Home About us Current issue Back issues Submission Instructions Advertise Contact Login   

Search Article 
  
Advanced search 
 
Saudi Journal of Kidney Diseases and Transplantation
Users online: 446 Home Bookmark this page Print this page Email this page Small font sizeDefault font size Increase font size 
 


 
Table of Contents   
ORIGINAL ARTICLE  
Year : 2019  |  Volume : 30  |  Issue : 1  |  Page : 45-52
Study of contrast-induced oxidative stress in nondiabetic patients undergoing coronary angiography


1 Department of Biochemistry, Sri Venkateswara Institute of Medical Sciences, Tirupati, Andhra Pradesh, India
2 Department of Biostatistics, Christian Medical College, Vellore, Tamil Nadu, India
3 Department of Cardiology, Sri Venkateswara Institute of Medical Sciences, Tirupati, Andhra Pradesh, India
4 Department of Nephrology, Sri Venkateswara Institute of Medical Sciences, Tirupati, Andhra Pradesh, India

Click here for correspondence address and email

Date of Submission02-Dec-2017
Date of Acceptance30-Dec-2017
Date of Web Publication26-Feb-2019
 

   Abstract 


Administration of iodinated contrast media is associated with serious complications such as acute kidney injury (AKI). Oxidative stress is implicated as a major mechanism underlying the production of contrast-induced AKI (CI-AKI). There are very few human studies on oxidative stress occurring after contrast administration. Twenty-seven patients scheduled for coronary angiography were recruited. An average of 22.2 mL low-osmolal nonionic contrast was administered. Plasma conjugated dienes (CD), lipid hydroperoxides (LOOH), malondialdehyde (MDA), protein carbonyl (PC), protein thiols (PTs), ferric reducing ability of plasma (FRAP), erythrocyte super oxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase were estimated before, 30 min, 2 and 4 h after contrast administration. CD, LOOH, MDA, and PC increased (P <0.001), whereas PTs, FRAP, SOD, CAT (P <0.001), and GPx (P = 0.013) decreased in the first 4 h. Estimated glomerular filtration rate (eGFR) showed inverse association with MDA and positive association with GPx. The study provides evidence for oxidative stress following contrast administration even in the absence of predisposing factors. Association of eGFR with MDA and GPx indicate kidney as the source of oxidative stress. Hence, antioxidant therapy before contrast administration helps to prevent the development of oxidative stress, thereby reducing the risk of CI-AKI.

How to cite this article:
Manda P, Srinivasa Rao P V, Bitla AR, Vinapamula KS, Jeyaseelan L, Rajasekhar D, Vishnubhotla S. Study of contrast-induced oxidative stress in nondiabetic patients undergoing coronary angiography. Saudi J Kidney Dis Transpl 2019;30:45-52

How to cite this URL:
Manda P, Srinivasa Rao P V, Bitla AR, Vinapamula KS, Jeyaseelan L, Rajasekhar D, Vishnubhotla S. Study of contrast-induced oxidative stress in nondiabetic patients undergoing coronary angiography. Saudi J Kidney Dis Transpl [serial online] 2019 [cited 2019 Jul 20];30:45-52. Available from: http://www.sjkdt.org/text.asp?2019/30/1/45/252932



   Introduction Top


Iodinated contrast medium is widely used in various imaging techniques including coronary angiography. One of the serious complications of administration of iodinated contrast media is contrast-induced acute kidney injury (CI-AKI), which is also the third most common pre-renal cause of AKI. Risk factors for CI-AKI include pre-existing renal disease, hypotension, heart failure, anemia, old age, diabetes mellitus, and concomitant nephrotoxic medications.[1] Oxidative stress plays an important role in the generation of CI-AKI.[2] The basis for this concept is indirect and mainly comes from animal studies[3],[4],[5] and the preventive role of drugs having antioxidant properties.[5],[6],[7] There are very few human studies on oxidative stress following administration of contrast.[8],[9] More information on the effects of contrast on oxidative stress will help in better prevention strategies for CI-AKI. Hence, this study was taken up to understand the time course changes in the markers of oxygen radical production and markers of antioxidant defense following contrast administration.


   Materials and Methods Top


Patients scheduled to undergo coronary angiography (CAG) in the Department of Cardiology, Sri Venkateswara Institute of Medical Sciences, Tirupati, from December 2014 to March 2015 were considered for recruitment into the study. Six hundred and fifty-seven patients underwent elective coronary angiography during this period. Of these, 30 consecutive patients who met the inclusion criteria and willing to participate were recruitted for the study. Patients with normal serum creatinine levels who were found suitable and scheduled for elective CAG by the cardiologist were included in the present study. Patients with history of preexisting renal disease, diabetes mellitus, hypertension, hypotension, thyroid disorders, cardiogenic shock, those on glucocorticoid therapy, or allergic to contrast media and patients unwilling to participate were excluded from the study. The study was approved by the Institutional Ethics Committee. Written informed consent was obtained from all the patients. Patients were advised liberal oral fluids before angiography as part of patient management protocol of the department. The contrast used was low-osmolal nonionic contrast agent, Iohexol (Omnipaque 350 mg I/mL, Wipro GE Healthcare, China). Of the 30 patients included, three patients who developed CI-AKI as per the modified European Society of Urogenital Radiology guidelines[10] were not included for data analysis to maintain homogeneity of the study groups which will strengthen the conclusions of the study. The average dose of the contrast administered was of 22.2 mL (range 15–50 mL). CAG was performed by using standard techniques.

Five mL of peripheral venous blood sample was collected in nitrogen filled heparinized tubes before, 30 min, 2 and 4 h after administration of contrast. One sample for creatinine was collected at the end of 48 h for all the patients. Plasma was separated and hemolysate obtained by washing erythrocytes four times with saline before lysing with equal volume of cold distilled water. The separated plasma and hemolysate were stored at -80°C until analysis. The oxidant and antioxidant markers were assayed using the following methods:

Conjugated dienes (CD) with alternating double and single bonds between carbon atoms absorb wavelengths of 230–235 nm in the UV region and the value is expressed as optical density.[11] Lipid hydroperoxides (LOOH) which are products of fatty acid oxidation abstract hydrogen from a methylene group, forming a carbon-centered radical that reacts with molecular oxygen to form lipid peroxide that is detected spectrophotometrically using Fox reagent at 560 nm.[12] Malondialdehyde (MDA), which is another indicator of lipid peroxidation, was measured spectrophoto-metrically as thiobarbituric acid reactive substances (TBARS) after precipitating the proteins with trichloroacetic acid (TCA).[13] Protein carbonyl (PC) content was measured based on the method described by Levine et al, in which 2, 4-dinitro phenyl hydrazine reacts with PCs forming a Schiff base to produce the corresponding hydrazone that is analyzed spectrophotometrically at 370 nm.[14] Assay for thiols was performed using Ellman's reagent or 5, 5'-dithiobis (2-nitrobenzoic acid) (DTNB). The procedure is based on the reaction of thiols with DTNB to give a mixed disulfide and 2-nitro-5-thiobenzoic acid (TNB) which is quantified by the absorbance of the anion (TNB) at 412 nm.[15] Ferric reducing ability of plasma (FRAP) assay was performed as per the method described by Benzie and Strain. The antioxidant power reduces ferric ions to ferrous ions at low pH resulting in the formation of a colored ferrous-tripyridyltriazine complex. FRAP values are obtained by comparing the change in absorbance of test reaction mixtures at 593 nm with ferrous standards.[16] Super oxide dismutase (SOD) activity measurement is based on the ability of SOD to inhibit the oxidation of adrenaline to form a pink-colored adrenochrome. SOD activity was determined by monitoring the rate of adrenochrome formation spectrophotometrically at 470 nm. One unit of SOD was defined as the amount of enzyme required to cause 50% inhibition of adrenaline oxidation.[17] Catalase (CAT) activity was measured by continuous spectrophotometric rate determination. One unit of CAT will decompose 1.0 μM of H2O2 per min at pH 7. The rate of disappearance of H2O2 is observed as rate of decrease in absorbance at 240 nm.[18] Glutathione peroxidase (GPx) activity was estimated by an enzymatic reaction initiated by the addition of terbutyl hydroperoxide (TBHP) to a reaction mixture containing reduced glutathione (GSH), reduced nicotinamide adenine dinucleotide (NADPH) and glutathione reductase (GR). The change in absorbance was monitored spectrophotometrically at 340 nm.[19] Activities of holo and apo forms of glutathione reductase (GR) in hemolysate are measured with and without addition of FAD and by spectrophotometric determination of NADP formed.[20] All the analyses were performed using Lambda 25 UV-visual double beam spectrophotometer (Perkin Elmer, Singapore). Within-run imprecision for the parameters ranged from 0.9% to 3.9%. Creatinine was estimated by standard methods using comercial kits on CX5 autoanalyser (Synchron Cx5 Beckman Coulter, Inc., Galway, Ireland). The estimated glomerular filtration rate (eGFR) was calculated using Cockcroft Gault formula.[21]


   Statistical Analysis Top


Data were expressed as mean ± standard deviation. Data were converted to percentage of baseline value before performing analysis of variance (ANOVA) for repeated measures with bonferroni post hoc testing. Generalized estimating equations were used to study the association between repeated measures variables. Analysis of the data was performed using Microsoft Excel 2007 for Windows (Microsoft Corporation, Redmond, WA, USA), Statistical Package for the Social Sciences (SPSS) version 16.0 (SPSS Inc., Chicago, IL, USA). A value of P <0.05 was considered as significant.


   Results Top


The demographic and base line characteristics of patients are presented in [Table 1]. The changes in the oxidant markers observed are presented in [Table 2] and [Figure 1]. CD, LOOH, MDA and PC showed a significant increase (P < 0.001) in the first 4 h indicating production of reactive oxygen species (ROS) due to the administration of the contrast. There was a significant decrease in thiols, FRAP, SOD, CAT (P <0.001) and GPx (P = 0.013) demonstrating a decrease in antioxidant defense [Table 3] and [Figure 2]. eGFR was significantly lower in cases and showed significant inverse association with MDA (B = -2.398, P = 0.015) and positive association with GPx (P = 0.814, P<0.001) [Figure 3].
Table 1: Demographic and baseline characteristics of the patients (n=27).

Click here to view
Table 2: Time course changes in the oxidant markers studied.

Click here to view
Figure 1: Percentage time course changes in the oxidant markers studied.
A: ANOVA for repeated measures (0–4 h), B: 0 h versus 30 min, C: 0 h versus 2 h, D: 0 h versus 4 h, E: 30 min versus 2 h, F: 30 min versus 4 h, G: 2 h versus 4 h. ANOVA was performed after transforming the data to percentages of baseline (0 h) value to include only the changes observed during the study period in the analysis.


Click here to view
Table 3: Time course changes in the antioxidant parameters studied.

Click here to view
Figure 2: Percentage time course changes in the antioxidant parameters studied.
A: ANOVA for repeated measures (0–4 h), B: 0 h versus 30 min, C: 0 h versus 2 h, D: 0 h versus 4 h, E: 30 min versus 2 h, F: 30 min versus 4 h, G: 2 h versus 4 h. ANOVA was performed after transforming the data to percentages of baseline (0 h) value to include only the changes observed during the study period in the analysis.


Click here to view
Figure 3: Regression curve analysis of eGFR with MDA (A) and GPx (B).
eGFR: estimated glomerular filtration rate, MDA: Malondialdehyde, GPx: Glutathione peroxidase eGFR showed inverse association with MDA and positive association with GPx.


Click here to view



   Discussion Top


Oxidative stress has been implicated in CI-AKI.[1],[2] AKI is one of the important risk factors for progression to chronic kidney disease and hence needs to be identified and managed early. Experimental findings showed that administration of iodinated contrast induces renal hypoxia which enhances ROS production, mainly in the medullary thick ascending limb that has dense mitochondrial population. The protective effect of ROS scavenging drugs[6],[7],[8] supports this. As the patients undergoing coronary angiography will be having CAD, the baseline oxidative stress that may be present due to CAD is likely to confound the conclusions drawn on oxidative stress that occurs after the administration of contrast. To avoid this, the data were converted into percentages with baseline as 100% before inclusion into statistical analysis. CD and LOOH, followed by MDA are the products of action of free oxygen radicals on lipids. In the present study, plasma levels of CD and LOOH showed significant increase during the first 4 h after the administration of contrast (P <0.001). The levels increased in the first 2 h followed by a decline but were still higher than the baseline value at the end of 4 h [Table 2]. These findings provide evidence for production of ROS immediately after the contrast administration. There are no studies reporting on changes in CD, which is the earliest of the lipid peroxidation products, after contrast administration. Saitoh et al[8] reported increased urinary LOOH at 2 h after CAG, which was completely abolished with administration of glutathione prior to the procedure. This study also supports our findings. Animal studies[3] reported increased kidney tissue MDA levels after administration of contrast. In the present study, plasma MDA increased more than threefold during the first 4 h after contrast administration. PCs, the product of action of free oxygen radicals on proteins, showed 74% increase (P <0.001). Similar findings are reported by others for MDA[9] and PCs.[5]

As the increase in these initial lipid peroxidation products was observed at 30 min, we collected blood samples at 0 and 10, 15 and 20 min for another five patients inquisitive to know how early ROS production starts. We could find a significant increase in CD at 15 min (0.35 ± 0.02 vs. 0.36 ± 0.02; 2.6 % increase; P = 0.002) and in LOOH at 20 min (3.12 ± 1.7 vs. 4.18 ± 2.13; 9.5% increase; P = 0.028), whereas MDA levels showed no change (1.06 ± 0.4 vs. 1.06 ± 0.4; P = 0.228) when compared to baseline levels. Thus our findings of the present study provide strong evidence for ROS production due to contrast administration.

Plasma FRAP levels which reflect the overall antioxidant capacity decreased reaching 59.8% of the base line by the 4th h (P <0.001). Earlier studies reported a decrease[22] or no change[23] in FRAP levels after contrast administration. However, these were either animal studies or studies on the effect of antioxidants in patients undergoing interventions using contrast media. Protein thiol (PT), another marker of anti-oxidant defense, decreased reaching 65% of the baseline levels (P <0.001) at the end of 4 h after contrast administration. This finding is in line with that of Deng et al,[5] who reported a decrease in PTs after contrast administration that is prevented by prior administration of antioxidants.

There was a continuous fall in the activity of erythrocyte SOD resulting in 7% of the baseline levels (P <0.001) at the end of 4 h after contrast administration. Studies conducted on rats,[3],[4] and in humans[22] have shown a similar decrease in the SOD activity. However, Akgüllü et al[7] did not find change in SOD levels. Erythrocyte CAT activity and GPx activity also decreased during the study period after contrast administration resulting in 19% and 5% of the baseline levels respectively (P <0.001 and P <0.013 for CAT and GPx, respectively) at the end of 4 h after contrast administration. Earlier studies have also reported a decrease in CAT[3],[4] and GPx[3],[6] activity after contrast administration. There are no studies on GR in CI renal injury. In the present study also, erythrocyte GR did not alter significantly after contrast administration.

The degree of renal injury following contrast administration is dependent on baseline renal function, presence of risk factors such as diabetes, volume, and type of contrast used. All our patients had normal baseline renal function, were nondiabetic and received low-osmolal, nonionic contrast medium in low volumes (15–50 mL). Finding of oxidative stress even in these minimal renal injury conditions without any additional risk factors provide a strong evidence for oxidative stress as the main mechanism of renal injury due to contrast administration. Further supporting this concept, we observed significant negative association between eGFR and MDA (B = -2.398, P = 0.015) and positive association with GPx (B = 0.814, P <0.001) indicating that the source of oxidative stress is kidney. Findings of the present study indicate that contrast administration results in oxidative stress which is likely to contribute to the production of AKI. Our findings strengthen the concept of administration of antioxidants before contrast administration to reduce the iatrogenic renal injury and the associated morbidity and mortality.

Acknowledgment

We would like to thank Sri Venkateswara Institute of Medical Sciences and Tirumala Tirupati Devasathanams, Tirupati for providing financial support under Sri Balaji Aarogya Vara Prasadini Scheme (GRANT NO: SBAVP-RG/MD/22).

Conflicts of interest:

None declared.



 
   References Top

1.
Thomson VS, Narayanan K, Singh JC. Contrast induced nephropathy in urology. Indian J Urol 2009;25:437-45.  Back to cited text no. 1
[PUBMED]  [Full text]  
2.
Seeliger E, Sendeski M, Rihal CS, Persson PB. Contrast-induced kidney injury: Mechanisms, risk factors, and prevention. Eur Heart J 2012;33:2007-15.  Back to cited text no. 2
    
3.
Devrim E, Cetin M, Namuslu M, Ergüder IB, Cetin R, Durak I. Oxidant stress due to non ionic low osmolar contrast medium in rat kidney. Indian J Med Res 2009;130:433-6.  Back to cited text no. 3
[PUBMED]  [Full text]  
4.
Tasanarong A, Kongkham S, Itharat A. Antioxidant effect of Phyllanthus emblica extract prevents contrast-induced acute kidney injury. BMC Complement Altern Med 2014; 14:138.  Back to cited text no. 4
    
5.
Deng J, Wu G, Yang C, Li Y, Jing Q, Han Y. Rosuvastatin attenuates contrast-induced nephropathy through modulation of nitric oxide, inflammatory responses, oxidative stress and apoptosis in diabetic male rats. J Transl Med 2015;13:53.  Back to cited text no. 5
    
6.
Buyuklu M, Kandemir FM, Ozkaraca M, Set T, Bakirci EM, Topal E. Protective effect of curcumin against contrast induced nephropathy in rat kidney: What is happening to oxidative stress, inflammation, autophagy and apoptosis? Eur Rev Med Pharmacol Sci 2014;18:461-70.  Back to cited text no. 6
    
7.
Akgüllü Ç, Hekim T, Eryılmaz U, Boyacıoğlu M, Güngör H, Meteoğlu İ. The usefulness of carvedilol and nebivolol in preventing contrast nephropathy in rats. Ren Fail 2015;37:511-7.  Back to cited text no. 7
    
8.
Saitoh T, Satoh H, Nobuhara M, et al. Intravenous glutathione prevents renal oxidative stress after coronary angiography more effectively than oral N-acetylcysteine. Heart Vessels 2011;26:465-72.  Back to cited text no. 8
    
9.
Yılmaz M, Aydınalp A, Okyay K,. Comparison of carvedilol and metoprolol for preventing contrast-induced nephropathy after coronary angiography. Cardiorenal Med 2015;5:199-207.  Back to cited text no. 9
    
10.
Waikar SS, Bonventre JV. Creatinine kinetics and the definition of acute kidney injury. J Am Soc Nephrol 2009;20:672-9.  Back to cited text no. 10
    
11.
Buege JA, Aust SD. Microsomal lipid peroxidation. Methods Enzymol 1978;52:302-10.  Back to cited text no. 11
    
12.
Jiang ZY, Woollard AC, Wolff SP. Lipid hydroperoxide measurement by oxidation of fe2+ in the presence of xylenol orange. Comparison with the TBA assay and an iodometric method. Lipids 1991;26:853-6.  Back to cited text no. 12
    
13.
Sangeetha P, Das UN, Koratkar R, Suryaprabha P. Increase in free radical generation and lipid peroxidation following chemotherapy in patients with cancer. Free Radic Biol Med 1990;8:15-9.  Back to cited text no. 13
    
14.
Levine RL, Garland D, Oliver CN, et al. Determination of carbonyl content in oxidatively modified proteins. Methods Enzymol 1990;186:464-78.  Back to cited text no. 14
    
15.
Motchnik PA, Frei B, Ames BN. Measurement of antioxidants in human blood plasma. Methods Enzymol 1994;234:269-79.  Back to cited text no. 15
    
16.
Benzie IF, Strain JJ. The ferric reducing ability of plasma (FRAP) as a measure of “antioxidant power”: The FRAP assay. Anal Biochem 1996;239:70-6.  Back to cited text no. 16
    
17.
Winterbourn CC, Hawkins RE, Brian M, Carrell RW. The estimation of red cell superoxide dismutase activity. J Lab Clin Med 1975;85:337-41.  Back to cited text no. 17
    
18.
Aebi H. Catalase in vitro. Methods Enzymol 1984;105:121-6.  Back to cited text no. 18
    
19.
Wendel A. Glutathione peroxidase. Methods Enzymol 1981;77:325-33.  Back to cited text no. 19
    
20.
McCormicfc DB, Greene HL. Vitamins. In: Burtis CA, Ashwood ER, editors. Tietz Textbook of Clinical Chemistry. 3rd ed. Philadelphia: W.B. Saunders Company; 1999. p. 1015-16.  Back to cited text no. 20
    
21.
Botev R, Mallié JP, Couchoud C, et al. Estimating glomerular filtration rate: Cockcroftgault and modification of diet in renal disease formulas compared to renal inulin clearance. Clin J Am Soc Nephrol 2009;4:899-906.  Back to cited text no. 21
    
22.
Kongkham S, Sriwong S, Tasanarong A. Protective effect of alpha tocopherol on contrast-induced nephropathy in rats. Nefrologia 2013;33:116-23.  Back to cited text no. 22
    
23.
Onbasili AO, Yeniceriglu Y, Agaoglu P, et al. Trimetazidine in the prevention of contrast-induced nephropathy after coronary procedures. Heart 2007;93:698-702.  Back to cited text no. 23
    

Top
Correspondence Address:
P V. L. N Srinivasa Rao
Department of Biochemistry, Sri Venkateswara Institute of Medical Sciences, Tirupati - 517 507, Andhra Pradesh
India
Login to access the Email id


DOI: 10.4103/1319-2442.252932

PMID: 30804266

Rights and Permissions


    Figures

  [Figure 1], [Figure 2], [Figure 3]
 
 
    Tables

  [Table 1], [Table 2], [Table 3]



 

Top
   
 
 
    Similar in PUBMED
    Search Pubmed for
    Search in Google Scholar for
    Email Alert *
    Add to My List *
* Registration required (free)  
 


 
    Abstract
   Introduction
    Materials and Me...
   Statistical Analysis
   Results
   Discussion
    References
    Article Figures
    Article Tables
 

 Article Access Statistics
    Viewed306    
    Printed7    
    Emailed0    
    PDF Downloaded43    
    Comments [Add]    

Recommend this journal