Supplementary Table 1 | Findings indicative of iron-related kidney injury in haem-related diseases
Disease or insult
Findings
Human studies
Red blood cell aplasia
↑Urinary haemosiderin (method not specified); ↑CMJ iron (MRI); not haemolysing1
Chronic haemolytic
anaemia
Urinary haemosiderin ↑ (Prussian reaction of Perls); kidney iron ↑ (Turnbull iron reaction); hepatic iron↔1,2
Antibody haemolytic
anaemia
Kidney iron↑ (Prussian blue staining); histological kidney damage3
HO1 deficiency
Proximal tubule iron ↑ (Prussian blue staining); histological kidney damage; HO1 caused haemolytic anaemia4
HIV–HUS
Urinary iron↑ (ferrozine staining); glomerular and tubular damage; transferrin, haemopexin, haptoglobin, lactotransferrin, NGAL↑5
Thalassemia major,
mechanical haemolysis, pernicious anaemia
Urinary iron ↑ (benzidine method of Crosby and Furth and reaction with dipyridyl); pernicious
anaemia is a type of megaloblastic anaemia6
Sickle cell disease
Urinary iron ↑ (benzidine method of Crosby and Furth and reaction with dipyridyl); proximal tubule iron MRI ↑ (MRI); histological kidney damage6-8
PNH
Urinary haemosiderin ↑ (Prussian reaction of Perls, benzidine method of Crosby and Furth and
reaction with dipyridyl); kidney iron ↑ (Turnbull iron reaction, MRI Prussian blue staining); proteinuria2,6,9-12
Hereditary spherocytosis
Urinary haemosiderin ↑ (Prussian reaction of Perls); kidney iron ↑ (Turnbull iron reaction)2
Haemolysis after
cardiac surgery
Kidney iron ↑ (Prussian blue staining); histological kidney damage (electron microscopy); AKI13
Rhabdomyolysis
Myoglobin in urine ↑ (immunoassay); myoglobinuria predicted kidney failure after rhabdomyolysis14-16
-thalassaemia
Kidney iron ↑ (MRI)17
Patients who complied badly with chelation therapy had higher serum ferritin levels and kidney
dysfunction than did patients who complied well
Animal studies
Haemolytic anaemia
in mice
Kidney iron ↑ (Prussian blue staining)18
Glycerol-induced
myoglobinuric AKI
(rhabdomyolysis)
Urinary iron ↑ (bleomycin assay); kidney iron ↑ (bleomycin assay, immunoperoxidase method to
detect myoglobin); glomerular and tubular damage (histologic, malondialdehyde, PAS); renal cortical haemopexin mRNA ↑;
AKI in rat by plasma creatinine and plasma nitrogen urea, partly in vitro
Deferoxamine infusion attenuated kidney dysfunction and oxidative stress19-25
Haem protein infusion in rat
Large haem-filled endolysosomes in proximal tubule; haem casts in distal nephron; proximal tubule damage (histologic)26
HO1 deficiency leading to haemolytic
anaemia
Proximal tubule iron ↑ (Prussian blue staining); histological kidney damage; CD163, ferritin light
chain, ferritin heavy chain, ferroportin mRNA ↑; haemopexin protein ↑27
In vitro studies
Cells exposed to
haem
DNA damage in distal tubule (MCDK) cells but not proximal tubule (LLC-PK1) cells20
Proximal tubule cells
exposed to myoglobin
Cell damage (LDH); HK-2 cells and extracted proximal tubule cells used28
1
Proximal tubule cells
exposed to haemoglobin
Cell damage (MTT assay); human proximal tubule cells used29
Abbreviations: ↑, increased; ↔, unchanged; AKI, acute kidney injury; AKF, acute kidney failure; CMJ, corticomedullary
junction; HK-2, human kidney 2; HO1, haem oxygenase 1; HUS, haemolytic uremic syndrome; LDH, lactate dehydrogenase;
LLC–PK1, pig kidney epithelial cells; MDCK, Madin–Darby canine kidney; MTT, PAS, pernicious anaemia staining; PNH, paroxysmal nocturnal haemoglubinuria.
1. Rakow-Penner, R., Glader, B., Yu, H. & Vasanawala, S. Adrenal and renal corticomedullary junction iron
deposition in red cell aplasia. Pediatr. Radiol. 40, 1955–1957 (2010).
2. LEONARDI, P. & RUOL, A. Renal hemosiderosis in the hemolytic anemias: diagnosis by means of needle
biopsy. Blood 16, 1029–1038 (1960).
3. Fervenza, F. C. et al. Induction of Heme Oxygenase-1 and Ferritin in the Kidney in Warm Antibody Hemolytic
Anemia. Am. J. Kidney Dis. 52, 972–977 (2008).
4. Yachie, A. et al. Oxidative stress causes enhanced endothelial cell injury in human heme oxygenase-1
deficiency. J. Clin. Invest. 103, 129–135 (1999).
5. Soler-García, Á. A., Johnson, D., Hathout, Y. & Ray, P. E. Iron-related proteins: Candidate urine biomarkers in
childhood HIV-associated renal diseases. Clin. J. Am. Soc. Nephrol. 4, 763–771 (2009).
6. Sears, D. A., Anderson, P. R., Foy, A. L., Williams, H. L. & Crosby, W. H. Urinary iron excretion and renal
metabolism of hemoglobin in hemolytic diseases. Blood 28, 708–725 (1966).
7. Schein, A., Enriquez, C., Coates, T. D. & Wood, J. C. Magnetic resonance detection of kidney iron deposition in
sickle cell disease: A marker of chronic hemolysis. J. Magn. Reson. Imaging 28, 698–704 (2008).
8. Buckalew Jr, V. M. & Someren, A. Renal manifestations of sickle cell disease. Arch. Intern. Med. 133, 660–669
(1974).
9. Hsiao, P. J., Wang, S. C., Wen, M. C., Diang, L. K. & Lin, S. H. Fanconi Syndrome and CKD in a Patient With
Paroxysmal Nocturnal Hemoglobinuria and Hemosiderosis. Am. J. Kidney Dis. 55, e1-e5 (2010).
10. Suzukawa, K. et al. Demonstration of the deposition of hemosiderin in the kidneys of patients with paroxysmal
nocturnal hemoglobinuria by magnetic resonance imaging. Intern. Med. 32, 686–690 (1993).
11. Nath, K. A. et al. Heme protein-induced chronic renal inflammation: Suppressive effect of induced heme
oxygenase-11. Kidney Int. 59, 106–117 (2001).
12. Ballarín, J. et al. Acute renal failure associated to paroxysmal nocturnal haemoglobinuria leads to intratubular
haemosiderin accumulation and CD163 expression. Nephrol. Dial. Transplant. 26, 3408–3411 (2011).
13. Qian, Q., Nath, K. A., Wu, Y., Daoud, T. M. & Sethi, S. Hemolysis and acute kidney failure. Am. J. Kidney Dis.
56, 780–784 (2010).
14. Holt, S. G. & Moore, K. P. Pathogenesis and treatment of renal dysfunction in rhabdomyolysis. Intensive Care
Med. 27, 803–811 (2001).
15. Al-Ismaili, Z., Piccioni, M. & Zappitelli, M. Rhabdomyolysis: Pathogenesis of renal injury and management.
Pediatr. Nephrol. 26, 1781–1788 (2011).
16. Khan, A. A. & Quigley, J. G. Control of intracellular heme levels: Heme transporters and heme oxygenases.
Biochim. Biophys. Acta 1813, 668–682 (2011).
17. Hashemieh, M., Azarkeivan, A., Akhlaghpoor, S., Shirkavand, A. & Sheibani, K. T2-star (T2*) magnetic
resonance imaging for assessment of kidney iron overload in thalassemic patients. Arch. Iran. Med. 15, 91–94
(2012).
18. Veuthey, T., D’Anna, M. C. & Roque, M. E. Role of the kidney in iron homeostasis: Renal expression of
Prohepcidin, Ferroportin, and DMT1 in anemic mice. Am. J. Physiol. Renal Physiol. 295, F1213-F1221 (2008).
19. Baliga, R., Zhang, Z., Baliga, M. & Shah, S. V. Evidence for cytochrome P-450 as a source of catalytic iron in
myoglobinuric acute renal failure. Kidney Int. 49, 362–369 (1996).
20. Nath, K. A. et al. Intracellular targets in heme protein-induced renal injury. Kidney Int. 53, 100–111 (1998).
21. Zager, R. A., Foerder, C. & Bredl, C. The influence of mannitol on myoglobinuric acute renal failure: functional,
biochemical, and morphological assessments. J. Am. Soc. Nephrol. 2, 848–855 (1991).
22. Wei, Q., Hill, W. D., Su, Y., Huang, S. & Dong, Z. Heme oxygenase-1 induction contributes to renoprotection by
G-CSF during rhabdomyolysis-associated acute kidney injury. Am. J. Physiol. Renal Physiol. 301, F162-F170
(2011).
23. Paller, M. S. Hemoglobin- and myoglobin-induced acute renal failure in rats: role of iron in nephrotoxicity. Am.
J. Physiol. 255, (1988).
24. Boutaud, O. et al. Acetaminophen inhibits hemoprotein-catalyzed lipid peroxidation and attenuates
rhabdomyolysis-induced renal failure. Proc. Nat. Acad. Sci. USA 107, 2699–2704 (2010).
25. Zager, R. A., Johnson, A. C. M. & Becker, K. Renal cortical hemopexin accumulation in response to acute
kidney injury. Am. J. Physiol. Renal Physiol. 303, F1460-F1472 (2012).
26. Zager, R. A. Rhabdomyolysis and myohemoglobinuric acute renal failure. Kidney Int. 49, 314–326 (1996).
27. Kovtunovych, G., Eckhaus, M. A., Ghosh, M. C., Ollivierre-Wilson, H. & Rouault, T. A. Dysfunction of the heme
recycling system in heme oxygenase 1-deficient mice: Effects on macrophage viability and tissue iron
2
distribution. Blood 116, 6054–6062 (2010).
28. Zager, R. A. & Burkhart, K. Myoglobin toxicity in proximal human kidney cells: Roles of Fe, Ca2+, H2O2, and
terminal mitochondrial electron transport. Kidney Int. 51, 728–738 (1997).
29. Sheerin, N. S., Sacks, S. H. & Fogazzi, G. B. In vitro erythrophagocytosis by renal tubular cells and tubular
toxicity by haemoglobin and iron. Nephrol. Dial. Transplant. 14, 1391–1397 (1999).
3