Study on the concentration of urinary neutrophil gelatinase associated lipocalin in patients occuring acute kidney failure in intensive care unit

Objective: To evaluate urinary neutrophil gelatinase-associated lipocalin (NGAL) concentration and its relation with causes, categories, stages and biochemical indexes of acute kidney injury (AKI) patients. Subjects and methods: A prospective, cross-sectional study in 121 patients with AKI who admitted to a general Intensive Care Unit (ICU), Trungvuong Hospital, Hochiminh City from 12 - 2013 to 01 - 2017 and a control group of 51 healthy people. Urinary NGAL had done in all 116 patients and healthy people.

Journal of military pharmaco-medicine no7-2017 STUDY ON THE CONCENTRATION OF URINARY NEUTROPHIL GELATINASE - ASSOCIATED LIPOCALIN IN PATIENTS OCCURING ACUTE KIDNEY FAILURE IN INTENSIVE CARE UNIT Pham Ngoc Huy Tuan*; Nguyen Trung Kien**; Le Viet Thang** SUMMARY Objective: To evaluate urinary neutrophil gelatinase-associated lipocalin (NGAL) concentration and its relation with causes, categories, stages and biochemical indexes of acute kidney injury (AKI) patients Subjects and methods: A prospective, cross-sectional study in 121 patients with AKI who admitted to a general Intensive Care Unit (ICU), Trungvuong Hospital, Hochiminh City from 12 - 2013 to 01 - 2017 and a control group of 51 healthy people Urinary NGAL had done in all 116 patients and healthy people Results: All of the AKI patients (100%) had urinary NGAL elevation The average concentration of urinary NGAL in our study group (434.06 ng/mL) was significantly higher than in control group (10.74 ng/mL) with p < 0.001 There was no significant difference in concentration of uNGAL in terms of causes The concentration of urinary NGAL was significantly higher in oliguria group than non-oliguria group (571.70 ng/mL compared to 355.95 ng/mL) with p < 0.005 Patients’ uNGAL concentration at the time of ICU admission was significantly related to their KDIGO stage (p < 0.001) Urinary NGAL had a moderate positive correlation with serum urea concentration (r = 0.567, p < 0.001) and a strong positive linear correlation with serum creatinine concentration (r = 0.850, p < 0.001) Conclusion: Urinary NGAL elevation was common in AKI patients The concentration of urinary NGAL depended on category and stage of AKI It had a moderate positive correlation with serum urea and strong positive correlation with creatinine concentration * Keywords: Acute kidney injury; Urinary neutrophil gelatinase-associated lipocalin; Intensive care unit INTRODUCTION Acute kidney injury is a common and devastating problem with in-hospital mortality of 40% to 80% in the intensive care setting [10] The traditional blood (creatinine, blood urea nitrogen) and urine markers of kidney injury (casts, fractional excretion of sodium, urinary concentrating ability) that have been used for decades in clinical studies in diagnosis and prognosis of AKI are insensitive and nonspecific and not directly reflect injury to kidney cells Therefore, early recognition of renal injury is important and may help prevent further renal damage and functional impairment Neutrophil gelatinase-associated lipocalin is a small, 23 kDa protein that is an early biomarker for ischemic, septic, or nephrotoxic kidney injury It is normally * 103 Military Hospital ** Trung Vuong Hospital Corresponding author: Pham Ngoc Huy Tuan (bshuytuantv@yahoo.com.vn) Date received: 03/07/2017 Date accepted: 18/07/2017 71 Journal of military pharmaco-medicine No7-2017 produced at low levels by the epithelial 2017 and a control group of 51 healthy cells of the kidney, but it is quickly people upregulated in the thick ascending limb * Excluding criteria: patients with (TAL) of the loop of Henle and the chronic kidney failure, anuria, did not fit collecting ducts within three hours of with diagnostic criteria, did not have tubular epithelial injury enough test results or did not agree to Urinary NGAL (uNGAL) has been evaluated as an early biomarker of renal tubular damage in a acute clinical settings such as the operating room, ICU and emergency department, and in high-risk procedures such as cardiac surgery, radio-contrast injection and after adult and pediatric kidney and liver transplantation [1, 6, 7, 8, 9] There is considerable evidence that compared to increases in serum creatinine, NGAL better detects early or subclinical kidney injury, and better predicts dialysis requirement and mortality [1] In Vietnam, there are lack of studies on the role of uNGAL in AKI diagnosis and prognogsis in patients who admitted to general ICU Therefore, we conducted this research for the aim: Evaluation of participate in the study Methods * Study design: A prospective, crosssectional descriptive study * Urinary NGAL measurement: 24- hour urine was collected After that, the volume of urine was measured before collecting mL sample for testing purpose uNGAL was measured by the sandwich ELISA method using NGAL monoclonal antibody in the NGAL kit After that, the sample will be analyzed by Achitech System of Abbott, America to measure uNGAL concentration * Diagnostic criteria: KDIGO definition and classification of AKI [5] * Diagnostic criteria for AKI: serum creatinine ≥ 136.5 µmol/L within 48h the uNGAL concentration and its relation * AKI degree: with causes, categories, stages and some + AKI stage I: serum creatinine from biochemical indexes of AKI patients SUBJECTS AND METHODS Subjects The study was conducted with a study 136.5 - 220 µmol/L + AKI stage II: serum creatinine from 220 - 353.6 µmol/L + AKI stage III: ≥ 353.6 µmol/L group of 121 AKI patients who admitted to * Statistical analysis: a general ICU, Trungvuong Hospital, Statistical analyses were conducted Hochiminh City from 12 - 2013 to 01 72 using SPSS 20.0 Journal of military pharmaco-medicine no7-2017 RESULTS AND DISCUSSIONS Table 1: Urinary NGAL concentration in study group Index Urinary NGAL (ng/mL) Control group (n = 51) Study group (n = 116) p X ± SD 10.74 ± 5.18 434.06 ± 333.14 < 0.001 Max 20.28 1292.38 Min 3.32 69.63 The average concentration of urinary NGAL in our study group was 434.06 ng/mL, which was significantly higher than in control group (10.74 ng/mL) with p < 0.001 The maximum and minimum concentration of urianay NGAL was 1292.38 and 69.63 ng/mL respectively With the refference range of urinary NGAL from 43.62 to 114.66 ng/mL, all of the AKI patients (100%) had urinary NGAL elevation Study of Au V also showed that the mean immediate postoperative urinary NGAL levels in patients who developed sustained AKI were 204.8 ng/mL, and significantly higher than those who had normal renal function (31.9 ± 113 ng/mL) with p < 0.001 [1] This result was similar to other studies of Geus H.R, Makris K, Zappitelli M: there was a significant higher of urinary NGAL concentration in patients who submitted AKI compare to non-AKI patients with p < 0.05 [6, 7, 8] These differences in uNGAL concentration are expected because kidney injury associated with primary renal insults may be more severe than that in most patients included in our study But our patients were probably more severely ill, with a higher proportion having sepsis than healthy people In current clinical practice, the gold standard for identification and classification of AKI is dependent on serial serum creatinine measurements, which are especially unreliable during acute changes in kidney function We identified uNGAL as one of the most upregulated genes in the kidney soon after ischemic injury NGAL protein was also markedly induced in kidney tubule cells and easily detected in the plasma and urine in animal models of ischemic and nephrotoxic AKI The expression of uNGAL protein was also dramatically increased in kidney tubules of humans with ischemic, septic, and posttransplant AKI Importantly, NGAL in the urine was found to be an early predictive biomarker of AKI in a variety of acute clinical settings Emerging experimental and clinical evidence indicated that in the early phases of AKI from diverse etiologies, NGAL accumulates within two distinct pools, namely, a renal and a systemic pool Gene expression studies in AKI had clearly demonstrated rapid and massive upregulation of NGAL mRNA in the thick ascending limb of Henle's loop and the collecting ducts, with resultant synthesis of NGAL protein in the distal nephron (the renal pool) and secretion into the urine where it comprises the major fraction of urinary NGAL This finding also confirms the need for future research to evaluate uNGAL in different renal disease subgroups, in order to understand fully how best to use uNGAL to diagnose AKI 73 Journal of military pharmaco-medicine No7-2017 Table 2: Relation between urinary NGAL concentration and the causes of AKI Causes n % Serum NGAL (ng/mL) Sepsis (1) 67 57.8 447.28 ± 321.45 Dehydration (2) 18 15.5 341.75 ± 312.55 Cirrhosis (3) 6.0 690.17 ± 467.48 Shock (4) 11 9.5 451.19 ± 339.97 Toxic (5) 3.45 187.48 ± 98.69 Urinary obstruction (4) 4.3 259.29 ± 285.91 Others (7) 3.45 597.91 ± 330.90 p > 0.05 In our study, sepsis was the most common causes with the proportion of 57.8% There was no significant difference between these causes with p > 0.05 Our result was similar to study of Vaidya D.S: there was no significant difference between urinary NGAL concentration and several causes of AKI in these studies (p > 0.05) [10], but was different with other studies of Di Nardo M and Geus H.R (there was a significant higher concentration of urinary NGAL in septic AKI patients than nonseptic AKI patients with p < 0.001 [4, 6] Lipoproteins also have strong affinity for Toll-like receptors (TLRs) that trigger an innate immune response Therefore, it could be postulated that these circulating ligands which are linked to tubular epithelial TLR activation are responsible for the increased uNGAL concentrations that we observed in patients who had sepsis, but showed no increases in their serum creatinine levels However, recent studies in patients with sepsis, septic shock, and systemic inflammatory response syndrome had reported contradictory findings A possible explanation for this difference is the variability of the subject inclusion time (up to 48 h after ICU admission) Intensive resuscitation and the administration of antibiotics may have already occurred before study inclusion, therefore most likely inducing rapid changes of uNGAL values Table 3: Relation between urinary NGAL concentration and AKI category Categories n % Urinary NGAL (ng/mL) Non-anuria 74 63.8 355.95 ± 302.05 Anuria 42 36.2 571.70 ± 344.14 p < 0.005 In our study, category of anuria occupied in 36.2% of all AKI patients The concentration of urinary NGAL was significantly higher in anuria group than non-anuria group (571.70 ng/mL compared with 355.95 ng/mL) with p < 0.005 Our findings highlight the mechanistic insights of NGAL levels based on the specimens being measured 74 Journal of military pharmaco-medicine no7-2017 Urine NGAL is proposed to derive predominantly from local renal synthesis of NGAL in the thick ascending limb of the loop of Henle and the collecting ducts when under inflammatory and oxidative stress Therefore, the concentration of urinary NGAL was directly related to the renal tubule injury in AKI patients as well as urine excretion ability Table 4: Relation between serum NGAL concentration and stage of AKI AKI stages (KDIGO) n % Urinary NGAL (ng/mL) 81 69.8 244.07 ± 156.79 26 22.4 818.01 ± 153.01 7.8 1034.83 ± 160.49 < 0.001 p1-2, p1-3 < 0.001, p2 -3 < 0.005 pANOVA Following to the KDIGO classification, the stage AKI in our study made up the highest proportion (69.8%) Stage and occupied smaller proportion (22.4% and 7.8%, respectively) Our results also pointed that patients’ uNGAL concentrations at the time of ICU admission were significantly related to their KDIGO stage (p < 0.001) This result was similar to the study of Geus H.R (p < 0.0001) and Zapittelli M (p < 0.0002) when research on the relation between uNGAL and RIFFLE stage [6, 8] NGAL fulfills a central role in regulating epithelial neogenesis, and in iron chelation and delivery after ischemic or toxic insults to the renal tubular epithelium After kidney injury, NGAL is rapidly expressed on the apical epithelial membranes of the distal nephron NGAL is excreted in the urine through exocytosis and has local bacteriostatic and proapoptotic effects Therefore, uNGAL concentration had a positive relation with the level of renal damage which exhibited throughout the high stage of KDIGO classification Table 5: Correlation between serum NGAL and urea, creatinine concentration Indexes Serum NGAL Correlation equation r p Urea 0.567 < 0.001 uNGAL = 18,89*urea + 165.63 Creatinine 0.850 < 0.001 uNGAL = 2.63*creatinine - 142.30 In our study, urinary NGAL had a moderate positive correlation with serum urea concentration (r = 0.567, p < 0.001) and a strong positive linear correlation with serum creatinine concentration (r = 0.850, p < 0.001) The correlation equation was created (uNGAL = 18,89*Urea + 165.63; uNGAL = 2.63*creatinine 142.30) Boglignano D also pointed that a significant correlation was also found between serum creatinine and uNGAL (r = 0.399, p < 0.001) [2] NGAL has mainly been studied in the setting of acute renal failure Patients who experienced 75 Journal of military pharmaco-medicine No7-2017 acute renal dysfunction showed a marked increase in urinary NGAL levels, which preceded the increase in serum creatinine by a day In a single case of acute tubular necrosis due to heart failure induced hypotension, NGAL tubular expression was reported to be strongly increased [3] Hence, measurements of NGAL may serve as a very early marker of worsening renal function Urinary (or plasma) NGAL levels could therefore be used to adjust therapy, to anticipate and possibly prevent expected renal injury, even before a peak in serum creatinine occurs This potential of NGAL needs to be explored further in future studies uNGAL uNGAL = 18.89*Urea + 165.63 2000 1500 1000 500 0 20 40 Urea 60 80 Chart 1: Correlation between urinary NGAL and urea concentration uNGAL uNGAL = 2,63*creatinine - 142.30 2500 2000 1500 1000 500 0 200 400 600 800 1000 Creatinine Chart 2: Correlation between urinary NGAL and creatinine concentration CONCLUSIONS In our study, all of the AKI patients (100%) had urinary NGAL elevation The average concentration of urinary NGAL in our study group (434.06 ng/mL) was significantly higher than that in control group (10.74 ng/mL) with p < 0.001 There was no significant difference in concentration of uNGAL in terms of causes The concentration of urinary NGAL was significantly higher in oliguria 76 group compared to non-oliguria group (571.70 ng/mL compared to 355.95 ng/mL) with p < 0.005 Patients’ uNGAL concentrations at the time of ICU admission were significantly related to their KDIGO stage (p < 0.001) Urinary NGAL had a moderate positive correlation with serum urea concentration (r = 0.567, p < 0.001) and a strong positive linear correlation with serum creatinine concentration (r = 0.850, p < 0.001) Journal of military pharmaco-medicine no7-2017 REFFERENCES Au V et al Urinary neutrophil gelatinaseassociated lipocalin distinguishes sustained from transient acute kidney injury after general surgery KI reports 2016, (1), pp.39 Bolignano D.et al Neutrophil gelatinaseassociated lipocalin as a marker of kidney damage American Journal of Kidney Diseases 2008, 52 (3), pp.595-605 Damman K et al Urinary neutrophil gelatinase associated lipocalin (NGAL), a marker of tubular damage, is increased in patients with chronic heart failure European Journal of Heart Failure 2008, 10 (10), pp.997-1000 Geus H R H D et al Neutrophil gelatinase-associated lipocalin at ICU admission predicts for acute kidney injury in adult patients American Journal of Respiratory and Critical Care Medicine 2011, 183 (7), pp.907-914 Makris K et al Urinary neutrophil gelatinase-associated lipocalin as an early marker of acute kidney injury in critically ill multiple trauma patients Clinical Chemistry and Laboratory Medicine 2009, p.79 Zappitelli M et al Urine neutrophil gelatinase-associated lipocalin is an early marker of acute kidney injury in critically ill children: a prospective cohort study Critical Care 2007, 11 (4), p.R84 Di Nardo M et al Impact of severe sepsis on serum and urinary biomarkers of acute kidney injury in critically Ill children: An observational study Blood Purification 2013, 35 (1-3), pp.172-176 Chertow G.M et al Acute kidney injury, mortality, length of stay, and costs in hospitalized patients J Am Soc Nephrol 2005, 16 (11), pp.3365-3370 Disease, K Improving global outcomes (KDIGO) acute kidney injury work group: KDIGO clinical practice guideline for acute kidney injury Kidney Int Suppl 2012, 2, pp.1-138 10 Vaidya V.S et al Urinary biomarkers for sensitive and specific detection of acute kidney injury in humans Clin Transl Sci 2008, (3), pp.200-208 77 ... urinary NGAL and creatinine concentration CONCLUSIONS In our study, all of the AKI patients (100%) had urinary NGAL elevation The average concentration of urinary NGAL in our study group (434.06... local renal synthesis of NGAL in the thick ascending limb of the loop of Henle and the collecting ducts when under inflammatory and oxidative stress Therefore, the concentration of urinary NGAL was... synthesis of NGAL protein in the distal nephron (the renal pool) and secretion into the urine where it comprises the major fraction of urinary NGAL This finding also confirms the need for future