Assessment of Renal Function

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Assessment of Renal Function

• Iohexol clearance is a simple, reliable and accurate method for estimating GFR

David R Mole Philip D Mason

may be expressed as P x C, where P is the plasma concentration and C is a ‘virtual’ volume of blood, completely cleared by the kidney, of solute per unit time termed the ‘renal clearance’. The equation is expressed as UV = PC, and rearrangement gives C = UV/P (Figure 1). Following filtration, water and solutes may be reabsorbed, secreted, synthesized, or metabolized in the renal tubule. Hence, both tubular function and glomerular filtration govern renal clearance, but GFR can be estimated by measuring the clearance of a solute that is neither secreted nor reabsorbed in the tubule. Candidate molecules should also be freely filtered by the glomeruli and be present at a stable plasma concentration. The solute can be either naturally occurring (e.g. urea, creatinine, cystatin C) or exogenous (e.g. inulin, DTPA), but there are no ‘perfect’ solutes and all estimates are an approximation. Urea and creatinine are most commonly used as measures of renal function. However, urea undergoes passive tubular reabsorption and creatinine is actively secreted in the proximal tubule; as a result, urea clearance underestimates and creatinine clearance overestimates GFR. These relatively minor tubular effects become significant sources of error at low GFR. Although inulin clearance and other accurate methods (see below) are necessary for physiological studies and clinical trials, they are cumbersome for daily clinical use. Simpler methods are generally used, each of which has advantages and disadvantages.

Assessment of renal function is usually considered to involve some measure of glomerular filtration; this contribution focuses on the methods by which glomerular filtration may be assessed with particular emphasis on those used in clinical practice. Such measures are of importance not only in helping to detect and monitor renal disease, but also in calculating dose adjustments in prescribing. However, it should be remembered that the kidney has many other roles, including tubular functions such as concentrating ability, hydrogen ion excretion and regulation of electrolytes, and the endocrine functions of 1α-hydroxylation of vitamin D and production of erythropoietin and renin. One-fifth of the cardiac output flows through the two kidneys, which weigh about 250 g (i.e. a flow rate of 1000–1200 ml/minute or 4 ml/g/minute). This blood flow is distributed among about 2 million glomeruli with a combined filtering area of about 1 m2. The rate of filtration across this membrane is governed by multiple factors including renal blood flow, intrarenal blood distribution, glomerular capillary pressure and glomerular capillary permeability. These in turn are partly controlled by afferent and efferent arteriolar tone, and by other intraglomerular factors such as mesangial cell tone. Normal glomerular filtration rate (GFR) is about 130 ml/minute (180 litre/day or 2 ml/second).

Measurement of clearance Concept of ‘clearance’ The renal excretion of any substance per unit time equates to its urinary concentration (U) multiplied by the volume of urine per unit time (V). When the given solute is in a steady state, renal excretion equates to the removal of solute from the blood and

Inulin clearance Inulin (a fructose polymer of molecular weight 500 Da) is almost a perfect solute, being neither secreted nor reabsorbed by tubules. Its clearance is usually regarded as the gold standard for estimation of GFR. The original method of estimating GFR by inulin clearance requires infusion of inulin to produce a steady-state concentration. Urinary excretion (requiring an accurately timed urine collection)

The concept of solute clearance • • • •

David R Mole is Specialist Registrar in Nephrology at the Churchill Hospital, Oxford, UK. He qualified from the University of Cambridge and the University of Southampton, and trained in general medicine and nephrology in Stoke-on-Trent, Reading and Oxford.

U, urine concentration; V, urine volume per unit time; P, plasma concentration; C, clearance (volume per unit time)

Philip D Mason is Consultant Nephrologist at the Churchill Hospital, Oxford, UK. He qualified from Guy’s Hospital, London, and trained in nephrology at the Royal Postgraduate Medical School, Hammersmith Hospital, London. His research interest is transplantation medicine.

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Renal excretion of solute per unit time = UV Removal of solute from the blood = PC Renal excretion = removal from blood (i.e. UV = PC) Clearance = UV/P

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is then measured. A single bolus injection followed by sequential measurements of disappearance of inulin from plasma avoids the need for complicated infusions, errors in timing urine collection and problems of incomplete bladder emptying. This method measures plasma rather than renal clearance, but these are equivalent in the absence of non-renal excretion or redistribution into non-plasma compartments. Disappearance of inulin from plasma follows first-order kinetics and levels decrease exponentially. It is essential to know the precise time interval between the taking of blood samples, but accurate estimates of GFR may be obtained when the technique is performed carefully. When GFR is low, a longer time interval may be required between samples. The inulin clearance method is now seldom performed, and inulin is no longer readily available in the UK.

merulus and is produced at a relatively constant rate. Its levels are independent of tubular function, and are therefore closely correlated with GFR. The cystatin C concentration range seems to differ little between men and women, and correction for body surface area does not appear to be necessary. Studies suggest that plasma cystatin C levels correlate better with radioisotope estimates of GFR than do creatinine levels and are more sensitive to mild renal impairment, but there are contrary views. Furthermore, there is a lack of prospective studies, and malignancy and corticosteroid treatment may affect the usual range. Although use of cystatin C appears promising as a measure of GFR, further studies are necessary before it can be recommended widely. Creatinine clearance Creatinine production is relatively constant and therefore its clearance is commonly used to estimate GFR. The most common source of error in determining creatinine clearance is the urine collection. • It is important to tell the patient to discard the first sample of urine, so the collection begins with an empty bladder. • The patient should note the time and collect all urine produced for the next 24 hours, ending with complete emptying of the bladder at the same time the following day. • When the urine volume is found to be very low, or the same as that of the container, the patient may have misunderstood the instructions or may have needed a second bottle. • 24-hour creatinine excretion is constant and depends on age, gender and lean body mass, and may therefore be estimated to check the adequacy of urine collection. Alternatively, the creatinine clearance obtained can be compared with that calculated from the Cockroft–Gault equation (see below). Although excretion of creatinine is predominantly by glomerular filtration, additional tubular secretion occurs such that measurement of creatinine clearance tends to overestimate GFR by 10–20%. Fortuitously, at high GFR, this balances errors in the measurement of plasma (but not urine) creatinine concentration, so creatinine clearance gives a good approximation of GFR. As plasma creatinine rises, however, increased tubular (and additionally bowel) secretion leads to progressive overestimation of GFR. More accurate estimates of GFR may be obtained by competitively blocking tubular secretion of creatinine with the H2-receptor antagonist cimetidine, or by averaging creatinine and urea clearance, but these methods are seldom performed. Clinical decisions are

Other clearance methods Isotope methods: when labelled with radioactive chromium-51, EDTA can be easily and precisely measured in the blood. A single bolus injection of 51Cr-EDTA with subsequent blood sampling is commonly used to estimate GFR and correlates well with inulin clearance. The technique must be performed carefully, with accurate timing and avoidance of extravasation of the bolus at the injection site. Accuracy can be improved at low GFR by collecting urine, but this introduces problems of obtaining accurately timed collections of radioactive urine. Disadvantages include radiation exposure (typically 4% of that of a chest radiograph) and the cost of the isotope chelate. Dynamic renography: technetium-99-DTPA and 99Tc-MAG3 are used for dynamic renal scanning. GFR can be estimated from dynamic scans by the rate of disappearance of the isotope, and the accuracy can be improved if blood and/or urine samples are collected. Iohexol: use of iohexol (a low-osmolality, non-ionic contrast medium) has recently gained popularity because it is nonradioactive and can be measured with high-performance liquid chromatography. There is a good correlation with single-bolus inulin techniques. Iohexol clearance techniques estimated from finger-prick samples obtained by the patient and sent back to the hospital are being developed. Cystatin C is an endogenous cysteine protease inhibitor. It is a low molecular weight protein that is freely filtered by the glo-

Cockroft–Gault equation for estimating creatinine clearance Men Creatinine clearance = 1.23 x (140 – age) x (weight in kg) serum creatinine (µmol/litre) Women Creatinine clearance = 1.04 x (140 – age) x (weight in kg) serum creatinine (µmol/litre) 2

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Changes in creatinine clearance with age

Interpreting creatinine and urea

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Reduced • Small muscle mass • Pregnancy • Raised antidiuretic hormone

Reduced • Liver disease • Starvation/anabolic state • Pregnancy • Raised antidiuretic hormone

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Changes with age: the number of glomeruli in each individual is determined at birth. GFR increases during childhood with growth of individual nephrons and improved blood flow. During adult life, the number of nephrons decreases and renal blood flow declines progressively, resulting in deterioration of renal function. This may be masked by a concomitant decrease in muscle mass, such that creatinine levels increase to a lesser degree. Figure 3 gives a guide to the effect of age and gender on creatinine clearance (as predicted by the Cockroft–Gault equation derived from a largely Caucasian Belgian population). A creatinine clearance of 50 ml/minute is normal for a 75-year-old woman, but inappropriate for a 30-year-old man. In view of the wide normal range, it is helpful to obtain any previous data available on patients’ renal function, to determine whether they have fallen below their centile curve.

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Age (years) The lines are centile plots of creatinine clearance, calculated by the Cockroft–Gault equation

Measurement of urea and creatinine levels

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Interpreting urea and creatinine concentrations The simplest measure of renal function is the serum level of urea or creatinine. When interpreting such values, the ‘hole-in-thebucket’ analogy must be considered. At steady state, the level of water in the bucket depends not only on the size of the hole, but also on the rate at which the bucket is topped up from the tap. By analogy, the level of urea or creatinine depends not only on the rate of glomerular filtration, but also on the rate of production of each solute (Figure 4).

not made on the basis of minor changes in creatinine clearance at such low levels; patients in whom creatinine clearance is less than 15 ml/minute must be considered for dialysis. Cockroft–Gault equation: several formulae have been derived to estimate creatinine clearance, to overcome the difficulty and lack of immediacy involved in obtaining an accurate urine collection. The most widely used is the method of Cockroft and Gault (Figure 2), which is based on the patient’s age, sex, weight and blood creatinine. The formula was derived experimentally in a Canadian population of unspecified racial origin aged 18–92 years. The formula should therefore not be used in patients below 18 years of age, in other racial groups, in patients who are grossly obese, oedematous or cachectic, in those with ascites when muscle mass is reduced (e.g. paraplegics), or in pregnant women.

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Causes of misleading urea/creatinine levels Catabolism: urea is the main nitrogenous waste product derived from protein breakdown. At steady state, urea production rate is determined by dietary protein intake. Malnutrition (a common feature of chronic renal failure, CRF) is therefore associated with artificially low urea levels, whereas a large protein meal (e.g. a bowel full of haemoglobin following an upper gastrointestinal haemorrhage) falsely elevates blood urea concentration. Urea is

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Serum creatinine and glomerular filtration rate

Substances that interfere with the measurement of creatinine Common drugs • Cephalosporins • 5-fluorocytosine • Methanol metabolites

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Endogenous substances • Protein • Ketones • Glucose • Bilirubin • Fatty acids • Urate • Urea

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also artificially elevated in catabolic patients with muscle breakdown caused by sepsis or corticosteroids. All of these aberrations are be more apparent at lower GFRs. Muscle mass: creatinine is formed by non-enzymatic degradation of muscle creatine. Its production is consequently proportional to muscle mass and dietary meat intake; thus, for the same GFR, creatinine levels are higher in a young, muscular man than in a slight, elderly woman. In addition to age, sex, and weight, muscle mass and hence creatinine levels are also influenced by racial origin. Large amounts of creatinine may be produced during the muscle necrosis seen in rhabdomyolysis, leading to a disproportionate increase in creatinine.

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Glomerular filtration rate (ml/minute) Creatinine clearance is about 100 ml/minute in a 24-year-old man weighing 70 kg with serum creatinine 100 µmol/litre. A rise in serum creatinine from 100 µmol/litre to 150 µmol/litre represents a loss of 33% of renal function. A further rise from 150 µmol/litre to 200 µmol/litre is a loss of a further 17%, and from 200 µmol/litre to 250 µmol/litre a further 10%; creatinine clearance is then 40 ml/minute.

Filtration rate: at low GFR (e.g. in dehydration), tubular function becomes more apparent. Urea is highly diffusible (unlike creatinine) and is therefore reabsorbed to a greater extent at the low flows associated with dehydration; in addition to the already raised urea levels resulting from low GFR, this accounts for the disproportionate increase seen in dehydration. Conversely, tubular excretion of creatinine effectively lessens the increase in plasma creatinine concentration. In intrinsic renal disease, glomeruli and tubules are often lost in proportionate numbers.

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Reciprocal creatinine plots A single measure of plasma creatinine does not provide a reliable indicator of GFR. However, in an individual patient, the rate of production of creatinine is more constant than that of urea. Therefore, serial measurements of creatinine, in which all of the above factors are assumed to remain constant, provide a means of monitoring changes in renal function. Because the relationship between creatinine and GFR is exponential (Figure 6), the reciprocal of creatinine concentration is linearly proportional to GFR (P = UV/C or C = UV/P). An important consequence of this relationship is that a significant proportion of renal function may be lost before creatinine levels rise above the normal range, particularly in small, non-muscular, vegetarian individuals. At high GFRs, even large changes in renal function can lead to minor changes in creatinine levels. Conversely, at low GFRs, a large change in creatinine can result from small changes in renal function (Figure 6). In many forms of CRF, the decrease in GFR occurs at a constant rate, and a plot of the reciprocal of creatinine concentration against time should therefore give a straight line (with scatter).

Drugs: certain drugs (e.g. cimetidine, trimethoprim, probenecid, amiloride, spironolactone, triamterene) block creatinine secretion in the tubules. Administration of these drugs in CRF therefore leads to an increase in creatinine without a change in GFR. Their effect is more pronounced at low GFR. Other substances: several endogenous and exogenous substances interfere with the creatinine assay (Figure 5). Creatinine is usually measured by the Jaffe reaction, in which creatinine reacts with alkaline picrate to form an orange-red Janovsky complex. Other chromagens that react to give a similar or identical colour are present in plasma (but not urine), resulting in overestimation of plasma creatinine concentration with consequent underestimated creatinine clearance. The effect is most pronounced at low creatinine levels (i.e. within the normal range).

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Reciprocal creatinine plots in four patients In many forms of chronic renal failure, glomerular filtration rate decreases at a constant rate, such that a plot of the reciprocal of serum creatinine against time is a straight line. Any deviation from this line gives important information, as in these examples. Prediction of the anticipated date for commencement of dialysis

Progression of renal failure may be slowed by good blood pressure control

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Addition of an angiotensin-converting enzyme inhibitor causes worsening of renal function in the presence of renal artery stenosis Ð stopping the angiotensin-converting enzyme inhibitor quickly may allow a return to the original rate of progression

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The approximate date when dialysis will be required can be predicted by the extrapolated intercept with the x-axis. Furthermore, any deviation from the predicted plot may give an early indication of an intercurrent additional renal insult (e.g. obstruction from papillary necrosis in a patient with diabetic nephropathy), prompting further investigation. Figure 7 gives some examples.

kidney using a gamma-camera, and uptake of activity can then be used to determine the relative contribution of each kidney to overall GFR. The greatest potential source of error is the difference in depth of the two kidneys; each 1 cm difference accounts for a 10% difference in the activity count. The technique is therefore relatively inaccurate, but shows major differences in differential function.

Differential renal function

Tests of tubular function

In certain situations (particularly when planning surgery), it is important to know the relative contribution of each kidney to the overall renal function. This may be determined reliably using 99 Tc-DMSA, which localizes to proximal tubular cells. Activity– time curves corrected for background are generated for each

The properties of the tubule vary along its length and solutes may be modified by different factors at each point. Various tubular mechanisms could theoretically be investigated, but, in clinical practice, tests of tubular function are required less often than those of glomerular function.

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In the less common proximal (type II) RTA, bicarbonate reabsorption is defective; the urine may still be acidic if bicarbonate entry to the tubule is reduced by severe systemic acidosis and/or low GFR. Assessment of such patients tests the ability to conserve a bicarbonate load. The diagnosis of type II RTA may be obvious in those with a generalized tubular leak of glucose and amino acids (Fanconi’s syndrome). u

Urinary concentration and dilution Inability to concentrate the urine to more than 600 mOsmol/litre indicates diabetes insipidus. This can be demonstrated using a water-deprivation test, in which plasma and urine osmolality are measured repeatedly until the urine osmolality remains constant or exceeds 600 mOsmol/litre; care is required to avoid excessive volume depletion. Cranial diabetes insipidus can be distinguished from nephrogenic diabetes insipidus by a subsequent response to administration of antidiuretic hormone (ADH) in those in whom urine concentration fails to increase in response to fluid restriction alone. Specific protocols are available in more detailed texts. This test is indicated in the investigation of polyuria in patients with normal renal function, but seldom in patients with reduced GFR, because they are likely to have impaired tubular function and may need to avoid dehydration. The ability to dilute urine to achieve maximum free water clearance is also often impaired in patients with CRF, though seldom tested. It can be assessed by administering 1000–1500 ml of water (20 ml/kg body weight). More than 75% of the water load should be excreted within 3 hours and urinary osmolality should fall below 100 mOsmol/kg. Renal clearance of osmoles is assessed, as is the clearance of any other solute (Cosm = Uosm.V/Posm). Free water clearance can then be determined by subtracting Cosm from V. Ability to excrete free water declines with age, and care should be taken when administering large volumes of 5% dextrose to elderly patients, or in states of raised ADH (e.g. postoperative, pain, nausea).

FURTHER READING Cockroft D W, Gault M H. Prediction of Creatinine Clearance from Serum Creatinine. Nephron 1976; 16: 31–41. Davison A M, Cameron J S, Grünfeld J-P et al. Oxford Textbook of Clinical Nephrology. 2nd ed. Oxford: Oxford University Press, 1998. Elseviers M M, Verpooten G A, De Broe M E et al. Interpretation of Creatinine Clearance. Lancet 1987; i: 457. Rose B D, Black R M. Manual of Clinical Problems in Nephrology. Boston: Little, Brown, 1988. Sweny P, Varghese Z. Clinical Tests: Renal Disease. London: Wolfe, 1988. Whitworth J A, Lawrence J R. Textbook of Renal Disease. 2nd ed. Edinburgh: Churchill Livingstone, 1994.

Urinary electrolytes and other tests of tubular function 24-hour sodium output may be helpful in determining whether a patient is complying with a low-sodium diet and in the management of those with salt-losing nephropathy. Low urinary sodium (< 10 mM) is associated with pre-renal failure but should not determine management, because patients with higher levels may still respond to appropriate filling. Assessment of distal chloride handling may be helpful in identifying patients with Bartter’s syndrome. Tests of proximal tubular function may be required in the investigation of Fanconi’s syndrome or isolated proximal tubular defects (e.g. urate clearance). Bicarbonate, glucose, phosphate and amino acids are all normally reabsorbed in the proximal tubule. Their presence in the urine is abnormal, and though formal methods to measure maximal reabsorption are available, they are seldom necessary. β2-microglobulin is reabsorbed by the proximal tubule, and its urinary excretion is nonspecifically increased by diseases of the proximal tubule.

Practice points

Urinary acidification tests Type I and type II renal tubular acidosis (RTA) cause hypokalaemic hyperchloraemic metabolic acidosis (a picture more commonly caused by diarrhoea). In distal (type I) RTA, patients are unable to generate an acid urine (pH < 5.5) because hydrogen ion excretion is defective; the diagnosis is confirmed if low urine pH is not achieved in the presence of systemic acidosis. Oral acidification tests (usually with ammonium chloride) are clinically indicated only in partial RTA, in which patients are hypokalaemic but not acidotic as a result of increased hydrogen ion levels with ammonia.

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• Urea and creatinine are the simplest indicators of GFR • Creatinine clearance is the best practical measure • Reciprocal creatinine plots are the most sensitive longitudinal measure • Inulin clearance and radioisotope studies are the most accurate at low GFR • Renal function should be interpreted with regard to age and sex

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