Heavy Metal Suicide

Am J Physiol Renal Physiol 313: F959–F960, 2017; First published May 31, 2017; doi:10.1152/ajprenal.00273.2017. EDITORIAL FOCUS Heavy metal suicide X Andreas Linkermann,1 Brent Stockwell,2 and Tom Vanden Berghe3,4 1 Division of Nephrology, Department of Internal Medicine III, University Hospital Carl Gustav Carus at the Technische Universität Dresden, Dresden, Germany; 2Department of Chemistry, Columbia University, New York, New York; 3VIB Inflammation Research Center, Ghent, Belgium; and 4Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium “. . . wheel of fortune, Sally Ride, heavy metal suicide . . .” (AKI), the majority of dying tubular cells succumbs to an iron-dependent form of regulated necrosis, referred to as ferroptosis. Ferroptosis is essentially mediated by iron-catalyzed lipid peroxidation on GPX4 dysfunction. Heme oxygenase-1 (HO-1) is a master regulator of intracellular free iron because of the conversion of heme to iron, carbon monoxide, and biliverdin (12), and therefore represents a potential regulator of ferroptotic cell death. In a recent issue of AJP-Renal Physiology, Adedoyin et al. (1) demonstrate that the lack of HO-1 sensitizes renal tubular cells to ferroptosis. Agarwal and colleagues incubated primary renal proximal tubular cells (PTCs) with different concentrations of the class I ferroptosis-inducing compound (FIN) erastin, or the class II FIN RSL3, which directly targets and inactivates glutathionperoxidase 4 (GPX4) (16). Whereas 10 ␮M RSL3 clearly induced profound ferroptosis within 16 h, approximately 35% of PTCs died following treatment with 1 ␮M erastin. Cleavage of capsase-3 remained absent over the investigated time of the experiment, ruling out the involvement of apoptotic cell death. Instead, the iron chelator desferoxamine (DFO), N-acetylcysteine (NAC), or the ferrostatin (Fer-1) were effective in preventing cell death, which underscores that both FINs induced ferroptosis in PTCs. Interestingly, both FINs upregulated the expression of HO-1 during ferroptotic cell death, which was reduced on treatment with the ferroptosis inhibitors DFO, NAC, and Fer-1. Functional characterization of HO-1 knockout PTCs also revealed an increased sensitivity to ferroptosis, suggesting a protective role of HO-1 in ferroptosis. In line with these data, it was previously demonstrated that genetic absence of HO-1 sensitizes mice to cisplatin-induced AKI, and that the induction of HO-1 expression by addition of the molecule hemin to human renal tubules reduced necrotic damage (13). Also in hepatocellular carcinoma cells, a protective role for the NRF2-HO-1 pathway was observed in response to erastin- and sorafenib-induced ferroptosis (14). The extraordinary role of ferroptosis in renal tubules is highlighted by three hallmark observations. First, inducible deletion of GPX4 in renal tubules results in massive tubular necrosis that is prevented by the ferroptosis inhibitor liproxIN ACUTE KIDNEY INJURY Address for reprints and other correspondence: A. Linkermann, Div. of Nephrology, Dept. of Internal Medicine III, University Hospital Carl Gustav Carus at Technische Universität Dresden, Fetscherstr. 74, 01307 Dresden, Germany (e-mail: [email protected]). http://www.ajprenal.org 1931-857X/17 Copyright © 2017 the American Physiological Society F959 Downloaded from http://ajprenal.physiology.org/ by 10.220.33.3 on November 3, 2017 We Didn’t Start the Fire (Billy Joel) statin-1 (3). Second, all renal clear cell carcinoma cell lines in the NCI tumor panel were sensitive to RSL3-mediated ferroptosis (16). And third, prevention of ferroptosis in murine models of AKI resulted in remarkable preservation of renal function, even on otherwise lethal ischemic damage (9). In contrast to other clinically relevant forms of regulated necrosis, such as necroptosis or pyroptosis, ferroptosis exhibits the only known cell death pathway that is capable of inducing synchronized regulated necrosis in functional units of interconnected cells in a noncell autonomous manner (8). Importantly, this effect does not appear to be limited to renal tubules. Recently, cell culture models have been employed to detect similar effects that are mediated by ferroptosis (6). With this growing mechanistic understanding of ferroptosis and its regulation, novel methodological approaches may allow more detailed research. In this sense, ACSL4 was recently demonstrated to mediate ferroptotic cell death (2, 5), providing the opportunity to generate tubular cell specific knockout of ACSL4. Several obstacles remain related to the mechanisms of ferroptosis during AKI. One major question to address is the measurement of free or labile iron inside the cellular cytoplasm. This is of particular importance because the reaction that is catalyzed by HO-1 releases free iron (Fe2⫹) from heme and should increase the labile iron pool, which was postulated to contribute to the cytotoxicity already in 1999 (15). More recently, this idea was further supported by the finding that HO-1 is accelerating erastin-induced ferroptosis in fibroblasts (7). This cytotoxic vs. cytoprotective action of HO-1 might be related to the iron buffering capacity of the cells (such as ferritin) or the expression levels of the iron export protein ferroportin. Should this capacity be interfered with, e.g., by genetic targeting, tubular cells could no longer keep control about their free iron content, which may result in detrimental necrosis and systemic inflammation. Therefore, the results presented in the current work, together with previously published knowledge about HO-1 biology (11), suggest a faint and tightly regulated balance between ferritin, free iron, and HO-1 activity. It will be important to investigate these balances in more detail to understand ferroptotic tubular necrosis and its inevitably associated inflammatory response—not only for AKI, but also for kidney transplantation and possibly ischemic injury in many other organ systems. In addition, the work by Adedoyin et al. provides an important in vitro link between HO-1 and ferroptosis. In vivo, HO-1 therefore should control local necroinflammation (10) and indirectly regulate systemic inflammation and distant organ effects in AKI (4). F960 DISCLOSURES No conflicts of interest, financial or otherwise, are declared by the author(s). AUTHOR CONTRIBUTIONS 7. A.L., B.R.S., and T.V.B. analyzed data; A.L., B.R.S., and T.V.B. interpreted results of experiments; A.L., B.R.S., and T.V.B. drafted manuscript; A.L., B.R.S., and T.V.B. edited and revised manuscript; A.L., B.R.S., and T.V.B. approved final version of manuscript. 8. 9. REFERENCES 10. 11. 12. 13. 14. 15. 16. AJP-Renal Physiol • doi:10.1152/ajprenal.00273.2017 • www.ajprenal.org Downloaded from http://ajprenal.physiology.org/ by 10.220.33.3 on November 3, 2017 1. Adedoyin O, Boddu R, Traylor AM, Lever JM, Bolisetty S, George J, Agarwal A. Heme oxygenase-1 mitigates ferroptosis in renal proximal tubule cells. Am J Physiol Renal Physiol. doi:10.1152/ajprenal.00044. 2017. 2. Doll S, Proneth B, Tyurina YY, Panzilius E, Kobayashi S, Ingold I, Irmler M, Beckers J, Aichler M, Walch A, Prokisch H, Trümbach D, Mao G, Qu F, Bayir H, Füllekrug J, Scheel CH, Wurst W, Schick JA, Kagan VE, Angeli JP, Conrad M. ACSL4 dictates ferroptosis sensitivity by shaping cellular lipid composition. 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