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StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2024 Jan-.

StatPearls [Internet].

Renal Function Tests

Verena Gounden ; Harshil Bhatt ; Ishwarlal Jialal .

Authors

Affiliations

Last Update: July 27, 2024 .

Introduction

The kidneys play a vital role in excreting waste products and toxins, such as urea, creatinine, and uric acid. They also regulate extracellular fluid volume, serum osmolality, and electrolyte concentrations and produce hormones such as erythropoietin, 1,25 dihydroxy vitamin D, and renin. The functional unit of the kidney is the nephron, which consists of the glomerulus, proximal and distal tubules, and collecting duct. Assessing renal function is crucial in treating patients with kidney disease or pathologies affecting renal function. Renal function tests are useful for identifying the presence of renal disease, monitoring the response of kidneys to treatment, and determining the progression of renal disease. According to the National Institutes of Health, the overall prevalence of chronic kidney disease (CKD) is approximately 14%. Globally, the most common causes of CKD are hypertension and diabetes.[1][2][3][4] This resource provides an update on the relevant biochemical tests for assessing renal function.

Specimen Collection

Specimen collection requirements depend on the procedure or test requested. Generally, no additional patient preparation is required for serum creatinine and blood urea nitrogen (BUN) levels, and a random blood sample is sufficient. However, recent high protein ingestion may significantly increase serum creatinine and urea levels. In addition, hydration status can have a considerable impact on BUN measurement.

For timed urine collections such as the 24-hour urine creatinine clearance, accurate collection over the entire period is crucial, as under- or over-collection can significantly affect the results. Hence, a 5- to 8-hour timed collection may be preferable to a 24-hour collection, especially outside a hospital setting.[5][6][7]

Collecting midstream urine for urine analysis is preferred as this sample is less likely to be contaminated by epithelial cells and commensal bacteria.

Procedures

Assessment of Renal Function

Several clinical laboratory tests help investigate and evaluate kidney function. Clinically, the most practical tests for assessing renal function are those that estimate the glomerular filtration rate (eGFR) and quantify proteinuria (albuminuria).

Glomerular Filtration Rate

The best overall indicator of the glomerular function is the glomerular filtration rate (GFR). GFR is the rate in milliliters per minute at which substances in plasma are filtered through the glomerulus; in other words, the clearance of a substance from the blood. The normal GFR for an adult male is 90 to 120 mL/min. However, this number varies significantly by age. Some studies suggest a decrease of 7.5 mL/min/1.73m 2 after 30 years due to aging processes. Therefore, an otherwise healthy 70-year-old individual may have a GFR of 60 mL/min/1.73m 2 .[8][9][8]

The characteristics of an ideal marker of GFR are as follows:

As no such endogenous marker currently exists, exogenous markers of GFR are used. Assessment of GFR using inulin, a polysaccharide, is considered the reference method for estimating GFR. This method involves the infusion of inulin and then the measurement of blood levels after a specified period to determine the inulin clearance rate. Other exogenous markers used are radioisotopes such as chromium-51 ethylenediaminetetraacetic acid (51 Cr-EDTA) and technetium-99-labeled diethylenetriaminepentaacetic acid (99 Tc-DTPA). The most promising exogenous marker is the non-radioactive contrast agent, iohexol, especially in children.

The inconvenience associated with the use of exogenous markers, specifically that the testing has to be performed in specialized centers with the ability to assay these substances, has encouraged the use of endogenous markers.

The most commonly used endogenous marker for assessing glomerular function is creatinine. The calculated creatinine clearance is used to provide an indicator of GFR. This process involves collecting urine over 24 hours or over another accurately timed period of 5 to 8 hours, as 24-hour collections are unreliable. Creatinine clearance is calculated using the equation:

C = (U_Cr × V) / P_Cr

C=clearance, U=urinary concentration (mg/dL), V=urinary flow rate (volume/time in mL/min), and P=plasma concentration (mg/dL)

Creatinine clearance should be corrected for body surface area. Improper or incomplete urine collection is a major issue affecting the accuracy of this test; hence, accurately timed collection is essential. Furthermore, due to tubular secretion, creatinine overestimates GFR by around 10% to 20%.

Creatinine is the by-product of creatine phosphate in muscle, and it is produced at a constant rate by the body. For the most part, creatinine is cleared from the blood entirely by the kidney. Decreased clearance by the kidney results in increased blood creatinine. The amount of creatinine produced per day depends on muscle mass. Thus, there are different creatinine ranges in males and females and lower creatinine values in children and those with decreased muscle mass. Dietary factors also influence creatinine levels. Creatinine can change as much as 30% after ingesting red meat. During pregnancy, GFR increases, leading to lower creatinine levels in pregnant women. In addition, serum creatinine indicates renal impairment at a relatively late stage—renal function is decreased by up to 50% before a rise in serum creatinine is observed.

Serum creatinine is also used in GFR estimating equations such as the Modified Diet in Renal Disease (MDRD) and the Chronic Kidney Disease-Epidemiology Collaborative Group (CKD-EPI) equations. The CKD-EPI group has also developed complex equations incorporating serum creatinine and cystatin C, using a population comprising healthy individuals and CKD patients; these equations are preferred when estimating GFRs in multi-ethnic populations due to their reduced bias.[10][11][12] A recent review by Inker et al presented new equations using cystatin C and creatinine without race that show an improved correlation between measured and calculated GFR. Further details of these complex equations can be found in this study and supplementary materials.[13]

Some support the use of race as a qualitative factor to estimate muscle mass, but the trend is toward removing race from GFR calculations. These equations were generally formulated for White Americans with a modification factor for Black Americans, supposing increased GFR for the same creatinine level. Race can be categorized as Black or non-Black, or additional race categories can be considered, such as Asian, Hispanic, or Native American. Some studies have found that Blacks in Europe do not have a significantly elevated GFR for a given creatinine level, and other studies have found that adding a race variable to creatinine-based equations in Africans or Asians does not improve the correlation between estimated and calculated GFR.[14][15][16][17] No clear consensus is present, and many guidelines suggest using cystatin C as a marker instead of creatinine, as cystatin C is not muscle-mass dependent. In addition, further research is required to identify additional compounds consistent across age, sex, and race to estimate GFR.[16][18][19]

The CKD-EPI formulas have undergone several iterations.[20] Although some experts may prefer eGFR equations using cystatin or both cystatin and creatinine, in practice, cystatin is not widely measured. Therefore, the National Kidney Foundation (NKF) and the American Society of Nephrology (ASN) Task Force recommend using the CKD-EPI creatinine-based equation.[15] This formula has been adopted by most major laboratories for estimated GFR, including Quest and Labcorp.

The CKD-EPI equation: eGFR = 142 × min(SCr/κ,1) α × max(SCr/κ,1) −1.200 × 0.9938 Age × 1.012 (if female)

(Abbreviations/units: SCr is serum creatinine in mg/dL, κ=0.7 for females and 0.9 for males, α=−0.329 for females and −0.411 for males, min=the minimum of S/κ or 1, and max=the maximum of S/κ or 1)

For estimating GFR in children, the Chronic Kidney Disease in Children Study (CKiD or Schwartz bedside) equation is commonly used, which uses serum creatinine (mg/dL) and the child's height (cm).[21] Another formula, the Schwartz-Lyon equation, has also been used for children (younger than 18) and is believed to be more accurate compared to CKD-EPI when measured GFR is lower than 75 mL/min/1.72 m 2 .[22][23] The CKD-EPI equation cannot be used in children, and it overestimated GFR in young adults aged 18 to 39. Modifications to the CKD-EPI formula using sex-specific creatinine growth curves for children and adults aged 18 to 40 allow a well-validated improvement of eGFR; this formula is also referred to as CKD-EPI40.[22]

GFR is classified into the following stages based on kidney disease. Stage 1 CKD refers to patients with a normal GFR but other signs of possible renal structural damage, such as proteinuria.

Kidney Disease Improving Global Outcomes (KDIGO) stages of CKD are as follows:

These stages provide an easier estimation of GFR without requiring urine collection or the use of exogenous materials. However, as they use serum creatinine, they are also affected by the issues around serum creatinine measurement; hence, the correction for different variables is required. In addition, these equations using a single serum creatinine presuppose a stable creatinine; therefore, they are often inaccurate in the frequent situation of rapidly changing creatinine levels.

Blood Urea Nitrogen

Urea, or BUN, is a nitrogen-containing compound formed in the liver as the end product of protein metabolism and the urea cycle. About 85% of urea is eliminated through the kidneys, whereas the rest is excreted through the gastrointestinal (GI) tract. Serum urea levels increase in conditions where renal clearance decreases, such as acute and chronic renal failure or impairment. Urea may also increase in other conditions not related to renal diseases, such as upper gastrointestinal bleeding, dehydration, catabolic states, and high protein diets. Urea may be decreased in starvation, low-protein diet, and severe liver disease. Serum creatinine is a more accurate assessment of renal function compared to urea; however, urea is increased earlier in renal disease.

When BUN levels are increased, the BUN-to-creatinine ratio can be useful to differentiate pre-renal from renal causes. In pre-renal disease, the ratio is close to 20:1, whereas in intrinsic renal disease, it is closer to 10:1. Upper gastrointestinal bleeding can be associated with a very high BUN to creatinine ratio (sometimes >30:1).

Cystatin C is a low-molecular-weight protein that functions as a protease inhibitor produced by all nucleated cells in the body. Cystatin C is formed at a constant rate and freely filtered by the kidneys. Serum levels of cystatin C are inversely correlated with the GFR. In other words, high values indicate low GFRs, whereas lower values indicate higher GFRs, similar to creatinine. The renal handling of cystatin C differs from creatinine. Although glomeruli freely filter both, once cystatin C is filtered, it is reabsorbed and metabolized by proximal renal tubules, unlike creatinine. Thus, under normal conditions, cystatin C does not enter the final excreted urine to any significant degree. Cystatin C is measured in serum and urine. The advantages of cystatin C over creatinine are that it is not affected by age, muscle bulk, or diet, and many reports have indicated that it is a more reliable marker of GFR compared to creatinine, particularly in early renal impairment when creatinine levels are within normal range. Cystatin C has also been incorporated into eGFR equations, such as the combined creatinine-cystatin KDIGO CKD-EPI equation.

Cystatin C concentration may be affected by the presence of cancer, thyroid disease, and smoking. Evidence suggests that thyroid hormones increase cystatin C production.[24]

Albuminuria and Proteinuria

Albuminuria refers to the abnormal presence of albumin in the urine. The term microalbumin is now considered obsolete as there is no such biochemical molecule; the condition is now referred to only as increased urine albumin. Albuminuria is used as a marker to detect incipient nephropathy in diabetics. Albuminuria is an independent marker for cardiovascular disease as it connotes increased endothelial permeability, and it is also a marker for chronic renal impairment. Urine albumin may be measured in 24-hour urine collections or early morning or random specimens as an albumin/creatinine ratio. The presence of albuminuria on 2 occasions, excluding a urinary tract infection, indicates glomerular dysfunction. The presence of albuminuria for 3 or more months is indicative of CKD. Frank proteinuria is defined as greater than 300 mg/d of protein. Normal urine protein is up to 150 mg/d (approximately 30% to 40% albumin, 10% to 20% globulins, and 50% uromodulin [Tamm-Horsfall protein]).[25]

Increased amounts of protein in the urine may be due to the following:

Glomerular proteinuria: Caused by defects in the selectivity of the glomerular filtration barrier to plasma proteins, such as glomerulonephritis and nephrotic syndrome

Tubular proteinuria: Caused by incomplete tubular reabsorption of proteins, such as interstitial nephritis

Overflow proteinuria: Caused by increased plasma concentration of proteins, such as Bence-Jones protein and myoglobinuria

Urine protein may be measured using either a 24-hour urine collection or a random urine protein-to-creatinine ratio (an early morning sample is preferred as it is a close representative of the 24-hour sample).

The KDIGO classification defines 3 stages of albuminuria:

In nephrotic syndrome, urine protein excretion exceeds 3.5 g/d and is associated with edema, hypoalbuminemia, and hypercholesterolemia.

Tubular Function Tests

The renal tubules play a crucial role in reabsorbing electrolytes and water and maintaining acid-base balance. Electrolytes such as sodium, potassium, chloride, magnesium, phosphate, and glucose can be measured in urine. Measurement of urine osmolality allows for assessment of the concentrating ability of urine tubules. A urinary osmolality higher than 750 mOsm/kg H₂O implies a normal concentrating ability of tubules. A water deprivation test can be used to exclude nephrogenic diabetes insipidus. In distal renal tubular acidosis, an ammonium chloride test can be used to confirm the diagnosis of distal renal tubular acidosis with failure to acidify the urine to a pH of less than 5.3. In Fanconi syndrome, aminoaciduria, glycosuria, phosphaturia, and bicarbonate wasting (proximal renal tubular acidosis) are present.

Urine Analysis

Urine analysis involves evaluating urine characteristics to aid in disease diagnosis, through physical observation, chemical, and microscopic examination. The physical inspection involves assessing color and clarity. Normal urine is straw-colored, whereas darker urine may indicate dehydration. Red urine may indicate hematuria or porphyria or could represent the dietary intake of food such as beets. Cloudy urine may be observed in pyuria due to urinary tract infection. Specific gravity indicates renal concentrating ability, which can be measured using refractometry or, chemically, a urine dipstick. The physiological range for specific gravity is 1.003 to 1.030. Specific gravity is increased in concentrated urine and decreased in dilute urine.

The urine dipstick provides qualitative analysis of different analytes in urine using chemical analysis.

The urine dipstick uses dry chemistry methods to detect protein, glucose, blood, ketones, bilirubin, urobilinogen, nitrite, and leukocyte esterase and can be performed as a point-of-care test. The color changes following the urine's interaction with the chemical reagents impregnated on the dipstick paper are compared to a color chart guide to interpret the results.

In healthy urine specimens, urine protein is negative. Bilirubin is not detected in normal urine. Glucose is not detected in healthy patients but may be observed in diabetes mellitus, pregnancy, and renal glycosuria. Ascorbic acid (vitamin C) supplements may cause false-negative results for hemoglobin, leukocyte esterase, nitrite, and glucose.[26]

Blood may be present after renal tract injury or infection, with ascorbic acid causing a falsely negative result. The urine dipstick detects the globin portion of hemoglobin and thus cannot detect the difference between myoglobin and hemoglobin in the urine.

In addition, both intact red blood cells (RBCs) and hemoglobinuria are detected. The presence of blood on a urine dipstick test with normal RBC indicates rhabdomyolysis and can help differentiate it from hematuria, where RBCs are also detected on the urine dipstick. In normal urine, RBC per high-power field is between 0 and 3, and white blood cells (WBCs) are between 0 and 5. Ketones are present in fasting, severe vomiting, and diabetic ketoacidosis. Urine dipstick only detects acetoacetate and acetone, not the ketone beta-hydroxybutyrate. Bilirubin is detected in the presence of conjugated hyperbilirubinemia. Urobilinogen may typically be present, but it is absent in conjugated hyperbilirubinemia and increased in the presence of prehepatic jaundice and hemolysis. Nitrite and leucocyte esterase are indicators of urinary tract infection. Some bacteria, for example, Enterobacteriaceae, convert nitrates to nitrites.

The microscopic urinalysis involves a wet-prep analysis to identify cells, casts, crystals, and microorganisms. RBC casts typically denote glomerulonephritis, whereas WBC casts are consistent with pyelonephritis. The presence of WBCs and WBC casts indicates infection; RBCs indicate renal injury; and RBC casts indicate tubular damage or glomerulonephritis. Hyaline casts consist of protein and may occur in glomerular disease. Fatty casts are observed in nephrotic syndrome. Crystals may also be identified in urine and are indicative of the following conditions:

Triple phosphate crystals resemble a coffin lid found in alkaline urine and urinary tract infections.

Oxalate crystals are envelope-shaped and are present in ethylene glycol poisoning or primary and secondary hyperoxaluria.

The best specimen for urine analysis is a freshly voided midstream urine sample. Midstream urine is less likely to be contaminated by commensal bacteria and epithelial cells.

Acute Versus Chronic Renal Impairment

Acute renal impairment or acute kidney injury refers to the sudden onset of kidney injury within a period of a few hours or days. CKD is caused by long-term diseases such as hypertension and diabetes. Causes of acute kidney injury can be divided into the following:

Causes that result in decreased blood flow to the kidneys (pre-renal causes), such as hypotensive and cardiogenic shock, dehydration, and blood loss from major trauma

Causes that result in direct damage to the kidneys (renal/intrinsic causes), such as damage to kidneys by nephrotoxic medications and other toxins, sepsis, cancers such as myeloma, autoimmune diseases or conditions that cause inflammation, or damage to the kidney tubules

Causes that result in blockage of the urinary tract, such as bladder, prostate, or cervical cancer, large kidney stones, and blood clots in the urinary tract

Importantly, pre-renal kidney injury may progress to acute tubular necrosis and cause intrinsic renal injury.

Urine output is a useful tool for evaluating kidney function and is used in guidelines to define acute kidney injury. Patients with acute kidney injury present with oliguria, defined as less than 400 mL/d or less than 0.5 mL/kg/h for more than 6 hours.[27] The RIFLE classification (risk, injury, failure, loss of kidney function, and end-stage kidney disease) is based on serum creatinine, GFR changes, and urine output determinants. The Acute Kidney Injury Network (AKIN) classification criteria for acute kidney injury also uses serum creatinine changes and urine output; however, it does not rely on GFR changes and does not require a baseline serum creatinine.

Investigations that assist in determining if the renal injury is pre-renal, renal, or post-renal include measuring urine specific gravity, which is increased (greater than 1.020) in dehydration and pre-renal causes. The presence of WBCs and RBCs, tubular epithelial cells, casts, or crystals in the urinary sediment under light microscopy can assist in the differential diagnosis.

Fractional excretion of sodium (FeNa) is useful in distinguishing acute tubular necrosis from pre-renal uremia and requires the measurement of creatinine and sodium in spot urine specimens. Fractional excretion is calculated using the following formula:

FeNa = 100 × (urinary sodium × serum creatinine)/(serum sodium × urinary creatinine).

A value of less than 1% indicates a pre-renal cause, and values greater than 2% indicate intrinsic causes. Spot urine sodium concentrations of less than 20 mmol/L indicate pre-renal acute kidney injury. Fractional excretion of urea calculated similarly to FeNa using serum urea and urine urea instead of sodium can also be used to determine the presence of pre-renal versus intrinsic acute kidney injury, with values less than 35% suggesting pre-renal injury. In general, urine osmolality of more than 500 mOsm/kg is associated with pre-renal causes, whereas an osmolality similar to serum (approximately 300 mOsm/kg) reflects an intrinsic cause. However, in patients receiving diuretic therapy, the FeNa is not reliable.[28]

Novel Biomarkers

Several new biomarkers have been reported to be useful for determining acute kidney injury and have utility in differentiation between acute kidney injury and stable CKD and pre-renal and intrinsic acute kidney injury. These biomarkers include low-molecular-weight proteins and fall into 2 main categories as follows:

Recently recognized filtration markers pseudouridine, acetylthreonine, myoinositol, phenylacetylglutamine, and tryptophan [14]