Ironing Out the Pathogenesis of Acute Kidney Injury

Acute kidney injury (AKI) is a serious and common complication with a reported incidence varying from 19 to 70/1,000 hospitalized patients. Despite recent advances in clinical care, the mortality associated with AKI remains high, ranging from 20% to 60% in hospitalized patients.1 Moreover, recent data emphasize that even in survivors, recovery of kidney function often is incomplete, and chronic kidney disease and even end-stage renal disease may ensue.2

How can the outcomes in patients with AKI be improved? An important initial step would be to develop better tools for the identification and surveillance of at-risk patients and prognosis in the disease evolution.3 Current definitions of AKI rely on a change in serum creatinine level as a glomerular filtration rate marker. However, changes in creatinine level, if not in equilibrium, cannot be used to accurately monitor kidney function. Even in the steady state, creatinine has limitations as a filtration marker in patients with alterations in creatinine generation, such as malnutrition, muscle wasting, hypercatabolism, cancer, elderly age, and those in whom third-space fluid accumulation and vigorous volume resuscitation results in dilution of the creatinine pool.4, 5, 6, 7 Thus, the field of AKI is ripe for the development of new biomarkers that identify at-risk patients, and major efforts to accomplish this are well underway. However, AKI is a complex and heterogeneous process, and the identification of biomarkers should not be limited to initial injury alone, but should include markers of risk, injury propagation, and resolution of injury. It can be argued that markers of early resolution will be equally as important as markers of initial injury. Identifying markers of risk, injury, protection, and repair in human studies may also help us better understand the pathogenesis of AKI, thereby informing future therapeutic efforts.

In this issue of the American Journal of Kidney Diseases, Ho et al8 apply proteomics to the analysis of urine for biomarker discovery in a prospective cohort study of individuals undergoing cardiopulmonary bypass surgery. There were several notable findings in the study. First, regardless of whether AKI developed, all subjects had early evidence of tubular dysfunction and stress, shown by early β2-microglobulinuria. Systemic markers of inflammation, such as the proinflammatory cytokine interleukin 6 and several chemokines, were also increased in both groups at each time measured. Second, in patients who went on to develop AKI, the urinary proteome became more complex, suggesting either a second phase of injury or progression of injury. In these individuals, α1-microglobulin and neutrophil gelatinase-associated lipocalin (NGAL) were identified as early as 1 hour into the bypass procedure and remained increased at postoperative days 3 to 5 in patients with AKI compared with patients who did not develop AKI. Third, and perhaps most interesting, proteomic analysis identified 2 high-intensity peaks for which the appearance postoperatively was associated with lack of development of AKI. One of these peaks was subsequently determined to be hepcidin-25, which was preferentially found in urine on postoperative day 1 of patients who did not go on to develop AKI. Because hepcidin is a master regulator of iron homeostasis, these data again invoke the importance of free iron in the pathogenesis of ischemic and toxic AKI.

Two decades ago, seminal studies suggested that free iron has a role in models of ischemic and toxic AKI. In a rat model of gentamicin toxicity, rats treated with a hydroxyl radical scavenger or an iron chelator (desferoxamine) had significantly lower serum urea nitrogen levels and improved histological characteristics compared with untreated rats, suggesting a role for iron and generation of hydroxyl radicals in toxic AKI.9 Similarly, in a rat model of ischemia-reperfusion injury induced by clamping the renal artery, pretreatment with desferoxamine can prevent the development of AKI. In that study, urinary levels of free iron increased 10- to 20- fold during reperfusion with no change in plasma levels. These studies each support a role for iron in mediating AKI through the generation of oxygen free radicals and suggest that this injury takes place in the urinary space.10 Human studies have shown excess iron accumulation in proximal tubule lysosomes in biopsy specimens from patients with AKI.11

Hepcidin-25, the 25–amino acid bioactive form of the peptide hormone made in the liver, regulates iron homeostasis through downregulation of ferroportin, an iron exporter located in hepatocytes, enterocytes, and macrophages. Hepcidin production is suppressed by anemia and hypoxia and induced by infection and inflammation.12 In the kidney, hepcidin is strongly expressed in the thick ascending limb of the cortex and connecting tubules. Moderate expression is found in the collecting ducts and thick ascending limb of the medulla.13 One could postulate that upregulation of hepcidin in these segments of the nephron after renal ischemia leads to downregulation of the ferroportin exporter, preventing the further release of free iron into tissues and resulting in perpetuation of tubular injury.

It is of considerable interest in the study by Ho et al8 that urinary NGAL and hepcidin, both proteins that bind or regulate iron, had opposing predictive effects for the development of AKI. It may turn out to be the ratio of NGAL to hepcidin, rather than each marker in isolation, that best predicts the development or lack thereof of AKI. NGAL, which binds siderophore iron, is expressed in the kidney, where it may be localized in the loop of Henle and collecting ducts. However, circulating NGAL released from inflammatory cells probably accounts for most of the NGAL found in the proximal tubule and presumably in urine in the setting of kidney injury.14 To further complicate the picture, it has recently been reported that bone morphogenetic protein signaling, with hemojuvelin as a coreceptor, regulates the expression of hepcidin in the liver.15 Because bone morphogenetic protein signaling in the kidney may be important in reparative processes, it is possible that if this pathway is operative in the ischemic kidney, it may account for the observed association of increased urinary hepcidin with protection from AKI. The kidney is also the mouse tissue in which iron regulatory protein 1 is most highly expressed and is localized to the proximal tubule,16, 17 and the role of multiple participants in kidney iron regulation still needs to be carefully examined for their relevance in the pathogenesis of AKI.

Although of great interest and potential importance, there are a number of concerns with the study by Ho et al8 that must be addressed in future research. First, the surface-enhanced laser desorption/ionization time-of-flight mass spectrometric proteomics platform used in this study is largely qualitative and not ideal for definitive quantitation or even structural identification of analytes. Second, validation of the findings in other cohorts is necessary given the high potential for false detection rates in proteomics studies. New studies examining the hypothesis that hepcidin is an important protective molecule in renal ischemic/toxic injury should be facilitated now that a serum enzyme-linked immunosorbent assay for hepcidin has been developed and validated in healthy volunteers.18