Exposure to low- vs iso-osmolar contrast agents reduces NADPH-dependent reactive oxygen species generation in a cellular model of renal injury

doi:10.1016/j.freeradbiomed.2013.11.016

Free Radical Biology and Medicine

Volume 68, March 2014, Pages 35-42

https://doi.org/10.1016/j.freeradbiomed.2013.11.016 Get rights and content

Highlights

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An in vitro model of contrast medium (CM) toxicity in renal tubular cells is proposed.
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Iso-osmolar CM iodixanol shows less cytotoxicity than low-osmolar CM iopamidol and iohexol.
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Some cytotoxic and proapoptotic effects of iodinated CMs are due to intracellular ROS generation and cell apoptosis.
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Activation of NADPH oxidase Nox4 is a key mechanism in the induction of cell toxicity and apoptosis after CM exposure.
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Nox4 inhibition is a potential pharmacological target to block direct CM toxicity in renal tubular cells.

Abstract

Contrast-induced nephropathy represents the third cause of hospital-acquired acute renal failure. This study investigated the effects of low- vs iso-osmolar contrast medium (CM) exposure on NADPH-dependent reactive oxygen species (ROS) generation by tubular cells. X-ray attenuation of iohexol, iopamidol, and iodixanol was assessed at equimolar iodine concentrations and their effects on human renal proximal tubular cells (PTCs) were evaluated with equally attenuating solutions of each CM. Cytotoxicity, apoptosis, and necrosis were investigated by trypan blue exclusion, MTT assay, and annexin V/propidium iodide assay, respectively. ROS production was assessed by DCF assay, NADPH oxidase activity by the lucigenin-enhanced chemiluminescence method, and Nox4 expression by immunoblot. Yielding the same X-ray attenuation, CM cytotoxicity was assessed in PTCs at equimolar iodine concentrations. More necrosis was present after incubation with iohexol and iopamidol than after incubation with equal concentrations of iodixanol. Iohexol and iodixanol at low iodine concentrations induced less cytotoxicity than iopamidol. Moreover, both iohexol and iopamidol induced more apoptosis than iodixanol, with a dose-dependent effect. ROS generation was significantly higher with iopamidol and iohexol compared to iodixanol. NADPH oxidase activity and Nox4 protein expression significantly increased after exposure to iopamidol and iohexol, with a dose-dependent effect, compared with iodixanol. CM-induced Nox4 expression and activity depended upon Src activation. In conclusion, at angiographic concentrations, iodixanol induces fewer cytotoxic effects on cultured tubular cells than iohexol and iopamidol along with a lower induction of Nox4-dependent ROS generation. This enzyme may, thus, represent a potential therapeutic target to prevent iodinated CM-related oxidative stress.

Section snippets

Contrast media

Three CMs were used: nonionic low-osmolar iohexol (672 mOsm/kg H₂O; Omnipaque-300, GE Healthcare, Princeton, NJ, USA), nonionic low-osmolar iopamidol (616 mOsm/kg H₂O; Iopamiro-300, Bracco Imaging, Milan, Italy), and nonionic iso-osmolar iodixanol (290 mOsm/kg H₂O; Visipaque-320, GE Healthcare, Little Chalfont, UK). All contrast agents were used in ready-to-use formulations.

Definition of equally attenuating CM concentrations

One-milliliter syringes filled with 20, 50, 125, and 250 mg iodine(mg I)/ml concentrations of iohexol, iopamidol, and

X-ray attenuation does not differ among contrast agents

We first compared the radiopacity of the three CMs selected for this study: low-osmolar CMs iohexol and iopamidol and iso-osmolar CM iodixanol. To this aim 1-ml syringes were filled with dilutions of ready-to-use 250, 125, 50, and 20 mg I/ml concentrations of each CM and X-ray attenuation was measured at 70–80 kV (data not shown). Quantitative analysis of digitally recorded images did not show statistically significant differences among X-ray attenuation of the different CMs at the concentrations

Discussion

In this study we demonstrate that the lower cytotoxic and proapoptotic effects of the iso-osmolar CM iodixanol, compared to the low-osmolar CMs iohexol and iopamidol, on human kidney cells are mediated by a differential activation of the NADPH oxidase isoenzyme Nox4, which leads to intracellular ROS generation and cell apoptosis, and that these events are mediated by CM-induced Src activation.

The pathogenesis of contrast-induced nephropathy is poorly understood. Classically the osmolality of CM

Conclusion

The pivotal role of NADPH oxidase in the upregulation of ROS in a cellular model of contrast-induced nephropathy makes this enzyme a potential target for therapeutic intervention in iodinated contrast medium-related oxidative stress.

Acknowledgments

This work was funded by MERIT Program RBNE08BNL7 and by PRIN2008 Program 2008M9WSJX_002, both from the Italian Ministry of Education and University (granted to G.S.N.) and by an unrestricted grant from GE Healthcare (granted to L.G.). C.P. is the recipient of a MERIT Program fellowship.

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In addition, in animal models, IOCM is associated with a lower induction of Nox4-dependent reactive oxygen species generation. Furthermore, IOCM also exerts fewer vasoconstriction effects than LOCM.53 Indeed, evidence suggests that AKI is more frequent when using LOCM, but may not occur when using IOCM together with an adequate prophylaxis.
Patients with cancer are subjected to several imaging examinations which frequently require the administration of contrast medium (CM). However, it has been estimated that acute kidney injury (AKI) due to the injection of iodinated CM accounts for 11% of all cases of AKI, and it is reported in up to 2% of all computed tomography (CT) examinations. Remarkably, the risks of developing AKI are increased in the elderly, in patients with chronic kidney disease or diabetes, and with dehydration or administration of nephrotoxic chemotherapeutics. Given the common occurrence of post-contrast acute kidney injury (PC-AKI) in clinical practice, primary care physicians and all specialists involved in managing patients with cancer should be aware of the strategies to reduce the risk of this event. In 2018, a panel of five experts from the specialties of radiology, oncology and nephrology were speakers at the annual meeting of the Italian Society of Medical Radiology (Società Italiana di Radiologia Medica e Interventistica), with the aim of commenting on existing evidence and providing their experience on the incidence and management of PC-AKI in patients with cancer. The discussion represented the basis for this white paper, which is intended to be a practical guide organised by statements describing methods to reduce renal injury risks related to CM-enhanced CT examinations in patients with cancer.
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It is also well recognized that high osmolar contrast media (HOCM) are more nephrotoxic than low (LOCM) or isoosmolar contrast media (IOCM) [12–14]. Trial done by Netti et al. [15] showed a significant benefit of IOCM over LOCM but other trials have failed to show this [16–19]. Current data are insufficient to justify specific recommendations about IOCM and LOCM, and specific guidelines on use of different CM especially in patients with higher stages of CKD have been lacking [20].
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A total of 2839 patients were included in 10 trials, in which 1430 patients received IOCM and 1393 received LOCM. When compared to LOCM, IOCM was not associated with significant benefit in preventing CIN (OR = 0.72, [CI: 0.50–1.04], P = 0.08, I2 = 59%). Subgroup analysis revealed non-significant difference in incidence of CIN based on baseline use of N-acetylcystine (NAC), diabetes status, ejection fraction, and whether percutaneous coronary intervention vs coronary angiography alone was performed. The difference between IOCM and LOCM was further attenuated when restricted to studies with larger sample size (> 250 patients) (OR = 0.93; [CI: 0.66–1.30]) or when compared with non-ionic LOCM (OR = 0.79, [CI: 0.52–1.21]).
In patients with CKD stage 3 and above undergoing coronary angiography, use of IOCM showed overall non-significant difference in incidence of CIN compared to LOCM. The difference was further attenuated when IOCM was compared with non-ionic LOCM.
Post-contrast acute kidney injury in cancer patients
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ESUR 10 does not refer to differences between "low osmolarity" contrast injection (577–823 mOsm / kg H 2 O) and isoosmolar (290 mOsm / kg H 2 O).6 However, the isoosmolar iodixanol CM induces less cytotoxic effects in cultured tubular cells and the production oxididated radicals is less than "low" osmolarity iohexol and iopamidol.29 In addition, in high-risk patients, the iodixanol isoosmolar dimer is associated with less nephrotoxic effects than the "low" osmolarity.30
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Original ContributionExposure to low- vs iso-osmolar contrast agents reduces NADPH-dependent reactive oxygen species generation in a cellular model of renal injury

Highlights

Abstract

Section snippets

Contrast media

Definition of equally attenuating CM concentrations

X-ray attenuation does not differ among contrast agents

Discussion

Conclusion

Acknowledgments

Am. J. Kidney Dis.

Kidney Int.

Am. J. Kidney Dis.

Kidney Int.

Biochim. Biophys. Acta

Curr. Opin. Cell Biol.

FEBS Lett.

J. Biol. Chem.

Kidney Int.

J. Biol. Chem.

Acute kidney injury, mortality, length of stay, and costs in hospitalized patients

J. Am. Soc. Nephrol.

Contrast-induced nephropathy

Am. J. Roentgenol

Contrast-induced nephropathy: a clinical and evidence-based approach

Circulation

Incidence and prognostic importance of acute renal failure after percutaneous coronary intervention

Circulation

Renal failure after percutaneous coronary intervention is associated with high mortality

Catheter Cardiovasc. Interv.

Metaanalysis of the relative nephrotoxicity of high- and low-osmolality iodinated contrast media

Radiology

Nephrotoxic effects in high-risk patients undergoing angiography

N. Engl. J. Med.

Nephropathy induced by contrast medium

N. Engl. J. Med.

Contrast-agent-induced acute renal dysfunction: is iodixanol the answer?

N. Engl. J. Med.

Contrast media-induced nephrotoxicity: questions and answers

Br. J. Radiol.

Prophylaxis of iodinated contrast media-induced nephropathy: a pharmacological point of view

Invest. Radiol.

Effects of iodinated X-ray contrast media on renal epithelial cells in culture

Invest. Radiol.

Direct toxic effect of the radiocontrast agent diatrizoate on renal proximal tubule cells

Am. J. Physiol.

Comparison of the toxicity of the radiocontrast agents, iopamidol and diatrizoate, to rabbit renal proximal tubule cells in vitro

J. Pharmacol. Exp. Ther.

Original Contribution
Exposure to low- vs iso-osmolar contrast agents reduces NADPH-dependent reactive oxygen species generation in a cellular model of renal injury