Neutrophil gelatinase-associated lipocalin (NGAL) levels in response to unilateral renal ischaemia in a novel pilot two-kidney porcine model.

OBJECTIVES To test a novel porcine two-kidney model for evaluating the effect of controlled acute kidney injury (AKI) related to induced unilateral ischaemia on both renal units (RUs) To use neutrophil gelatinase-associated lipocalin (NGAL) and physiological serum and urinary markers to assess AKI and renal function. METHODS Twelve female Yorkshire pigs had bilateral cutaneous ureterostomies placed laparoscopically with identical duration of pneumoperitoneum for all cases. An experimental group (n =9) underwent induced unilateral renal ischaemia with left hilar clamping of timed duration (15, 30, 60 min) and a control group (n =3) had no induced renal ischaemia. Urine was collected and analysed from each RU to assess creatinine and NGAL concentration preoperatively and at multiple postoperative time points. Serum was collected and analysed daily for creatinine and NGAL levels. Statistical comparisons were made using the rank-sum and sign-rank tests. RESULTS Three pigs were excluded because of intra-operative and postoperative complications. In the RUs that experienced renal ischaemia (n=7),the median urine volume was lower (P=0.04) at 6, 12, 24 and 48 h and the median NGAL concentration was higher (P=0.04) at 12 and 48 h compared with the RUs of control pigs that experienced no renal ischaemia (n=2). When comparing the ischaemic (left) RU of the pigs in the experimental group with their contralateral non-ischaemic (right) RU, ischaemic RUs had a lower median cumulative urine volume at 6, 12, 24 and 48 h (P = 0.05) and a higher median NGAL concentration at 12, 24 and 48 h (P<0.05). At 48 h, no significant increase was found in serum NGAL in pigs in the experimental group compared with controls (P=0.2). Creatinine clearance (CC) was lower in ischaemic RUs compared with non-ischaemic RUs 1 day after surgery (P=0.04) with decreasing CC as the duration of ischaemia increased. CONCLUSIONS We have developed a promising novel small-scale pilot surgical model that allowed the evaluation of bilateral RU function separately during and after unilateral renal ischaemia. The induction of unilateral renal ischaemia corresponds with physiological changes in both the ischaemic and contralateral RU. AKI as measured by increases in NGAL and decreased renal function as measured by decreases in CC, are specific to the RU exposed to ischaemia. Keywords: human NGAL protein, ischaemia, acute kidney injury, models, animal, biomarkers INTRODUCTION Iatrogenic human renal ischaemia in the setting of partial nephrectomy (PN) occurs when a clamp is applied to the renal artery or renal hilum to reduce the bleeding that may occur when resecting a portion of one renal unit (RU). To investigate the impact of such ischaemia on renal function, experiments have been performed in solitary kidney models to reduce the impact from the contralateral RU [1–4]. The results of these experiments are informative but fail to adequately account for the compensatory mechanisms of the contralateral RU, and hence may not adequately reproduce endogenous physiological reactions to ischaemia. Acute kidney injury (AKI) is defined as a rapid decline in renal function. The markers most commonly used to measure AKI in the setting of PN are serum creatinine (SCr) alone or its derived formula, estimated GFR (eGFR). Changes in SCr, and thus eGFR, are delayed by 24–48 h from the time of AKI [5–7] and SCr level or eGFR may also be influenced by a variety of clinical factors such as hydration status or changes in filtration by the contralateral RU [8, 9]. Additionally, eGFR shows how much renal function remains after an insult, and shows only indirectly the amount of renal damage incurred. An improved marker of AKI should allow a more rapid detection of injury and, in turn, the initiation of interventions that can mitigate renal damage and the subsequent loss of renal function. Elevated neutrophil gelatinase-associated lipocalin (NGAL) has been shown to correlate with AKI and to precede a rise in SCr level in a variety of clinical settings, such as cardiac surgery [7, 10, 11], receipt of i.v. contrast [10, 12], renal transplant [10, 13, 14] and in the emergency room [15]. Because urine and serum NGAL levels are independent of SCr, they may be less influenced by clinical factors that affect eGFR. In addition, recent studies have shown than NGAL may be more sensitive to AKI and may be a better predictor of morbidity than SCr [16]. Large animal models using pigs have evaluated the role of NGAL to detect renal injury in renal transplant models [17, 18,19], but no-one has yet evaluated NGAL levels in response to induced unilateral renal ischaemia in an animal with intact bilateral kidneys. Furthermore, there is currently no documented evidence in the literature that NGAL production is specific to the side of injury or if there is a dose–response relationship between unilateral NGAL levels and the duration of renal ischaemia. We developed a novel two-kidney porcine model of AKI to evaluate independently the effects of unilateral renal ischaemia on each individual RU as well as to elucidate the utility of NGAL as a marker of AKI in a kidney surgery model.