Difference between revisions of "Kidney"

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Introduction

Homeostasis can be defined as the maintenance of a constant internal environment within an organism, whether unicellular or multicellular. Sufficient cellular function can only be maintained within a narrow range of conditions, thereby validating the undeniable significance of homeostatic mechanisms in life. The concept of homeostasis was first pioneered in the 19th century by the French physiologist Claude Bernard. He famously stated in 1857 that “La fixité du milieu intérieur est la condition de la vie libre”, which can be translated as “the constancy of the internal environment is necessary for free life” . Light can be shed on the underlying meaning of this maxim by comparing an organism with a wide array of sensitive and efficient homeostatic mechanisms, to one with none whatsoever. Whilst the latter can only inhabit an environment in which the external conditions are relatively constant, the former can exploit many more environments- hence Bernard’s “vie libre”, or “free life”- due to its ability to maintain a constant internal environment.

The mammalian kidney plays a pivotal role in maintaining homeostasis. This organ mediates not just one, but several aspects of homeostasis , including the excretion of certain metabolites and excess water and ions; the regulation of electrolyte concentration in the plasma and interstitial fluid; osmoregulation; the control of blood pressure; the maintenance of an equilibrium in pH; erythropoeisis, the process by which red blood cells are produced; and glucose homeostasis. Consequently, the kidney is of vital importance to the survival and efficient functioning of cells. Its role in maintaining the constancy of such a large number of conditions suggests the need to understand the kidney’s profound role in the body. As a result, this essay will focus on the ways, and associated mechanisms, in which the kidney aids in the regulation a constant internal environment.

The nephron: Ultrafiltration, reabsorption and secretion

To begin, the kidney is crucial in the excretion of excess water, ions and waste products, such as urea, uric acid and creatinine. The removal of excess water and ions is required to ensure that the solute potential (ψs) of interstitial fluid, the fluid that perfuses the tissues in the body, and of the plasma, remain relatively constant. Fluctuations of the solute potential of the tissue fluid can result in too many water molecules moving by osmosis either into or out of cells across their cell surface membranes, thus causing osmotic damage. More specifically, the former scenario, caused by hypotonic interstitial fluid, can lead to cell lysis , whereas the latter, which is induced by a hypertonic fluid, can lead to crenation3. In order to understand the intricacies of this filtration and excretion process, an overview of the anatomy of the kidney is first necessary.

A transverse section of a kidney will show that it consists of three major regions: the outer cortex (covered by a fibrous capsule), the inner medulla, composed of the renal pyramids, and the collecting duct system, consisting of the renal pelvis . Located within the kidney are microscopic tubules, the nephrons. These are responsible for the filtration of the blood, the reabsorption of substances useful to the body back into the blood, and the removal and subsequent excretion of metabolites and excess water and ions. The ‘head’ of these nephrons is the renal corpuscle, or Malpighian body, which consists of a Bowman’s capsule and the glomerulus. The glomerulus consists of a ‘knot’ of capillaries enclosed by the capsule . The Malpighian body is found in the cortex, as is the first segment of the nephron, the proximal convoluted tubule. The tubule then descends into the medulla, where it forms the loop of Henle. The loop of Henle consists of a descending limb and an ascending limb, forming a U-shaped loop whereby the descending loop undergoes a hairpin bend in the medulla to form the ascending limb. The third segment of the tubule is the distal convoluted tubule, which is again found in the cortex. The DCT subsequently joins with a collecting duct; the collecting ducts finally converge at the renal pelvis and transport the fluid into the ureter, through which the solution, now called urine, is carried to the bladder for ejection from the body.

In terms of the renal vasculature, the kidney is supplied by the renal artery, and the blood is removed via the renal vein. Arterioles originating from the renal artery, the afferent arterioles, supply the glomerulus, and efferent arterioles carry the blood away from the corpuscle. Subsequently, the efferent arteriole develops into peritubular capillaries and the vasa recta , whose purpose will be discussed in more detail later. The peritubular capillaries are responsible for the reabsorption of useful substances from the luminal fluid in the nephron into the bloodstream.

The nephron is the fundamental unit of the kidney, the parenchyma; it has three significant and distinct roles: filtration, reabsorption and secretion. Nephrons, of which there are approximately 106 in each human kidney , are highly specialised to carry out these roles. The first of these, filtration, occurs at the Malpighian body in the renal cortex. To gain an insight into this filtration mechanism, a microscopic view of the renal corpuscle, as well as the way in which the Bowman’s capsule and glomerular capillaries are adapted for this purpose, are necessary. The filter itself consists of three layers: the epithelial cells of the Bowman’s capsule, the endothelial cells of the capillaries, and a basement membrane that lies between these two cell layers . The epithelial cells, called podocytes are unique in their structure. They have ‘feet’, or primary processes, which themselves split to form smaller secondary processes. These secondary processes extend down and attach to the basement membrane ; the miniscule gap formed between two adjacent processes, a filtration slit, is an essential feature in the filtration process. In addition, the presence of fenestrated endothelia in the glomerular capillaries selectively allows molecules to pass through in relation to their size, shape and charge . Moreover, the basement membrane, which acts as an effective dialysing membrane, further restricts the permeability of the capillary wall by preventing molecules greater than a particular size passing through. The basement membrane is the most restrictive of the three layers of the Malpighian body10. The overall effect achieved by the three layers is that certain blood proteins (such as albumin, due to its negative charge) and red blood cells, due to their size, are retained in the capillaries, are therefore not present in the glomerular filtrate. Only uncharged molecules with an effective radius of 1.8nm or under are able to pass through freely, whereas molecules with an effective radius of greater than 4.0nm are completely restricted .

The end result is that the renal corpuscle essentially acts as a molecular sieve. This process by which the blood is filtered at the molecular level under pressure is called ultrafiltration. There are, however, certain forces acting within the renal corpuscle that must be taken into consideration. The colloid osmotic pressure produced by the blood proteins and red blood cells that have been retained in the capillaries is one such force. The retention of these components ensures that the water potential of the blood in the capillaries is lower than that of the glomerular filtrate, thereby drawing fluid out of the space in the Bowman’s capsule by osmosis . Similarly, the hydrostatic pressure produced by the glomerular filtrate also tends to prevent the entry of fluid into the capsule space. In order to overcome the combined effect of these forces, the hydrostatic pressure of the plasma in the glomerular capillaries must be significantly higher, thus ensuring the entry of fluid into the capsule space at a sufficient rate, and, therefore, the eventual excretion of metabolites. This high hydrostatic pressure is achieved by the fact that the lumen of the efferent arteriole is markedly narrower than that of the afferent arteriole8. By restricting blood flow thus, hydrostatic pressure is augmented. A relatively high and constant glomerular filtration rate (GFR) is maintained by a process called auto regulation, whereby the muscle tone in the arterioles is controlled in response to fluctuations in blood pressure. In humans, approximately 125 cm3 of plasma in the glomerular capillaries is filtered out in the Malpighian body per minute . To illustrate the importance of the high hydrostatic pressure produced in the blood in the glomerulus, the table below (reproduced from Biology: Principles and Processes, page 371) conveys the magnitude of the forces involved within the renal corpuscle:

Pressure Value (kPa) Hydrostatic pressure of blood in glomerulus 8.0 Hydrostatic pressure of fluid in capsule 2.4 Osmotic pressure of blood in glomerulus 4.3 Net filtration pressure 1.3

The glomerular filtrate now proceeds into the proximal convoluted tubule. The epithelial cells in the PCT are highly specialised for their main purpose: the reabsorption of useful substances into the peritubular capillaries that run adjacent to the nephrons. The PCT, couple with the DCT, are so effective that in a healthy kidney, approximately 99.5% of water and ions are reabsorbed2. One important substance that is entirely reabsorbed by the PCT (in a healthy kidney) is glucose. To begin, glucose is transported from the lumen of the tubule into the epithelial cells through the apical membrane via a Na+-glucose symporter (particularly SGLT-2, which is responsible for most of the glucose reabsorption at the apical membrane ). Both sodium cations and glucose are both simultaneously transported into the epithelial cells in a 2:1 molar ratio (SGLT-1) or a 1:1 molar ratio (SGLT-2) . The sodium-glucose cotransporter exploits the downhill electrostatic gradient of sodium ions in order to move glucose against its concentration gradient, rendering the system an example of secondary active transport. Indeed, this results in glucose accumulating in the epithelial cells. As a result, glucose can now move into the interstitial fluid surrounding the nephron via facilitated diffusion14, as achieved by the protein glucose permease (GLUT), and into the intercellular spaces and basal channels due to the establishment of a concentration gradient. Subsequently, the glucose moves into the peritubular capillaries, once again down a concentration gradient. The sodium ions, however, are reabsorbed into the capillaries via various intrinsic proteins in the basolateral membrane, including Na+-K+ ATPase and Na+-3HCO3- cotransporters, whose role in bicarbonate reabsorption will be discussed later. The sodium ions accumulate in the intercellular spaces in a similar fashion to glucose and finally diffuse into the capillaries. Chloride (Cl-) ions follow these sodium ions into the bloodstream passively, down an electrostatic gradient. The reabsorption of water is osmotically induced. The movement of ions and glucose into the peritubular capillaries reduces the solute potential of the plasma; on account of this, water moves into the capillaries by osmosis, via aquaporins situated in the cell surface membranes of the epithelial cells. In addition, low relative molecular mass proteins (less than 60,000Da ), as well as low concentrations of albumin , that were able to penetrate the layers in the renal corpuscle are removed by pinocytosis, followed by degradation by hydrolytic enzymes in lysosomes . Transepithelial reabsorption of amino acids occurs at the proximal convoluted tubule, as does that of organic acids, such as lactate and acetate, and of sulphate and phosphate anions by cotransport with Na+.

The PCT carries out most of the reabsorption, whereas the DCT is needed for the ‘fine control’ of electrolyte concentration and balance, in association with the endocrine system. In order to undertake reabsorption at such a large scale, therefore, the epithelial cells of the PCT are specialised in several ways. The cytoplasm contains a high density of mitochondria, which provide the ATP necessary for active transport; for example, the movement of sodium ions at the basolateral membrane is dependent by Na+-K+ ATPase on the hydrolysis of ATP. Furthermore, the presence of microvilli at the apical membrane greatly augments the surface area, thus facilitating the reabsorption of substances . There is a variety of enzymes at the brush border, such as carbonic anhydrase, whose role will be explored later.

Much of the urea, creatinine and uric acid are removed from the blood by ultrafiltration at the Malpighian body. Some creatinine is actively secreted into the lumen of the proximal convoluted tubule. The excretion of these metabolites is vital to keep the sufficient functioning of cells. Hyperuricemia, or elevated levels of serum uric acid, can lead to diseases such as gout, and is often associated to the Lesch-Nyhan syndrome . This problem is compounded by the fact that humans and other primates lack the enzyme urate oxidase, which is responsible for the breakdown of uric acid . The excretion of urea is required to maintain nitrogen equilibrium, which can be off-set by an excess intake of protein in the diet. Urea, CO(NH2)2, is synthesised in the liver in two stages: first, the deamination of amino acids to produce ammonia, and second, the use of this ammonia in the ornithine cycle to form urea. In an adult human, approximately 30g of urea is synthesised every day on an average diet , thus illustrating the importance of excreting this large amount of substance. Moreover, several substances are secreted by the distal convoluted tubule, including ammonia, hydrogen (H+) ions and penicillin.

Osmoregulation

The kidney’s role in the control of osmotic pressure in the body is of undeniable significance. For one, maintaining a constant water potential of tissue fluid prevents osmotic damage to cells, either by cell lysis or crenation. Further, a constant osmotic pressure maintains a constant hydrostatic pressure in the blood; it also ensures that the tissues are perfused by a sufficient volume of interstitial fluid, whilst preventing oedema. As the amount of water determines the concentration of ions and other substances in the interstitial fluid, it is crucial to regulate the concentration of water molecules in order to ensure the efficient function of cells. In addition, it preserves the necessary concentration gradients required for the efficient transport of ions across the phospholipid bilayer of cell membranes by the various intrinsic proteins. Electrolyte balance is essential as the relative concentrations of the different types of cations and anions in both intracellular and extracellular fluid is necessary for the execution of many essential processes. For example, the intracellular and extracellular concentrations of Na+ and K+ ions is significant in neurones, as they play a vital role in the establishment of resting potentials, and for depolarisation and subsequent repolarisation. Sufficient water is necessary as it is also a metabolite: it plays a crucial role in many reactions and pathways, such as respiration and hydrolysis (for example, of glycogen and proteins). In addition, a suitable amount of water is necessary due to its ability to act as an effective temperature buffer to the body, on account of its relatively high specific heat capacity.

The first important feature in the kidney that helps to regulate a constant osmotic pressure is the loop of Henle. The loop of Henle employs a countercurrent multiplier mechanism in the renal medulla, which causes urine to be concentrated. Consequently, this is, in essence, a water preserving mechanism, as water from the urine is reabsorbed into the bloodstream. The loop itself consists of three segments: the descending limb, the thin ascending limb and the thick ascending limb. The ascending and descending portions of the loop of Henle differ markedly in terms of their structure and properties, and it is this difference, coupled with the hair-pin bend of the loop in the medulla, that allows for this countercurrent mechanism to occur.

The descending limb is highly permeable to water, on account of the expression of the water channel aquaporin-1 (AQP1) & , and moderately permeable to ions and urea. In contrast, however, the ascending limb is impermeable to water. The apical membrane of epithelial cells in the thick segment of the ascending limb contains a Na+/K+-2Cl- cotransport system , which involves the transport of sodium, potassium and chloride ions across the membrane in a 1:1:2 molar ratio. Na+-K+ ATPase at the basolateral membrane maintains a low concentration of sodium cations within the epithelial cells, thus establishing the concentration gradient for sodium ions necessary for the functioning of the cotransport system. The result is that sodium chloride is transported into the medullary interstitium, causing the interstitial fluid to become hypertonic. Further, the expression of urea transporter 2 (UT-A2) in the epithelial cells of the thin descending limb26 facilitate the movement of urea out into the interstitial fluid. Thus, this contributes to establishing a hypertonic interstitium in the medulla. Water from the luminal fluid in the ascending limb does not follow by osmosis, as it is impermeable to water. Conversely, the descending limb is permeable to water, thereby resulting in water molecules diffusing from the lumen of the nephron into the interstitial fluid, down the osmotic gradient created by the sodium chloride removed from the ascending limb. The water is subsequently reabsorbed into the capillaries. This removal in water results in an increase in the osmolarity of the luminal fluid in the descending limb.

Approximately 20% to 25% of the sodium, chloride and potassium filtered out into the Bowman’s capsule are reabsorbed in the thick ascending limb28. The continuous influx of sodium chloride from the proximal convoluted tubule ensures that a high concentration of sodium chloride in the medullary interstitium is maintained. Whilst the luminal fluid in the descending limb becomes increasingly hyperosmotic down the length of the limb, the fluid in the ascending limb becomes increasingly hypotonic, as sodium, potassium and chloride are being removed with the aid of the Na+/K+-2Cl- cotransport system in the apical membrane. Although the difference in the osmolarity of the fluid in adjacent portions of the ascending and descending limbs is slight, the effect is additive. This countercurrent multiplier mechanism, therefore, is termed as it is because the concentrating effect of each segment of the loop of henle is “multiplied” along the length of the loop, hence multiplier; and because the filtrate in the descending limb is flowing in an opposite direction to that in the ascending limb, hence countercurrent.

The result of the countercurrent multiplier mechanism employed by the loop of Henle is that the osmolarity reaches a maximum of 1200mOsm kg-1H2O at the papillary tip. The medullary interstitium becomes increasingly hypertonic, whereby the osmolarity augments from 280mOsm kg-1H2O to 1200mOsm kg-1H2O at the apex of the loop . It should be noted, however, that of the two classified types of nephron- the cortical (superficial) and the juxtamedullary- only the juxtamedullary nephrons are responsible for the enhanced osmolarity of the interstitial fluid. This is because their loops of Henle extend deep down into the medulla, allowing the interstitial fluid to be made increasingly hypertonic right to the tips of the papilla. The loops of Henle of cortical nephrons do not extend sufficiently deep down into the renal medulla. In addition, the vasa recta are important in maintaining the high osmolarity in the interstitium. By employing a similar countercurrent multiplier mechanism , as enabled by their hair-pin arrangement, the vasa recta prevents dissipation of the solutes in the interstitium. Due to the increasing osmolarity of the interstitium as the vasa recta proceeds deeper into the medulla, a growing volume of water is osmotically drawn out of the descending vasa recta. This continues until at the apex of the loop, the osmolarity of the plasma equilibrates with that of the surrounding interstitial fluid. The ascending vasa recta, however, regains the water, and a large proportion of the solutes in the plasma is lost and retained in the interstitium. The augmented viscousity of the blood towards the apex reduces blood flow in the vasa recta, which further prevents dissipation of solutes.

The reabsorption of the water drawn out from the descending limb is one of the two aforementioned water preservation mechanisms. The establishment of a markedly hypertonic medullary interstitium is closely linked with the second of these: the collecting ducts and the action of arginine vasopressin, AVP . As the collecting tubule descends further into the medulla, an increasing amount of water is lost from the duct via osmosis on account of the increasing osmolarity of the interstitial fluid in the medulla. This water is subsequently reabsorbed into the capillaries under the influence of oncotic pressure . The permeability of the collecting ducts, however, is largely influenced by the hormone AVP, whose secretion is, in turn, regulated by a classical negative feedback loop involving the pituitary gland and osmoreceptors in the hypothalamus.

When an increase in the plasma osmolarity is detected by the osmoreceptors, AVP is secreted from the posterior pituitary gland into the blood. Once it reaches the kidney, the hormone induces a markedly enhanced permeability of the collecting ducts and the distal convoluted tubule to water. AVP achieves this by activating the receptor V2 , a subtype of vasopressin receptor that is exclusively expressed in the kidney, found on the basolateral membrane of the tubular cells. Upon activation, the action of AVP is mediated by cyclic AMP (cAMP), which acts as the secondary messenger33. Cyclic AMP is synthesised from ATP via the enzyme adenylyl cyclase, which is in turn activated by G-proteins coupled to the V2 receptors. The subsequent cAMP cascade (achieved through the activation of protein kinase A) results in aquaporin-2 (AQP 2) found in storage vesicles in the cell to be inserted into the apical membrane by exocytosis . In addition, the activity of urea transporter A1 (UT-A1) in the inner medullary collecting ducts is stimulated by the action of AVP . This causes urea to diffuse into the medullary interstitium, thereby raising its concentration in the area and rendering the interstitial fluid even more hypertonic. As a result, the efflux of water down the osmotic gradient is further enhanced. The ameliorated diffusion of water down an osmotic gradient and into the bloodstream brings about a decrease in the osmotic pressure of the plasma. This is detected by the osmoreceptors, which causes secretion of AVP to be inhibited. Similarly, in response to an unusually low plasma osmotic pressure, AVP secretion by the posterior pituitary will be inhibited. The necessity of AVP and of the role it plays in the kidney can be best demonstrated by a condition in which the action or secretion of AVP is somehow interrupted: diabetes insipidus. Diabetes insipidus results in excessive thirst and production of copious amounts of dilute urine . The causes of this condition can vary; nephrogenic diabetes insipidus, for example, can be caused by a defect in the V2 receptor protein.

The regulation of blood pressure: The Renin-Angiotensin-Aldosterone System

The maintenance of a constant blood pressure, which is closely associated with fluid balance, is equally essential to the survival of mammalian organisms. Hypotension, the condition where the arterial pressure is abnormally low, and hypertension, when the arterial pressure is abnormally high, can both have a significant impact on the body. The causes for these conditions are numerous, ranging from haemorrhage and desanguination (inducing a decrease in blood volume) for the former, to obesity, sleep apnea and high sodium content in the diet for the latter. In terms of capillary haemodynamics, the effects of an abnormal blood pressure are clear. Hypotension will result in the hydrostatic pressure at the arterial end being too low. Not only will the markedly lower hydrostatic pressure at the arterial end cause less tissue fluid to perfuse the cells, but also the effect of the oncotic pressure at the venous end will be far greater than usual. The result is that there is insufficient perfusion of the tissue by interstitial fluid, which in turn prevents efficient cellular function. Conversely, an abnormally high blood pressure results in a high hydrostatic pressure in the capillaries; as a result, interstitial fluid accumulates in the tissues, eventually causing oedema. In addition, the damage caused to capillary walls, and hence to organs that contain a large amount of capillaries, can create severe problems. Indeed, on a larger scale, hypertension can bring about stroke, heart failure and chronic renal disease.

The kidney is integral in regulating blood pressure, and more specifically in responding to hypotension. It achieves this through two primary mechanisms: the action of AVP on the collecting tubule, and the renin-angiotensin-aldosterone system (RAAS). In the case of the former, fluid volume is associated with blood pressure. By enhancing the permeability of the collecting tubules to water and urea, AVP ameliorates the reabsorption of water into the blood; this, in turn, augments fluid volume and restores blood pressure to normal levels. The modus operandi of the latter mechanism, RAAS, is centered around the principle of solute potential and the tubular reabsorption of Na+ cations, and on the many effects of its main effector, angiotensin II. An increased concentration of Na+ in the plasma will draw water from the luminal fluid back into systemic circulation, on account of the reduced solute potential, or ψs value. Consequently, blood pressure will be raised to within the normal limits.

The RAAS is the mechanism by which the reabsorption of sodium at the distal tubule in response to an abnormally low blood pressure is initiated. As can be inferred from the name itself, the RAAS is composed of three principle parts: the secretion of the enzyme renin, the generation of the oligopeptide angiotensin II, and subsequently the secretion of the hormone aldosterone. The juxtaglomerular cells (those associated with the afferent arteriole) are where the expression and storage of renin in the kidney predominantly occurs. Upon reduced perfusion of the juxtaglomerular apparatus due to a decrease in blood pressure, renin, which was stored in dense core secretory granules, is released, thus activating the RAAS . Renin is secreted into the interstitium, where it enters the bloodstream. Renin enters the lumen of nephrons by three means: 1) Via filtration at the Malpighian body, 2) production in the tubule itself, and 3) transport from the interstitium. Although renin itself does not have a direct effect on blood pressure, it is responsible for the cleavage of angiotensinogen, an inactive peptide synthesised in the liver, to produce angiotensin I . Renin that remains in circulation can act as an endocrine messenger to other parts of the body by activating local renin-angiotensin systems in other tissues.

Angiotensin I is subsequently converted into angiotensin II by the angiotensin-converting enzyme (ACE); although angiotensin I does play a minor role in the regulation of blood pressure, angiotensin II is by far the main effector of the system. The detection of angiotensinogen mRNA in the cells of the proximal tubule suggests localised synthesis of the peptide, though the liver firmly remains the primary site for angiotensinogen production . Secretion of angiotensinogen via the apical membrane, in addition to freely-circulating angiotensinogen from the liver that has filtered out at the renal corpuscle, are converted into angiotensin I. Moreover, high concentrations of ACE at the brush border of the apical membrane catalyse the conversion of angiotensin I to angiotension II54. It should be noted that there are other sites where ACE is expressed, and hence where angiotensin II can be generated, such as the pulmonary capillaries.

The local RAS in the kidney is unique on account of the way in which angiotension II effects an increase in blood pressure. Tubular angiotensin II acts on both the apical and basolateral membranes of the proximal tubule through AT 1 receptors, which are the major route by which angiotensin II affects the proximal tubule . By stimulating apical Na+-H+ antiporters and basolateral Na+-3HCO3- cotransporters, sodium reabsorption is greatly enhanced . More water from the tubular fluid, therefore, will follow by osmosis. Similarly, it can be inferred from the presence of AT receptors in the distal segments of nephrons that angiotensin II also mediates distal sodium reabsorption. Stimulation of sodium channel activity and the enhanced synthesis of Na+ channels in the cortical collecting duct, the latter induced by chronic action of Ang II,56 are two ways in which Ang II increase sodium reabsorption in more distal regions of the nephron. Furthermore, Ang II in the renal interstitium regulates blood pressure by improving tubular sodium reabsorption and controls microvasculature through vasoconstriction. Ang II also mediates the vasoconstriction of the afferent arteriole, and has a markedly greater effect on it than the efferent arteriole ; the result is that glomerular hydrostatic pressure is augmented, and the GFR is maintained.

The second major mechanism involves the hormone aldosterone. Ang II stimulates the secretion of aldosterone from the adrenal cortex , and more specifically the zona glomerulosa, the site where the enzyme aldosterone synthase acts. Aldosterone itself promotes sodium reabsorption and potassium secretion in the cortical collecting tubule . Although it is responsible for only a small proportion of Na+ ions that is reabsorbed, the role of aldosterone must nonetheless not be underestimated. The fine control over the concentration of sodium ions in the plasma can have a marked effect on blood pressure, and is therefore a pivotal variable to control when regulating blood pressure. Aldosterone acts on the distal segments by increasing the permeability of the apical membrane, by recruiting more Na+ channel proteins, and by activating basolateral Na+/K+ ATPase and increasing the number of pumps .

The pathway of the renal RAAS can be summarised below:

                                      Renin                                       ACE                                 Action on adrenal cortex 

Angiotensinogen Angiotensin I Angiotensin II Aldosterone

The action of angiotensin II is not restricted to just the kidney. Amongst other effects, Ang II causes the secretion of AVP from the supraoptic region (found in mammalian brains) of the hypothalamus; is a potent dipsogen (stimulates the sensation of thirst) in an attempt to increase fluid volume; and results in the release of adrenocorticotropic hormone (ACTH) from the anterior pituitary gland. ACTH also induces an increase in aldosterone levels in the blood by direct action on glomerulosa cells . These mechanisms are designed to increase the fluid volume of the plasma in order to return blood pressure to normal levels. An understanding of the renin-angiotensin system has clear clinical significance. For example, the use of ACE inhibitors, such as Captopril, and of direct renin inhibitors, such as aliskiren, which obviates the binding of angiotensinogen to renin , are important ways in which hypertension can be treated.

Erythropoeisis

The maintenance of a constant hematocrit, the proportion of the volume of blood that consists of erythrocytes (red blood cells), is a significant aspect of homeostasis that is, in part, regulated by the kidney. The production of erythrocytes in the bone marrow is mainly mediated by the hormone erythropoietin (EPO) , a protein that is mainly synthesised in the kidneys. Erythropoiesis, the process of producing mature red blood cells, is of great importance on account of the vital role erythrocytes play in mammals. Each red blood cell in circulation contains several molecules of haemoglobin, a metalloprotein consisting of four globular protein subunits (α and β chains) and a heme group, consisting of a central Fe2+ cation covalently bonded to a heterocyclic ring, porphyrin, which acts as a ligand. It is this molecule that is responsible for the transport of oxygen, the molecule crucial in aerobic respiration in cells, and more specifically as the final electron acceptor in oxidative phosphorylation in mitochondria. In areas where there is a high partial pressure of oxygen, namely the lungs, haemoglobin absorbs oxygen. By contrast, haemoglobin unloads oxygen where it is required: at respiring tissue, where there is a low partial pressure of O2 and a high partial pressure of CO2.

The capacity and precision with which erythrocytes are produced in the bone marrow is great. The bone marrow has the ability to increase the rate of erythrocyte production from the normal by eight times66; in addition, the sensitivity of the EPO mechanism is such that the loss in the number of red blood cells can be precisely compensated for by a corresponding increase in erythrocyte production. In humans, erythrocytes are responsible for approximately 45% of the volume of blood, and the typical rate of production of erythrocytes in an adult human is around 200 billion cells a day . A large number of situations, ranging from haemorrhage and menstruation to increased altitude, where there is an augmented oxygen tension, can stimulate erythrocyte production, in order to maximally satisfy the oxygen demand of the body. A failure to supply a particular region of the body with sufficient oxygen results in tissue hypoxia, which in turn can lead to necrosis.

The EPO mechanism, as mentioned previously, is dependent on the degree of oxygen tension. The kidney essentially acts as a “critmeter” , responding to a low ambient partial pressure of oxygen, and hence a low hematocrit, by secreting erythropoietin. EPO messenger RNA detected in the peritubular fibroblasts of the renal cortex and outer medulla has confirmed that these interstitial cells are indeed the site of EPO secretion . The kidney detects reduced oxygen availability by the presence of hypoxia sensitive units, known as hypoxia-inducible factors, in these fibroblasts. Hypoxia-inducible factors, of which there are two types, HIF-1 and HIF-2, collectively known as HIFs, are the key in the mechanism through which the detection and response to hypoxia is effected. HIFs are heterodimeric transcription factors , meaning that they consist of two subunits: an oxygen-sensitive α−subunit, which is the fundamental to the EPO mechanism, and a β−subunit. Upon the heterodimerisation of the α−subunit with HIF-β, the transcription factor, classified as one of a group of heterodimeric basic helix-loop-helix (bHLH) transcription factors, is activated. Subsequently, the genes controlled by this transcription factor, which includes the EPO gene, are upregulated.

The formation of HIF relies on the stabilisation of the α−subunit, followed by its translocation to the nucleus. Under normoxia, the α−subunit is hydroxylated (the process by which the compound is oxidised via the introduction of –OH, or hydroxyl, groups) by a group of cytoplasmic enzymes, known as HIF prolyl-4-hydroxylases. This hydroxylation allows the HIF-α to bind to an ubiquitin ligase complex, the process of polyubiquination, which in turn labels it for proteasomal degradation. Ergo, this obviates activation of the transcription factor, as the heterodimerisation of HIF-α and HIF-β is prevented. In mammals, three types of HIF prolyl-hydroxylases that have been identified: PHD1 (prolyl-hydroxylase domain), PHD2 and PHD3. PHD2 seems to be responsible for the hydroxylation, and hence the proteolysis of, HIF-α, PHD3 appears to hydroxylate the α-subunit during reoxygenation.

During hypoxia, however, inactivation of the prolyl-hydroxylases occurs, resulting in the stabilisation of HIF-α on account proteasomal degradation of the subunit being inhibited. The consequent accumulation of HIF-α usually occurs as a result of the oxygen concentration falling below 5% . The stabilization of the α−subunit is essential as it is subsequently translocated to the nucleus, where it heterodimerises with HIF-β. The transcription factor is formed, which binds onto the DNA at a specific base sequence (RCGTG). Furthermore, a second regulatory step is also present in order to upregulate expression of EPO during oxygen deprivation: FIH. FIH, or factor-inhibiting HIF, is an asparaginyl hydroxylase that mediates the hydroxylation of an asparagine residue in HIF; this, in turn, disrupts recruitment of the transcription cofactor CBP/p300 . In hypoxic conditions, however, the activity of FIH is inhibited, thus enabling recruitment of the cofactor and enhanced transcription of genes targeted by HIF to occur. It is important to note the other genes controlled by HIF, which range from those concerned with energy metabolism, such as glucose transporter-1 (GLUT1), to those mediating angiogenesis. It is also of significance to recognise that there are forms of signaling which are independent from hypoxia-mediated HIF transcriptional activity. Angiotensin II , the final effector of the RAS, and nitric oxide (NO) are examples of substances that have shown to augment transcription of genes controlled by HIFs. In addition, mitochondrial reactive oxygen species (ROS), nitrogen oxide and certain oncogenes can inhibit prolyl-hydroxylase activity on HIFs70, thereby facilitating HIF-α stabilisation.

Upon the secretion of erythropoietin by the peritubular fibroblasts into systemic circulation, the hormone travels in the bloodstream until it reaches the bone marrow. EPO stimulates the production of mature erythrocytes by mediating the survival, proliferation and differentiation of unipotent erythroid progenitor cells , called erythroblasts. When EPO binds onto erythropoietin receptors in the bone marrow, the JAK2 (Janus Kinase 2) cascade is initiated, thereby protecting the erythroid progenitor cells from apoptosis . Thus, the viability of the erythroblasts is preserved, and this is of great importance because these progenitor cells are the precursors to the mature erythrocytes. Consequently, the erythroblast can undergo maturation. The progenitor cell loses its nucleus, and most of its intracellular membranes are degraded; the now non-nucleated form of the cell, termed a reticulocyte, then loses its ribosomes, thus preventing any further synthesis of haemoglobin . At this stage, the cell is an erythrocyte and can enter circulation. It is clear, therefore, that reduced EPO secretion can lead to anaemia. During chronic renal disease, EPO secretion is significantly surpressed. The application of genetic engineering and the use of recombinant DNA, however, have allowed the in vitro production of EPO, known as Human Recombinant EPO (rhEPO). rhEPO has shown great promise in treating and preventing anaemia in patients suffering from chronic renal disease.

Glucose homeostasis

The regulation of the concentration of glucose in the blood is of utmost importance. Glucose is used as an energy source in tissue, whereby it is used to produce ATP via aerobic or anaerobic respiration. In particular, the brain is highly dependent on glucose as its energy source, being an obligate glucose consumer; neuroglycopenia, the condition where there is an inadequate supply of glucose to the brain, can affect function and may even to lead to coma. This underlies the need to prevent hypoglycaemia. Furthermore, a change in the concentration of free glucose in circulation can have a marked effect on the water potential of the blood. This, in turn, can bring about an imbalance in the amount of water moving into or out of tissues by osmosis. In this sense, modulating glucose concentration is closely linked with osmoregulation, further outlining the importance of preventing hypo- and hyperglycaemia. Previously, the kidney is thought to have contributed to glucose homeostasis in a minor way. This observation has been based on evidence derived from experiments that have employed a “net balance” approach: by measuring the net uptake or release of glucose by a particular organ. The inherent flaw in employing this approach is that it does not take into account that glucose uptake and release may be occurring simultaneously; the two processes may be compartmentalised, so uptake and release may be occurring in different locations within the same organ and may be differentially mediated. Consequently, the “net balance” approach may tend to underestimate the contribution of an organ such as the kidney to glucose homeostasis.

New techniques, such as those based on isotopic glucose dilution, in combination with the “net balance” approach, can illustrate with greater accuracy the role of the kidney in glucose homeostasis , which has been much underestimated. The claim that the liver is the only source of glucose in a normal postabsorptive state (namely, not after a period of prolonged fasting or in acidotic conditions) has been falsified. In addition to being an important source of glucose following prolonged fasting and in conditions of acidosis, studies have demonstrated that kidneys also release a significant amount of glucose in a normal postabsorptive state. The process by which this glucose is released is gluconeogenesis, a metabolic pathway that produces free glucose from certain substrates, such as pyruvate, lactate and glucogenic amino acids .

In terms of the physiology of the kidney, the sites of glucose uptake and glucose release are partitioned. Whereas glucose uptake primarily occurs in the renal medulla, glucose release is exclusively effected by the cortex. This compartmentalisation is due to the fact that different enzymes are expressed in the two different regions. Whilst cells in the renal cortex may contain the enzymes necessary for gluconeogenesis, there is an absence of gluconeogenic enzymes in the medulla, but contain enzymes that mediate glucose-phosphorylation and glycolysis. As a result, the gluconeogenic enzyme activity in cortical cells confers the ability to produce free glucose that can be released into circulation, yet the renal medulla is responsible for glucose uptake. These functionally compartmentalised sites, as previously mentioned, are differentially regulated. During hypoglycaemia or following a period of prolonged fasting, for example, glucose release via gluconeogenesis increases whereas renal glucose uptake decreases. One study on humans has shown that during hypoglycaemia (with the blood glucose concentration at 3.6mmol/l) the amount of glucose released by the kidney had doubled, and its contribution to total glucose release in the body had augmented from 22% to 36% . It is clear from this and many other studies that the kidney’s role in glucose homeostasis is significant and should not be underestimated. Although there is a marked reduction in the renal uptake of glucose during hypoglycaemia, in absolute terms this contribution to the overall reduction in glucose uptake in the body is less significant.

Moreover, it has been shown that renal glucose release can be hormonally modulated. Insulin, for example, reduces glucose release by suppressing gluconeogenesis whilst enhancing glucose uptake. It appears that the suppression of gluconeogenesis is primarily not brought about by simply reducing substrate availability, but rather by another yet unknown mechanism. For example, the insulin reduces fatty acid uptake and could therefore indirectly inhibit gluconeogenesis, as fatty acids have shown to stimulate gluconeogenesis in vitro; alternatively, the insulin could be shifting away the precursors of the gluconeogenic pathway . By contrast, adrenaline seems to trigger an increase in glucose release through gluconeogenesis; once again, the mechanism through which gluconeogenesis is further stimulated is unknown, and could also involve free fatty acids. Glucagon appears to have no effect on either glucose uptake or release in the kidney.

Another aspect of glucose homeostasis that the kidney contributes to is the body in a postprandial state. The increase in glucose uptake, in absolute terms, did not contribute as greatly to the overall increase in glucose uptake, as was the case with the reduction in renal uptake of glucose during hypoglycaemia. One study has shown that when a 75g load of glucose was ingested, the kidney only accounted for approximately 10% of the glucose removed . The main way in which the kidney responds to an increased glucose concentration, rather counterintuitively, is by increasing glucose release. Although the mechanism through which this is induced is unknown, it may be brought about by increased availability of precursors of the gluconeogenic metabolic pathway and via enhanced stimulation by the sympathetic nervous system. This release of glucose facilitates the decrease in blood glucose concentration is by allowing the inhibition of hepatic glucose release, permitting the liver to produce glycogen more efficiently79.

The nephron itself has a nominal role to play in glucose homeostasis. The efficiency with which glucose is reabsorbed in the proximal tubule is an essential way to prevent hypoglycaemia. When hyperglycaemia occurs, however, any excess glucose that cannot be reabsorbed is excreted in the urine. This occurs on account of the fact that the protein transport mechanisms have a maximum rate at which it can facilitate transepithelial movement, which is indeed the limiting factor in glucose reabsorption. When the glucose concentration reaches a threshold value, which is determined to be approximately 180-200 mg/dL in humans , the apical sodium-glucose symporters become “saturated”. The rate at which the substrates of the symporter can bind onto their respective domains on the protein is at a maximum, meaning that if glucose concentration is augmented above the threshold value, the excess glucose is simply excreted from the body.