Acute kidney injury (AKI) secondary to ischemia–reperfusion injury (IRI) is a major cause of patient morbidity and mortality in the perioperative period. It can lead to new onset of chronic kidney disease and accelerate its progression. Patients with risk factors undergoing cardiac, vascular, and liver transplantation surgeries, which may inevitably involve IRI, are more susceptible to AKI. Anesthetic agents have been postulated to possess renoprotective properties. Thus, exploring the utilization of selective perioperative anesthetic agents with renoprotective properties may be a promising avenue to reduce the risk of AKI. This review discusses the effects and mechanisms of dexmedetomidine, inhalational and intravenous anesthetics, and xenon-mediated renoprotection. Although the renoprotective effects of these agents obtained in the laboratory are promising, much work especially via clinical trials is required to determine the translational value from the bench to the bedside.
anesthetics; ischemia–reperfusion injury; mechanism; renal injury;
Acute kidney injury (AKI) secondary to ischemia–reperfusion injury (IRI) is a key contributor to morbidity and mortality in the perioperative period, presenting a major health care and economic burden.1, 2, 3, 4 It also leads to new onset of chronic kidney disease (CKD) while concomitantly expediting its progression. The term AKI that is used throughout this review has gained recognition to replace the previously used term, acute renal failure (ARF), in order to achieve two main objectives: first, to establish a conventional terminology that enables comparison of data more efficiently; and second, to illustrate the wide range of disease states the term incorporates.5, 6 Delayed graft function, which can be defined as a patient needing dialysis within 7 days of renal transplantation,7 is an unfortunate extension of AKI in the transplant process that is associated with a higher incidence of acute rejection and poorer long-term graft survival.8 In this review, we discuss the epidemiology, cause, and mechanisms of AKI together with dexmedetomidine, inhalational and intravenous anesthetics, and xenon gas-mediated renoprotection and their potential clinical applications.
2. Epidemiology of AKI
The Risk, Injury, Failure, Loss, and End-stage renal disease classification (RIFLE) is now used as a valuable criteria for diagnosis and monitoring prognosis of AKI (Table 1).6 A meta-analysis identified that the overall mortality rate from AKI remained unchanged at 50% for the past 5 decades.9Approximately 1% of surgical patients develop AKI, however, perioperative AKI mortality can be as high as 83% in cardiac surgery.10 In a large retrospective study of 2672 patients undergoing coronary artery bypass grafting (CABG), perioperative AKI was found to be associated with a 14-fold increase in mortality.11 It is also linked to vascular intervention12 and surgery,13 as well as urology, kidney, and liver transplantation.14 A prospective multicenter cohort study of 29,269 intensive care unit (ICU) patients demonstrated an incidence of 5.7% and mortality of 60.3%.15 Its high incidence is comparable with acute lung injury and severe sepsis.16 In addition, AKI significantly prolonged the length of hospitalization by up to 50%.17 As a consequence of advancing techniques, the surgical population is steadily ageing with increasing comorbidities, causing AKI to become a worsening problem, which continues to be a substantial burden on the health care system.18 Moreover, there is evidence to suggest that AKI can not only predispose but compound the effects of an existing diagnosis of CKD to increase the risk of end-stage renal disease amongst the elderly.19
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3. Causes of AKI and risk factors
Causes of AKI have traditionally been classified as prerenal (occurring in 30–60% of cases), intrinsic renal (20–40%), or postrenal (10%).5 Prerenal causes are associated with hypoperfusion and acute tubular necrosis (ATN), which is linked to trauma, hemorrhage, cardiogenic shock, sepsis, systemic inflammatory response syndrome (SIRS), and IRI in surgical patients. IRI is a major cause of perioperative AKI, involving the inevitable temporary cessation and restoration of blood supply during surgeries such as cardiopulmonary bypass, renal artery angioplasty, and liver and kidney transplantation.5 The short-term hypoxia is detrimental to the kidneys. Up to 50% of donor kidneys are involved in short-term hypoxia in renal transplantation, further diminishing the donor rate and leading to a 10% failure rate of primary grafts in those transplanted. The resurgence of blood flow may commence the recovery of the injured tissues from the ischemic phase yet also paradoxically induce further damage. Patients with high risk factors, such as advanced age, hypertension, diabetes, decreased cardiac function, and pre-existing renal disease20 undergoing cardiac,21 vascular, and liver transplantation surgeries, which inevitably involve IRI, are more susceptible to AKI and higher mortality.22
4. Mechanisms of IRI-induced AKI
IRI is defined as “the cellular damage after reperfusion of previously viable ischemic tissues”.23 Its pathogenesis is a multifactorial interplay between biochemical, cellular, vascular endothelial, and tissue-specific factors, with inflammation being a common feature.24 Renal hypoperfusion triggers vasoconstriction, alteration of the ultrafiltration coefficient, tubular obstruction, and vascular congestion.25 However, recent experimental work has challenged this established “decreased perfusion paradigm”. It has been postulated that intrarenal circulation changes such as modification in efferent arteriolar function and intrarenal shunting may be more likely drivers of AKI than changes in global blood flow.26
Hypoxia as a result of ischemia and subsequent reperfusion is associated with increased reactive oxygen species (ROS) production and dysfunction of the antioxidant system, resulting in tubular cell injury and death.27 Inflammatory cytokines and chemokines are secreted, initiating the innate immune response. There is induction of toll-like receptors, activation of polymorphonuclear cells, with neutrophil infiltration following expression of adhesion molecules27 causing microvascular plugging and promoting local tissue destruction. Complement activation via the alternative pathway is also involved.28 Local inflammation causes ATN, which accounts for 75% of AKI. The necrotic debris acts as a danger signal and stimulates further inflammatory response in a vicious cycle. The molecular mechanisms of AKI are summarized in Fig. 1.
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5. The renoprotective effects of anesthetics
There is a distinct lack of effective strategies to obviate the number of surgical cases of patients developing AKI. Therefore, formulating effective strategies to prevent and treat AKI, which especially occur during the perioperative period, are urgently needed. Utilization of selective perioperative anesthetic agents with renoprotective properties may be a promising avenue. The most recent publications in this regard are shown in Table 2 and the details of these publications are discussed below.
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5.1. Sedative dexmedetomidine
Gamma-aminobutyric acid (GABA)-based medication such as benzodiazepines (usually midazolam) and propofol are commonly used sedative agents in the ICU but have potentially fatal adverse effects including respiratory depression and other acute brain dysfunction disorders.29 As a result, other classes of drugs are often trialed as alternatives or adjuncts to the traditional GABA-based agents, one of which are the α2-adrenoceptor agonists. α2-Adrenoceptor agonists act on the locus coeruleus and spinal cord to exert sedative, analgesic, and anxiolytic effects,30 and one example of such an agent is dexmedetomidine. In addition to its indicated use in the ICU, dexmedetomidine has also been investigated for its potential renoprotective effects in the surgical setting.
AKI can occur peri- or postoperatively following hemodynamic changes leading to renal vasoconstriction, hypoperfusion,31 and reduced urine output. α2-Adrenoceptors are known to be present in the human kidney32 and animal models have shown that these adrenoceptors are important in mediating several renal functions including reducing renin release32 and increasing water and sodium excretion.33 During surgery, dexmedetomidine has been shown to decrease catecholamine plasma levels,33, 34 maintain hemodynamic stability,35, 36 and also increase urinary output in both humans37 and animals.38, 39 Canine models have shown that this increase in urinary output may be a result of aqueous diuresis secondary to central inhibition of vasopressin secretion.38 Dexmedetomidine may also have a role in tubular regeneration as one study found that dexmedetomidine stimulated the α2B-adrenoceptors in the proximal tubule cells of rats, which activated the extracellular signal-regulated kinase (ERK) pathway leading to enhanced mitogen-activated protein kinase (MAPK) activity, translocation of ERK2 to the nucleus, and increased proliferation of epithelial cells in the proximal tubules.40 Therefore, several studies have looked at whether dexmedetomidine may have a renoprotective effect in certain settings; the two most commonly used settings are IRI and cardiothoracic surgery.
IRI is injury to tissue due to its reperfusion after being ischemic for a period of time as the return of blood to the ischemic tissue carries with it immune cells that release inflammatory mediators in response to the tissue damage caused by the ischemia and production of oxygen free radicals. IRI is a common cause of perioperative AKI25 because transient deprivation of blood flow to the kidneys results in severe, continuous damage to the renal parenchyma when blood flow is restored. Dexmedetomidine has been shown to attenuate renal injury in murine models of IRI41, 42, 43 with normal glomeruli found in mice treated with dexmedetomidine whereas the untreated mice had tubular cell swelling, medullary congestion, and renal cell necrosis on histology.43 The mechanism by which dexmedetomidine exerts its renoprotective effects is unclear, but it is thought to activate the cell survival signal phosphoinositide-3-kinase (PI3)–AKT, which upregulates anti-apoptotic proteins such as Bcl-2 and Bcl-xl and inhibits the inflammatory-associated high-mobility group protein B1 (HMGB-1)/Toll-like receptor 4 (TLR-4)41 and Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathways44 via its actions on the α2-adrenoceptors, thus reducing renal cell death and preserving tubular structure. Si et al44 explored the link between dexmedetomidine and the JAK/STAT pathway. Evidence has shown that the control group exhibited significant increase in expression of phosphorylated JAK2 (p-JAK2) protein in comparison to the sham-operated group of Wistar rats, whereas the dexmedetomidine-treated group and a second group treated with AG490 (JAK2 inhibitor tyrphostin) both showed a comparatively significant reduced level of p-JAK2 and its downstream effectors p-STAT1 and STAT3. This suggested a similarity in their mechanisms and further establishes a relationship between dexmedetomidine and its inhibition on the JAK/STAT signaling pathway. Evidence from Yang et al45 further demonstrated that preconditioning with AG490 in rats resulted in a better histological profile with a lower tubular necrosis score and reduced cell apoptosis in epithelial cells compared with the vehicle-alone control group. Dexmedetomidine has also been shown to have a protective effect over lung injuries induced by renal IRI through both α2-adrenoceptor-dependent and -independent mechanisms.41
The renoprotective effects of dexmedetomidine have also been investigated in CABG because it may attenuate renal function by stimulating the sympathetic nervous system and influencing the body's hemodynamics and renal blood flow.46 The first human study in this area was a double-blinded, placebo-controlled randomized controlled trial (RCT) of 80 patients undergoing CABG who were given either a saline placebo or dexmedetomidine before induction of anesthesia and then perioperatively until the end of the operation.37 The authors found that dexmedetomidine reduced intraoperative sympathetic tone and attenuated hemodynamic instability with reduced incidence of tachycardia and decreased variation of systolic arterial pressure.37 Renal function, however, was not analyzed. Subsequent RCTs46, 47, 48 have shown no significant improvement or worsening renal function using creatinine clearance or serum creatinine (SCr) as a measure of outcome. However, these contrasting results were due to using lower than normal dosages,47 and either recruiting patients who had normal renal function and SCr and hence were at relatively lower risk of AKI,46 or recruiting patients with a greater incidence of preoperative renal failure into the dexmedetomidine group.49 Additionally, one of the RCTs did notice a mean 74% increase in urinary output in patients given dexmedetomidine.46 A third RCT50 investigating dexmedetomidine effects on perioperative renal function during a thoracotomy did find improved renal function in terms of creatinine clearance and SCr. However, the study was performed on a small cohort of patients. Hence further trials are warranted to clarify if dexmedetomidine improves renal function in patients undergoing cardiac surgery. Currently, one study in Korea is recruiting patients undergoing valvular heart surgery to investigate the incidence of AKI postoperatively after receiving perioperative dexmedetomidine or placebo (http://clinicaltrials.gov/ct2/show/NCT01886079).
5.2. Volatile anesthetics
Isoflurane is a volatile anesthetic commonly used in general anesthesia to maintain loss of consciousness, analgesia, and muscle relaxation. Its mechanism of action as an anesthetic agent is still incompletely understood but it is thought to bind to the GABA,51 glutamate,52 and glycine53 receptors among many others to exert its anesthetic effects.
Volatile anesthetics have also been known to confer some protection against cardiac IRI and this is demonstrated for some volatile anesthetics in renal IRI such as isoflurane.54 Renal IRI mice anesthetized with isoflurane have significantly lower plasma creatinine,55 reduced migration of neutrophils and macrophages into the kidneys,56 reduced expression of adhesion and proinflammatory molecules,56 and attenuated necrotic and apoptotic injury57as compared to renal IRI mice anesthetized with pentobarbital.
Isoflurane may exert its renoprotective effects by inducing, via preconditioning, an acute56, 58 and delayed59 protective effect 24–72 hours later, known as delayed protection. This protective effect includes attenuating the renal tubular apoptotic and renal dysfunction response to IRI, and is mediated by cellular signals, for example, by hypoxia-inducible factor-1α (HIF-1α). HIF-1α is a transcription factor that is expressed and activated in response to low oxygen concentrations to protect against oxygen deprivation and has been shown to confer a renoprotective effect against IRI.60 Other studies have also shown that isoflurane may protect against renal IRI via other pathways that include the release of transforming growth factor-β1 (TGF-β1), which generates adenosine, a potent cytoprotective molecule, via ecto-5′-nucleotidase (CD73),57 and sphingosine-1-phosphate (S1P) via sphingosine kinase (SK).55, 61 It is therefore likely that the renoprotective effects of isoflurane stem from some, if not all, of its downstream effectors.
It has been shown that sevoflurane may also orchestrate anti-inflammatory and anti-apoptotic effects via activation of ERK and AKT as well as a host of heat shock proteins in the HSP family.62 There is further evidence that the signaling pathway is initiated by the externalization of phosphatidylserine on the plasma membrane and the subsequent induction and release of TGF-β1.63In addition, isoflurane has been shown to be linked to the activation of the lysophospholipid S1P signaling pathway, catalyzed by the activation of the SK enzyme; S1P receptor antagonist and SK enzyme inhibitors reversed the effects of renoprotection mediated by isoflurane.55 Song et al64 found that isoflurane causes an increase in caveolae formation on the plasma membrane that contains a number of key components that include ERK and SK1, which is important in the S1P pathway, and receptors for TGF-β1 in human kidney proximal tubule (HK-2) cells.
Desflurane confers the same level of renoprotection in living kidney transplantation recipients as isoflurane and sevoflurane in a retrospective study looking at serum markers of worsening renal function; this analysis included SCr levels and estimated glomerular filtration rate (eGFR).65 The eGFR values taken in patients treated with desflurane were consistently better compared to the isoflurane or the sevoflurane group until 7 days postsurgery.54 However, in a murine model study with induced renal ischemia followed by 3-hour reperfusion with continuous inhalational anesthetic of choice—volatile (desflurane, halothane, isoflurane, or sevoflurane) versus intravenous (pentobarbital or ketamine)—the desflurane-treated group had lower plasma creatinine compared to the intravenous group, but was significantly higher compared to the other volatiles species. This finding is consistent with the histological features observed in the desflurane-treated group, where its renoprotective effects were less compared to the other volatile groups in terms of glomerular tubular swelling, dilatation, and necrosis. Overall, the data achieved from laboratory studies as mentioned are promising but clinical trials are required to elucidate if these findings can be translated to the bedside.
5.3. Intravenous anesthetics
Propofol is the most widely used intravenous anesthetic agent for induction and maintenance of general anesthesia. It is thought to principally target the GABAA receptor to potentiate its inhibitory activity66, 67, 68, 69, 70 and only recently a plausible binding site has been suggested for its action on the receptor71 to induce and maintain anesthesia.
Propofol has also been investigated for its renoprotective effects in IRI-induced AKI after several studies showed propofol can ameliorate IRI in other organs including the brain,72, 73 heart,74, 75 lungs,76 lower limbs,48 and testicles.77 Wang et al78 found that giving propofol 30 minutes before induction of renal ischemia until 30 minutes after reperfusion reduced the increase in blood urea nitrogen and SCr levels in the IRI rats. There was also histological improvement in the cortical tubules of the treated group compared to the untreated group.
The exact molecular mechanism behind the protective effect of propofol towards renal IRI is unclear, but it is thought to involve the upregulation of heme oxygenase-1 (HO-1) expression.78, 79 HO is an enzyme that catalyzes the conversion of heme to biliverdin, carbon monoxide (CO), and free ferrous iron, which may have anti-inflammatory and antioxidant effects. Biliverdin and bilirubin, from the former's conversion by biliverdin reductase, have antioxidant activity and could attenuate the oxidative stress damage induced by IRI.80 CO is often used as a signaling molecule in the body and has been shown to have anti-inflammatory and antioxidant effects too. Its anti-inflammatory effects involve the downregulation of proinflammatory cytokines production such as tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) and upregulation of anti-inflammatory cytokines via the mitogen-activated protein kinase kinase-3 (MAPKK-3)/p38 MAPK pathway.81, 82 Its antioxidant effects, by contrast, are via a pathway that involves ERK1/2, p38 MAPK, and AKT to remove free radicals.83 HO-1 is an inducible isoform of HO, and is strongly induced by various stimuli such as hypoxia, oxidative stress, heavy metals, cytokines, hormones, endotoxins, heat shock, and IRI.78 Hence, propofol may attenuate IRI by increasing the expression of HO-1 in response to the injurious stimuli.
Adenosine triphosphate-sensitive potassium (KATP) channels may also have a part in mediating the protective effect of propofol in renal IRI. Pretreatment with propofol significantly attenuates IRI-induced apoptosis of renal cells84and this has been observed in myocardial and endothelial cells also.85, 86However, the mechanism by which KATP channels contribute to this effect is still incompletely understood and warrants follow-up study. Propofol also contains a phenol hydroxyl group that has an intrinsic antioxidant effect,87either by reacting with lipid peroxyl radicals to form the stable propofol phenoxyl radical88 and/or by scavenging peroxynitrite, a highly reactive oxidant.89 Hence propofol could have a direct protective effect against oxidative stress-induced damage of renal tissue in IRI.
Propofol has also been investigated for its renoprotective effects in sepsis-mediated kidney injury. Sepsis is a systemic inflammatory response in the presence of an infection that leads to cytokine release, neutrophil recruitment, and free radical generation contributing to AKI. Lipopolysaccharide (LPS) is a common endotoxin found in Gram-negative bacteria that stimulates an inflammatory response in the kidney via the nuclear factor-κB (NF-κB) pathway and TNF-α release, an important mediator of inflammation.90 Hsing et al87 found that the inflammatory response in LPS-induced kidney injury is characterized by increased expression of proinflammatory mediators, adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1), and pro-apoptotic molecules such as caspase-3 and Bcl-2-associated X protein (Bax). Hence inhibiting NF-κB activation and TNF-α release could have a protective effect in sepsis-mediated AKI.
In murine models of sepsis, preconditioning with propofol has been shown to reduce TNF-α and ICAM-1 expression, neutrophil infiltration of the renal cortex, and necrotic and apoptotic renal cell death with significantly reduced levels of caspase-3 and Bax but increased expression of Bcl-2, an anti-apoptotic protein.87, 91 Propofol also decreased free radical production in a dose-dependent manner and conferred a protective effect against oxidative stress-induced kidney injury due to its antioxidant effects.87, 89, 92 These effects of propofol are thought to be mediated in part by inhibiting the NF-κB pathway87, 93 and by increasing bone morphogenetic protein-7 (BMP-7) expression levels in the kidneys.87 BMP-7 is important for nephrogenesis during development and also helps maintain renal architecture and function.87 In immunoglobulin A (IgA)-induced nephropathy, BMP-7 has been shown to repress TNF-α-mediated inflammation and protect the kidney from injury by inhibiting NF-κB activity and translocation into the nucleus to prevent transcription of proinflammatory genes.94 Hence, propofol may increase BMP-7 levels to inhibit the NF-κB pathway repressing TNF-α-mediated inflammation.
Thiopental was used often for the induction of anesthesia. It is a barbiturate and hence binds to GABAA receptors to prolong its postsynaptic inhibitory effect in the central nervous system. Thiopental is another intravenous anesthetic whose potential renoprotective effects have been investigated albeit less extensively in renal IRI models. Wistar rats given thiopental at subanesthetic doses 15 minutes before the reperfusion phase had less renal damage compared to the control group in terms of significantly reduced levels of malondialdehyde (MDA), a marker of oxidative stress, and less severe histopathological damage.95 Additionally, this effect appeared to be enhanced when thiopental was given at clinically high doses.96 Thiopental has antioxidant and cytoprotective properties demonstrated by its ability to inhibit lipid peroxidation, and hypoxic-induced and glucose-deprivation cell death.97 Thiopental has also been shown to significantly suppress neutrophil functions such as chemotaxis, phagocytosis, and free radical production at clinically relevant concentrations.98 Hence thiopental may exert its renoprotective effects, at least in part, by reducing the free radical-induced damage of the kidney during IRI.
5.4. Anesthetic gas xenon
Xenon is a noble gas that has an inert chemical and toxicological profile. Studies have demonstrated that xenon provides organoprotective properties in acute injuries of the brain, heart, and kidneys.99, 100 Rats that were treated with gentamicin with 70% xenon administration showed better renal function with protected SCr and blood urea nitrogen in comparison to the gentamicin-only treatment group after days of intermittent administration. Morphologically, Jia et al101 demonstrated that less structural damage and signs of apoptosis were seen in the xenon-treated group against gentamicin-induced nephrotoxicity in male Wistar rats. The attenuation of gentamicin-induced nephrotoxicity by xenon has been suggested to be related to its antioxidative effects, because MDA was significantly lower in the xenon-treated mice compared with control.101 However, the lack of difference in the TNF-α concentration in the serum between the experimental groups suggests that xenon may not have a role to play in the reduction of gentamicin-induced inflammation. Measurements of HIF-2α and vascular endothelial growth factor (VEGF) showed significant increases 4 days post-last administration of gentamicin in the xenon-treated group compared to the controls, but this difference was not detected prior to this period. Although this study did not report any changes in HIF-1α expression, Ma et al102 demonstrated that 70% xenon preconditioning of adult rat kidney for 2 hours induced upregulation of HIF-1α at 2–24 hours postexposure, which caused a substantial increase in erythropoietin (EPO) and VEGF. This was demonstrated in HK-2 cells with similar results.
Xenon might also have a role in transplantation in ameliorating the results of kidney injury from warm and cold ischemia–reperfusion,101, 103 principally by improving tubular cell proliferation and dampening inflammatory-associated cell apoptosis.104 Bcl-2 is able to enhance the survival of HK-2 cells,103, 105 and thus limit the oxidative damage in mitochondria and the nuclei. Zhao et al103 showed that xenon can attenuate the effects of renal ischemia in renal grafts ex vivo by concomitant enhancement of Bcl-2 and HSP-70 in western blot analysis, and reduction in cytochrome c and apoptosis-inducing factor (AIF) by mitochondrial sequestration, therefore limiting the potential for damage in the tubular model. Zhao and colleagues103 also showed in a separate paper that 70% xenon dampens the immune response in HK-2 cells by inhibiting the translocation process of HMGB-1 by nuclei sequestration. This was further evidenced in the study of tubular renal cells in a Lewis model, where reduced translocation of HMGB-1 into the cytosol resulted in reduced downstream effectors of TLR-4 and NF-κB expressions in vivo.103 Additionally, xenon has a role in attenuating the effects of hypothermic storage conditions, which can induce IRIs in renal grafts.106 Overall, these changes confer renoprotective properties by reducing proinflammatory mediators and suggest that xenon could be useful in transplant settings for improved clinical outcome.
6. Clinical applications
Broadly speaking, most studies investigating the role of anesthetic agents in renoprotection have been in vitro and in vivo studies (Table 2). However, human studies have been performed investigating the role of dexmedetomidine. Leino et al46 conducted a double-blind RCT involving 87 patients with normal renal function scheduled for CABG, randomized to either placebo or a dexmedetomidine infusion. The trial found that intravenous dexmedetomidine did not influence renal function but increased urinary output by 74%. Ji et al49 performed a retrospective analysis of cardiac surgery patients, some of whom (n = 568) had received dexmedetomidine postoperatively in the ICU. There was a greater incidence of postoperative renal failure in patients who received dexmedetomidine, however this was attributed to a greater incidence of pre-existing, preoperative renal failure in the dexmedetomidine group. Thus clinical results are equivocal and further clinical trials are required to elucidate the clinical implications of the use of such anesthetic agents in renoprotection.
Currently, AKI is associated with high mortality during the perioperative period. Selective use of certain anesthetics may confer significant benefit to patients, including better postischemic renal function following surgical procedures in which renal ischemia can be inevitable. However, our understanding of the role of anesthetics in renoprotection is still in its infancy and much work, especially within clinical trials, is required to elucidate the translational value from the bench to the bedside.
This work was supported by grants from the Medical Research Council, London, UK and the European Society of Anesthesiology, Brussels, Belgium.