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Analysis of human urine reveals metabolic changes related to the development of acute kidney injury following cardiac surgery

Helena U. Zacharias1*, Gunnar Schley2*, Jochen Hochrein1, Matthias S. Klein1, Carmen Köberle2, Kai-Uwe Eckardt2, Carsten Willam2, Peter J. Oefner1 and Wolfram Gronwald1

1Institute of Functional Genomics, University of Regensburg, Josef-Engert-Str. 9, 93053 Regensburg, Germany

2Department of Nephrology and Hypertension, Friedrich-Alexander-University Erlangen-Nuremberg, Ulmenweg 18, 91054Erlangen, Germany

*Both authors contributed equally to this work

Supplemental Table S1: Clinical characteristics and outcome of all patients enrolled.

Non-AKI / AKI / P-value
Number of patients / 72 / 34 / -
Age [years] / 68 ± 10 / 73 ± 8 / 0.011a
Sex, male / 58 (80.6%) / 23 (67.6%) / 0.131a
BMI [kg/m2] / 28 ± 4 / 29 ± 5 / 0.481a
Serum creatinine [mg/dL] / 1.16 ± 0.41 / 1.36 ± 0.53 / 0.057a
eGFR 13-60 mL/min per 1.73 m2 / 27 / 23 / 0.006b
eGFR > 60 mL/min per 1.73 m2 / 45 / 11 / 0.006b
Comorbid disease, n [%]
Diabetes mellitus
Hypertension
Cardiac insufficiency
Hypercholesterolemia
Acute myocardial infarction
COPD
PAVD
Cardiovascular disease
CKD
Valvular heart disease
AKI in the past
RRT in the past
IABP pre-op / 26 (36.1%)
70 (97.2%)
57 (79.2%)
67 (93.1%)
25 (34.7%)
12 (16.7%)
10 (13.9%)
24 (33.3%)
17 (23.6%)
36 (50.0%)
2 (2.8%)
2 (2.8%)
1 (1.4%) / 18 (52.9%)
34 (100%)
30 (88.2%)
30 (88.2%)
15 (44.1%)
7 (20.6%)
6 (17.7%)
15 (44.1%)
22 (64.7%)
25 (73.5%)
3 (8.8%)
0 (0%)
2 (5.9%) / 0.139b
1.000b
0.293b
0.464b
0.395b
0.601b
0.772b
0.291b
0.00008b
0.034b
0.325b
1.000b
0.240b
Preoperative medication, n [%]
Statin
ACE inhibitor
Beta-blocker
Other antihypertensive drugs
Insulin
Oral anti-diabetic medication
NSAID
Diuretic / 53 (73.6%)
56 (77.8%)
53 (73.6%)
12 (16.7%)
9 (12.5%)
15 (20.8%)
1 (1.4%)
43 (59.7%) / 25 (73.5%)
27 (79.4%)
27 (79.4%)
11 (32.4%)
7 (20.6%)
11 (32.4%)
0 (0%)
23 (67.6%) / 1.000b
1.000b
0.631b
0.081b
0.383b
0.230b
1.000b
0.522b
Type of surgery
CABG
Aortic valve surgery
Mitral valve surgery
CABG + aortic valve surgery
CABG + mitral valve surgery
Thoracic aortic surgery / 54 (75%)
6 (8.3%)
2 (2.8%)
5 (6.9%)
1 (1.4%)
4 (5.6%) / 16 (47.1%)
7 (20.6%)
2 (5.9%)
7 (20.6%)
1 (2.9%)
1 (2.9%) / 0.008b
0.110b
0.592b
0.051b
0.541b
1.000b

aP-values were calculated using a two-sided t-test assuming unequal variance. bP-values were calculated using Fisher’s exact test. Abbreviations: ACE, angiotensin-converting-enzyme; AKI, acute kidney injury; BMI, body mass index; CABG, coronary artery bypass grafting; CKD, chronic kidney disease; COPD, chronic obstructive pulmonary disease; eGFR, estimated glomerular filtration rate; IABP, intra-aortic balloon pump; NSAID, non-steroidal anti-inflammatory drug; PAVD, peripheral arterial vascular disease; RRT, renal replacement therapy.

Supplemental Table S2: Classification performance on randomly permuted data. Given are the total accuracy, area under the ROC curve (AUC), number of selected features, optimal cost and gamma parameters, sensitivity and specificity. Given are the mean values and corresponding standard deviations averaged over 10 nested cross-validation runs with randomly permuted data.

Total accuracy [%] / AUC / Number of selected features / Optimized cost parameter / Optimized gamma parameter / Sensitivity / Specificity
59.4±0.8 / 0.51±0.05 / 1.1±0.3 / 1.0±0.0 / 0.125±0.0 / 0.2±0.7 / 98.9±1.3

Supplemental Table S3: Spectral positions and P-values of features that discriminated AKI from non-AKI patients. The 106 specimens studied were collected 24 hours post-operatively. A) Identified compounds. The first three features were used by the radial SVM for classification. B) Non-identified compounds. Additionally, the corresponding multiplet structures are listed.

A false discovery rate (FDR) below 5% was applied. The FDR was adjusted according to the method of Benjamini and Hochberg (BH). In case more than one compound contributed to one significant bin, all possibly corresponding names are annotated.

A

Spectral
position [ppm] / P-value
unadjusted / P-value
BH adjusted / Metabolite
2.475 / 9.67e-11 / 5.04e-08 / Carnitine
2.465 / 1.44e-10 / 5.04e-08 / 2-Oxoglutaricacid/carnitine
1.395 / 3.93e-08 / 9.19e-06 / Tranexamic acid
2.485 / 8.22e-08 / 1.15e-05 / Acetyl-L-carnitine
2.245 / 4.52e-07 / 5.28e-05 / Acetone
2.225 / 2.96e-06 / 2.96e-04 / Acetone
7.675 / 5.39e-06 / 4.36e-04 / Pseudouridine
3.045 / 6.22e-06 / 4.36e-04 / Creatinine
7.435 / 7.24e-06 / 4.62e-04 / Phenylacetylglutamine
7.615 / 1.21e-05 / 6.50e-04 / Unknown/hippuricacid
3.295 / 1.66e-05 / 8.30e-04 / Myo-inositol
2.365 / 2.51e-05 / 1.10e-03 / 3-Hydroxyisovalericacid
1.385 / 3.61e-05 / 1.36e-03 / Tranexamic acid
1.655 / 4.45e-05 / 1.53e-03 / Tranexamic acid
7.425 / 4.60e-05 / 1.53e-03 / Phenylacetylglutamine
7.625 / 5.51e-05 / 1.76e-03 / Hippuricacid
1.845 / 6.33e-05 / 1.87e-03 / Tranexamic acid
1.045 / 7.22e-05 / 2.03e-03 / Tranexamic acid
3.205 / 8.36e-05 / 2.25e-03 / Acetyl-L-carnitine
1.025 / 9.32e-05 / 2.42e-03 / Tranexamic acid
1.825 / 1.26e-04 / 3.16e-03 / Tranexamic acid
1.945 / 1.39e-04 / 3.37e-03 / Tranexamic acid, N-acetyl-L-glutamine
1.065 / 1.75e-04 / 4.08e-03 / Tranexamic acid
1.055 / 1.85e-04 / 4.19e-03 / Tranexamic acid
4.065 / 2.70e-04 / 5.90e-03 / Creatinine
1.275 / 2.86e-04 / 6.08e-03 / 3-Hydroxyisovalericacid
7.395 / 3.71e-04 / 7.64e-03 / Phenylalanine, Unknown
2.015 / 4.20e-04 / 8.41e-03 / N-Acetyl-L-groups
3.195 / 6.81e-04 / 1.19e-02 / Acetyl-L-carnitine
2.025 / 7.43e-04 / 1.24e-02 / N-Acetyl-groups
1.635 / 7.86e-04 / 1.28e-02 / Tranexamic acid
2.035 / 9.29e-04 / 1.48e-02 / N-Acetyl-groups
1.375 / 1.10e-03 / 1.71e-02 / Tranexamic acid
1.405 / 1.18e-03 / 1.80e-02 / Tranexamic acid
2.455 / 1.32e-03 / 1.88e-02 / Carnitine, 2-oxoglutaricacid, glutamine
1.955 / 1.34e-03 / 1.88e-02 / Tranexamic acid, N-acetyl-L-glutamine
1.835 / 1.40e-03 / 1.92e-02 / Tranexamic acid
2.385 / 1.49e-03 / 2.01e-02 / L-Pyroglutamicacid
3.235 / 1.55e-03 / 2.05e-02 / Carnitine
2.635 / 1.72e-03 / 2.23e-02 / Acetyl-L-carnitine
1.075 / 1.77e-03 / 2.25e-02 / Tranexamic acid
7.385 / 1.80e-03 / 2.26e-02 / Phenylacetylglutamine
6.515 / 1.95e-03 / 2.35e-02 / Noise
2.625 / 2.18e-03 / 2.58e-02 / Acetyl-L-carnitine
2.165 / 2.21e-03 / 2.58e-02 / Paracetamol-glucuronide
1.645 / 2.29e-03 / 2.63e-02 / Tranexamic acid
1.665 / 2.38e-03 / 2.66e-02 / Tranexamic acid
2.045 / 2.45e-03 / 2.66e-02 / N-Acetyl-groups
1.925 / 2.46e-03 / 2.66e-02 / Tranexamic acid, N-acetyl-L-glutamine
1.355 / 3.27e-03 / 3.32e-02 / Tranexamic acid, 2-hydroxyisobutyricacid
2.125 / 3.69e-03 / 3.64e-02 / Tranexamic acid, N-acetyl-L-glutamine
7.635 / 4.03e-03 / 3.91e-02 / Unknown, hippuricacid
1.935 / 4.07e-03 / 3.91e-02 / Tranexamic acid, N-acetyl-L-glutamine
7.415 / 4.22e-03 / 3.98e-02 / Phenylacetylglutamine
7.155 / 4.75e-03 / 4.38e-02 / Paracetamol-glucuronide
8.195 / 5.25e-03 / 4.71e-02 / Hypoxanthine
7.135 / 5.43e-03 / 4.82e-02 / Paracetamol-glucuronide
3.265 / 5.52e-03 / 4.84e-02 / Betaine, taurine, trimethylamine-N-oxide

B

Spectral
position [ppm] / Multiplet structure2 / P-value
unadjusted / P-value
BH adjusted / Metabolite
8.325 / s / 5.40e-08 / 9.46e-06 / Unknown1
2.355 / nr / 5.72e-06 / 4.36e-04 / Overlap of several compounds
2.195 / s / 8.63e-06 / 5.04e-04 / Unknown
4.415 / s / 1.83e-05 / 8.57e-04 / Unknown
2.615 / s / 3.66e-05 / 1.36e-03 / Unknown
7.605 / nr / 3.67e-05 / 1.36e-03 / Overlap of several compounds
2.495 / mp / 6.39e-05 / 1.87e-03 / Unknown
3.135 / s / 4.76e-04 / 9.26e-03 / Unknown
8.045 / nr / 5.72e-04 / 1.07e-02 / Overlap of several compounds
2.345 / nr / 5.81e-04 / 1.07e-02 / Overlap of several compounds
3.605 / nr / 6.18e-04 / 1.11e-02 / Overlap of several compounds
4.015 / nr / 6.99e-04 / 1.20e-02 / Overlap of several compounds
8.085 / mp / 1.23e-03 / 1.84e-02 / Unknown
8.095 / mp / 1.30e-03 / 1.88e-02 / Unknown
3.595 / nr / 1.86e-03 / 2.28e-02 / Overlap of several compounds
2.605 / nr / 2.46e-03 / 2.66e-02 / Overlap of several compounds
3.585 / nr / 2.51e-03 / 2.67e-02 / Overlap of several compounds
8.075 / mp / 2.60e-03 / 2.72e-02 / Unknown
7.185 / nr / 3.16e-03 / 3.25e-02 / Overlap of several compounds
2.305 / s / 3.50e-03 / 3.50e-02 / Unknown
7.595 / mp / 4.26e-03 / 3.98e-02 / Unknown
7.735 / nr / 4.83e-03 / 4.39e-02 / Overlap of several compounds

1Unkown compound of probably endogenous origin

2s, singlet; mp, complex multiplet; nr, not resolved (due to overlap)

1