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. 2021 Dec 17:9:802502.
doi: 10.3389/fcell.2021.802502. eCollection 2021.

Neuraminidase1 Inhibitor Protects Against Doxorubicin-Induced Cardiotoxicity via Suppressing Drp1-Dependent Mitophagy

Yating Qin  1 , Chao Lv  1 , Xinxin Zhang  1 , Weibin Ruan  1 , Xiangyu Xu  2 , Chen Chen  3 , Xinyun Ji  1 , Li Lu  1   4 , Xiaomei Guo  1
Affiliations

Neuraminidase1 Inhibitor Protects Against Doxorubicin-Induced Cardiotoxicity via Suppressing Drp1-Dependent Mitophagy

Yating Qin et al. Front Cell Dev Biol. .

Abstract

Anthracyclines, such as doxorubicin (DOX), are among the effective chemotherapeutic drugs for various malignancies. However, their clinical use is limited by irreversible cardiotoxicity. This study sought to determine the role of neuraminidase 1 (NEU1) in DOX-induced cardiomyopathy and the potential cardio-protective effects of NEU1 inhibitor oseltamivir (OSE). Male Sprague-Dawley (SD) rats were randomized into three groups: control, DOX, and DOX + OSE. NEU1 was highly expressed in DOX-treated rat heart tissues compared with the control group, which was suppressed by OSE administration. Rats in the DOX + OSE group showed preserved cardiac function and were protected from DOX-induced cardiomyopathy. The beneficial effects of OSE were associated with the suppression of dynamin-related protein 1 (Drp1)-dependent mitochondrial fission and mitophagy. In detail, the elevated NEU1 in cardiomyocytes triggered by DOX increased the expression of Drp1, which subsequently enhanced mitochondrial fission and PINK1/Parkin pathway-mediated mitophagy, leading to a maladaptive feedback circle towards myocardial apoptosis and cell death. OSE administration selectively inhibited the increased NEU1 in myocardial cells insulted by DOX, followed by reduction of Drp1 expression, inhibition of PINK1 stabilization on mitochondria, and Parkin translocation to mitochondria, thus alleviating excessive mitochondrial fission and mitophagy, alleviating subsequent development of cellular apoptotic process. This work identified NEU1 as a crucial inducer of DOX-induced cardiomyopathy by promoting Drp1-dependent mitochondrial fission and mitophagy, and NEU1 inhibitor showed new indications of cardio-protection against DOX cardiotoxicity.

Keywords: cardiotoxicity; doxorubicin; dynamin-related protein 1 (Drp1); mitophagy; neuraminidase1; oseltamivir.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
Doxorubicin (DOX) exposure resulted in elevated neuraminidase 1 (NEU1) activity and expression in adult rats. (A) The chemical structure of NEU1 inhibitor oseltamivir (OSE) and the schematic diagram of this study. (B–E) The content of NEU1 and its regulated downstream metabolite N-acetylneuraminic acid (Neu5AC) in animal plasma or heart tissue measured by commercial ELISA assays (n = 8 per group). (F) Representative immunohistochemical images of NEU1 expression in rat hearts (n = 5 per group, scale bar = 50 μm). (G) The expression of NEU1 in rat hearts analyzed by western blot (n = 6 per group). (**p < 0.01 vs. control group, ***p < 0.001 vs. control group. #p < 0.05 vs. DOX group, ##p < 0.01 vs. DOX group, ###p < 0.001 vs. DOX group).
FIGURE 2
FIGURE 2
NEU1 inhibitor improved DOX-induced cardiac dysfunction in rats. (A) The tendency of body weight change of rats in three groups (n = 12 per group). (B) The survival curve of rats in different groups. (C) Representative images of M-mode echocardiography in different rat groups at the endpoint of the study. (D) Statistical analyses of echocardiographic parameters including ejection fraction (EF), fractional shortening (FS), left ventricular end-systolic dimension (LVEDs), and left ventricular end-diastolic dimension (LVEDD) of rats in the three groups at the end of the study. (E) Representative images of hematoxylin and eosin (HE) staining, Masson’s trichrome staining, and wheat germ agglutinin (WGA) staining of heart tissues in the three different groups, which display the extent of cardiac atrophy, myocardial fibrosis, and cell sizes, respectively (scale bar = 50 μm). (F) Concentrations of cardiac injury markers including cardiac troponin T (cTnT), creatine kinase isoenzyme-MB (CK-MB), lactate dehydrogenase (LDH), and aspartate aminotransferase (AST) in plasma of rats in different groups. (**p < 0.01 vs. control group, ***p < 0.001 vs. control group. #p < 0.05 vs. DOX group, ##p < 0.01 vs. DOX group, ###p < 0.001 vs. DOX group).
FIGURE 3
FIGURE 3
NEU1 inhibitor modulated autophagy, mitochondrial fission, and mitophagy in DOX-treated rat hearts. (A) Representative images of LC3II expression in heart tissues of the three groups (scale bar = 50 μm). (B) The expression levels of LC3I/II, ATG5, Beclin 1, and P62 in heart tissues of the different groups by western blot (n = 6 per group). (C) Representative electron microscopy images of heart tissues in the three groups; the red arrows direct autophagic vacuoles containing cargos (scale bar = 1 μm). (D) The expression levels of dynamin-related protein 1 (Drp1), PINK1, and Parkin in heart tissues of the three groups by western blot (n = 6 per group). (E) Representative immunohistochemical images of Drp1, PINK1, and Parkin expressions in heart tissues of the three groups (n = 5 per group, scale bar = 50 μm). (**p < 0.01 vs. control group, ***p < 0.001 vs. control group. #p < 0.05 vs. DOX group, ##p < 0.01 vs. DOX group, ###p < 0.001 vs. DOX group).
FIGURE 4
FIGURE 4
NEU1 inhibitor effectively prevented apoptosis in DOX-treated rat hearts. The concentrations of endogenous antioxidants including total antioxidant capacity (T-AOC) (A), glutathione (GSH) (B), superoxide dismutase (SOD) (C), and endogenous oxidative factor hydrogen peroxide (H2O2) (D) in blood serum measured by commercial assays. (E) Representative immunohistochemical images of cleaved caspase-3 abundance in heart tissues of the three groups (n = 5 per group, scale bar = 50 μm). The expression levels of cleaved caspase-3 and cleaved caspase-9 (F) and Bax, Bcl-2, and Bad (G) in heart tissues of the three groups by western blot (n = 6 per group). (H) Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining of heart tissues in three groups; positive nuclei were stained in green while nuclei were stained in blue (n = 5 per group, scale bar = 50 μm). (**p < 0.01 vs. control group, ***p < 0.001 vs. control group. ##p < 0.01 vs. DOX group, ###p < 0.001 vs. DOX group).
FIGURE 5
FIGURE 5
DOX induced the upregulation of NEU1 and dysfunction of autophagy and mitophagy in H9C2 cells in a time-dependent manner. Representative immunofluorescent images of NEU1 (A) and Drp1 (B) expressions in H9C2 cells that suffered from DOX exposure for 0, 3, 6, and 12 h. Western blot analysis showed that prolonged DOX exposure gradually increased expressions of NEU1 (C) and Dpr1 (D) in H9C2 cells. (E) Prolonged DOX exposure increased the expressions of autophagic markers including ATG5, Beclin 1, and LC3I/II, while it decreased the expression of P62 in H9C2 cells. (F) Prolonged DOX exposure enhanced the expressions of mitophagy-related markers like PINK1 and Parkin in H9C2 cells. (G) Prolonged DOX exposure activated apoptotic markers including caspase-3 and caspase-9 in H9C2 cells. (*p < 0.05, **p < 0.01, ***p < 0.001).
FIGURE 6
FIGURE 6
OSE effectively blunted excessive mitochondrial fission and mitophagy in DOX-exposed H9C2 cells. (A) OSE suppressed the elevated NEU1 expression induced by DOX in H9C2 cells in a dose-dependent manner. (B) OSE suppressed the elevated Drp1 expression induced by DOX in H9C2 cells in a dose-dependent manner. Representative immunofluorescent images of NEU1 expression (C) or Drp1 expression (D) after OSE treatment in DOX-induced H9C2 cells. (E) Western blot analysis revealed that OSE suppressed the excessive autophagy activation induced by DOX in H9C2 cells in a dose-dependent manner. (F) Western blot analysis showed that OSE attenuated mitophagy activity in DOX-treated H9C2 cells in a dose-dependent manner through inhibiting PINK1 accumulation, Parkin recruitment, and ubiquitination on mitochondria. (G) OSE suppressed apoptotic activity in DOX-treated H9C2 cells in a dose-dependent manner. (*p < 0.05 vs. control group, **p < 0.01 vs. control group, ***p < 0.001 vs. control group. ##p < 0.01 vs. DOX group, ###p < 0.001 vs. DOX group).
FIGURE 7
FIGURE 7
Drp1 inhibitor Mdivi-1 showed similar effects with OSE in DOX-exposed H9C2 cells. (A) Mdivi-1 decreased the elevated Drp1 expression and suppressed the enhanced autophagic activity in DOX-treated H9C2 cells, similarly like OSE. (B) Mdivi-1 reduced PINK1 accumulation on mitochondria and prevent Parkin translocation from cytosol to mitochondria in DOX-induced H9C2 cells. (C) Consistent with the effects of OSE, Mdivi-1 suppressed apoptotic activity in DOX-induced H9C2 cells. (D) Representative TUNEL staining images showing that Mdivi-1 suppressed the enhanced apoptosis in DOX-induced H9C2 cells, similarly like OSE.
FIGURE 8
FIGURE 8
The potential molecular mechanisms of OSE against DOX-induced cardiotoxicity.
See this image and copyright information in PMC

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References

    1. Ambrosone C. B., Zirpoli G. R., Hutson A. D., McCann W. E., McCann S. E., Barlow W. E., et al. (2020). Dietary Supplement Use during Chemotherapy and Survival Outcomes of Patients with Breast Cancer Enrolled in a Cooperative Group Clinical Trial (SWOG S0221). J. Clin. Oncol. 38 (8), 804–814. 10.1200/jco.19.01203 - DOI - PMC - PubMed
    1. Ashrafi G., Schwarz T. L. (2013). The Pathways of Mitophagy for Quality Control and Clearance of Mitochondria. Cell Death Differ 20 (1), 31–42. 10.1038/cdd.2012.81 - DOI - PMC - PubMed
    1. Cardinale D., Colombo A., Bacchiani G., Tedeschi I., Meroni C. A., Veglia F., et al. (2015). Early Detection of Anthracycline Cardiotoxicity and Improvement with Heart Failure Therapy. Circulation 131 (22), 1981–1988. 10.1161/circulationaha.114.013777 - DOI - PubMed
    1. Catanzaro M. P., Weiner A., Kaminaris A., Li C., Cai F., Zhao F., et al. (2019). Doxorubicin‐induced Cardiomyocyte Death Is Mediated by Unchecked Mitochondrial Fission and Mitophagy. FASEB j. 33 (10), 11096–11108. 10.1096/fj.201802663R - DOI - PMC - PubMed
    1. Chen Q.-Q., Ma G., Liu J.-F., Cai Y.-Y., Zhang J.-Y., Wei T.-T., et al. (2021). Neuraminidase 1 Is a Driver of Experimental Cardiac Hypertrophy. Eur. Heart J. 42, 3770–3782. 10.1093/eurheartj/ehab347 - DOI - PubMed