leads to reduced ATP level.

Conclusion

The protective role of spermine on energy metabolism disorder is based on higher CaSR protein level and lower gp78 activation, pointing to the possibility that spermine can be a target for the prevention and treatment of DCM.

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Availability of Data and Materials

The used and/or analyzed datasets are available from the corresponding author on reasonable request.

Abbreviations

AMFR:

autocrine motility factor receptor

ATP:

adenosine triphosphate

ATP5F1:

ATP synthase F(0) complex subunit B1

COX5A:

cytochrome c oxidase subunit Va

Cx43:

connexin-43

DCM:

diabetic cardiomyopathy

Drp1:

dynamin-related protein 1

ERAD:

endoplasmic reticulum related degradation

Fis1:

fission 1

GJIC:

gap junction intercellular communication

GSK-3β:

glycogen synthase kinase 3 beta

HG:

high glucose

Hsp70:

heat-shock protein 70

Mfn:

mitofusin

ND1:

NADH dehydrogenase subunit 1

ODC:

ornithine decarboxylase

p-β-catenin:

phospho-β-catenin

p-Cx43:

phospho-connexin-43

p-GSK-3β:

phospho-glycogen synthase kinase 3 beta

SDHA:

succinate dehydrogenase complex flavoprotein subunit A

SLDT:

scrape-loading dye transfer technique

SSAT:

spermine-N1-acetyltransferase

TBST:

tris-buffered saline-Tween 20

UQCRQ:

ubiquinol-cytochrome c reductase

VDAC:

voltage-dependent anion channel

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Funding

This research was supported by the National Natural Science Foundation of China (No. 81800260), University Nursing Program for Young Scholars with Creative Talents in Heilongjiang Province (UNPYSCT), Harbin Medical University Innovation and Entrepreneurship Training Project for College Students (No.201910226010 and No. 201910226448) and Heilongjiang Postdoctoral Fund (No. LBH-Z17103).

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Author notes

  1. Yuehong Wang and Yuwen Wang contributed equally to this work.

Authors and Affiliations

  1. Department of Pathophysiology, Harbin Medical University, Baojian Road, Harbin, 150081, China

    Yuehong Wang, Fadong Li, Xinying Zhang, Hongzhu Li, Changqing Xu & Can Wei

  2. Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin, 150000, China

    Yuwen Wang

  3. Departemnt of Chemistry and Biochemistry, Laurentian University, Sudbury, P3E 2C6, Canada

    Guangdong Yang

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  1. Yuehong Wang
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Contributions

CW, YHW and YWW designed the research and drafted the manuscript; YHW, YWW, FDL, XXL, and XYZ completed the experiment and data analysis; HZL, GDY, and CQX revised the paper and gave some suggestions. All authors reviewed the results and approved the final version of the manuscript.

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Correspondence to Can Wei.

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Wang, Y., Wang, Y., Li, F. et al. Spermine Protects Cardiomyocytes from High Glucose-Induced Energy Disturbance by Targeting the CaSR-gp78-Ubiquitin Proteasome System. Cardiovasc Drugs Ther 35, 73–85 (2021). https://doi.org/10.1007/s10557-020-07064-z

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