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The Body’s water
1. 2. 3. 4. Body water and its determination Nature and composition of ECF and ICF Electrolyte balance pH balance
1. BODY WATER AND ITS DERMINATION
1.1 INTRODUCTION 1.2 CALCULATION OF BODY WATER 1.3 DETERMINATION OF BODY WATER
1.1 INTRODUCTION Body water is all of the water content of the human body. A significant fraction of the human body is water. Lean muscle tissue contains about 75% water. Blood contains 95% water, body fat contains 14% water and bone has 22% water. The human body is about 60% water in adult males and 55% in adult females. In diseased states where body water is affected, the compartment or compartments that have changed can give clues to the nature of the problem. Body water is regulated by hormones, including antidiuretic hormone (ADH), aldosterone and atrial natriuretic peptide. There are many methods that can be used to determine body water. One way to get a simple estimate is by calculation. 1.2 CALCULATION OF BODY WATER In individuals of normal weight, water is abundant in most parts of the body, except in adipose tissue (fat). These calculations are for adults of average build, and are inappropriate for obese or overly muscular people. These proportions are very simplified and use round numbers for quick calculation. The largest component of the body is water. Water makes up between 45 and 75% of body weight, with the variability due primarily to differences in body fat. While most tissues including muscle, skin, and visceral organs are over 70% water, adipose tissue contains less than 10% water. The percentage of body weight that is water therefore varies inversely with body fat. In the average lean adult male around 60% of the body weight is water. The remaining body weight consists of 16-18% fat with 22-24% protein, carbohydrate and other solids. In the female the percentage of body weight that is water is lower due to a relatively greater amount of subcutaneous fat. Body water is broken down into the following compartments: Intracellular fluid (2/3 of Body Water) Extracellular fluid (1/3 of Body Water) o Plasma (1/5 of Extracellular fluid) o Interstitial fluid (4/5 of Extracellular fluid) o Transcellular fluid (normally ignored in calculations) Contained inside organs, such as the gastrointestinal, cerebrospinal, and ocular fluids. The simplest calculation is the 60-40-20 rule.
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Total Body Water = 60% of Body Weight Intracellular fluid = 40% of Body Weight Extracellular fluid = 20% of Body Weight This is consistent with the above relations between total body water and the compartmental fluids. 1.3 DETERMINATION OF BODY WATER A) Dilution and equilibration Total body water can be determined using Flowing afterglow mass spectrometry FA-MS measurement of deuterium abundance in breath samples from individuals. A known dose of deuterated water (Heavy water, D2O) is ingested and allowed to equilibrate within the body water. The FA-MS instrument then measures the deuterium-to-hydrogen (D:H) ratio in the exhaled breath water vapour. The total body water is then accurately measured from the increase in breath deuterium content in relation to the volume of D2O ingested. Different substances can be used to measure different fluid compartments: total body water: tritiated water or deuterium extracellular fluid: inulin blood plasma: Evans blue
Fig 1. Schematic diagram of the FA-MS method The principle of the FA-MS technique is that it exploits ion chemistry coupled with fast flow tube techniques and quantitative mass spectrometry. However, FA-MS is specifically used for the online determination of the deuterium content of water vapour in exhaled breath and the vapour above aqueous liquids. Thus, for example, the deuterium content of single breath exhalations can be measured following the ingestion of a small known amount of deuterium oxide when, following the principle of isotope dilution, the total body water of the subject can be obtained, which is proving to be particularly useful in nephrology. B) Bioelectrical impedance analysis Another method of determining total body water percentage (TBW%) is via Bioelectrical Impedance Analysis (BIA). In the traditional BIA method, a person lies on a cot and spot
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electrodes are placed on the hands and bare feet. Electrolyte gel is applied first, and then a current of 50 kHz is introduced. BIA has emerged as a promising technique because of its simplicity, low cost, high reproducibility and noninvasiveness. BIA prediction equations can be either generalized or population-specific, allowing this method to be potentially very accurate. Selecting the appropriate equation is important to determining the quality of the results. For clinical purposes, scientists are developing a multi-frequency BIA method that may further improve the method's ability to predict a person's hydration level. New segmental BIA equipment that uses more electrodes may lead to more precise measurements of specific parts of the body. ***
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2. NATURE AND COMPOSITION OF ECF AND ICF
2.1 INTRODUCTION 2.2 EXTRA CELLULAR FLUID 2.3 INTRA CELLULAR FLUID 2.1 INTRODUCTION Water accounts for 60% of body mass in males, i.e. over 40 L in a 70 kg adult. Total body water is divided between intracellular (two-thirds of total body fluid) and extracellular compartments (onethird of total body fluid), with plasma forming a major sub-compartment (one-quarter of ECF) within the extracellular space. Water can diffuse freely between these fluid spaces and this exchanges ins controlled by pressure gradients across the cell membrane (which separates ECF and ECF spaces) and the capillary wall (which separates plasma from interstitial fluid). In each case the distribution of fluid depends on the net effect of hydrostatic and osmotic forces. To solutes controlling osmosis at the two sites, 2.1 EXTRA CELLULAR FLUID-nature and composition Extracellular fluid a general term for all the body fluids outside the cells, comprising about one third of the body fluids. Extracellular fluid circulates in the spaces between the cells and brings to the cells the nutrients and other substances needed for their functioning. The extracellular fluid also includes the transcellular fluid; making up only about 2.5 percent of the ECF. In humans, the normal glucose concentration of extracellular fluid that is regulated by homeostasis is approximately 5 mM. The pH of extracellular fluid is tightly regulated by buffers around 7.4. The volume of ECF is typically 15 L (of which 12 L is interstitial fluid and 3 L as plasma) In some animals, including mammals, the extracellular fluid can be divided into two major subcompartments, interstitial fluid and intravascular fluid (blood plasma). Interstitial fluid is the extracellular fluid bathing most tissues, constituting the environment of body cells, excluding the fluid within the lymph and blood vessels. It is low in protein, is formed by filtration through the capillaries, and drains away as lymph. Intravascular fluid a term sometimes used to refer to that part of the extracellular fluid that is within the blood vessels, in other words, the plasma.
Ionic composition of ECF The extracellular concentrations of a number of important ions are commonly determined suing plasma from venous blood samples. National values are summarized in Table.1. Although the total charge carried by all the cations and anions must be equal, a significant fraction of the anion loaded is not accounted for in routine measurements. It can be seen that the sum of the measures cations exceeds the sum of the measured anions, i.e. [Na+] + [K+] + [Ca+] = 142 mmol L-1 [Cl-] [HCO3-] = 130 mmol L-1 (103 + 27) The difference between these totals is called the anion gap; (17 mmol L-1 in this case). This represents the contribution made by anions which are not normally measured in the clinical laboratory, e.g. sulphate, phosphate, proteins and other organic anions of this sort, e.g. the anions
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released by dissociation of keto-acids in diabetes mellitus, will be assosicated with an increase in the anion gap.
2.2 INTRA CELLULAR FLUID-nature and composition The cytosol or intracellular fluid (or cytoplasmic matrix) is the liquid found inside cells. In eukaryotes this liquid is separated by cell membranes from the contents of the organelles suspended in the cytosol, such as the mitochondrial matrix inside the mitochondrion. The entire contents of a eukaryotic cell, minus the contents of the cell nucleus, are referred to as the cytoplasm. The ICF is a complex mixture of substances dissolved in water. Although water forms the large majority of the ICF, its structure and properties within cells is not well understood. The concentrations of ions such as sodium and potassium are different in the ICF than in the extracellular fluid; these differences in ion levels are important in processes such as osmoregulation and cell signaling. The ICF also contains large amounts of macromolecules, which can alter how molecules behave, through macromolecular crowding. Normal human ICF pH ranges between 7.3 - 7.5, depending on the cell type involved, whereas the pH of the extracellular fluid is 7.4. The viscosity of cytoplasm is roughly the same as pure water, although diffusion of small molecules through this liquid is about four-fold slower than in pure water, due mostly to collisions with the large numbers of macromolecules in the cytosol. Ionic composition of ICF Most of the ICF is water, which makes up about 70% of a typical cell by volume. The concentrations of the other ions in ICF are quite different from those in extracellular fluid and the ICF also contains much higher amounts of charged macromolecules such as proteins and nucleic acids than the outside of the cell. The major anions and cations are different from those found outside the cell.
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Intra cellular [Na+] is low (about 10 mmol L-1) and [K+] is high (about 160 mmol L-1). Intracellular [Cl-] is very low (2 mmol L-1) while proteins, phosphate and sulphate account for the majority of intracellular cations. In contrast to extracellular fluid, ICF has a high concentration of potassium ions and a low concentration of sodium ions. This difference in ion concentrations is critical for osmoregulation, since if the ion levels were the same inside a cell as outside, water would enter constantly by osmosis - since the levels of macromolecules inside cells are higher than their levels outside. Instead, sodium ions are expelled and potassium ions taken up by the Na⁺/K⁺-ATPase, potassium ions then flow down their concentration gradient through potassium-selection ion channels, this loss of positive charge creates a negative membrane potential. To balance this potential difference, negative chloride ions also exit the cell, through selective chloride channels. The loss of sodium and chloride ions compensates for the osmotic effect of the higher concentration of organic molecules inside the cell.
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3. ELECTROLYTE BALANCE 3.1 INTRODUCTION 3.2 ELECTROLYTE BALANCE 3.1 INTRODUCTION Electrolytes are minerals in your body that have an electric charge. They are in your blood, urine and body fluids. Maintaining the right balance of electrolytes helps your body's blood chemistry, muscle action and other processes. Sodium, calcium, potassium, chlorine, phosphate and magnesium are all electrolytes. You get them from the foods you eat and the fluids you drink. Levels of electrolytes in your body can become too low or too high. That can happen when the amount of water in your body changes. Causes include some medicines, vomiting, diarrhea, sweating or kidney problems. Problems most often occur with levels of sodium, potassium or calcium. To function normally, the body must keep fluid levels from varying too much in the areas of the body that contain fluid (called compartments). The three main compartments are Fluid within cells Fluid in the space around cells Blood Electrolytes, particularly sodium, help the body maintain normal fluid levels in these compartments (called fluid balance), because how much fluid a compartment contains depends on the concentration of electrolytes in it. If the electrolyte concentration is high, fluid moves into that compartment. If the electrolyte concentration is low, fluid moves out of that compartment. To adjust fluid levels, the body can actively move electrolytes in or out of cells. Thus, having electrolytes in the right concentrations (called electrolyte balance) is important in maintaining fluid balance among the compartments. The kidneys help maintain electrolyte concentrations by filtering electrolytes from blood, returning some electrolytes, and excreting any excess into the urine. Thus, the kidney help maintain a balance between daily consumption and excretion. If the balance of electrolytes is disturbed, disorders can develop. An electrolyte imbalance can result from the following: Becoming dehydrated Taking certain drugs Having certain heart, kidney, or liver disorders Being given intravenous fluids or feedings in inappropriate amounts 3.2 BALANCE OF ELECTROLYTES Individual cells work in conjunction with the kidneys and lungs to regulate and maintain normal water and electrolyte balance within the body. The kidneys have the largest responsibility for maintaining blood chemistry, and in concert with the lungs are responsible for regulating the acid-base balance within the blood. These processes involve both electrolytes and water.
Fig.2. Electrolyte balance in cells by Na pump mechanism Dr. M. Estari
3.2.1 Electrolyte balance In Cells (Fig. 2) Sodium (Na+) is the major ion, or electrolyte, outside of the cell. Potassium (K+) is the major ion inside the cell. Since cell membranes are permeable to Na+, it diffuses into the cell where its concentration is lower. To maintain the low intracellular Na+ concentration, the cell quickly pumps Na+ back into the extracellular fluid. Sodium pumps located in the cell membrane pump Na+ out of and K+ into the cell, maintaining normal osmotic gradients. 3.2.2 Electrolyte balance In Kidneys (Fig 3) The kidneys filter the blood to remove harmful metabolic acids and wastes and reabsorb those substances the body needs. They help control plasma ion concentration and maintain pH by removing strong ions such as Na+ and Cl- from plasma into urine. By removing more Na+ than Cl-, for example, the kidneys lower the plasma Strong Ion Difference [SID]. SID is simply the difference between the concentrations of positive and negative strong ions (Fig 3). As Na+ is removed, the number of positive ions decreases causing a relative increase in the concentration of negative ions. To offset the increase of negative charges created by Na+ removal, the concentration of hydrogen ions, [H+], increases. As [H+] increases, pH decreases, making plasma more acidic. On the other hand, removing more Cl- than Na+ raises plasma [SID], which lowers [H+] and raises pH.
Fig 3. Electrolyte balance in kidney tubules
The kidneys also control blood volume by regulating the amount of water in extracellular fluid. (Fig 3). Two hormones, aldosterone and antidiuretic hormone (ADH) help the kidneys control the fluid volume of blood. When water is lost from the body, blood volume decreases. This leads to increased production of aldosterone and ADH. Elevated aldosterone increases Na+ pump activity in the kidney tubules. As a result, more Na+ is removed from the distal tubules and concentrated within the kidney rather than being excreted. This high concentration of Na+ in kidney tissue creates an osmotic gradient that pulls water out of the tubules. Elevated ADH works in conjunction with aldosterone by increasing the permeability of the tubules to water, allowing water to follow Na+ out of the tubules, thereby reducing urinary water loss. 3.2.3 Electrolyte balance In Lungs The lungs play an important role in regulating plasma CO2 and pH. Since CO2 is a gas, the term partial pressure, PCO2, is used to describe its concentration in liquids. Changes in respiration rate relate to changes in the partial pressure of CO2. For example, CO2 is produced during exercise and
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is removed from muscle tissue by the blood, increasing PCO2. To quickly rid the blood of excess CO2, respiration rate increases. In this process, CO2 from the muscle combines with water to form carbonic acid in the blood, which then dissociates to H+ and bicarbonate ions (HCO3-). Bicarbonate is the primary storage and transportation form of CO2 in plasma. In the lungs, this process is reversed with CO2 and water being exhaled.
Fig 4. Electrolyte balance by Co2 transport in blood
As the rate of CO2 production increases, PCO2, [HCO3-] and respiration rate increase. If CO2 production outstrips the lungs’ ability to convert HCO3- to water and CO2, plasma [HCO3-] will continue to rise. Plasma [H+] will also rise, offsetting these, causing plasma pH to decrease. Acidosis = ↑PCO2 , ↑HCO3- , ↓pH Alkalosis = ↓PCO2 , ↓HCO3- , ↑pH ***
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4. pH BALANCE (ACID-BASE BALANCE) 4.1 INTRODUCTION 4.2 pH BALANCE BY METABOLIC REGULATION 4.3 pH BALANCE BY RESPIRATORY REGULATION 4.1 INTRODUCTION By regulating blood chemistry, the kidneys and lungs maintain an electrically neutral environment. The processes involved in maintaining electrical neutrality are referred to as acid-base balance. The power of the lungs to excrete large quantities of carbon dioxide enables them to compensate rapidly, while the smaller capacity of the kidneys corresponds to a relatively slower rate of compensation through metabolic means. Both must work in concert to maintain an acid-base/pH balance. 4.2 pH BALANCE BY METABOLIC REGULATION Strong ions are fully dissociated from each other in aqueous solutions such as plasma and form charged water complexes. For example, Na+ and Cl- do not associate with each other to form NaCl in plasma. Instead, Na+ associates with the O- component of water while Cl- associates with the H+ component. The orientation of water molecules to a strong ion counterbalances and isolates the ion’s charge, and exposes either the positive or negative portion of the water molecules. Charged water complexes of Na+ have an overall positive charge that attracts OH-, while negative charges associated with Clwater complexes attract H+. By selective removal of Na+ or Cl-, the kidneys adjust the relative proportion of H+ to OH- in plasma. As Na+ is removed, the amount of OH- required to offset positive charged water complexes decreases. Consequently, the amount of OH- in the solution decreases. As OH- decreases, the relative amount of H+ increases, bringing about a reduction in pH. On the other hand, removal of negative charged water complexes reduces the amount of H+ needed in the solution to offset negative charges. As the relative amount of OH- increases, the solution becomes more basic and pH rises. These processes are summarized below.
Fig 5. Electroneutrality maintained by kidney
4.3 pH BALANCE BY RESPIRATORY REGULATION Since bicarbonate (HCO3-) is the plasma transport form of CO2, its regulation falls under the jurisdiction of the lungs, not the kidneys. As previously discussed, the lungs quickly adjust the partial pressure of carbon dioxide in plasma (PCO2) by either increasing or decreasing respiration rate. As a result, plasma bicarbonate either decreases or increases. Combined Effect of Respiratory & Metabolic Regulation : Although the lungs and the kidneys have their own regulatory processes, they work together to maintain the electrical neutrality of plasma. As an example, consider a baby calf that is undergoing the common summertime problem of heat stress. In an attempt to get rid of extra heat, the calf’s respiration rate increases. Although rapid breathing rids the body of some excess heat, it also causes a loss of CO2, which lowers plasma PCO2. This lowers [HCO3-] and reduces the associated [H+] required to offset the negative charges associated with bicarbonate. As plasma
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[H+] decreases, the relative [OH-] increases, causing plasma to become more alkaline. Plasma pH rises. This situation is generally referred to as respiratory alkalosis. Figure 6 summarizes the changes that occur in the calf as a result of increased respiration rate due to heat stress.
Fig 6. Effect of increased respiration rate
To compensate, the kidneys remove Na+. As Na+ is removed, the [OH-] required to offset the positive charge of sodium is reduced. This reduction in plasma [OH-] increases the relative [H+], bringing plasma pH down to normal. Figure 7 summarizes the kidney response to the calf’s respiratory alkalosis caused by heat stress.
Fig 7. Respiratory alkalosis
Through their combined actions the lungs and kidneys may have averted a couple of potentially life-threatening situations. Nevertheless, the calf has lost body water during the heat stress and the removal of Na+ has lowered the plasma [SID] below normal in order to maintain electrical neutrality. Oral electrolyte therapy is an obvious remedy for both the water and electrolyte loss. If administered early enough, the electrolyte treatment could avert or at least lessen the heat stress and resulting physiological changes in the calf. pH in Body fluids : The pH of intracellular fluid is about 7.0 and about 7.4 for extracellular fluid. At normal body temperature, the pH of a solution is 6.8. Therefore, body fluids are actually maintained at a slightly alkaline pH. The pH range of physiological solutions is small, with a pH change of 1.0 being fatal. Ion movements in body fluids cause changes in pH, making it a dependant variable. As demonstrated above, significant changes can occur in plasma chemistry that result in virtually no change in pH. Furthermore, equal changes in [H+] and [OH-] in physiological solutions do not bring about equal changes in pH. To summarize, a change in pH indicates a problem. It does not, however, indicate what is causing the problem or what needs to be corrected. pH is not a very sensitive measure for evaluating the acid-base status or changes in status of body fluids. ***
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Chapter – 2
5. 6. 7. 8. Formation, composition and functions of blood Mechanism of blood coagulation Actions and uses of anticoagulants Thrombolytic therapy and Anti-coagulant therapy
5. FORMATION, COMPOSITION AND FUNCTIONS OF BLOOD 5.1 INTRODUCTION 5.2 FORMATION OF BLOOD 5.3 COMPOSITION AND FUNCTIONS OF BLOOD 5.1 INTRODUCTION Blood is a specialized bodily fluid that delivers necessary substances to the body's cells—such as nutrients and oxygen—and transports waste products away from those same cells. In vertebrates it is composed of blood cells suspended in a liquid called blood plasma. Plasma, which comprises 55% of blood fluid, is mostly water (90% by volume), and contains dissolved proteins, glucose, mineral ions, hormones, carbon dioxide (plasma being the main medium for excretory product transportation), platelets and blood cells themselves. The blood cells present in blood are mainly red blood cells (also called RBCs or erythrocytes) and white blood cells, including leukocytes and platelets (also called thrombocytes). Haemopoietic cells (those which produce blood) first appear in the yolk sac of the 2-week embryo. By 8 weeks, blood making has become established in the liver of the embryo, and by 12-16 weeks the liver has become the major site of blood cell formation. It remains an active haemopoietic site until a few weeks before birth. The spleen is also active during this period, particularly in the production of lymphoid cells, and the foetal thymus is a transient site for some lymphocytes. The highly cellular bone marrow becomes an active blood making site from about 20 weeks gestation and gradually increases its activity until it becomes the major site of production about 10 weeks later. At birth, active blood making red marrow occupies the entire capacity of the bones and continues to do so for the first 2-3 years after birth. The red marrow is then very gradually replaced by inactive, fatty, yellow, lymphoid marrow. The latter begins to develop in the shafts of the long bones and continues until, by 20-22 years, red marrow is present only in the upper ends of the femur and humerus and in the flat bones of the sternum, ribs, cranium, pelvis and vertebrae. However, because of the growth in body and bone size that has occurred during this period, the total amount of active red marrow (approximately 1000-1500 g) is nearly identical in the child and the adult. Adult red marrow has a large reserve capacity for cell production. In childhood and adulthood, it is possible for blood making sites outside marrow, such as the liver, to become active if there is excessive demand as, for example, in severe haemolytic anaemia or following haemorrhage. In old age, red marrow sites are slowly replaced with yellow, inactive marrow. Red marrow forms all types of blood cell and is also active in the destruction of red blood cells. Red marrow is, therefore, one of the largest and most active organs of the human body, approaching the size of the liver in overall mass although as mentioned it is distributed in various parts of the body.
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About two-thirds of its mass functions in white cell production (leucopoiesis), and one-third in red cell production (erythropoiesis). However as we have already seen there are approximately 700 times as many red cells as white cells in peripheral blood. This apparent anomaly reflects the shorter life span and hence greater turnover of the white blood cells in comparison with the red blood cells. 5.2 FORMATION OF BLOOD Hemopoiesis (hematoiesis) is the process that produces the formed elements of the blood. Hemopoiesis takes place in the red bone marrow found in the epiphyses of long bones (for example, the humerus and femur), flat bones (ribs and cranial bones), vertebrae, and the pelvis. Within the red bone marrow, hemopoietic stem cells (hemocytoblasts) divide to produce various ―blast‖ cells. Each of these cells mature and becomes a particular formed element. It is now generally accepted that all blood cells are made from a relatively few 'uncommitted' cells which are capable of mitosis and of differentiation into 'committed' precursors of each of the main types of blood cell.
Fig. 5.2 Hematopoiesis: The formation of the different types of blood cells from a common source the stem cell 5.2.1 Erythropoiesis (Formation of red blood cells) The production of red blood cells is referred to as erythropoiesis. Mature red blood cells develop from haemocytoblasts. This development takes about 7 days and involves three to four mitotic cell divisions, so that each stem cell gives rise to 8 or 16 cells. The various cell types in erythrocyte development are characterised by the gradual appearance of haemoglobin and disappearance of ribonucleic acid (RNA) in the cell the progressive degeneration of the cell's nucleus which is eventually extruded from the cell the gradual loss of cytoplasmic organelles, for example mitochondria a gradual reduction in cell size
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The young red cell is called a retlculocyte because of a network of ribonucleic acid (reticulum) present in its cytoplasm. As the red cell matures the reticulum disappears. Between 2 and 6% of a new-born baby's circulating red cells are reticulocytes, but this reduces to less than 2% in the healthy adult. However, the reticulocyte count increases considerably in conditions in which rapid erythropoiesis occurs, for example following haemorrhage or acute haemolysis of red cells. A reticulocyte normally takes about 4 days to mature into an erythrocyte. In health, erythropoiesis is regulated so that the number of circulating erythrocytes is maintained within a narrow range. Normally, a little less than l% of the body's total red blood cells are produced per day and these replace an equivalent number that have reached the end of their life span. However that still represents a huge 200,000,000,000 cells. Erythropoiesis is stimulated by hypoxia (lack of oxygen). However, oxygen lack does not act directly on the haemopoietic tissues but instead stimulates the production of a hormone, erythropoietin. This hormone then stimulates haemopoietic tissues to produce red cells. Erythropoietin is a glycoprotein. It is inactivated by the liver and excreted in the urine. It is now established that erythropoietin is formed within the kidney by the action of a renal erythropoietic factor erythrogenin on plasma protein, erythropoietinogen. Erythrogenin is present in the juxtaglomerular cells of the kidneys and is released into the blood in response to hypoxia in the renal arterial blood supply. In anaemia there is a reduction in blood haemoglobin concentration due to a decrease in the number of circulating erythrocytes and/or in the amount of haemoglobin they contain. Anaemia occurs when the erythropoietic tissues cannot supply enough normal erythrocytes to the circulation. In anaemias due to abnormal red cell production, increased destruction and when demand exceeds capacity, plasma erythropoietin levels are increased. However, anaemia can also be caused by defective production of erythropoietin as, for example, in renal disease. 5.2.2 Monocyte formation Monocytes are produced in the bone marrow, developing from nucleated precursors, the monoblast and promonocyte. Mature cells have a life in blood of approximately 3-8 hours and, like granulocytes, there is a circulating and marginating pool. Fig. 5.2.2 Formation of monocytes from phagocytic cells Monocytes are actively phagocytic (engulf other cells) and, on migration into the tissues, they mature into larger cells called macrophages (Derives from the Ancient Greek: macro = big, phage = eat), which can survive in the tissues for long periods. These cells form the mononuclear phagocytic cells of the mononuclear phagocytic system (reticuloendothelial system) in bone marrow, liver, spleen and lymph nodes. Tissue macrophages (sometimes called histiocytes) respond more slowly than neutrophils to chemotactic stimuli. They engulf and destroy bacteria, protozoa, dead cells and foreign matter. They also function as modulators of the immune response by processing antigen structure and
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facilitating the concentration of antigen at the lymphocyte's surface. This function is essential in order that full antigenic stimulation of both T and B lymphocytes can take place. 5.2.3 Granulocyte formation (Granulopoiesis/Leucopoiesis) Granulocytes are the collective name given to three types of white blood cell. Namely these are neutrophils, basophils and eosinophils. In terms of their formation (granulopoiesis) they all derive from the same type of committed stem cells called myeloblasts. After birth and into adulthood granulopoiesis occurs in the red marrow. The process of producing granulocytes is characterised by the progressive condensation and lobulation of the nucleus, loss of RNA and other cytoplasmic organelles, for example mitochondria, and the development of cytoplasmic granules in the cells involved. The development of a polymorphonuclear leukocyte make take a fortnight, but this time can be considerably reduced when there is increased demand, as, for example, in bacterial infection. The red marrow also contains a large reserve pool of mature granulocytes so that for every circulating cell there may be 50-100 cells in the marrow. Mature cells pass actively through the endothelial lining of the marrow sinusoid into the circulation. In the circulation, about half the granulocytes adhere closely to the internal surface of the blood vessels. These are called marginating cells and are not normally included in the white cell count. The other half circulate in the blood and exchange with the marginating population. Within 7 hours, half the granulocytes will have left the circulation in response to specific requirements for these cells in the tissues. Once a granulocyte has left the blood it does not return. It may survive in the tissues for 4 or 5 days, or less, depending on the conditions it meets. The turnover of granulocytes is, therefore, very high. Dead cells are eliminated from the body in faeces and respiratory secretions and are also destroyed by tissue macrophages (monocytes). 5.2.4 Lymphocyte formation Lymphocytes are round cells containing large round nuclei. The cytoplasm stains pale blue and appears non-granular under light microscopy. However, some cytoplasmic granules and organelles are present. Morphologically, lymphocytes can be divided into two groups: the more numerous small lymphocytes, with a diameter of 7-10 mm; and large lymphocytes, which have a diameter of 10-14 mm. Lymphocytes are produced in bone marrow from primitive precursors, the lymphoblasts and prolymphocytes. Immature cells migrate to the thymus and other lymphoid tissues, including that found in bone marrow, and undergo further division, processing and maturation. 5.2.5 Platelete formation (Thrombopoeisis) Platelets are produced in bone marrow by a process known as thrombopoiesis. They are formed in the cytoplasm of a very large cell, the megakaryocyte. The cytoplasm of the megakaryocyte fragments at the edge of the cell. This is called platelet budding. Megakaryocytes mature in about 10 days, from a large stem cell, the megakaryoblast. It is likely that there are thrombopoietic feedback mechanisms as the platelet count remains fairly constant in health, and platelet production is reduced following an infusion of platelets and increased following removal of platelets. However, these feedback mechanisms have not been discovered yet.
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At any one time, about two-thirds of the body's platelets are circulating in the blood and one-third are pooled in the spleen. There is constant exchange between the two populations. The life span of platelets is between 8 and 12 days. They are destroyed by macrophages, mainly in the spleen and also in the liver. 5.3 COMPOSITION AND FUNCTIONS OF BLOOD The average human being has 5 liters of blood in his body. 55% of the blood is made up of plasma constituting the fluid part of the blood. The cells and platelets that are present in our blood make up the other 45%. 5.3.1 Blood plasma Plasma is a pale yellowish fluid with a total volume of 2-3 liters in a normal adult. Its contents are : Water 90.0% Protein Inorganic Ions Organic Substances 8.0% 0.9% 1.1%
Plasma Proteins: Plasma proteins are made up of 3 main types. Serum Albumin Serum Globulin Fibrinogen Prothrombin Most of the proteins are produced by the liver apart from the Serum Globulin which is produced by the body's immune system. Serum Albumin composed of 60% of the total plasma protein. It is the smallest of the 4 plasma proteins and can pass through capillary walls. Hence this will usually lead to a small leakage into the intestinal fluid. Nonetheless, this has been exploited by medical science to be an effective medium of testing for abnormalities in the body and damaged organs or diseases. Serum Globulins make up 36% of the total plasma protein. This can be further split into 3 fragments, the alpha, beta and gamma. Globulins aid in the inflammatory response of the body. Fibrinogen and prothrombin are important in the clotting process of blood. Protein Functions Include: Transportation of insoluble substances around the body by allowing them to bind to protein molecules. Protein reserve for the body Blood clotting Responses in accordance to disease (inflammatory response) Protection from infection the gamma globulins function Striking balance for the pH of the blood Inorganic Ions : Inorganic ions play a very important role in the blood. ION Sodium
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CONCENTRATION (mmol/l) 135-146
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Potassium Calcium Chloride Phosphate
K+ Ca++ ClPO4--
3.5-5.2 2.1-2.7 98-108 23-31 0.7-1.4
Hydrogen Carbonate HCO3-
Without sodium, the body will lack extracellular fluid and might affect the blood pressure, leading to insufficient circulation of oxygen causing drowsiness, nausea etc. Lack of potassium will result in muscle abnormalities and weakness, leading to vomiting and diarrhoea. Hence all the inorganic ions facilitate in a very important role as do other cells within the blood. Organic Substances: Blood plasma carry organic substances such as nutrients. They may include digested food substances like glucose, sucrose and amino acids. Other nutrients in transit in the plasma include glycerol, triglycerides, cholesterol and vitamins. Waste products of the body are also transmitted in the blood plasma. They include urea and cellular waste that will be excreted out of the body. Hormones, such as cortisol and thyroxine are also transported around the body in plasma attached to plasma proteins. Medicine and drugs also circulate within the plasma. 5.3.2 Red Blood Cells Red blood cells are the most common cells found in blood. There are about 5 million red blood cells in each cubic millimeter of blood or approximately 250 million red blood cells in every drop of blood. This number varies with individuals in accordance to heredity, gender and state of health. These cells are produced by the bone marrow and have a lifespan of 3-4 months. When they die, they are destroyed by macrophages in the liver and spleen. This process releases iron to be stored in the liver and bile pigments to be excreted. Structure of A Red Blood Cell : Red blood cells have a bi-concave shape with a flattened center. It has a diameter less than 0.01 millimeters and do not have a nucleus. Red blood cells contain a protein chemical known as haemoglobin, which gives it the red color. Haemoglobin contains iron, which can easily transport gases such as oxygen and carbon dioxide. Red blood cells are highly elastic, rendering it able to squeeze through capillary walls bigger than itself. Functions of Red Blood Cells: Red blood cells are important in the process of respiration. Gases involved in respiration are carried around the body through these cells. Oxygen readily combines with haemoglobin to form oxy-haemoglobin in the lungs where there is high concentration of oxygen. However, oxy-haemoglobin is an unstable compound and will break down to release oxygen when there is low concentration of oxygen in the surroundings. Hence there will be an even distribution of oxygen to all parts of the body. Red blood cells also carry part of the carbon dioxide waste from the cells through most is transmitted through plasma as soluble carbonates.
Fig. 5.3.2 Red blood cells and White blood cells
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5.3.3 White Blood Cells (Leucocytes) White blood cells are responsible for the defense system in the body. There are approximately 6,000 white blood cells per millimeter of blood or a million white blood cells in every drop of human blood. White blood cells fight infections and protect our body from foreign particles, which includes harmful germs and bacteria. White blood cells, the red blood cells are formed from the stem cell of the bone marrow. It has a life-span of a couple of days. When they die, they are destroyed by surrounding white blood cells and replaced with new ones. Structure of White Blood Cells : White blood cells are colorless without haemoglobin. It contains a nucleus and has an irregular shape. Though there are fewer white blood cells than red blood cells, they are much bigger in size. They can change their shape easily and this allows them to squeeze through walls of the blood vessels into the inter-cellular spaces. Types of White Blood Cells: Unlike the Red blood cells or platelets, there are 5 different types of white blood cells, each serving a different purpose in our body’s immune system. Neutrophils Eosinophils Basophils Monocytes Lymphocytes Neutrophils : Neutrophils make up 55%-70% of the total white blood count in the bloodstream. They have a segmented nuclei and it is said to be C shaped. Neutrophils can be most commonly found near sites of infection or injury where they will stick to the walls of the blood vessels and engulf any foreign particles that try to enter the bloodstream. They can also be found in the pus of wounds. Eosinophils: Eosinophils make up 2%-5% of the total blood count and mainly attacks parasites and any antigen complexes. These cells are also responsible for allergic response within the blood. Basophils: Basophils make up less than 1% of the total white blood count. They secrete anticoagulant and antibodies, which mediate hypersensitivity reactions within the blood. They are known to have phagocytory features though they are more often related to immediate immune reaction against external germs and diseases. Monocytes: Monocytes, though having only 5%-8% in the total white blood count, are the largest of the 5 types of white blood cells. They act as tissue macrophages and remove foreign particles and prevent the invasion of germs which cannot be effectively dealt with by the neutrophils. They have been known to have phagocytic function Lymphocytes: Lymphocytes produce anti-bodies against toxins secreted by bacteria and infecting germs. These antibodies will be excreted into the plasma to kill bacteria in the blood as well as act as anti-toxins. These anti-bodies will cause the foreign particles to cluster together, rendering them easily engulfed by the phagocytes. However, the nature of lymphocytes is highly specific and they can only recognize certain antigens 5.3.4 Blood platelets Normal platelet count in a healthy person is between 150,000 and 400,000 per mm³ of blood (150– 400 x 109/L). Blood Platelets are granular non-nucleated fragments of cytoplasm in the form of
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oval discs. A platelet consists of two parts, a clear outer ground susbstance occupying the greater part of the platelet and a central part that contains granules. Functions of Blood Platelets: They secrete a hormone called serotonin which constricts torn blood vessels. They also have a major role in accumulating at sites of injury sticking together to plug gaps in broken blood vessels. They are rich in a certain activator that activates some proteins found in plasma. These proteins are thrown out in the form of fibers as a network. This network traps the escaping RBCs and forms a clot that will seal the cut blood vessels and so bleeding is stopped.
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6. MECHANISM OF BLOOD COAGULATION 6.1 INTRODUCTION 6.2 MECHANISM OF BLOOD COAGULATION 6.1 INTRODUCTION Coagulation is a complex process by which blood forms solid clots. It is an important part of hemostasis (the cessation of blood loss from a damaged vessel) whereby a damaged blood vessel wall is covered by a platelet- and fibrin-containing clot to stop bleeding and begin repair of the damaged vessel. Disorders of coagulation can lead to an increased risk of bleeding and/or clotting and embolism. Coagulation is highly conserved throughout biology; in all mammals, coagulation involves both a cellular (platelet) and a protein (coagulation factor) component. The system in humans has been the most extensively researched and therefore the best understood. Coagulation is initiated almost instantly after an injury to the blood vessel damages the endothelium (lining of the vessel). Platelets immediately form a hemostatic plug at the site of injury; this is called primary hemostasis. Secondary hemostasis occurs simultaneously—proteins in the blood plasma, called coagulation factors, respond in a complex cascade to form fibrin strands which strengthen the platelet plug. 6.2 MECHANISM OF BLOOD COAGULATION The mechanism of blood coagulation completed in three steps: Production of Thromboplastin by extrinsic and intrinsic pathways Formation of Thrombin Formation of Fibrin clot 6.2.1 Production of Thromboplastin Thromboplastin is produced by extrinsic and intrinsic pathway. Extrinsic pathway (Tissue factor pathway): The extrinsic pathway is initiated at the site of injury in response to the release of tissue factor (factor III). Tissue factor is a cofactor in the factor VIIa-catalyzed activation of factor X. Factor VIIa, a gla residue containing serine protease, cleaves factor X to factor Xa in a manner identical to that of factor IXa of the intrinsic pathway. The activation of factor VII occurs through the action of thrombin or factor Xa. The ability of factor Xa to activate factor VII creates a link between the intrinsic and extrinsic pathways. An additional link between the two pathways exists through the ability of tissue factor and factor VIIa to activate factor IX. The formation of complex between factor VIIa and tissue factor is believed to be a principal step in the overall clotting cascade. Evidence for this stems from the fact that persons with hereditary deficiencies in the components of the contact phase of the intrinsic pathway do not exhibit clotting problems. A major mechanism for the inhibition of the extrinsic pathway occurs at the tissue factor--factor VIIa--Ca2+--Xa complex. Intrinsic pathway: The intrinsic pathway requires the clotting factors VIII, IX, X, XI, and XII. Also required are the proteins prekallikrein (PK) and high-molecular-weight kininogen (HK or HMWK), as well as calcium ions and phospholipids secreted from platelets. Each of these pathway constituents leads to the conversion of factor X (inactive) to factor Xa (―a‖ signifies active).
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Initiation of the intrinsic pathway occurs when prekallikrein, high-molecular-weight kininogen, factor XI and factor XII are exposed to a negatively charged surface. The assemblage of contact phase components results in conversion of prekallikrein to kallikrein, which in turn activates factor XII to factor XIIa. Factor XIIa can then hydrolyze more prekallikrein to kallikrein, establishing a reciprocal activation cascade. Factor XIIa also activates factor XI to factor Xia. In the presence of Ca2+, factor XIa activates factor IX to factor IXa. Factor IX is a proenzyme that contains vitamin K-dependent g-carboxyglutamate (gla) residues, whose serine protease activity is activated following Ca2+ binding to these gla residues. Several of the serine proteases of the cascade (II, VII, IX, and X) are gla-containing proenzymes. Active factor IXa cleaves factor X at an internal arg-ile bond leading to its activation to factor Xa. The activation of factor Xa requires assemblage of the tenase complex (Ca2+ and factors VIIIa, IXa and X) on the surface of activated platelets. One of the responses of platelets to activation is the presentation of phosphatidylserine (PS) and phosphatidylinositol (PI) on their surfaces. The exposure of these phospholipids allows the tenase complex to form. The role of factor VIII in this process is to act as a receptor, in the form of factor VIIIa, for factors IXa and X. Factor VIIIa is termed a cofactor in the clotting cascade. At the presence of calcium ions VIIIa factor produced Thromboplastin. 6.2.2 Production of Trombin The common point in both pathways is the activation of factor X to factor Xa. Factor Xa activates prothrombin (factor II) to thrombin (factor IIa). Thrombin, in turn, converts fibrinogen to fibrin. The activation of thrombin occurs on the surface of activated platelets and requires formation of a prothrombinase complex. This complex is composed of the platelet phospholipids, phosphatidylinositol and phosphatidylserine, Ca2+, factors Va and Xa, and prothrombin. Factor V is a cofactor in the formation of the prothrombinase complex, similar to the role of factor VIII in tenase complex formation. Like factor VIII activation, factor V is activated to factor Va by means of minute amounts and is inactivated by increased levels of thrombin. Factor Va binds to
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specific receptors on the surfaces of activated platelets and forms a complex with prothrombin and factor Xa. Prothrombin is a 72,000-Dalton, single-chain protein containing ten gla residues in its N-terminal region. Within the prothrombinase complex, prothrombin is cleaved at 2 sites by factor Xa. This cleavage generates a 2-chain active thrombin molecule containing an A and a B chain which are held together by a single disulfide bond. 6.2.3 Formation of fibrin clot Fibrinogen (factor I) consists of 3 pairs of polypeptides ([A-a][B-b][g])2. The 6 chains are covalently linked near their N-terminals through disulfide bonds. The A and B portions of the A-a and B-b chains comprise the fibrinopeptides, A and B, respectively. The fibrinopeptide regions of fibrinogen contain several glutamate and aspatate residues imparting a high negative charge to this region and aid in the solubility of fibrinogen in plasma. Active thrombin is a serine protease that hydrolyses fibrinogen at four arg-gly bonds between the fibrinopeptide and the a and b portions of the protein. The initial event in mechanism of fibrin polymerization is the thrombin activation at only one of the Aa-chains in fibrinogen. The resulting highly reactive intermediate is the true fibrin monomer and it rapidly, and irreversibly, self-associates to form the stable fibrin dimer (s20.w=12S). Fibrin dimer possesses the N-terminal pattern alanine/glycine/tyrosine (1:1:2) per 340000 molecular weight. The fibrin dimer is a soluble inert molecule, but additional thrombin activation of its remaining intact Aa-chains leads to new associations into larger inert soluble fibrin polymers.
Factor Prekallikrein (PK)
Trivial Name(s) Fletcher factor
Characteristic Functions with HMWK and factor XII Co-factor in kallikrein and factor XII activation, necessary in factor XIIa activation of XI, precursor
High molecular weight kininogen (HMWK)
contact activation cofactor; Fitzgerald, Flaujeac Williams factor
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for bradykinin (a potent vasodilator and inducer of smooth muscle contraction I II III IV V VI (same as Va) VII Fibrinogen Prothrombin Tissue Factor Calcium Proaccelerin, labile factor, accelerator (Ac-) globulin Accelerin Proconvertin, serum prothrombin conversion accelerator (SPCA), cothromboplastin Antihemophiliac factor A, antihemophilic globulin (AHG) Christmas Factor, antihemophilic factor B,plasma thromboplastin component (PTC) Stuart-Prower Factor Plasma thromboplastin antecedent (PTA) Hageman Factor Protransglutaminase, fibrin stabilizing factor (FSF), fibrinoligase Both Both Extrinsic Both Both Contains N-term. gla segment Protein cofactor This is Va, redundant to Factor V Endopeptidase with gla residues Protein cofactor
Endopeptidase with gla residues Endopeptidase with gla residues Endopeptidase Endopeptidase Transpeptidase
X XI XII XIII
Both Intrinsic Intrinsic Both
Table: Different coagulation factors and its characterisitics ***
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7. ACTION AND USES OF ANTICOAGULANTS An anticoagulant is a substance that prevents coagulation; that is, it stops blood from clotting. A group of pharmaceuticals called anticoagulants can be used in vivo as a medication for thrombotic disorders. Some chemical compounds are used in medical equipment, such as test tubes, blood transfusion bags, and renal dialysis equipment. Anticoagulants are given to people to stop thrombosis (blood clotting inappropriately in the blood vessels). This is useful in primary and secondary prevention of deep vein thrombosis, pulmonary embolism, myocardial infarctions and strokes in those who are predisposed. 7.1 VITAMIN-K ANTAGONISTS The oral anticoagulants are a class of pharmaceuticals that act by antagonizing the effects of vitamin K. Examples include warfarin. It is important to note that they take at least 48 to 72 hours for the anticoagulant effect to develop fully. In cases when any immediate effect is required, heparin must be given concomitantly. Generally, these anticoagulants are used to treat patients with deep-vein thrombosis (DVT), pulmonary embolism (PE), atrial fibrillation (AF), and mechanical prosthetic heart valves. 7.1.1 Adverse effects Patients aged 80 years or more may be especially susceptible to bleeding complications with a rate of 13 bleeds per 100 person-years. These oral anticoagulants are used widely as poisons for mammalian pests, especially rodents. 7.1.2 Available agents Warfarin (Coumadin) This is the main agent used in the U.S. and UK Acenocoumarol and phenprocoumon This is used more commonly outside the U.S. and the UK Brodifacoum Rat poison, not used medically Phenindione 7.2 HEPARIN AND DERIVATIVE SUBSTANCES Heparin is a biological substance, usually made from pig intestines. It works by activating antithrombin III, which blocks thrombin from clotting blood. Heparin can be used in vivo (by injection), and also in vitro to prevent blood or plasma clotting in or on medical devices. Vacutainer brand test tubes containing heparin are usually colored green. 7.2.1 Low molecular weight heparin Low molecular weight heparin is a more highly processed product that is useful as it does not require monitoring of the APTT coagulation parameter (it has more predictable plasma levels) and has fewer side effects. 7.2.2 Synthetic pentasaccharide inhibitors of factor Xa Fondaparinux is a synthetic sugar composed of the five sugars (pentasaccharide) in heparin that bind to antithrombin. It is a smaller molecule than low molecular weight heparin. Ex: Idraparinux 7.2.3 Major pharmaceutical Heparin In March 2008 major recalls of Heparin were announced by pharmaceuticals due to a suspected and unknown contamination of the raw Heparin stock imported from China. The U.S. Food and Drug Administration was quoted as stating that at least 19 deaths were believed linked to a raw
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Heparin ingredient imported from the People's Republic of China, and that they had also received 785 reports of serious injuries associated with the drug’s use. According to the New York Times: 'Problems with heparin reported to the agency include difficulty breathing, nausea, vomiting, excessive sweating and rapidly falling blood pressure that in some cases led to life-threatening shock'. 7.3 DIRECT THROMBIN INHIBITORS Another type of anticoagulant is the direct thrombin inhibitor. Current members of this class include argatroban, lepirudin, bivalirudin, and dabigatran. An oral direct thrombin inhibitor, ximelagatran (Exanta) was denied approval by the Food and Drug Administration (FDA) in September 2004 and was pulled from the market entirely in February 2006 after reports of severe liver damage and heart attacks.
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8. THROMBOLYTIC AND ANTICOAGULANT THERAPY 8.1 THROMBOLYTIC THERAPY 8.2 ANTICOAGULANT THERAPY 8.1 THROMBOLYTIC THERAPY 8.1.1 Definition Thrombolytic therapy is the use of drugs that dissolve blood clots. 8.1.2 Purpose When a blood clot forms in a blood vessel, it may cut off or severely reduce blood flow to parts of the body that are served by that blood vessel. This can cause serious damage to those parts of the body. If the clot forms in an artery that supplies blood to the heart, for example, it can cause a heart attack. A clot that cuts off blood to the brain can cause a stroke. Thrombolytic therapy is used to dissolve blood clots that could cause serious, and possibly life-threatening, damage if they are not removed. Research suggests that when used to treat stroke, thrombolytic therapy can prevent or reverse paralysis and other problems that otherwise might result.Thrombolytic therapy also is used to dissolve blood clots that form in tubes put into people's bodies for medical treatments, such as dialysis or chemotherapy. 8.1.3 Description Thrombolytic therapy uses drugs called thrombolytic agents, such as alteplase (Activase), anistreplase (Eminase), streptokinase (Streptase, Kabikinase), urokinase (Abbokinase), and tissue plasminogen activator (TPA) to dissolve clots. These drugs are given as injections, only under a physician's supervision. 8.1.4 Recommended dosage The physician supervising thrombolytic therapy decides on the proper dose for each patient. He or she will take into account the type of drug, the purpose for which it is being used, and in some cases, the patient's weight. 8.1.5 Precautions For thrombolytic therapy to be effective in treating stroke or heart attack, prompt medical attention is very important. The drugs must be given within a few hours of the beginning of a stroke or heart attack. However, this treatment is not right for every patient who has a heart attack or a stroke. Thrombolytic therapy may cause bleeding. Usually this is not serious, but severe bleeding does occur in some people. This is especially likely in older people. To lower the risk of serious bleeding, people who are given this drug should move around as little as possible and should not try to get up on their own unless told to do so by a health care professional. Following all the instructions of the health care providers in charge is very important. Thrombolytic therapy may be more likely to cause serious bleeding in people who have certain medical conditions or have recently had certain medical procedures. Before being given a thrombolytic agent, anyone with any of these problems or conditions should tell the physician in charge about it: blood disease or current or past bleeding problems in any part of the body heart or blood vessel disease stroke (recent or in the past) high blood pressure
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After being treated with thrombolytic therapy, women who are breastfeeding should check with their physicians before starting to breastfeed again. 8.1.6 Side effects Anyone who has fever or who notices bleeding or oozing from their gums, from cuts, or from the site where the thrombolytic agent was injected should immediately tell their health care provider. People who are given thrombolytic therapy should also be alert to the signs of bleeding inside the body and should check with a physician immediately if any of the following symptoms occur: blood in the urine, blood or black, tarry stools, constipation, coughing up blood, vomiting blood or material that looks like coffee grounds, nosebleeds, unexpected or unusually heavy vaginal bleeding, dizziness, sudden, severe, or constant headaches and Pain or swelling in the abdomen or stomach.
Fig. 8.1 Thrombolytic therapy 8.2 ANTICOAGULANT THERAPY 8.2.1 Definition Anticoagulant therapy is the use of Anticoagulant drugs to prevent clot formation or to prevent a clot that has formed from enlarging. They inhibit clot formation by blocking the action of clotting factors or platelets. Anticoagulant drugs fall into three categories: inhibitors of clotting factor synthesis, inhibitors of thrombin and antiplatelet drugs. 8.2.2 Purpose Anticoagulant drugs reduce the ability of the blood to form clots. Although blood clotting is essential to prevent serious bleeding in the case of skin cuts, clots inside the blood vessels block the flow of blood to major organs and cause heart attacks and strokes. Although these drugs are sometimes called blood thinners, they do not actually thin the blood.
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Anticoagulant drugs are used for a number of conditions. The four most common conditions for which anticoagulant therapy is prescribed are atrial fibrillation, deep vein thrombosis, pulmonary embolism, and mechanical heart valves. 8.2.3 Description Anticoagulant drugs, also called anticlotting drugs or blood thinners, are available only with a physician's prescription. They come in tablet and injectable forms. They fall into three groups: Inhibitors of clotting factor synthesis. These anticoagulants inhibit the production of certain clotting factors in the liver. One example is warfarin (brand name: coumadin). Inhibitors of thrombin. Thrombin inhibitors interfere with blood clotting by blocking the activity of thrombin. They include heparin, lepirudin (Refludan). Antiplatelet drugs. Antiplatelet drugs interact with platelets, which is a type of blood cell, to block platelets from aggregating into harmful clots. They include: aspirin, ticlopidine (Ticlid), clopidogrel (Plavix), tirofiban (Aggrastat), and eptifibatide (Integrilin). 8.2.4 Recommended dosage The recommended dosage depends on the type of anticoagulant drug and the medical condition for which it is prescribed. The prescribing physician or the pharmacist who filled the prescription can provide information concerning the correct dosage. Usually, the physician will adjust the dose after checking the patient's clotting time. Anticoagulant drugs must be taken exactly as directed by the physician. Larger or more frequent doses should not be taken, and the drug should also not be taken for longer than prescribed. Taking too much of this medication can cause severe bleeding. Anticoagulants should also be taken on schedule. A record of each dose should be kept as it is taken. 8.2.5 Precautions Persons who take anticoagulants should see a physician regularly while taking these drugs, particularly at the beginning of therapy. The physician will order periodic blood tests to check the blood's clotting ability. The results of these tests will help the physician determine the proper amount of medication to be taken each day. Time is required for normal clotting ability to return after anticoagulant treatment. During this period, patients must observe the same precautions they observed while taking the drug. The length of time needed for the blood to return to normal depends on the type of anticoagulant drug that was taken. The prescribing physician will advise as to how long the precautions should be observed. Alcohol can change the way anticoagulant drugs affect the body. Anyone who takes this medicine should not have more than one to two drinks at any time and should not drink alcohol every day. 8.2.6 Side effects The most common minor side effects are bloating or gas. These problems usually go away as the body adjusts to the drug and do not require medical treatment. More serious side effects may occur, especially if excessive anticoagulant is taken. If any of the following side effects occur, a physician should be notified immediately: bleeding gums, sores or white spots in the mouth or throat, unusual bruises or purplish areas on the skin, cloudy or dark urine, painful or difficult urination or sudden decrease in amount of urine, black, tarry, or bloody stools, yellow eyes or skin and severe or continuing headache. ***
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Chapter – 3
CEREBROSPINAL FLUID (CSF)
9. Composition and functions of CSF 10. Composition and functions of Lymph
9. COMPOSITION AND FUNCTIONS OF CSF 9.1 INTRODUCTION 9.2 COMPOSITION OF CSF 9.3 FUNCTIONS OF CSF 9.1 INTRODUCTION Cerebrospinal fluid (CSF), Liquor cerebrospinalis, is a clear bodily fluid that occupies the subarachnoid space and the ventricular system around and inside the brain. Essentially, the brain "floats" in it. More specifically the CSF occupies the space between the arachnoid mater (the middle layer of the brain cover, meninges) and the pia mater (the layer of the meninges closest to the brain). Moreover it constitutes the content of all intra-cerebral (inside the brain, cerebrum) ventricles, cisterns and sulci (singular sulcus), as well as the central canal of the spinal cord. It is produced in the brain by modified ependymal cells in the choroid plexus. It circulates from the choroid plexus through the interventricular foramina (foramen of Monro) into the third ventricle, and then through the mesencephalic duct (cerebral aqueduct) into the fourth ventricle, where it exits through two lateral apertures (foramina of Luschka) and one median aperture (foramen of Magendie). It then flows through the cerebromedullary cistern down the spinal cord and over the cerebral hemispheres. It is an approximately isotonic solution and acts as a "cushion" or buffer for the cortex, providing also a basic mechanical and immunological protection to the brain inside the skull. 9.2 COMPOSITION OF CSF The cerebrospinal fluid is produced at a rate of 500 ml/day. Since the brain can only contain from 135-150 ml, large amounts are drained primarily into the blood through arachnoid granulations in the superior sagittal sinus. This continuous flow into the venous system dilutes the concentration of larger, lipoinsoluble molecules penetrating the brain and CSF. The CSF contains approximately 0.3% plasma proteins, or 15 to 40 mg/dL, depending on sampling site. CSF pressure ranges from 60 - 100 mmH2O or 4.4 - 7.3 mmHg, with most variations due to coughing or internal compression of jugular veins in the neck. Fig. 9.1 Cerebrospinal fluid 9.3 FUNCTIONS OF CSF Protection The CSF protects the brain from damage by "buffering" the brain. In other words, the CSF acts to cushion a blow to the head and lessen the impact.
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Buoyancy Because the brain is immersed in fluid, the net weight of the brain is reduced from about 1,400 gm to about 50 gm. Therefore, pressure at the base of the brain is reduced. Excretion of waste products The one-way flow from the CSF to the blood takes potentially harmful metabolites, drugs and other substances away from the brain. Endocrine medium for the brain The CSF serves to transport hormones to other areas of the brain. Hormones released into the CSF can be carried to remote sites of the brain where they may act.
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10. COMPOSITION AND FUNCTIONS OF LYMPH 10.1 INTRODUCTION 10.2 COMPOSITION OF LYMPH 10.3 FUNCTIONS OF LYMPH 10.1 INTRODUCTION Lymph is the fluid that is formed as the interstitial fluid enters the lymph vessels by filtration. The lymph then travels to at least one lymph node before emptying ultimately into the right or the left subclavian vein, where it mixes back with blood. The lymph is a transparent, usually slightly yellow, often opalescent liquid found within the lymphatic vessels, and collected from tissues in most parts of the body and returned to the blood via the lymphatic system. Lymph is the name given to interstitial fluid which enters the lymphatic vessels. Lymphatic capillaries are present in nearly all tissues. Significant exceptions are the central nervous system and bone. Small interstitial channels are present in the brain and the fluid flows into the CSF and then passes back into the circulation via the arachnoid villi. The lymph capillaries are blind-ending and possess flap valves between adjacent lymphatic endothelial cells. These functional valves permit entry of ISF but prevent its return to the interstitium. The pressure inside the lymph capillary is about 1 mmHg at rest and the flap valves are closed. The lymph capillaries interconnect and join together to form lymph venules, and then large lymph veins which drain via lymph nodes into the thoracic duct (on the left) and the right lymphatic duct. By these two final pathways, lymph returns into the circulation. 10.2 COMPOSITION OF LYMPH Lymph has a composition comparable to that of plasma, but it is different in various parts of the body depending upon the tissue drained. It is about 95% water; the remainder consists of plasma proteins and other chemical substances contained in the blood plasma, but in a slightly smaller percentage than in plasma. In particular, the lymph that leaves a lymph node is richer in lymphocytes (about 8,000 per cubic millimeter), proteins, and fats. Likewise, the lymph formed in the digestive system called chyle is rich in triglycerides (fat), and looks white. 10.3 FUNCTIONS OF LYMPH Lymph plays an important part in the immune system and in absorbing fats from the small intestine. The three functions of the lymphatic system are: Return of protein and fluid from the ISF to the circulation to maintain a low interstitial fluid protein concentration and maintain the oncotic pressure gradient across the capillary membrane. Oedema will occur if ISF oncotic pressure is not kept low. Role in absorption and transport of fat from the small intestine. Immunological role -lymph glands & circulation of immune cells such as lymphocytes and dendritic cells, removal of bacteria. Lymph from most parts of the body usually has a low protein concentration. Liver lymph is different because:
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It normally has a high protein concentration (due to low reflection coefficient) It contributes more than half of all the thoracic duct lymph Consequently, the average lymph protein concentration in thoracic duct lymph is much higher than expected based on protein concentration in lymph from other body tissues. The thoracic duct carries about 80% of the total lymph flow. This total flow at rest is about 120 mls/hr. If interstitial hydrostatic pressure rises (ie becomes less negative) due to excess fluid filtration & accumulation, the total lymph flow can increase quite markedly. Chyle is lymph from the intestines which has a milky-while appearance due to the presence of large numbers of chylomicrons. Chylomicrons are 100nm diameter complexes of mostly triglycerides (containing the long chain fatty acids) enclosed in a hydrophobic protein coat. Chylomicrons enter the lymphatic lacteals in the villi, travel in the lymph and then enter the circulation via the thoracic duct.
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Chapter – 4
11. Formation, composition and functions of digestive secretions
11. FORMATION, COMPOSITION AND FUNCTIONS OF DIGESTIVE SECRETIONS 11.1 11.2 11.1 INTRODUCTION Digestive secretions are enzymes in the alimentary tract that break down food so that the organism can absorb it. The main sites of action are the oral cavity, the stomach, the duodenum and the jejunum. They are secreted by different glands: the salivary glands, the glands in the stomach, the pancreas, Liver and the glands in the small intestines. In the oral cavity, salivary glands secrete saliva, which digests starch into small segments of multiple sugars and into individual soluble sugars. The secretions that from the stomach are called gastric juice, which digest the proteins. The liver secretes bile juice, which emulsifies the fat substances. The pancreas at the region of duodenum secreted pancreatic juice, which digest the proteins, lipids and carbohydrates. The small intestine (Duedenum/jejunum) secretes succus entericus (Interstinal juice), which digest all type of food materials. 11.2 DIGESTIVE SECRETIONS The following are the digestive secretions which secreted from the different digestive glands like salivary glands, gastric glands of stomach, pancreas, liver and the glands in the small intestines. Saliva Gastric juice Bile juice Pancreatic juice Succus entericus (Intestinal juice) 11.2.1 Saliva: a) Formation of saliva The production of saliva is stimulated both by the sympathetic nervous system and the parasympathetic. The saliva stimulated by sympathetic innervation is thicker, and saliva stimulated parasympathetically is more watery. The formation of saliva is a multi-step process. Initially formed of an aqueous solution (water based solution) of electrolytes, proteins (mostly enzymes), and mucus, saliva undergoes several chemical changes before it is release from the glandular collecting ducts into the oral cavity. The sodium content is reduced and potassium levels increase along with the addition of bicarbonate ions that make the saliva alkaline. The average person produces approximately 700mL of saliva per day, which is much less than was once thought.
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INTRODUCTION DIGESTIVE SECRETIONS
b) Composition of saliva Produced in salivary glands, saliva is 98% water, but it contains many important substances, including electrolytes, mucus, antibacterial compounds and various enzymes. It is a fluid containing: Water Electrolytes: o 2-21 mmol/L sodium (lower than blood plasma) o 10-36 mmol/L potassium (higher than plasma) o 1.2-2.8 mmol/L calcium o 0.08-0.5 mmol/L magnesium o 5-40 mmol/L chloride (lower than plasma) o 25 mmol/L bicarbonate (higher than plasma) o 1.4-39 mmol/L phosphate Mucus. Mucus in saliva mainly consists of mucopolysaccharides and glycoproteins; Antibacterial compounds (thiocyanate, hydrogen peroxide, and secretory immunoglobulin A) Various enzymes. There are three major enzymes found in saliva. o α-amylase . Amylase starts the digestion of starch and lipase fat before the food is even swallowed. It has a pH optima of 7.4. o lysozyme. Lysozyme acts to lyse bacteria. o lingual lipase. Lingual lipase has a pH optimum ~4.0 so it is not activated till entering an acidic environment. o Minor enzymes include salivary acid phosphatases A+B, N-acetylmuramyl-L-alanine amidase, NAD(P)H dehydrogenase-quinone, salivary lactoperoxidase, superoxide dismutase, glutathione transferase, class 3 aldehyde dehydrogenase, glucose-6phosphate isomerase, and tissue kallikrein. Cells: Possibly as much as 8 million human and 500 million bacterial cells per mL. The presence of bacterial products (small organic acids, amines, and thiols) causes saliva to sometimes exhibit foul odor. Opiorphin, a newly researched pain-killing substance found in human saliva. c) Functions of saliva Digestion: The digestive functions of saliva include moistening food, and helping to create a food bolus, so it can be swallowed easily. Saliva contains the enzyme amylase that breaks some starches down into maltose and dextrin. Thus, digestion of food occurs within the mouth, even before food reaches the stomach. Salivary glands also secrete enzymes (salivary lipase) to start fat digestion. Disinfectants: A common belief is that saliva contained in the mouth has natural disinfectants, which leads people to believe it is beneficial to "lick their wounds". However, researchers find human saliva contains such antibacterial agents as secretory IgA, lactoferrin, and lactoperoxidase. It has not been shown that human licking of wounds disinfects them, but licking is likely to help clean the wound by removing larger contaminants such as dirt and may help to directly remove infective bodies by brushing them away. Therefore, licking would be a way of wiping off pathogens, useful if clean water is not available to the animal or person. Cleaning: Saliva is an effective cleaning agent used in art conservation. Cotton swabs coated with saliva are rolled across a paintings surface to delicately remove thin layers of dirt that may accumulate.
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11.2.2 Gastric secretion a) Formation of Gastric secretion Mucus, ions and water make up the bulk of the gastric juice secreted by the proximal and distal regions. The composition of the gastric secretions is not constant but varies with the flow rate. The basal secretion produced by the surface epithelial cells is plasma-like in composition built its modified extensively as gastric secretion increases through secretion of acid by the oxyntic cells of the proximal region. HCl is secreted by oxyntic cells. As rest these cells contain numerous tubulovesicles that are rich in H+, K+ -ATPase and the adjacent to canaliculi which open into the lumen of the gastric glands. When the cells are stimulated to secrete acid the vesicles fuse with the canaliculi, and at the same time there is an increase in canalicular membrance permeability to K+ and Cl-. The movement of K+ into the lumen activates the H+, K+, -ATPase so that intracellular H+ is exchanged with K+ and there is therefore a net secretion of HCl. The water accompanying the HCl is driven out of the cell by the efflux of ions. The composition of the acidic fluid secreted by the parietal cell Is therefore largely isosmotic HCl with a little KCl. For each mole of H+ secreted, an equivalent amount of base (OH-) is producec. In the oxynitic cell carbonic anhydrase catalyses the reaction that combines OH- with CO2 to give HCO3- , The HCO3- then passes out of the cell into the blood via a Cl-/HCO3exchanger. Tlhis provides the sourceof Cl- that accompanies the secretion of H+. Pepsinogens are synthesized in the chief cells, stored in granules and secreted by exocytosis is inactive precursors. This prevents digestion of the chief cells. Intrinsic factor is synthesized and secreted by the oxyntic cells and argentiferin cells. c) Composition and functions of Gastric secretion Mucus: The most abundant epithelial cells are mucous cells, which cover the entire lumenal surface and extend down into the glands as "mucous neck cells". These cells secrete a bicarbonate-rich mucus that coats and lubricates the gastric surface, and serves an important role in protecting the epithelium from acid and other chemical insults. Acid: Hydrochloric acid is secreted from parietal cells into the lumen where it establishes an extremely acidic environment. This acid is important for activation of pepsinogen and inactivation of ingested microorganisms such as bacteria. Proteases: Pepsinogen, an inactive zymogen, is secreted into gastric juice from both mucous cells and chief cells. Once secreted, pepsinogen is activated by stomach acid into the active protease pepsin, which is largely responsible for the stomach's ability to initiate digestion of proteins. In young animals, chief cells also secrete chymosin (rennin), a protease that coagulates milk protein allowing it to be retained more than briefly in the stomach. Hormones: The principal hormone secreted from the gastric epithelium is gastrin, a peptide that is important in control of acid secretion and gastric motility. 11.2.3 Bile juice a) Formation of Bile juice Bile is produced by hepatocytes in the liver, draining through the many bile ducts that penetrate the liver. During this process, the epithelial cells add a watery solution that is rich in bicarbonates that dilutes and increases alkalinity of the solution. Bile then flows into the common hepatic duct, which joins with the cystic duct from the gallbladder to form the common bile duct.
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The common bile duct in turn joins with the pancreatic duct to empty into the duodenum. If the sphincter of Oddi is closed, bile is prevented from draining into the intestine and instead flows into the gall bladder, where it is stored and concentrated to up to five times its original potency between meals. This concentration occurs through the absorption of water and small electrolytes, while retaining all the original organic molecules. Cholesterol is also released with the bile, dissolved in the acids and fats found in the concentrated solution. When food is released by the stomach into the duodenum in the form of chyme, the gallbladder releases the concentrated bile to complete digestion. The human liver can produce close to one litre of bile per day (depending on body size). 95% of the salts secreted in bile are reabsorbed in the terminal ileum and re-used. Blood from the ileum flows directly to the hepatic portal vein and returns to the liver where the hepatocytes resorb the salts and return them to the bile ducts to be re-used, sometimes two to three times with each meal. b) Composition of Bile juice Bile has various components, some of which are produced by hepatocytes (liver cells). Its constituents include: Cholesterol Phospholipids (mainly Lecithin) Bile pigments (bilirubin diglucoronoide) Bile salts (sodium glycocholate & sodium taurocholate) Bicarbonate ions and other ions Metabolized red blood cells The bile acids cholate and chenodeoxycholate are typically conjugated with taurine or glycine and are produced by the liver from cholesterol. Ordinarily the concentration of bile salts in bile is 0.8%, however the gall bladder removes water from the bile, concentrating it between meals. It concentrates it up to 5 times (increasing concentration to 4%), before contracting the walls and releasing it into the duodenum once chyme has entered the small intestine. c) Functions of Bile juice Role of Bile Acids in Fat Digestion and Absorption: Bile acids are derivatives of cholesterol synthesized in the hepatocyte. Cholesterol, ingested as part of the diet or derived from hepatic synthesis is converted into the bile acids cholic and chenodeoxycholic acids, which are then conjugated to an amino acid (glycine or taurine) to yield the conjugated form that is actively secreted into cannaliculi. Their amphipathic nature enables bile acids to carry out two important functions: Emulsification of lipid aggregates: Bile acids have detergent action on particles of dietary fat which causes fat globules to break down or be emulsified into minute, microscopic droplets. Emulsification is not digestion per se, but is of importance because it greatly increases the surface area of fat, making it available for digestion by lipases, which cannot access the inside of lipid droplets. Solubilization and transport of lipids in an aqueous environment: Bile acids are lipid carriers and are able to solubilize many lipids by forming micelles - aggregates of lipids such as fatty acids, cholesterol and monoglycerides - that remain suspended in water. Bile acids are also critical for transport and absorption of the fat-soluble vitamins. Role of Bile Acids in Cholesterol Homeostasis: Hepatic synthesis of bile acids accounts for the majority of cholesterol breakdown in the body. In humans, roughly 500 mg of cholesterol are
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converted to bile acids and eliminated in bile every day. This route for elimination of excess cholesterol is probably important in all animals, but particularly in situations of massive cholesterol ingestion. Besides its digestive function, bile serves as the route of excretion for the hemoglobin breakdown product (bilirubin) created by breakdown of erythrocytes, which are conjugated by glucuronidation in the liver ; it also neutralises any excess stomach acid before it enters the ileum, the final section of the small intestine. Bile salts are also bacteriocidal to the invading microbes that enter with food. 11.2.4 Pancreatic secretions a) Formation of Pancreatic secretions Pancreatic secretion is composed of two secretory products critical to proper digestion: digestive enzymes and bicarbonate. Bicarbonate ions are secreted by centro-acinar cells in higher concentrations than present in plasma, while Na+ and K+ concentrations are identical to that in the plasma. Since chloride concentration in the centro-acinar secretion is lower than in the plasma, it is exchanged for bicarbonate ions to maintain iso-osmolarity of the composite secretion. With increase in secretory rate, the available time for this exchange during the passage in the ducts diminishes considerably and hence the chloride concentration in the final secretion falls. This bicarbonate-chloride shift in responsible for the reciprocal relationship between the concentration of these two ions in the secretion. The enzymes are synthesized and stored as zymogen granules in the acinar cells. Under adequate stimuli, these zymogen granules are discharged into the central cavity by a process called emiocytosis (reverse pinocytosis). b) Composition and functions of Pancreatic secretion Pancreatic juice is a juice produced by the pancreas. It contains water (97.6%) and organic constituents like a variety of enzymes, including trypsinogen, chymotrypsinogen, elastase, carboxypeptidase, pancreatic lipase, and amylase (1.8%). Pancreatic juice is composed of two secretory products critical to proper digestion: digestive enzymes and bicarbonate. The enzymes are synthesized and secreted from the exocrine acinar cells, whereas bicarbonate is secreted from the epithelial cells lining small pancreatic ducts. Digestive Enzymes: 1. Proteases: Digestion of proteins is initiated by pepsin in the stomach, but the bulk of protein digestion is due to the pancreatic proteases. Several proteases are synthesized in the pancreas and secreted into the lumen of the small intestine. The two major pancreatic proteases are trypsin and chymotrypsin, which are synthesized and packaged into secretory vesicles as an the inactive proenzymes trypsinogen and chymotrypsinogen. As you might anticipate, proteases are rather dangerous enzymes to have in cells, and packaging of an inactive precursor is a way for the cells to safely handle these enzymes. The secretory vesicles also contain a trypsin inhibitor which serves as an additional safeguard should some of the trypsinogen be activated to trypsin; following exocytosis this inhibitor is diluted out and becomes ineffective - the pin is out of the grenade.
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Once trypsinogen and chymotrypsinogen are released into the lumen of the small intestine, they must be converted into their active forms in order to digest proteins. Trypsinogen is activated by the enzyme enterokinase, which is embedded in the intestinal mucosa. Once trypsin is formed it activates chymotrypsinogen, as well as additional molecules of trypsinogen. The net result is a rather explosive appearance of active protease once the pancreatic secretions reach the small intestine.
Trypsin and chymotrypsin digest proteins into peptides and peptides into smaller peptides, but they cannot digest proteins and peptides to single amino acids. Some of the other proteases from the pancreas, for instance carboxypeptidase, have that ability, but the final digestion of peptides into amino acids is largely the effect of peptidases on the surface of small intestinal epithelial cells. More on this later. 2. Pancreatic Lipase: A major component of dietary fat is triglyceride, or neutral lipid. A triglyceride molecule cannot be directly absorbed across the intestinal mucosa. Rather, it must first be digested into a 2-monoglyceride and two free fatty acids. The enzyme that performs this hydrolysis is pancreatic lipase, which is delivered into the lumen of the gut as a constituent of pancreatic juice. Sufficient quantities of bile salts must also be present in the lumen of the intestine in order for lipase to efficiently digest dietary triglyceride and for the resulting fatty acids and monoglyceride to be absorbed. This means that normal digestion and absorption of dietary fat is critically dependent on secretions from both the pancreas and liver.
Pancreatic lipase has recently been in the limelight as a target for management of obesity. The drug orlistat (Xenical) is a pancreatic lipase inhibitor that interferes with digestion of triglyceride and thereby reduces absorption of dietary fat. Clinical trials support the contention that inhibiting lipase can lead to significant reductions in body weight in some patients. 3.Amylase: The major dietary carbohydrate for many species is starch, a storage form of glucose in plants. Amylase (technically alpha-amylase) is the enzyme that hydrolyses starch to maltose (a glucose-glucose disaccharide), as well as the trisaccharide maltotriose and small branchpoints fragments called limit dextrins. The major source of amylase in all species is pancreatic secretions, although amylase is also present in saliva of some animals, including humans. i.
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Other Pancreatic Enzymes: In addition to the proteases, lipase and amylase, the pancreas produces a host of other digestive enzymes, including ribonuclease, deoxyribonuclease, gelatinase and elastase. Bicarbonates and Water Epithelial cells in pancreatic ducts are the source of the bicarbonate and water secreted by the pancreas. Bicarbonate is a base and critical to neutralizing the acid coming into the small intestine from the stomach. The mechanism underlying bicarbonate secretion is essentially the same as for acid secretion parietal cells and is dependent on the enzyme carbonic anhydrase. In pancreatic duct cells, the bicarbonate is secreted into the lumen of the duct and hence into pancreatic juice 11.2.5 Intestinal secretions (Succus entericus) a) Formation/secretion of intestinal secretions Enzymes, Mucus, water and electrolytes are the main constituents of intestinal secretion. The intestinal enzymes are secreted from small intestinal mucosa, these enzymes are: sucrase, maltase, lactase, lipases, amylases and erypsins. The secretion of enzymes in the small intestine does not occur in the usual manner. Instead, the digestive enzymes are formed in the epithelial cells lining the intestinal wall, and much of the digestive process occurs either inside these cells or on their surfaces. The another substance of intestinal secretion is mucus, which is secreted by Brunner’s glands, which are located within the duodenum and goblet cells located along the length of the intestinal epithelium and in the intestinal crypts, called the crypts of lieberkuhn. The final constituents of intestinal secretions are water and electrolytes. These are secreted by all the epithelial cells of the intestine. The watery secretion proves a solvent into which the products of digestion are dissolved. b) Composition and functions of intestinal secretions The small intestinal juice is a colorless, straw colored fluid. It is cloudy in appearance due to the shed epithelial cells and flecks of mucus. Due to rapid exchange of fluid in both directions of the intestinal mucosa, it has been difficult to determine the actual amount secreted. It is estimated that daily secretions may range between 1000 to 3000 ml. Succus entericus is composed of water, inorganic salts (1%) and organic material (0.6%). The organic matter consists mainly of enzymes, cellular debris and mucus. A number of the enzymes are located intracellularly (within the shed epithelial cells). Peptidases are the most important group of proteolytic enzymes in succus entericus. They bring about the final breakdown of polypeptides to amino acids. Di- and tripeptidases break up di- and tripeptides respectively, while aminopeptidases act on peptide linkages of terminal amino acids possessing free amino groups. Enteropeptidase or enterokinase activates trypsinogen. Its release in Succus entericus is triggered by the presence of pancreatic juice in the duodenum. Lipase is present in the intestinal juice along with a large groups of amylolytic enzymes mainly the disaccharides like lactase, sucrose, maltase and isomaltose. Most of the latter group of enzymes is located intracellularly. ***
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Chapter – 5
12. Formation of urine 13. Composition of urine
12. FORMATION OF URINE 12.1 12.2 12.3 12.4 12.1 INTRODUCTION Urine is composed of water, certain electrolytes, and various waste products that are filtered out of the blood system. Remember, as the blood flows through the body, wastes resulting from the metabolism of foodstuffs in the body cells are deposited into the bloodstream, and this waste must be disposed of in some way. A major part of this "cleaning" of the blood takes place in the kidneys and, in particular, in the nephrons, where the blood is filtered to produce the urine. Both kidneys in the body carry out this essential blood cleansing function. Normally, about 20% of the total blood pumped by the heart each minute will enter the kidneys to undergo filtration. This is called the filtration fraction. The rest of the blood (about 80%) does not go through the filtering portion of the kidney, but flows through the rest of the body to service the various nutritional, respiratory, and other needs that are always present. For the production of urine, the kidneys do not simply pick waste products out of the bloodstream and send them along for final disposal. The kidneys' 2 million or more nephrons (about a million in each kidney) form urine by three precisely regulated processes: 1. Glomerular Filtration also called "Ultra-filtration", 2. Tubular Reabsorption also called "Selective Re-Absorption" and 3. Tubular Secretion. 12.2 GLOMERULAR FILTRATION Blood enters the kidney via the renal artery. This seperates many times (Renal Artery -> Segmental Arteries -> Interlobar Arteries -> Arcuate Arteries -> Interlobular Arteries -> Afferent Arterioles), eventually forming many afferent arterioles, each of which delivers blood to an individual kidney nephron. The diameter of the afferent (incoming) arteriole is greater than the diameter of the efferent arteriole (by which blood leaves the glomerulus). The pressure of the blood inside the glomerulus is increased due to the difference in diameter of the incoming and out-going arterioles. This increased blood pressure helps to force the following components of the blood out of the glomerular capillaries: Most of the water; Most/all of the salts;
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INTRODUCTION FILTRATION REABSORPTION SECRETION
Most/all of the glucose; Most/all of the urea. The above are filtered in preference to other components of blood based on particle size. (Water and solutes of relative molecular mass less than 68,000 form the filtrate.) Blood cells and plasma proteins are not filtered through the glomerular capillaries because they are relatively larger in physical size. In a healthy person at rest almost 25% of cardiac output passes the two kidneys (1200 ml each min). The blood reaches the first part of the nephron through the afferent arteriole to the glomerular capillaries. The pressures governing the glomerular ultrafiltration rate (GFR) are called the Starling forces The filtration pressures are as follows: In the glomerular capillaries the hydrostatic pressure (Pgc) = +60 mmHg Hydrosatatic pressure within the tubule (Pt) = -20 mmHg Hydrostatic pressure gradient across the capillary wall (delta P) = Pgc - Pt Delta P = 60 – 20 = +40 mmHg Colloid osmotic pressure of the blood (Pco) = -30 mmHg Net filtration pressure (Pnet) = Pgc – Pco – Pt (Pnet) = +10 mmHg
The net ultrafiltration pressure (Pnet) is +10 mmHg through the glomerular capillaries, and provides the force for ultrafiltration of a fat- and protein- free fluid across the glomerular barrier into Bowmans space and flow through the renal tubules. 12.3 TUBULAR REABSORPTION Only about 1% of the glomerular fitrate actually leaves the body because the rest (the other 99%) is reabsorbed into the blood while it passes through the renal tubules and ducts. This is called tubular reabsorption and occurs via three mechanisms. They are:
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Osmosis Diffusion, and Active Transport. Reabsorption varies according to the body's needs, enabling the body to retain most of its nutrients. The processes of tubular reabsorption occur in the following order. a) In the PCT Most (80%) of the volume of the fitrate solution is reabsobed in the proximal convoluted tubule (PCT). This includes some water and most/all of the glucose (except in the case of diabetics). Most of the energy consumed by the kidneys is used in the reabsorption of sodium ions (Na+), which are solutes - that is, they are dissolved in the water component of the fitrate solution. Symporters simultaneously facilitate passage through the PCT membrane of both Na+ and another substances/solutes. Other such substances that are reabsorbed with Na+ in this way include glucose (an important type of sugar), amino acids, lactic acid, and bicarbonate ions (HCO3-). These then move on through cells via diffusion and/or other transport processes. A short way to summarize the above is to say that solutes are selectively moved from the glomular filtrate to the plasma by active transport. (However, almost all glucose and amino acids, and high but variable amounts of ions, are reabsorbed again later). Following the movement of solutes (including Na+ ), water is then also reabsorbed by osmosis. About 80% of the filtrate volume is reabsorbed in this way. As this part of the reabsorption process is not controlled by the proximal tubule itself, it is sometimes called obligatory water reabsorption. b) In the Loop of Henle The remaining water (together with the dissolved salts and urea) passes from the PCT into the descending limb of Henle. It then passes along the Loop of Henle, and up the ascending limb of Henle. The different permeability properties of the two limbs of the Loop of Henle, together with their counterflow arrangement, allows a countercurrent multiplication to generate a high solute concentration in the tissue fluid of the medulla (that is, outside of the tubules). The highest solute concentrations are generated deep in the medulla. This is explained as follows: i) Descending Limb of Loop of Henle The epithelium lining of the descending limb of Henle is relatively permeable to water but much much less permeable to the salts Na+ and Cl-, and to urea. Therefore water gradually moves from the descending limb and into the interstitium (surrounding the tubules) as fluid flows through this part of the system of renal tubules. ii) Thin Ascending Limb of Loop of Henle The thin ascending limb of Henle differs from the descending limb in that it is impermeable to water (so the water that is inside the tubule at this stage generally remains inside it), but is highly permeable to Na+ and Cl-, and somewhat permeable to urea. Therefore while the tubular fluid flows back towards the renal cortex, Na+ and Cl- (which are more concentrated in the tubular fluid than in the interstital fluid) diffuse from the tubules into the interstitium.
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Some urea also enters the tubules at this stage - but the loss of NaCl from the tubular fluid greatly exceeds the gain in urea.
iii) Thick Ascending Limb of Loop of Henle The thick ascending limb of Henle (and its continuation into the first part of the DCT), reabsorbs NaCl from the tubular fluid via a different transport process from that of the thin ascending limb of Henle. The overall effect of the processes outlined above is that the concentation of the fluid inside the renal tubules that form the Loop of Henle is highest at the deepest part of the renal medulla, and is less concentrated in the renal cortex. This is what is meant by the "concentration gradient" of the Loop of Henle. The term "counter-current" is also used in descriptions of the Loop of Henle - and refers to the tubular fluid flowing in opposite directions along the descending and ascending limbs (as indicated by the thin red arrows in the diagram above. c) In the DCT The water, urea, and salts contained within the ascending limb of Henle eventually pass into the distal convoluted tubule (DCT). The DCT reacts to the amount of anti-diuretic hormone (ADH) in the blood: The more ADH is present in the blood, the more water is re-absorbed into it. This happens because the presence of ADH in the blood causes the cells in the last section of the DCT (and associated tubules and collecting ducts) to become more permeable to water, therefore they allow more water to pass from the tubular fluid back into the blood. This results in more concentrated urine.
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12.4 TUBULAR SECRETION The third process by which the kidneys clean blood (regulating its composition and volume) is called tubular secretion and involves substances being added to the tubular fluid. This removes excessive quantities of certain dissolved substances from the body, and also maintains the blood at a normal healthy pH (which is typically in the range pH 7.35 to pH 7.45). Tubular secretion of H+ is important in maintaining control of the pH of the blood. When the pH of the blood starts to drop, more hydrogen ions are secreted. If the blood should become too alkaline, secretion of H+ is reduced. In maintaining the pH of the blood within its normal limits of 7.3–7.4, the kidney can produce a urine with a pH as low as 4.5 or as high as 8.5. The substances that are secreted into the tubular fluid (for removal from the body) include: Potassium ions (K+), Hydrogen ions (H+), Ammonium ions (NH4+), creatinine, urea, some hormones, and some drugs (e.g. penicillin). Tubular secretion occurs from the epithelial cells that line the renal tubules and collecting ducts. It is the tubular secretion of H+ and NH4+ from the blood into the tubular fluid (i.e. urine - which is then excreted from the body via the ureter, bladder, and urethra) that helps to keep blood pH at its normal level. The movement of these ions also helps to conserve sodium bicarbonate (NaHCO3). The typical pH of urine is about 6. Urine formed via the three processes outlined above trickles into the kidney pelvis. At this final stage it is only approx. 1% of the originally filtered volume but includes high concentrations of urea and creatinine, and variable concentrations of ions. ***
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13. COMPOSITION OF URINE 13.1 13.2 13.3 13.4 13.1 INTRODUCTION Urine is a transparent solution that is clear to amber in color, and usually is light yellow. It is the byproduct or waste fluid secreted by the kidneys, transported by the ureters to the urinary bladder where it is stored until it is voided through the urethra. Urine is made up of a watery solution of metabolic wastes (such as urea), dissolved salts and organic materials. Fluid and materials being filtered by the kidneys, destined to become urine, comes from the blood or interstitial fluid. The composition of urine is adjusted in the process of reabsorption when essential molecules needed by the body, such as glucose, are reabsorbed back into the blood stream via carrier molecules. The remaining fluid contains high concentrations of urea and other excess or potentially toxic substances that will be released from the body via urination. Urine flows through these structures: the kidney, ureter, bladder, and finally the urethra. Urine is produced by a process of filtration, reabsorption, and tubular secretion. Urine contains large amounts of urea, an excellent source of nitrogen for plants. As such it is a useful accelerator for compost. Urea is 10,000 times less toxic than ammonia and is a byproduct of deamination (2 NH3 molecules) and cellular respiration's. (1 CO2 molecule) products combining together. Other components include various inorganic salts such as sodium chloride (the discharge of sodium through urine is known as "natriuresis".) 13.2 CHEMICAL COMPOSITION OF URINE The composition of normal urine is: 1) Volume: 600- 2500 ml/24 hrs. Average: 1,200 ml. 2) Specific gravity: 1.003 - 1.030 3) Reaction: Acidic (pH: 4.7 - 7.5) Average pH: 6.0 4) Total solids: 30 - 70 g/liter. Urine is aprox. 95% water. The other components of normal urine are the solutes that are dissolved in the water component of the urine. These solutes can be divided into two categories according to their chemical structure (e.g. size and electrical charge). Organic molecules are electrically neutral and can be relatively large (compared with the 'simpler' ions - below). These include: Urea - Urea is an organic (i.e. carbon-based) compound whose chemical formula is: CON2H4 or (NH2)2CO. It is also known as carbamide. Urea is derived from ammonia and produced by the deamination of amino acids. The amount of urea in urine is related to quantity of dietary protein. Creatinine - Creatinine is a normal (healthy) constituent of blood. It is produced mainly as a result of the breakdown of creatine phosphate in muscle tissue. It is usually produced by the body at a fairly constant rate (which depends on the muscle mass of the body). Uric acid - Uric acid is an organic (i.e. carbon-based) compound whose chemical formula is: C5H4N4O3.
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INTRODUCTION FILTRATION REABSORPTION SECRETION
Due to its insolubility, uric acid has a tendency to crystallize, and is a common part of kidney stones. Other substances/molecules - Example of other substances that may be found in small amounts in normal urine include carbohydrates, enzymes, fatty acids, hormones, pigments, and mucins (a group of large, heavily glycosylated proteins found in the body). Ions are atoms or groups of atoms that have either, lost some outer electrons, hence have a positive electric charge, or have gained some outer electrons (to the atom or group of atoms), and hence have a negative electric charge. Even in the cases of ions formed by groups of atoms (they are ions due to the few lost or gained electrons), the groups are formed from only a small number of particles and therefore tend to be relatively small. These include: Individual elements: Sodium (Na+) : Amount in urine varies with diet and the amount of aldosterone (a steroid hormone) in the body. Potassium (K+) : Amount in urine varies with diet and the amount of aldosterone (a steroid hormone) in the body. Chloride (Cl-) : Amount in urine varies with dietart intake (chloride is a part of common salt, NaCl). Magnesium (Mg2+) : Amount in urine varies with diet and the amount of parathyroid hormone in the body. (Parathyroid hormone increases the reabsorption of magnesium by the body, which therefore decreases the quantity of magnesium in urine.) Calcium (Ca2+) : Amount in urine varies with diet and the amount of parathyroid hormone in the body. (Parathyroid hormone increases the reabsorption of calcium by the body, which therefore decreases the quantity of calcium in urine.) Small groups formed from a few different elements: Ammonium (NH4+) : The amount of ammonia produced by the kidneys may vary according to the pH of the blood and tissues in the body. Sulphates (SO42-) : Sulphates are derived from amino acids. The quantity of sulphates excreted in urine varies according to the quantity and type of protein in the person's diet. Phosphates (H2PO4-, HPO42-, PO43-) : Amount in urine varies with the amount of parathyroid hormone in the body - parathyroid hormone increases the quantity of phosphates in urine. Following inorganic constituents (listed in table) are excreted per 24 hours. Note: These values generally vary with diet. Constituent 1) Sodium 2) Potassium 3) Chlorides 4) Calcium 5) Inorganic phosphorus 6) Sulfur 7) Magnesium 8) Ammonia 9) Iodine 10) Arsenic
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Quantity excreted/24 hrs. 3-4g 1.5 – 2 g 9 - 16g 0.1 - 0.3 g 1 - 1.5 g 0.7 - 3.5 g 0.05 - 0.2 g 0.3 - 1.0 g 50 - 250 g less than 50 µg
Dr. M. Estari
11) Lead 12) Urea 13) Creatinine 14) Uric acid 15) Creatine 16) Hippuric acid 17) Purine bases 18) Ketone bodies 19) Oxalic acid 20) Indican 21) Allantoin 22) Coproporphyrins 23) Phenols 24) Vitamins, hormones, and enzymes
less than 50 µg 25 - 30 g 1 - 1.8 g 0.3 - 1.0 g 60 – 150 mg 0.1 - l.0 g 7 – 10 mg 3 - 15 mg 15 - 20 mg 4 - 2 mg 20 - 30 mg 60 - 280 µg 0.2 - 0.5 g Detected in small quantities
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Dr. M. Estari
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