Sarah Faubel

Neutrophil-independent mechanisms of caspase-1– and IL-18–mediated ischemic acute tubular necrosis in mice

Having recently described the injurious role of caspase-1-mediated production of the proinflammatory cytokine IL-18 in ischemic acute renal failure (ARF), we report here on the effect of the newly developed caspase inhibitor Quinoline-Val-Asp(Ome)-CH(2)-OPH (OPH-001) on caspase-1, IL-18, neutrophil infiltration, and renal function in ischemic ARF. C57BL/6 mice with ischemic ARF treated with OPH-001 had a marked (100%) reduction in blood urea nitrogen (BUN) and serum creatinine and a highly significant reduction in morphological acute tubular necrosis (ATN) score compared with vehicle-treated mice. OPH-001 significantly reduced the increase in caspase-1 activity and IL-18 and prevented neutrophil infiltration in the kidney during ischemic ARF. To evaluate whether this lack of neutrophil infiltration was contributing to the protection against ischemic ARF, a model of neutrophil depletion was developed. Neutrophil-depleted mice had a small (18%) reduction in serum creatinine during ischemic ARF but no reduction in ATN score despite a lack of neutrophil infiltration in the kidney. Remarkably, caspase-1 activity and IL-18 were significantly increased in the kidney in neutrophil-depleted mice with ARF. In addition, IL-18 antiserum-treated neutrophil-depleted mice with ischemic ARF had a significant (75%) reduction in serum creatinine and a significant reduction in ATN score compared with vehicle-treated neutrophil-depleted mice. These results suggest a novel neutrophil-independent mechanism of IL-18-mediated ischemic ARF.

Cisplatin-induced acute renal failure is associated with an increase in the cytokines interleukin (IL)-1beta, IL-18, IL-6, and neutrophil infiltration in the kidney.

We have demonstrated that caspase-1-deficient (caspase-1–/–) mice are functionally and histologically protected against cisplatin-induced acute renal failure (ARF). Caspase-1 exerts proinflammatory effects via the cytokines interleukin (IL)-1β, IL-18, IL-6, and neutrophil recruitment. We sought to determine the role of the cytokines IL-1β, IL-18, and IL-6 and neutrophil recruitment in cisplatin-induced ARF. We first examined IL-1β; renal IL-1β increased nearly 2-fold in cisplatin-induced ARF and was reduced in the caspase-1–/– mice. However, inhibition with IL-1 receptor antagonist (IL-1Ra) did not attenuate cisplatin-induced ARF. Renal IL-18 increased 2.5-fold; however, methods to inhibit IL-18 using IL-18 antiserum and transgenic mice that overproduce IL-18-binding protein (a natural inhibitor of IL-18) did not protect. Renal IL-6 increased 3-fold; however, IL-6-deficient (IL-6–/–) mice still developed cisplatin-induced ARF. We next examined neutrophils; blood neutrophils increased dramatically after cisplatin injection; however, prevention of peripheral neutrophilia and renal neutrophil infiltration with the neutrophil-depleting antibody RB6-8C5 did not protect against cisplatin-induced ARF. In summary, our data demonstrated that cisplatin-induced ARF is associated with increases in the cytokines IL-1β, IL-18, and IL-6 and neutrophil infiltration in the kidney. However, inhibition of IL-1β, IL-18, and IL-6 or neutrophil infiltration in the kidney is not sufficient to prevent cisplatin-induced ARF.

Acute Renal Failure after Bilateral Nephrectomy Is Associated with Cytokine-Mediated Pulmonary Injury

Clinical studies demonstrate that acute renal failure (ARF) is associated with increased mortality, which may be due to pulmonary complications. ARF may affect the lung via increased renal production or impaired clearance of mediators of lung injury, such as proinflammatory cytokines. Bilateral nephrectomy is a method to examine directly the deleterious systemic effects of absent renal clearance in ARF without the confounding effects that are associated with ischemia-reperfusion injury (e.g., ischemic ARF) or systemic toxicity (e.g., cisplatin-induced ARF). This study contrasts the effects of ischemic ARF and bilateral nephrectomy on serum cytokines and lung injury. It demonstrates that the acute absence of kidney function after both ischemic ARF and bilateral nephrectomy is associated with an increase in multiple serum cytokines, including IL-6 and IL-1beta, and that the cytokine profiles were distinct. Lung injury after ischemic ARF and bilateral nephrectomy was similar and was characterized by pulmonary vascular congestion and neutrophil infiltration. For investigation of the role of proinflammatory cytokines in pulmonary injury after ARF, the anti-inflammatory cytokine IL-10 was administered before bilateral nephrectomy. IL-10 treatment improved pulmonary architecture and was associated with a reduction in inflammatory markers, including bronchoalveolar lavage fluid total protein, pulmonary myeloperoxidase activity (a biochemical marker of neutrophils), and the chemokine macrophage inflammatory protein 2. These data demonstrate for the first time that the acute absence of kidney function results in pulmonary injury independent of renal ischemia and highlight the critical role of the kidney in the maintenance of serum cytokine balance and pulmonary homeostasis.

Interleukin-6 mediates lung injury following ischemic acute kidney injury or bilateral nephrectomy

Patients with acute kidney injury frequently have pulmonary complications. Similarly ischemic acute kidney injury or bilateral nephrectomy in rodents causes lung injury characterized by pulmonary edema, increased pulmonary capillary leak and interstitial leukocyte infiltration. Interleukin-6 is a pro-inflammatory cytokine that is increased in the serum of patients with acute kidney injury and predicts mortality. Here we found that lung neutrophil infiltration, myeloperoxidase activity, the neutrophil chemokines KC and MIP-2 and capillary leak all increased within 4 h following acute kidney injury in wild-type mice. These pathologic factors were reduced in interleukin-6-deficient mice following acute kidney injury or bilateral nephrectomy. The lungs of mutant mice had reduced KC but MIP-2 was similar to that of wild type mice. Wild-type mice, treated with an interleukin-6 inactivating antibody, had decreased lung myeloperoxidase activity and KC levels following acute kidney injury. Our study shows that interleukin-6 contributes to lung injury following acute kidney injury.

Serum Interleukin-6 and interleukin-8 are early biomarkers of acute kidney injury and predict prolonged mechanical ventilation in children undergoing cardiac surgery: a case-control study

IntroductionAcute kidney injury (AKI) is associated with high mortality rates. New biomarkers that can identify subjects with early AKI (before the increase in serum creatinine) are needed to facilitate appropriate treatment. The purpose of this study was to test the role of serum cytokines as biomarkers for AKI and prolonged mechanical ventilation.MethodsThis was a case-control study of children undergoing cardiac surgery. AKI was defined as a 50% increase in serum creatinine from baseline within 3 days. Levels of serum interleukin (IL)-1β, IL-5, IL-6, IL-8, IL-10, IL-17, interferon (IFN)-γ, tumor necrosis factor-α (TNF-α), granulocyte colony-stimulating factor (G-CSF), and granulocyte-macrophage colony-stimulating factor (GM-CSF) were measured using a bead-based multiplex cytokine kit in conjunction with flow-based protein detection and the Luminex LabMAP multiplex system in 18 cases and 21 controls. Levels of IL-6 and IL-8 were confirmed with single-analyte ELISA; IL-18 was also measured with single-analyte ELISA.ResultsIL-6 levels at 2 and 12 hours after cardiopulmonary bypass (CPB) and IL-8 levels at 2, 12 and 24 hours were associated with the development of AKI using the Wilcoxon rank-sum test and after adjustment for age, gender, race, and prior cardiac surgery in multivariate logistic regression analysis. In patients with AKI, IL-6 levels at 2 hours had excellent predictive value for prolonged mechanical ventilation (defined as mechanical ventilation for more than 24 hours postoperatively) by receiver operator curve (ROC) analysis, with an area under the ROC curve of 0.95. IL-8 levels at 2 hours had excellent predictive value for prolonged mechanical ventilation in all patients. Serum IL-18 levels were not different between those with and without AKI.ConclusionsSerum IL-6 and IL-8 values identify AKI early in patients undergoing CPB surgery. Furthermore, among patients with AKI, high IL-6 levels are associated with prolonged mechanical ventilation, suggesting that circulating cytokines in patients with AKI may have deleterious effects on other organs, including the lungs.

AKI Transition of Care: A Potential Opportunity to Detect and Prevent CKD

The incidence rate of AKI is increasing across the spectrum of hospitalized children and adults. Given the increased morbidity and mortality associated with AKI, significant research effort has been appropriately focused on standardizing AKI definitions, identifying risk factors, and discovering and validating novel, earlier structural biomarkers of kidney injury. In addition, a growing body of evidence demonstrates that AKI is a risk factor for the future development or accelerated progression of CKD. Unfortunately, prospective observational studies have not consistently followed survivors of episodes of AKI for longitudinal outcomes after hospital discharge, which could lead to ascertainment bias in terms of over- or underestimation of CKD development. Furthermore, data show that clinical follow-up of AKI survivors is low. This lack of systematic study and clinical follow-up represents a potential missed opportunity to prevent chronic disease after an acute illness and improve outcomes. Therefore, prospective study of transitions of care after episodes of AKI is needed to identify which patients are at risk for CKD development and to optimally target therapeutic interventions.

Caspases as drug targets in ischemic organ injury.

Caspases are intracellular cysteine proteases that mediate cell death and inflammation. Caspase-3 is a major mediator of both apoptotic and necrotic cell death. Caspase-1 mediates inflammation though the activation of the cytokines interleukin-1beta (IL-1beta) and interleukin-18 (IL-18). Increases in both caspase-1 and -3 have been described in ischemic injury to various organs including brain, heart and kidney. Both pharmacological inhibitors and genetic approaches have been used to inhibit caspases in vivo. Pancaspase inhibitors protect against ischemic injury in brain, heart and kidney. Pancaspase inhibition also reduces cold preservation injury due to apoptosis in liver endothelial cells and prolongs animal survival after orthotopic liver transplantation. Caspase-1 inhibition or caspase-1 deficiency protects against ischemic injury in brain, heart and kidney models of ischemia. Specifically, impaired IL-18 processing protects caspase-1-deficient mice from ischemic acute renal failure. This review focuses on studies of caspase-1 and pancaspase inhibition in ischemic injury to brain, heart and kidney. In addition, the studies of pancaspase inhibition in cold ischemic injury and organ preservation will be reviewed. The therapeutic potential of caspase inhibition in ischemic injury will be discussed.

Increased macrophage infiltration and fractalkine expression in cisplatin-induced acute renal failure in mice

Inflammatory mechanisms contribute to cisplatin-induced acute renal failure (CisARF). Our first aim was to determine renal macrophage infiltration in CisARF. A more than 2-fold increase in CD11b-positive macrophages in the kidney on day 2 preceded the increase in blood urea nitrogen (BUN) and serum creatinine (SCr). Our next aim was to determine the chemoattractant for macrophage infiltration in CisARF. Fractalkine (CX3CL1) is expressed on activated endothelial cells and is a potent chemoattractant for macrophages that express its receptor (CX3CR1). Immunoblotting showed that whole-kidney CX3CL1 expression on days 1, 2, and 3 after cisplatin administration was increased. On immunofluorescence, the intensity of renal endothelial staining of CX3CL1 in blood vessels was significantly increased on day 2. Circulating von Willebrand factor (vWF), a measure of systemic endothelial injury, was increased on day 2. Next we determined whether macrophages played an injurious role in CisARF. Macrophages were depleted with injections of liposome-encapsulated clodronate (LEC). LEC resulted in a decrease in renal CD11b-positive macrophages on day 3. However, LEC-treated mice were not protected from CisARF on day 3. To determine the role of CX3CR1, both a specific anti-CX3 CR1 antibody and CX3 CR1–/– mice were used. Administration of the CX3CR1 antibody and CX3 CR1–/– mice was not protected against CisARF. In summary, in CisARF, macrophage infiltration in the kidney, CX3CL1 expression in whole kidney and blood vessels, and the increase in circulating vWF precede BUN and SCr increase. However, inhibition of macrophage infiltration in the kidney or CX3CR1 blockade is not sufficient to prevent CisARF.

IL-33 Exacerbates Acute Kidney Injury

Inflammation contributes to the pathogenesis of acute kidney injury (AKI). IL-33 is a proinflammatory cytokine, but its role in AKI is unknown. Here we observed increased protein expression of full-length IL-33 in the kidney following induction of AKI with cisplatin. To determine whether IL-33 promotes injury, we administered soluble ST2 (sST2), a fusion protein that neutralizes IL-33 activity by acting as a decoy receptor. Compared with cisplatin-induced AKI in untreated mice, mice treated with sST2 had fewer CD4 T cells infiltrate the kidney, lower serum creatinine, and reduced acute tubular necrosis (ATN) and apoptosis. In contrast, administration of recombinant IL-33 (rIL-33) exacerbated cisplatin-induced AKI, measured by an increase in CD4 T cell infiltration, serum creatinine, ATN, and apoptosis; this did not occur in CD4-deficient mice, suggesting that CD4 T cells mediate the injurious effect of IL-33. Wildtype mice that received cisplatin and rIL-33 also had higher levels of the proinflammatory chemokine CXCL1, which CD T cells produce, in the kidney compared with CD4-deficient mice. Mice deficient in the CXCL1 receptor also had lower serum creatinine, ATN, and apoptosis than wildtype mice following cisplatin-induced AKI. Taken together, IL-33 promotes AKI through CD4 T cell-mediated production of CXCL1. These data suggest that inhibiting IL-33 or CXCL1 may have therapeutic potential in AKI.

Pulmonary Complications After Acute Kidney Injury

The development of respiratory failure in patients with AKI is a particularly devastating consequence that greatly increases patient mortality. When respiratory failure and AKI occur together, the mortality is greater than 80%. A clear understanding of the mechanisms leading to respiratory failure is of great clinical relevance to patients with AKI in order to prevent and treat this life-threatening complication. Pulmonary edema leading to respiratory failure has been a recognized complication of kidney failure since 1901. Remarkably, the pathogenesis of this complication remains elusive, despite over 100 years of clinical and experimental debate in the literature. A review of this literature suggests that there are 4 causes of pulmonary edema leading to respiratory failure in patients with AKI: (1) volume overload (cardiogenic edema), (2) left ventricular dysfunction (cardiogenic edema), (3) increased lung capillary permeability (noncardiogenic edema), and (4) acute lung injury (noncardiogenic edema with inflammation). In this review, these mechanisms are presented in historical context including the original descriptions of pathology and pathophysiology, recent epidemiologic data, and experimental studies in animals. Although volume overload is a well-accepted mechanism of pulmonary edema in patients with AKI, the purpose of this review was to highlight the evidence showing that noncardiogenic edema and acute lung injury also occur. By recognizing that the pulmonary complications of AKI are not simply from volume overload, specific treatment strategies may be discovered and used to improve outcomes in patients with the ominous and life threatening combination of AKI and respiratory failure.