Anat Cell Biol.  2018 Sep;51(3):189-199. 10.5115/acb.2018.51.3.189.

Exogenous spermidine ameliorates tubular necrosis during cisplatin nephrotoxicity

• Yoon SP
• Kim J

• KMID: 2447012
• DOI: http://doi.org/10.5115/acb.2018.51.3.189

Abstract

The hallmark of cisplatin-induced acute kidney injury is the necrotic cell death in the kidney proximal tubules. However, an effective approach to limit cisplatin nephrotoxicity remains unknown. Spermidine is a polyamine that protects against oxidative stress and necrosis in aged yeasts, and the present study found that exogenous spermidine markedly attenuated tubular necrosis and kidney dysfunction, but not apoptosis, during cisplatin nephrotoxicity. In addition, exogenous spermidine potently inhibited oxidative/nitrative DNA damage, poly(ADP-ribose) polymerase 1 (PARP1) activation and ATP depletion after cisplatin injection. Conversely, inhibition of ornithine decarboxylase (ODC) via siRNA transfection in vivo significantly increased DNA damage, PARP1 activation and ATP depletion, resulting in acceleration of tubular necrosis and kidney dysfunction. Finally, exogenous spermidine removed severe cisplatin injury induced by ODC inhibition. In conclusion, these data suggest that spermidine protects kidneys against cisplatin injury through DNA damage and tubular necrosis, and this finding provides a novel target to prevent acute kidney injury including nephrotoxicity.

Keyword

Cisplatin; Nephrotoxicity; Spermidine; Lipid peroxidation; Necrosis; Ornithine decarboxylase

MeSH Terms

Acceleration
Acute Kidney Injury
Adenosine Triphosphate
Apoptosis
Cell Death
Cisplatin*
DNA Damage
Kidney
Lipid Peroxidation
Necrosis*
Ornithine Decarboxylase
Oxidative Stress
Poly(ADP-ribose) Polymerases
RNA, Small Interfering
Spermidine*
Transfection
Yeasts
Adenosine Triphosphate
Cisplatin
Ornithine Decarboxylase
Poly(ADP-ribose) Polymerases
RNA, Small Interfering
Spermidine

Figure

• 

  Fig. 1 Exogenous spermidine reduces kidney dysfunction and tubular damage during cisplatin nephrotoxicity. The kidneys in C57BL/6 male mice were harvested 1, 2, or 3 days after cisplatin injection or non-injection (control). The mice were orally supplemented with either spermidine or vehicle twice daily from 24 hours before cisplatin injection up to the time that they were sacrificed (n=5 mice in each group). (A) The spermidine level in kidneys of mice treated with spermidine (10 mg/kg body weight) or vehicle 3 days after cisplatin injection. (B) The level of plasma spermidine in mice treated with spermidine (10 mg/kg body weight) or vehicle 3 days after cisplatin injection. (C) Periodic acid-Schiff (PAS) staining on kidney cortical sections of mice treated with spermidine (10 mg/kg body weight) or vehicle 3 days after cisplatin injection. Scale bars=50 µm. (D) Tubular injury score represented by PAS staining on kidney sections of mice treated with spermidine (1 to 10 mg/kg body weight) or vehicle 3 days after cisplatin injection. (E) Tubular injury score after cisplatin injection in mice treated with spermidine (10 mg/kg body weight) or vehicle 1, 2, and 3 days after cisplatin injection. (F) Plasma creatinine concentration at 3 days after cisplatin injection in mice treated with spermidine (1 to 10 mg/kg body weight) or vehicle. (G) Plasma creatinine concentration at 1, 2, and 3 days after cisplatin injection in mice treated with spermidine (10 mg/kg body weight) or vehicle. *P<0.05 vs. control, #P<0.05 vs. vehicle.

• 

  Fig. 2 Exogenous spermidine does not alter cleaved caspase-3 expression during cisplatin nephrotoxicity. The kidneys in C57BL/6 male mice were harvested 1, 2, or 3 days after cisplatin injection or non-injection (control). The mice were orally supplemented with either spermidine (10 mg/kg body weight) or vehicle twice daily from 24 hours before cisplatin injection up to the time that they were sacrificed (n=6 mice in each group). The expression of cleaved caspase-3 protein (A) was confirmed using a western blot analysis with anti-caspase-3 antibody in kidneys after cisplatin injection. Antibodies to β-actin were used as a loading control. The intensities of these protein bands (B) were quantified using the AzureSpot software (Azure Biosystems, Dublin, CA, USA). *P<0.05 vs. control.

• 

  Fig. 3 Exogenous spermidine diminishes oxidative and nitrosative stress during cisplatin nephrotoxicity. The kidneys in C57BL/6 male mice were harvested 1, 2, or 3 days after cisplatin injection or non-injection (control). The mice were orally supplemented with either spermidine (10 mg/kg body weight) or vehicle twice daily from 24 hours before cisplatin injection up to the time that they were sacrificed (n=5 mice in each group). (A) Lipid peroxidation represented by a concentration of lipid hydroperoxide in kidneys. (B) 8-OHdG concentration in kidneys. (C) 8-Nitroguanine concentration in kidneys. *P<0.05 vs. control, #P<0.05 vs. vehicle.

• 

  Fig. 4 Exogenous spermidine attenuates poly(ADP-ribose) polymerase 1 (PARP1) activation and ATP depletion during cisplatin nephrotoxicity. The kidneys in C57BL/6 male mice were harvested 1, 2, or 3 days after cisplatin injection or non-injection (control). The mice were orally supplemented with either spermidine (10 mg/kg body weight) or vehicle twice daily from 24 hours before cisplatin injection up to the time that they were sacrificed (n=5 mice in each group). (A) PARP1 activity in kidneys. (B) ATP concentration in kidneys. *P<0.05 vs. control, #P<0.05 vs. vehicle.

• 

  Fig. 5 Ornithine decarboxylase (ODC) siRNA exacerbates DNA oxidative/nitrative stresses, poly(ADP-ribose) polymerase 1 (PARP1) activation and ATP depletion during cisplatin nephrotoxicity. The kidneys in C57BL/6 male mice were harvested 1, 2, or 3 days after cisplatin injection or non-injection (control). The mice were orally supplemented with either spermidine (10 mg/kg body weight) or vehicle twice daily from 24 hours before cisplatin injection up to the time that they were sacrificed. ODC siRNA (siODC) or control siRNA (siControl) was injected at 48 and 24 hours before cisplatin injection (n=5 mice in each group). (A) The expression of ODC protein (left) was confirmed using a western blot analysis with anti-ODC antibody in kidneys after cisplatin injection. Antibodies to β-actin were used as a loading control. The intensities of these protein bands (right) were quantified using the AzureSpot software (Azure Biosystems, Dublin, CA, USA). (B) ODC expression in kidneys of control mice transfected with siODC or siControl, as represented by western blot. Antibodies to β-actin were used as loading control. (C) 8-OHdG concentration in kidneys. (D) 8-Nitroguanine concentration in kidneys. (E) PARP1 activity in kidneys. (F) ATP concentration in kidneys. *P<0.05 vs. control, #P<0.05 vs. vehicle, †P<0.05 vs. siControl.

• 

  Fig. 6 Ornithine decarboxylase (ODC) siRNA worsens tubular necrosis and kidney dysfunction during cisplatin nephrotoxicity. The kidneys in C57BL/6 male mice were harvested 1, 2, or 3 days after cisplatin injection or non-injection (control). The mice were orally supplemented with either spermidine (10 mg/kg body weight) or vehicle twice daily from 24 hours before cisplatin injection up to the time that they were sacrificed. ODC siRNA (siODC) or control siRNA (siControl) was injected at 48 and 24 hours before cisplatin injection (n=5 mice in each group). (A) Periodic acid-Schiff stain on kidney sections 3 days after cisplatin injection. Scale bars=50 µm. (B) Tubular necrosis score. (C) Plasma creatinine concentration. *P<0.05 vs. control, #P<0.05 vs. vehicle, †P<0.05 vs. siControl.

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