CHAPTER OUTLINE

The kidneys are located against the posterior abdominal wall on either side of the vertebral column. They lie anterior to the diaphragm and various muscles; their anterior surfaces are covered by parietal peritoneum. The left kidney is posterior to the stomach, pancreas, spleen and descending colon, and the right to the liver, the second part of the duodenum and the ascending colon. Their superior poles are covered by the suprarenal (adrenal) glands. In the adult, the kidneys are about 11–13 cm long, 6 cm wide and 4 cm thick. They weigh approximately 150 g each, yet together receive about 25% of the resting cardiac output. This blood is supplied by the renal arteries, which are branches of the aorta. Venous drainage is into the renal veins, which drain into the inferior vena cava. The kidneys are innervated by sympathetic nerves arising from the sympathetic chains and by parasympathetic fibres arising from the vagus nerve.

Each kidney is covered by a poorly distensible capsule. This limits the swelling that can occur during acute inflammation, and in such circumstances there is an increase in tissue pressure that tends to decrease the glomerular filtration rate (GFR).

Congenital absence of one kidney occurs in approximately 1 in 2400 individuals; this is only of significance if renal surgery is contemplated for any reason. The remaining kidney usually undergoes compensatory hypertrophy. Another common (approximately 1 in 10 000) congenital anomaly is the single ‘horseshoe’ kidney, in which the lower poles of the potential two kidneys are conjoined; renal function is normal but, occasionally, ureteric obstruction may predispose to calculus formation and infection.

• the loop of Henle, which has the configuration of a hairpin and extends into the deeper part of the kidney (the medulla), then doubles back on itself to return to the cortex

• the distal convoluted tubule, located in the cortex

• the collecting duct, which extends down through the medulla to the renal papillae, whence the urine drains into the renal pelvis. The renal pelvices are drained by the ureters, and the right and left ureters themselves drain into the urinary bladder, where urine is stored prior to voiding.

The glomerular filtrate is formed by the passage of fluid from the capillaries through fenestrations in the endothelial cells and through the basement membrane. This is covered on its epithelial aspect by the interdigitating foot processes of the epithelial cells, or podocytes, and the ultrafiltrate passes across a thin membrane (the epithelial slit diaphragm) into the filtration slits between the foot processes and then into the Bowman space, which is continuous with the lumen of the nephron. The selective retention of different constituents of the blood occurs at different stages. The endothelial fenestrations are too small to permit significant passage of erythrocytes, leukocytes or platelets; the major barrier to the filtration of proteins is mechanical and provided by the inner layer of the basement membrane, but the anionic surface layer also selectively retards anionic proteins. Finally, the foot processes and slit diaphragms also act as a mechanical barrier. The relative rate of progression of the constituents of the plasma into the Bowman space, therefore, depends on molecular mass, shape and charge. Glomeruli also contain mesangial cells. These stellate cells are contractile, are involved in the regulation of glomerular filtration and have phagocytic properties. The basement membrane does not extend between the mesangial cells and endothelial cells. This facilitates their ability to phagocytose large particles from the plasma. Their ability to ingest immune complexes plays an important part in the pathogenesis of certain forms of glomerular disease.

This structure is about 15 mm long and is composed of a single layer of inter-digitating epithelial cells united at their apices by tight junctions. The luminal surfaces of these cells have microvillous brush borders that provide the large surface area required for the absorptive function of the proximal tubule. The proximal tubule drains into a short straight segment directed towards the outer medulla and continuous with the descending limb of the loop of Henle.

The distal convoluted tubule is about 5 mm long and is made up of typical absorptive epithelial cells, although without a brush border. It passes close to the afferent and efferent arterioles of the same nephron. The tubular epithelial cells in this area have a special function in relation to the control of renin secretion and are known as the macula densa. Renin is secreted by myoepithelioid juxtaglomerular cells in the walls of the afferent arterioles. These cells, together with those of the macula densa and some other nearby cells (lacis cells), constitute the juxtaglomerular apparatus.

The distal convoluted tubule also contains intercalated (I) cells, responsible for the secretion of acid.

The collecting ducts are composed of I cells and also principal (P) cells, which become permeable to water under the influence of vasopressin. The ducts descend from the cortex down through the medulla, opening to the renal pelvis from the renal papillae.

In addition to the cells of the nephron and blood vessels, the kidneys contain medullary interstitial cells. These characteristically contain lipid droplets and secrete prostaglandins.

Blood leaving glomerular capillary tufts passes not into venules, but into efferent arterioles. Those draining the cortical glomeruli divide into a capillary plexus that enmeshes the renal tubules. Those draining the juxtamedullary glomeruli form the vasa recta; these are bundles of vessels that plunge into the medulla to varying depths, forming capillary plexi that envelop the loops of Henle and then rejoin to form the ascending vasa recta, which eventually drain into the renal veins. The vasa recta form a countercurrent exchange system (see below), which is essential to the formation of both concentrated and dilute (with respect to plasma) urine.

The principal functions of the kidneys are to excrete water-soluble (mostly nitrogenous) waste products and toxins from the body, and to control the volume and composition of the extracellular fluid (ECF). The kidneys also have important endocrine functions. It is perhaps surprising that the kidneys produce some 170 L of glomerular filtrate per 24 h, but only 1–2 L of urine. The bulk of the solvent and solute filtered is reabsorbed, but the control of the processes responsible gives considerable scope for the composition and volume of the urine to be adjusted to meet the physiological needs of the individual.

The physiology of the kidneys is well described in many standard physiology textbooks and will not be recounted in detail in this chapter. Rather, a brief summary is given and certain features that are pertinent to an understanding of the investigation of renal function and the mechanisms and consequences of renal impairment are emphasized.

At rest, 20–25% of the cardiac output flows through the kidneys (1.1–1.3 L/min). The bulk of this blood perfuses the cortex; indeed, the flow rate to the medulla is actually less than to most other tissues.

The process of glomerular filtration is essential to renal function and is dependent on an adequate renal perfusion pressure. Autoregulatory mechanisms allow maintenance of renal blood flow and GFR within narrow limits in the face of a wide variety of external variables, including arterial pressure, venous pressure, ureteric hydrostatic pressure and plasma oncotic pressure. Renal blood flow and GFR are independent of mean arterial blood pressure over the range 80–200 mmHg. This is achieved by intrinsic, and humorally and neurally mediated, alterations in the tone of the renal blood vessels and, in particular, of the efferent glomerular arterioles.

Renal hypoperfusion stimulates the release of renin from juxtaglomerular cells. This enzyme converts circulating angiotensinogen to angiotensin I, which is in turn converted by angiotensin-converting-enzyme to angiotensin II, a powerful vasoconstrictor. This contributes to the maintenance of systemic blood pressure and, in the kidney, by causing efferent arteriolar vasoconstriction, helps to maintain the intraglomerular pressure despite a reduction in perfusion pressure. Various other mechanisms are involved in the maintenance of systemic blood pressure, but in the poorly perfused kidney, release of prostaglandins causes vasodilatation which, at least in mild hypotension, helps to maintain renal blood flow. A perfusion pressure of at least 50–60 mmHg is required to overcome the combined hydrostatic and oncotic pressures that oppose filtration; if mean arterial pressure falls below 80 mmHg, renal blood flow and GFR decline rapidly.

A decrease in the area available for filtration can occur in many renal diseases in which there is glomerular damage. It can also result from contraction of mesangial cells mediated by agents such as angiotensin II, vasopressin, noradrenaline (norepinephrine), thromboxane A₂ and prostaglandin F₂; dopamine, natriuretic peptides and prostaglandin E₂ have opposite effects. A reduction in filtration area may be an important physiological mechanism for reducing GFR and thus conserving fluid.

The proximal tubules are responsible for the active reabsorption of the bulk of filtered solute, accompanied by an iso-osmotic amount of water. Thus, virtually all the filtered glucose, amino acids, bicarbonate and potassium are absorbed here, together with some two-thirds of the filtered sodium. Absorption of solutes is an active, energy-requiring process and is isotonic, so that an equivalent amount of water is also absorbed and the fluid entering the descending limb of the loop of Henle is isotonic with plasma.

This structure provides the countercurrent multiplier that generates the medullary hypertonicity that is essential for the regulation of water excretion and the production of concentrated urine.

Essentially, the process involves the active transport of solute (principally chloride and sodium ions) out of the thick ascending limb of the loop of Henle. This solute is not accompanied by water. As a result, the fluid within the lumen becomes hypotonic and the interstitial fluid surrounding the loop becomes hypertonic. Since water can diffuse out of the thin descending limb, but not the thick ascending limb, the net effect is that the fluid within the loop of Henle and the surrounding interstitial fluid become progressively hypertonic from the corticomedullary junction down into the medulla. There is further sodium and chloride absorption from the thick ascending limb. As water cannot diffuse out of the thick limb, the fluid entering the distal convoluted tubule is hypotonic with respect to plasma.

There is a reciprocal relationship between the rate of flow of fluid through the ascending limb of the loop of Henle and the first part of the distal convoluted tubule of individual nephrons, and the rate of glomerular filtration through the same nephron. Thus, a decrease in flow rate increases filtration and vice versa. This process tends to result in a constant load of solute being presented to the distal convoluted tubule. Feedback is mediated through constriction and dilatation of the afferent arteriole by factors such as renal prostaglandins, adenosine and the renin–angiotensin system, in response to changes in chloride absorption in the macula densa. This mechanism becomes more sensitive when ECF volume is decreased and vice versa.

Urea has an essential role in the generation of medullary hypertonicity. The thick ascending limb of the loop of Henle is permeable to urea (and sodium) but not to water. Movement of urea down its concentration gradient into the interstitium contributes significantly to medullary hypertonicity; the inner medullary portion of the collecting duct (see below) is also permeable to urea. The poor concentrating ability of the kidneys in the newborn is due to the decreased availability of urea to maintain medullary hypertonicity. In adults, renal concentrating power is dependent on an adequate protein intake (the source of urea) and is lower on a low protein diet.

This part of the nephron is responsible for the ‘fine tuning’ of urinary composition. Aldosterone stimulates sodium reabsorption, generating an electrochemical gradient that allows the secretion (and thus excretion) of potassium and hydrogen ions. Normal distal tubular function is essential to the maintenance of hydrogen ion homoeostasis.

These structures extend from the ends of the distal convoluted tubules to the renal papillae. Their main function is to permit the reabsorption of solute-free water and thus regulate the osmolality of the urine. This is achieved through the action of vasopressin (antidiuretic hormone). The cells of the collecting ducts are normally impermeable to water. Under the influence of vasopressin, aquaporin-2 water channels are inserted into the tubular wall, such that they become permeable, allowing water to move out of the hypotonic fluid in the tubular lumen into the hypertonic interstitium, so producing concentrated urine. In the absence of vasopressin, no water is removed and the urine is hypotonic.

The extremes of achievable urine osmolality are 30–1400 mmol/kg. When maximally dilute, as much as 13% of the filtered water is excreted and the rate of urine production is about 16 mL/min; when maximally concentrated, < 0.5% of filtered water is excreted and urine production is as little as 500 mL/24 h.

Some solute is also absorbed in the collecting ducts. In the cortical portion, sodium reabsorption takes place in exchange for potassium and hydrogen ions, as in the distal convoluted tubules. The medullary segment is partially permeable to urea, which moves passively out into the interstitium and helps to maintain the high osmolality of the medulla.

While the maximal urine flow rate when a water load is being excreted is approximately 16 mL/min (assuming an average solute load), higher rates are attainable if there are increased quantities of non-reabsorbed solutes in the tubules. This osmotic diuresis is due to the direct osmotic effect of the solutes and to a secondary effect on sodium reabsorption. The retention of water in the lumen of the proximal tubules increases the concentration gradient against which sodium must be reabsorbed; the same factor limits sodium reabsorption in the thick ascending part of the loop of Henle, thus interfering with the generation of medullary hypertonicity. This is the explanation for the diuresis characteristic of hyperglycaemia.

There are relatively few cardinal manifestations of renal disease and none is specific to any one underlying disorder. They include anuria, oliguria and polyuria; respectively a complete absence, decreased production and excessive production of urine. Polyuria may come to notice only by virtue of causing excessive production of urine at night (nocturia). These are usually symptoms of disordered renal function, although anuria and oliguria can be secondary to obstruction of the urinary tract and polyuria can be due to extrarenal disease that impairs the ability of the kidneys to concentrate the urine.

Urinary frequency (the passing of urine more frequently than normal, but in normal total 24 h volumes) must be distinguished from polyuria; it is often related to irritation of the urinary tract, for example by infection, but can also be due to prostatic enlargement. Irritation of the urinary tract is frequently accompanied by dysuria – pain on micturition. Patients with renal disease may experience loin pain or have tenderness in the loins. Renal or ureteric colic, an intermittent pain of considerable severity, is characteristic of the passage of urinary calculi.

Although patients with renal disease frequently present with an obvious functional abnormality, for example increased plasma creatinine or proteinuria, these are not specific to any one renal disease.

Many techniques are of value in the investigation of the kidneys and renal function. The major groupings of techniques are laboratory tests, particularly biochemical and immunological, radiological imaging and the histological examination of renal tissue usually obtained by percutaneous biopsy.

Imaging techniques: standard radiography, ultrasonography, computed tomography, magnetic resonance imaging, the use of radionuclides and duplex scanning, can provide valuable anatomical information. Such techniques can be used to diagnose, for example, structural abnormalities, tumours, hydronephrosis, polycystic disease and the scarring of pyelonephritis. Radionuclide techniques can also provide information concerning renal function, particularly divided function (the separate function of the two kidneys) and glomerular function. Duplex scanning is a good screening test to assess renal blood flow before proceeding to magnetic resonance angiography or renal angiography.

Useful immunological markers include the detection in serum of antiglomerular basement membrane antibodies (positive in Goodpasture syndrome), antineutrophil cytoplasmic antibodies (in vasculitis), antinuclear antibodies and double-stranded DNA antibodies (in systemic lupus erythematosus), complement components (in various types of glomerulonephritis) and antistreptolysin O antibodies (in post-streptococcal glomerulonephritis). Immunological techniques are also widely used in the histological examination of renal tissue. However, given the pivotal role of the kidneys in body fluid homoeostasis, it is not surprising that biochemical investigations that indicate derangements of homoeostasis are the most widely used tests of renal function. The remainder of this section covers such tests.

The assessment of renal function should start with examination of the urine. Simple observations on the urine still contribute to the investigation of patients with suspected renal disease. To be valid, they must be made on a freshly passed sample.

Turbidity in a fresh sample suggests infection, but can also be due to the presence of fat in patients with the nephrotic syndrome. The presence of chyle in the urine may cause turbidity so severe that the urine appears milky. Anything more than a scant amount of foaming when the urine is shaken, suggests proteinuria. The most frequent abnormal odour is that of ammonia, owing to the presence of urea-splitting microbes, either because the patient has a urinary infection or, in stale samples, because of contamination.

Both of these give an indication of the concentration of the urine. When this needs to be known accurately, for example in tests of renal concentrating ability, measurements of osmolality are essential. A reagent stick assessment of specific gravity may sometimes help in the interpretation of a screening test for proteinuria, since false negative results can occur with very dilute urine and vice versa. However, these reagent sticks detect ionic species only, and underestimate the specific gravity if other substances, for example glucose, are present.

Normal urine is acidic, except after meals. Measurements of urine pH are important in the investigation of renal stone formers and as part of a test of urinary acidification in the investigation of suspected renal tubular acidosis.

Proteinuria is an important feature of renal disease. Normal urine protein excretion is < 200 mg/24 h and is not detectable with standard reagent sticks (e.g. ‘Albustix’). Pathological proteinuria may be:

• glomerular (due to increased glomerular permeability to protein)

• tubular (due to decreased tubular reabsorption of filtered protein or saturated reabsorption)

• secreted (derived from the epithelium of the urinary tract).

Microscopic examination of the sediment obtained by centrifuging a freshly passed sample of urine frequently reveals a few cells (e.g. erythrocytes, polymorphonuclear leukocytes and cells derived from the kidney and urinary tract), hyaline (or rarely granular) casts composed of uromodulin (Tamm–Horsfall protein), and sometimes fat droplets, mucus threads and pigment granules.

Many other substances can be detected in the urine in appropriate circumstances, for example drugs or bilirubin. Their presence is due to the normal excretory function of the kidneys and is not a reflection of renal dysfunction, so they will not be considered further here.

Glomerular filtration is essential to renal function, and investigations designed to measure the GFR are the most frequently performed tests of renal function. The factors that determine the GFR have been discussed above. Its measurement is based on the concept of clearance – the determination of the volume of plasma from which a substance is completely removed (the plasma is ‘cleared’ of the substance) by glomerular filtration during its passage through the kidney. Clearance is a theoretical concept – no substance is cleared completely from the plasma in this way – but can nevertheless be used as a valid measure of the GFR.

For the clearance of a substance to equal the GFR, it must be freely filtered by the glomeruli and eliminated from the body solely by this route (e.g. it must not be metabolized by the liver or secreted into or reabsorbed from the urine). The substance should be non-toxic and accurate measurement should be easily available in a routine laboratory. The clearance of a substance requires measurements of the plasma concentration and urinary excretion rate:

where U is the concentration of the substance in the urine, is the rate of formation of urine and P the plasma concentration of the substance. It should be noted that is a rate; it has the dimensions (volume/time) and its determination requires the collection and measurement of the total amount of urine formed over a known period of time. The units of the various quantities are usually adjusted so that the clearance is expressed as mL/min.

Clearance can also be used to determine renal plasma flow, using a substance such as p-aminohippuric acid, which is almost completely cleared from the blood in a single passage through the kidneys by a combination of glomerular filtration and secretion.

Inulin is a plant polysaccharide that satisfies all the physiological criteria mentioned in the previous section. The measurement of inulin clearance remains the ‘gold standard’ for the estimation of GFR. However, it is a relatively complex procedure and, although test kits are commercially available, it is infrequently used in clinical practice. In essence, the test involves the injection of a bolus dose of inulin, followed by a maintenance infusion designed to produce a constant plasma concentration. Once this has been achieved, a series of timed urine samples is collected and blood is drawn for the measurement of inulin at the midpoints of the collection periods. The GFR is taken as the mean of the inulin clearances for each period.

Creatinine is an endogenous substance, a normal product of muscle metabolism. Its rate of production is fairly constant from day-to-day, being determined by muscle bulk rather than by activity. Creatinine is removed from the body mainly by glomerular filtration and its clearance can be measured as an index of GFR, although in the UK, it is much less frequently used than previously.

Creatinine is actively secreted into the urine, so that its clearance tends to overestimate the GFR. This effect is of little significance at normal filtration rates, the ratio (creatinine clearance/inulin clearance) being between 1.1 and 1.2, but in advanced renal disease the contribution of active secretion to the total amount of creatinine excreted in the urine becomes high in relation to the amount filtered, and creatinine clearance may significantly overestimate the true GFR. This is despite the fact that in established renal failure, bacterial degradation of creatinine secreted into the gut may contribute approximately 2 mL/min to the total clearance.

Inaccuracies arising from methodological problems with creatinine measurement have largely been overcome, but the major cause of inaccuracy in the determination of GFR by creatinine clearance is the accurate measurement of urine volume. Traditionally, patients have been required to collect urine over a 24 h period. This is a convenient time, but requires the patient to void their urine completely at the beginning of the 24 h period and collect all the urine passed over the ensuing 24 h, making a final collection at the end of this period. The possibilities for mistakes resulting in an incomplete collection are considerable and even under ideal conditions, for example in a metabolic unit with highly motivated patients, the coefficient of variation for repeated measurements in the same individual is > 10%. The critical difference (the amount by which two estimates must differ to give a 95% probability that there has been a true change in GFR) is therefore > 33%. The accuracy of estimates of the GFR by measurement of creatinine clearance may be improved by making two or more consecutive 24 h urine collections, but this is often not practical even if it is acceptable to the patient.

There is, however, nothing special about the 24 h period. The use of shorter periods, although more convenient for the patient, may reduce accuracy through inadequate bladder emptying, but a good compromise is to make a collection overnight. As long as the bladder was emptied at the beginning of the test (before retiring) and when the final collection was made (on rising), the urine production rate can be calculated and substituted in the formula.

It is essential that a reliable method is employed to measure plasma and urine creatinine concentrations. Colorimetric assays tend to overestimate creatinine since they detect non-creatinine chromogens. But even when reliable methods are used, it should be appreciated that the creatinine clearance is based on four measurements: plasma and urine creatinine concentrations, urine volume and time. Each has an inherent inaccuracy and the overall analytical variance will be the sum of the individual variances.

This is of critical importance to clinical practice, since in early renal disease (when it might be supposed that therapeutic intervention would be most beneficial), the plasma creatinine concentration may be ‘normal’ – that is, within the reference range – in spite of the GFR being reduced. Plasma creatinine concentration is thus a relatively insensitive index of mild renal functional impairment. At low values of GFR, however, it becomes extremely sensitive, although its accuracy declines because of the increased contribution of tubular secretion to overall renal excretion.

Other factors that must be taken into account in assessing the significance of plasma creatinine concentrations include muscle mass, diet (a recent meal including meat, particularly if stewed, may cause a transient increase in plasma creatinine concentration) and the presence in the plasma of substances that interfere with the assay. Ketones, bilirubin, cephalosporins and spironolactone have all been implicated, depending on the method used, but especially in Jaffé-(picric acid) based assay systems. Enzymatic assays for creatinine are said to be more specific than older colorimetric methods, but they are still prone to interference by certain drugs such as acetylcysteine and phenindione.

The importance of muscle mass is exemplified by the changes that occur with age. In healthy individuals, GFR, and thus creatinine clearance, declines with increasing age from about the end of the 4th decade, at a rate of approximately 1 mL/min per year. However, plasma creatinine concentration does not normally rise with age; this is because of a decrease in production rate, thought to reflect the tendency for muscle mass to fall with age.

Although comparison of a measured plasma creatinine concentration with a reference range can be misleading, the fact that intraindividual variation is less than interindividual variation means that the detection of a change in creatinine concentration in an individual provides a more sensitive indication of a change in renal function. Nevertheless, the biological and analytical variance (even using the best assays) results in a critical difference of about 17% (20 μmol/L at a plasma creatinine concentration of 120 μmol/L).