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Diagnostic Utility of Serial Microscopic Examination of the Urinary Sediment in Acute Kidney Injury
Vipin Varghese, Maria Soledad Rivera, Ali A. Alalwan, Ayman M. Alghamdi, Manuel E. Gonzalez and Juan Carlos Q. Velez
Kidney360. 2021 Feb; 2(2):182-191. Doi: 10.34067/KID.0004022020
Introduction
Acute Kidney Injury (AKI) is a common and costly outcome that occurs in 10-15% of hospitalized individuals and at least 50% of critically Ill patients (Ronco et al, Lancet, 2020). Of the many causes of AKI, Acute Tubular Injury (ATI) is among the most common (Liano et al, Kidney Int 1996). ATI (previously known as acute tubular necrosis) is a pattern of tubular cell damage and/or death that occurs from:
Ischemia
Toxin induced (e.g. heme, myoglobin, aminoglycoside, amphotericin, calcineurin inhibitors)
Sepsis
Once an insult is present, a drop in blood flow occurs which leads to an extension phase that includes ongoing hypoxia (from reduced blood flow) and/or an inflammatory response. A maintenance phase follows in which cellular repair, apoptosis, migration, and proliferation occurs thus setting up the kidney for a recovery phase, where tubular cellular polarity is reestablished and kidney function improves (Kunderant et al, Kidney Res Clin Prac 2018) Diagnosis typically relies on history, physical, and urine studies (classically, FENa >2%). However urine microscopy can clearly provide a diagnosis of ATI demonstrating renal tubular epithelial cells (RTEC), RTEC casts, granular casts, or muddy brown casts.
Figure B1. A-C: Muddy Brown Casts. D: Sternheimer-Malbin (SM) stained Granular Cast. Photo courtesy of Dr. Juan Carlos Velez’s 2019 Renal Fellow Network Post.
In an ideal world, the assessment of an individual with AKI, would include a diagnostic test that would both provide an etiology of the kidney injury as well as a prognostic outlook (i.e. initiation of dialysis or mortality). Serum creatinine (SCr) remains the most common marker for assessing severity of AKI, however it fails to provide any additional information on either diagnosis or prognosis. Efforts have been made to find the “renal troponin” that may allow for prognostication of progression of AKI, need for dialysis, or mortality. However there are significant limitations including cost, availability, and lack of large-scale trials that hinder the utility of known biomarkers (Kashani et al, Clin Chem Lab Med, 2016). Additionally, biomarkers provide no information as to the etiology of AKI (Perazella, AJKD 2015).
Fig B2. Urinary Biomarkers of Acute Kidney Injury and their location in the Nephron from Perazella, AJKD © National Kidney Foundation.
Enter urine microscopy: A widely available and relatively cheap test that can help make a diagnosis! Urine microscopy has been around since the mid-1800s and has long been used to determine the etiology of AKI (Perazella, AJKD 2015). Where novel biomarkers may be able to provide information regarding prognostication, examination of the urine sediment by a trained provider can yield the etiology of AKI (glomerulonephritis, ATI, crystal nephropathy, etc).
But what if urine microscopy could go one step further and help prognosis? This seems like a pretty tall task--especially given that, despite its usefulness and widespread availability, urine microscopy remains underutilized, possibly due to lack of training and significant interobserver variability in interpretation of results (Palssan et al, JNO 2020). So how do we approach correcting these deficiencies with such an available and capable test? One way, as demonstrated by both Perazella et al and Chawla et. al was to devise a standardized scoring method. And both studies successfully correlated a standardized score with progression of AKI.
However, there are many influencing factors that go into the interpretation of a diagnostic test (Charney et al, Diagnosis, 2019). Despite being fairly sensitive and specific for the distinguishing ATI from prerenal AKI (0.76 and 0.86 respectively for the Perazella Score), one of the limitations of the original Perazella and Chawla studies was that they based their findings on a single urine specimen, which may miss changes in the urine sediment over the course of the disease. The current study hypothesizes that serial examination of urine sediment using both the Perazella Score (PS) and the Chawla Score (CS) may provide additional diagnostic and prognostic information.
The Study
Methods
Design: Prospective, single center (Ochsner Medical Center)
Population: Urine samples were taken from hospitalized individuals with:
KDIGO Stage >2 AKI (Cr >2x baseline with < 0.5 mL/kg/hr urine output for >12 hours)
Seen by the Inpatient Nephrology Consultation Service.
Suspected intrinsic etiology of AKI.
Procedures:
1. Preparation of Urine Samples:
2. Evaluation of urine sediment occurred at three time intervals
3. At each time interval, urine cast scores (PS or CS) were assigned to each urine specimen.
A PS of 2-4 or a CS of 3-4 were considered diagnostic of ATI
4. Exposure variables that could influence conversion of the cast score from non-ATI to ATI were noted. These factors included:
Non-recovering AKI was defined as an increase in serum creatinine >0.1 mg/dL at the time of the second or third microscopy compared to the first.
Timing of AKI (Early: Day 1-6, Late: > Day 7)
Presumptive etiology of AKI based on history
Ischemic ATI
Toxic ATI
Acute GN
Prerenal
Hepatorenal
Outcomes
Odds Ratio and + Likelihood Ratio of conversion of conversion of Urine Sediment from Non-ATI to ATI by either Perazella Score or Chawla Score in terms of non-recovering AKI and timing of AKI
Use of a Serial Urinary Cast Scores as predictors of poor renal outcomes (initiation of RRT during AKI or >50% raise in serum Creatinine at discharge)
Statistics Simplified
Luckily, the statistics in this paper do not go terribly far beyond what I learned in my medical school basic biostatistics course, but just as a refresher, let’s talk briefly about the odds ratio (OR) and positive likelihood ratio (+LR).
The OR is a measure used to demonstrate an exposure and an outcome (i.e. what are the odds that something will occur with an exposure compared to without). Typically an OR > 1 implies that the odds of an event occurring are higher with the exposure, while the opposite is true with an OR < 1.
Likelihood ratios, on the other hand, are more of a measure of diagnostic accuracy. Ultimately, they determine whether the test result changes the probability that the disease state exists in patients with versus without a disease.
Funding
This paper was supported by the Ochsner Health System.
Results
Overall 497 microscopic examinations of urinary sediment were performed in 343 patients with AKI during the study out of which serial inspections were not performed in 222 patients as shown in the consort diagram below (Figure 3):
Figure 3. CONSORT diagram - the traditional figure 1
Out of 343 patients, the 222 excluded patients were those who did not receive serial microscopy due to anuria, discharge, death, commencement of hemodialysis, or logistic difficulties in collecting urine samples. The 121 patients with AKI had at least two serial microscopic examinations. Among them, 105 (86.7%) had a microscopic examination performed during the second time interval (days 2-3 after consultation), and 49 (40.4%) had microscopic examination performed during the third time interval (days 4–10 after consultation). Three serial examinations were performed on 33 (27%) patients.
The median age of the patients was 61(25-88)years, and were mainly white (64%) with over representation of male gender 64%(n=77). Non white race comprised 38% of the cohort. The mean serum creatinine at the time of initial urine microscopy was 3.36 ±1.9 mg/dl with 80% in stage 3 AKI. Around a third of cases had pre-existing CKD stage IIIa-V.
Based on the microscopic examination of the urinary sediment, ischemic ATI was the presumed etiology in 59% (n=71) of patients out of which 38% (n=46) needed dialysis during the course of the AKI (Table1).
Table1. Baseline characteristics of patients(n=121) in the cohort undergoing serial urine sediment microscopic exam
On day 1, the CS and PS scores revealed 64 (53% ) and 70 (58%) of patients respectively with non-ATI out of which 14 (22%) and 16 (23%) converted to ATI on second and third microscopy. Thus upon serial microscopy examinations,there was evidence of ATI by both scoring methods(20% by CS and 24% by PS as seen in Figure 4.
Figure 4: Serial urine microscopy identifies conversion from non-ATI to ATI cast scores
Over time, the microscopic examination of urinary sediment revealed ATI specimens throughout the 10 day period of consultation for AKI with highest number of patients within the 4-6th day of AKI interval (Figure 5)
Figure 5: Impact of timing of urine microscopy on probability of detecting ATI cast score
For patients with non recovering AKI, the PS changed from non-ATI to ATI (OR 5.8;95% CI ,1.7-19.3, p=0.005, with +LR , 2.0; 95%1.4-2.9). However in terms of influence of timing, there was no significant difference in rate of change in ATI category when performed early (<7 days ) versus at a later stage (>7 days) with PS having OR 1.0; 95% CI , 0.4-2.8, p=1.0, +LR 1.0; 95% CI, 0.7-1.5 and CS having OR 0.8;95%CI , 0.6-2.3, P=0.7, +LR 0.9; 95%CI,0.6-1.4 (Figure 6)
Figure 6: Predictive value of AKI course and timing for conversion to ATI cast score
Comparing the prognostic value of single versus serial microscopic inspections, the urine cast scores on initial microscopy did not predict AKI or AKI-RRT. On serial microscopy, for PS, OR decreased to 1.5 which was statistically non significant. For CS, OR and +LR increased mathematically (statistically not significant). With CS in the non-ESLD group, at initial microscopic examination, although the OR was 2.3 (95% CI 0.8-6.4, P=0.13) with the +LR being 1.4 (95% CI 0.9-2.2), however after serial microscopy the OR was reinforced to 3.3 (95% CI 1.0-10.3, p=0.04) with similar value of +LR at 1.4 (95% CI 1.0-2.0) (Figure7)
Figure 7: AKI prognosis assessed by single and serial urine microscopy
Continuing with the subgroup analysis, the cohort of patients with non recovering AKI ( n=66),when analysed for prognostication of AKD or AKI -RRT , on the initial microscopy PS had OR of 1.3;95% CI,0.4-4.9,p=0.66 with +LR 1.2;95% CI 0.6-2.3.
For CS the OR was 1.9;95%CI ,0.5-7.2,p=0.67 with +LR at 1.4;95%CI ,0.7-2.7. Serial microscopies revealed mathematical decrease in OR and +LR for PS and increase for CS data but was statistically non significant (Figure 8)
Figure 8. Utility of single versus serial exams
Discussion
Diagnostic relevance
This study by Verghese et al assessed the utility of serial urine examinations. They found 20%–25% of patients had ATI that was only revealed on serial exams. It was further observed that patients with non recovering AKI were likely to convert from non ATI to ATI category Figure 6. Performing serial urinary sediment microscopy may offer a specific diagnosis in cases where patients exhibit a stagnant or worsening clinical course.
The natural history of the urinary cast is poorly understood. Added to that is uncertainty around the best timing of the test which may affect the operating nephrologist behind the microscope to capture early changes in AKI. This report has highlighted the relationship between urine microscopy findings with time related changes in kidney due to injury showing a wide distribution of patients with ATI along with varied duration of AKI (Figure 5). Based on this observation, we should be encouraged to perform inspection of urine specimens at any time during the course of AKI and do serial examinations along the way. The evidence of ATI microscopically was seen mostly around 4-6 days from onset of AKI but this observation may just be due to higher number of examinations done coinciding with the timing of inpatient nephrology consultations.Another aspect worth highlighting based on the observational finding in this report was a small subgroup of patients who had evidence of ATI on initial microscopy but regressed to non ATI on subsequent serial examination(Figure 4). This shows that a bland sediment late in the course of AKI could be due to ATI. Having a high clinical suspicion during interpreting these findings can surely help risk stratification for meaningful outcomes in cases of AKI (Isaac et al.CJASN2011)
Prognostic relevance
Urine microscopic examination has been used for prognostication of renal failure in multiple studies quantifying cast per low power field Marcussen et al,Renal Failure 1995 or using the Addis method Addis T 1926 to actually developing a scoring system Chawla et al 2008 and Perazella et al 2008 CJASN in which higher number of casts/score predict more severe AKI and eventual need for RRT. Indeed Perazella et al accurately predicted the likelihood of final diagnosis of ATI using the pretest probability and number of RTECs and granular casts on initial microscopy.
Mark Perazella AJKD 2015 highlights the complementary nature of these urinary sediments with other AKI biomarkers e.g NGAL in predicting worse renal outcomes (Table D1 below) and also the pretty picture of cellular biomarkers by site of injury see Figure B)
Table 2. Urine Sediment Examination as Biomarker for Prognosis in AKI AJKD 2015
The study by Verghese et al represents a cohort with overrepresentation of ESLD with hepatorenal syndrome (HRS) type 1 patients compared to previous studies. Also the association of ATI scores with AKI or AKI-RRT based on urinary sediment microscopy in this study is not very robust (Figure 7). The logic behind this is that HRS1 progresses to anuric AKI demanding dialysis often leaving no trace of ATI on urine microscopy as well pointed out by Velez et al in Nat Review Nephrol2020 making ATI scores less of prognostic predictors in the cirrhotics. Actually these scores can be deceptive in this subgroup due to the presence of elevated bilirubin in the urine along with RTEC and RTECC which may not represent AKI all the time.
Figure 9. Bilirubin/Bile casts (containing RTECs – black arrows). Fresh and unstained urine sediment. Original magnification 400x. Bright field microscopy. From a RFN post by José Poloni.
Coming back to the study, the data was reanalysed by using a non ESLD group and a significant improvement in prognostic value of ATI score was seen (Figure 7). However the magnitude of OR was less than seen in Perazzela et al and Bagshaw et al due to more severe AKI included in the later studies as indicated by higher urine microscopy scores.
Limitations
There is operator dependency for interpretation of urine microscopy examinations and scoring of each specimen. A study of nephrologists found significant interobserver variability in the interpretation of urine sediment findings, even among experts.
As one out of the two operators was unblinded, there could be a possibility of observer bias.
There is heterogeneity in the results depending on the urine cast score used, namely PS versus CS.
This study has a higher percentage of intrinsic causes of AKI cases and hence need to be cautiously generalised to other AKI cohorts that are predominantly prerenal in etiology.
Lastly there has been no adjustment for confounding factors like the urine output although the authors accounted for serum creatinine and duration of AKI.
In summary, this study highlights the microscopic urine sediment examination to be a useful clinical tool for the practicing nephrologist guiding further therapeutic interventions. The study demonstrates that serial inspections may yield valuable clinical information compared to single exam at one point in time. This is perhaps more pertinent in patients with non recovering AKI.
If you are hungry for more urine sediment examination tips and examples, do check out some of these wonderful tags and sites! ( Renal Fellow Network’s Monthly Urine Sediment of the Month,#urinemicroscopy, #urinesediment, #urinalysis, #spinurine #FOAMed,#LiquidBiopsy #NephMadness #PisseProphets)
