MEDICINE AND SURGERY "F" Course of LABORATORY MEDICINE Disturbances of the metabolism of sugars and fat INHERITED DEFECTS OF SUGAR METABOLISM THE REGISTRATION OF ATTENDANCE TO THIS ON-LINE LECTURE IS ACTIVE!       To register your attendance please type in your surname and matricola number. Surname: Matricola: Notice that your attendance will be registered only if you completed the reading, questions, and audios, and that you cannot interrupt and resume the session (but you can repeat it as many times as you like). Remember to press the [send] button before leaving this page! A confirmation message will appear at the end of this page.       A comment section has been added at the end of this lecture. Adding a comment or question does not require registration with your matricola number, feel free to comment whenever you like. DISTURBANCES OF SUGAR METABOLISM       Favism       Favism results from autosomal, recessive inheritance of a poorly functional variant of the enzyme glucose-6-phosphate dehydrogenase (G6PDH). G6PDH catalyzes the initial step of the pentose phosphate pathway, converting glucose-6P to P-gluconolactone. Since the pathway is of fundamental importance for many tissues, notably for the erythrocytes, complete lack of G6PDH activity is incompatible with life. Reduced activity of the enzyme in the heterozygous state is asymptomatic; in the homozygous state causes a mild anemia and makes the patient extremely sensitive to oxidative stress. Ingestion of, or contact with, pro-oxidant substances causes a severe hemolytic crysis with marked hemoglobinuria; severe cases may result fatal. A dietary source of pro-oxidant compounds is the fava bean (but also its pollen!), that may contain alkaloids like divicine, whose formula is reported below.       In these patients the erythrocyte lifespan is reduced and a mild anemia is often present. In the mediterranean countries fava beans are a common food and the patient may experience repeated (hopefully minor) crises just because of pollen inalation, thus suggesting the diagnosis, which is confirmed by gene sequencing.       Favism is relatively common in (formerly) malaria endemic mediterranean countries because of a positive evolutionary selection. Indeed the malaria parasites metabolyzes hemoglobin and releases heme derivatives (hemozoin) in the sytoplasm of the infected erythrocyte. Hemozoin has pro-oxidant activity and causes the infected erythrocyte to rupture before the intraerythrocytic phase of malaria infection is completed, leading to an abortive infection. As a consequence the prevalence of favism in mediterranean population is surprisingly high. Further readings Luzzatto L. and Arese P. Favism and Glucose-6-Phosphate Dehydrogenase Deficiency N.E. J. Med. 2018, 378: 60-71.       Non diabetic melliturias       Non diabetic melliturias are a group of inherited diseases of monosaccharides that cause a sugar other than glucose to accumulate in the blood and in the urine. Since the only sugar in the blood of healthy subjects is glucose, these conditions only occur as a result of an inherited enzymatic defect. The most important cases are those of galactose, fructose, and xilulose.       Galactosemia and other disturbances of galactose catabolism. Galactose is a very important nutrient for newborns, because the milk sugar is lactose, a dimer of galactose and glucose. Galactose is a epimer of glucose, and requires conversion to glucose in order to be metabolized. The metabolic conversion of galactose to glucose is surprisingly complex and occurs via the Leloir pathway, as depicted below:       The pathway may be described as follows: 1) Galactose may be present in both stereoisomers of C1: αD galactose and βD galactose. Only αD galactose can be processed, but a specific mutarotase exists which rapidly interconverts the two isomers and allow all the sugar to be processed. 2) The enzyme galactokinase uses ATP to convert αD galactose to αD galactose-1-phosphate. Galactokinase deficiency is a quite severe disease of the newborn which is inherited as an autosomal recessive trait. Diagnosis is suspected because of stunted growth and presence of galactose in the blood and in the urine, and is confirmed by gene sequencing. If untreated, the disease leads to early cataract because of the accumulation of galactitol in the lens. Therapy is dietetic (artificial milk in which lactose is replaced by glucose). 3) The enzyme galactose-1-P uridyl transferase uses UDP-glucose as the second substrate and transfers galactose on the uridyl diphosphate moiety, releasing glucose. Deficiency of this enzyme, inherited as an autosomal recessive trait, leads to galactosemia, a very severe disease in which stunted growth and brain and liver damage are prominent. Early cataract may also develop. Diagnosis and treatment are similar to those described for galactokinase deficiency. 4) UDP-galactose is the substrate of a specific epimerase, whose product is UDP-glucose. Deficiency of UDP-galactose epimerase is inherited as an autosomal recessive trait. This condition is rare and relatively benign. Galactose-1-P accumulates in the erythrocytes and may leak in the blood and urine. Therapy is usually not required. The subsequent fate of UDP-glucose may be: incorporation into glycogen; glycolysis; or secretion in the blood.       The Leloir pathway occurs mainly in the liver, and impairment of galactose catabolism may be observed in prematurity or in conditions of liver failure; however, only in the diet of the newborn is galactose abundant enough that defects of its metabolism can cause significant symptoms.       Fructosemia, fructosuria and other disturbances of fructose catabolism.       Fructose is not a food for newborns, thus inherited defects of the metabolism of this sugar become apparent after weaning. Fructose is contained as such in some foods (e.g. honey), and is a constituent of sucrose (table sugar). Its metabolic usage is quite straightforward: the enzyme fructokinase uses ATP to convert fructose to fructose-1-phosphate; then phosphofructoaldolase decomposes fructose-1P into dihydroxyacetone phosphate (a metabolyte of glycolysis; no further conversion needed) and glyceraldehyde. Glyceradldehyde kinase converts glyceraldehyde to glyceraldehyde-3-phosphate that enters into glycolysis.       Fructosuria is caused by an inherited deficiency of fructokinase, an autosomal and recessive benign condition causing fructose to appear in the blood and in the urine. Formerly sugars in the urine were often confused with glucose, suggesting a diagnosis of diabetes, but nowadays the enzymatic tests used for glucose are specific enough to avoid these errors. The subject does not require therapy, but may be subject to frequent infections (cutaneous, urinary, etc.) thus a diet poor in fructose is advisable.       Hereditary fructose intolerance is a more severe clinical condition caused by a deficiency of phosphofructoaldolase activity. It is inherited as an autosomal and recessive trait. Accumulation of fructose-1P inhibits glyconeogenesis and glycogenolysis, resulting in severe crises of hypoglycemia, and possibly coma. The patient usually learns by himself or herself to avoid sucrose and fructose containing foods. The laboratory diagnosis relies on the finding of fructose in the blood and in the urine, which however the patient will try to keep at bay by his/her diet. The hypoglycemic crisis may be induced by intravenous administration of fructose, but this test is dangerous and may require prompt administration of glucose solutions. Gene sequencing confirms the diagnosis.       Inherited Deficiency of fructose-1,6P diphosphatase causes a blockade of glyconeogenesis and possible hypoglycemic crises; moreover accumulation of glyconeogenesis precursors (e.g. lactic acid or piruvic acid) may cause a metabolic acidosis. Diagnosis relies on gene sequencing, therapy is required only during the hypoglycemic crises, but a diet rich in glucose (starch) and poor in glyconeogenesis precursors (e.g. glycogenic aminoacids) is advisable.       Pentosuria is a rare, X-linked inherited disturbance of the metabolism of L-xylulose, a sugar that appears in the blood and urine. This condition is benign and no therapy is required. Diagnosis is by gene sequencing. Disturbances of the metabolism of monosaccharides disease enzyme/gene affected symptoms laboratory findings favism G6PDH hemolytic crises caused by oxidative stress (e.g. by fava beans) anemia, reduced G6PDH activity in the erythrocytes, gene sequencing galactokinase deficiency stunted growth, early cataract galactose in the blood and urine, gene sequencing galactosemia galactose-1-P uridyl transferase stunted growth, mental retardation, early cataract galactose in the blood and urine, gene sequencing deficiency of UDP-galactose epimerase asymptomatic gene sequencing fructosuria fructokinase essentially asymptomatic fructose in the blood and urine, gene sequencing hereditary fructose intolerance phosphofructoaldolase hypoglycemic crises, coma fructose in the blood and urine, hypoglycemia, gene sequencing fructose 1,6 diphosphatase deficiency hypoglycemic crises, coma hypoglycemia, metabolic acidosis, gene sequencing pentosuria L-xylulose dehydrogenase essentially asymptomatic L-xylulose in the blood and urine, gene sequencing       Glycogenoses       Glycogen is a glucose polymer used for storage of this important nutrient. It is present in the liver (circa 100 g under normal conditions) and in the striated muscles (a few grams). Liver glycogen is released under glucagon stimulation and serves to prevent fasting hypoglycemia. Muscle glycogen serves to allow the replenishment of oxaloacetate (produced via glycolysis and piruvate carboxylase) in the mitochondrion during strong muscular effort. It is imporant to underline that striated muscle uses mainly fatty acids for energy production. Some of the enzymes of glycogen metabolism are common to muscle and liver; of other enzymes there are tissue specific isoforms, thus inherited disturbances of glycogen metabolism may affect both liver and muscle or may be tissue specific. The figure below depicts the main steps of glycogen biosynthesis and degradation (biosynthetic reactions in black; cytoplasmic depolimerization reactions in blue; lysosomal depolimerization reactions in green; diseases in red).       Inherited disturbances of glycogen metabolism are collectively called glycogenoses. Their symptoms and laboratory findings differ in the case of glycogenoses affecting the liver or the muscle isoenzymes. Liver glycogenoses cause hypoglycemic crises, often severe, enlargement of the liver, and may lead to liver failure, with all associated laboratory signs. Muscle glycogenoses cause muscular weakness and fatigue, possibly heart enlargement and subsequent failure, but laboratory findings are scarce. Diagnosis is confirmed by gene sequencing. Glycogenoses disease enzyme/gene affected organ and symptoms laboratory findings 0: deficiency of glycogen synthase muscle and liver; kidney; hypoglycemic crises, liver steatosis fasting hypoglycemia, gene sequencing Ia: Von Gierke disease glucose-6-phosphatase liver and kidney; severe hypoglycemic crises, stunted growth, liver steatosis fasting hypoglycemia, metabolic acidosis, gene sequencing Ib glucose-6-P translocase similar to Ia, but less severe same as Ia II: Pompe disease lysosomal glycosidase affects all organs; enlargement and (later) failure of liver and heart biopsy reveals accumulation of glycogen in lysosomes; no laboratory signs; gene sequencing III: Forbes disease debrancher enzyme liver and muscle; fasting hypoglycemia fasting hypoglycemia, gene sequencing IV: Andersen disease brancher enzyme liver, muscle, heart enlargement and (later) failure gene sequencing V: Mc Ardle disease glycogen phosphorylase muscle; weakness, cramps on exercise gene sequencing VI: Hers disease glycogen phosphorylase liver; hypoglycemic crises fasting hypoglycemia, gene sequencing VII: Tarui disease phosphofructokinase muscle; cramps on exercise gene sequencing DISTURBANCES OF FAT METABOLISM       Inherited defects of cholesterol biosynthesis       Several disorders of cholesterol biosynthesis are described, the most important of which is mevalonic aciduria.       The transport of lipids across the placenta is complex: indeed, while water soluble metabolites (e.g. glucose, ammonia, urea, etc.) cross freely the placenta, triglycerides and cholesterol require receptor mediated transport because they are present in the maternal serum as lipoprotein complexes and lipoproteins do not cross the placenta (see Herrera et al. 2006). Triglycerides are uploaded by the placental cells, digested, and transferred to the foetal blood as free fatty acids, for the metabolic requirements of the foetus. This process occurs over the whole gestation. The placental uptake of cholesterol is significant during the early pregnancy, but becomes less and less relevant as gestation proceeds, and the foetus depends more and more on cholesterol biosynthesis. Thus, genetic defects of cholesterol biosynthesis cannot be compensated by the mother and cause developmental defects of the foetus, evident at birth. The newborn may present obvious physical (facial) malformations.       Several inheritable enzymatic defects of cholesterol biosynthesis are known. Since cholesterol is necessary for embrionic and fetal organogenesis these defects may be associated to multiple malformations (e.g. microcephaly) and hypocholesterolemia. The most important genetic defect of cholesterol biosynthesis is due to mutation of gene MVK encoding for the enzyme mevalonate kinase. Two different diseases are associated to defects of MVK, depending on the type of mutation and the extent of loss of enzymatic activity. The precursor of mevalonic acid is β-hydroxy, β-methyl glutarylCoA, which is also an intermediate in the catabolism of the aminoacid leucine; thus this aminoacid and its metabolytes should also be monitored in mevalonic aciduria. Mevalonic aciduria is the most severe form of cholesterol biosynthesis defect and is associated to mental retardation, hepatosplenomegaly, metabolic acidosis and various dismorfisms; hyperimmunoglobulinema D is the less severe form and is associated to recurrent fever. Mevalonic acid accumulates in the serum and in the urine and is easily detected by the clinical laboratory. Diagnosis is confirmed by gene sequencing.       Further readings:       Platt et al. Disorders of cholesterol metabolism and their unanticipated convergent mechanisms of disease. Annu Rev Genomics Hum Genet. 2014; 15: 173–194.       Gangliosidoses and other disturbances of complex lipid degradation       Triglycerides and phosphoglycerides are ubiquitous energy sources and cell components, and their (relatively simple) metabolism is too relevant to the cell survival to tolerate genetic diseases. More complex sphingolipids, however, are components of specific tissues and metabolic defects are compatible at least with fpetal life, resulting in newborns affected by inherited diseases, globally called gangliosidoses. Since gangliosides are chiefly metabolized inside cells, genetic defects of their metabolism do not release intermediates in the blood and the laboratory diagnosis of these defects mainly relies on gene sequencing. In particular the catabolism of gangliosides and cerebrosides occurs in the lysosomes of macrophages and other specialized cells and in the presence of genetic defects of the pertinent enzymes these cells enlarge significantly, and appear "foamy" in the microscope (thus biopsy is an important diagnostic procedure). The liver and spleen of the affected children may be very significantly enlarged, but this sign may require some years to fully develop. The bone marrow may also be involved leading to pancytopenia. The prognosis is usually poor. Some ethnic groups are particularly affected, notably Ashkenazi Jews. Gangliodidoses disease enzyme/gene affected symptoms and diagnosis Gaucher's disease glucocerebrosidase autosomal and recessive; massive hepato-splenomegaly; involvement of the bone marrow may lead to pancytopenia; mental retardation; demyelinization of long nervous fibers Niemann-Pick disease sphingomyelinase autosomal and recessive; massive hepato-splenomegaly; involvement of the bone marrow may lead to pancytopenia Fabry's disease α-galactosidase A (hydrolyzes galactosyl-ceramides) sex linked; renal or cardiac failure Tay-Sachs disease hexosaminidase A accumulation of GM2 gangliosides leads to brain damage and mental retardation Wolman's disease acid lipase accumulation of cholestryl-esters in the liver and spleen; survival beyond 6 months of age is uncommon   Other inherited defects of lysosomal enzymes Ceramide accumulation (Farber disease) ceramidase   Glycogenosis type II (Pompe disease) α-glicosidase (acid maltase/isomaltase)   Acid phosphatase deficiency acid phosphatase   Fucosidosis α-fucosidase   Mannosidosis α-mannosidase   Aspartylglucosaminuria Aspartylglucosamine glucosidase aspartylglucosamine peptides released in the blood and urine Mucopolysaccaridosis I (Hurler disease) α-iduronidase   Mucopolysaccaridosis II (Hunter disease) iduronosulfate sulfatase   Mucopolysaccaridosis III (Sanfilippo disease; several variants) heparansulfate hydrolases   Mucopolysaccaridosis IV (Morquio disease) N-actylgalactosamine sulfate sulfatase   Mucopolysaccaridosis VI (Moroteaux-Lamy disease) N-acetyl hexosamine sulfate sulfatase (arylsulfatase B)   Mucopolysaccaridosis VII β-glucuronidase   Multiple sulfatase deficiency arylsulfatases A, B, and C   Sialidosis sialidase (glycoprotein neuroaminidase)   Mucolipidosis I, II, III UDP-N-acetylglucosamine transferase   Mucolipidosis VI ganglioside neuroaminidase Questions and exercises: 1) Possible causes of hypoglycemia include: Insulinoma, liver glycogenosis, Cushing's disease Insulinoma, liver glycogenosis, Addison's disease Insulinoma, liver glycogenosis, favism 2) Laboratory findings in a case of favism may be: allergic reaction to fava bean alkaloids anemia, metabolic acidosis, glucose-6-phosphate in the urine anemia, reduced activity of G6PDH in the erythrocytes, homozygous mutation of G6PDH 3) Mevalonic aciduria is an inherited defect of cholesterol biosynthesis and leucine degradation an inherited defect of ganglioside metabolism an inherited defect of cholesterol catabolism 4) The main finding common to the majority of gangliosidoses is: enlargement of internal organs due to accumulation of non-metabolized sphingolipids mental retardation pancytpenia your score: 10 Attendance not registered because of insufficient score - Retry! You can type in a comment or question below (max. length=160 chars.); please specifiy that the subject of your comment or question is aminoacid metabolism: All comments posted on the different subjects have been edited and moved to this web page (for optimal reading try to have at least 80 characters per line)! Thank you Professor (lecture on bilirubin and jaundice). The fourth recorded part, the one on hyper and hypoglycemias is not working. Bellelli: I checked and in my computer it seems to work. Can you better specify the problem you observe? This Presentation (electrolytes and blood pH) feels longer than previous lectures Bellelli: it is indeed. Some subjects require more information than others. I was thinking of splitting it in two nest year. Bellelli in response to a question raised by email: when we compare the blood pH with the standard pH we do not mean to compare the "normal" blood pH (7.4) with the standard pH. Rather we compare the actual blood pH of the patient, with the pH of the same blood sample equilibrated under standard conditions. Thus, if we say that standard pH is lower than pH we mean that equilibriation with 40 mmHg CO2 has caused absorption of CO2 and has lowered the pH with respect to its value before equilibration. (Lipoproteins) Is the production of leptin an indirect cause of type 2 diabetes since it works as a stimulus to have more adipose tissue that produces hormones? Bellelli: in a sense yes, sustained increase of leptin causes the hypothalamus to adapt and to stop responding. Obesity ensues and this in turn may cause an increase in the production of resistin and other insulin-suppressing protein hormones produced by the adipose tissue. However, this is quite an indirect link, and most probably other factors contribute as well. (Urea cycle) what is the meaning of "dissimilatory pathway"? Bellelli: a dissimilatory pathway is a catabolic pathway whose function is not to produce energy, but to produce some terminal metabolyte that must be excreted. Dissimilatory pathways are necessary for those metabolytes that cannot be excreted as such by the kidney or the liver because they are toxic or poorly soluble. Examples of metabolytes that require transformation before being eliminated are heme-bilirubin, ammonia, sulfur and nitrogen oxides, etc. Talking about IDDM linked neuropathy can be the C peptide absence considered a cause of it?? Bellelli: The C peptide released during the maturation of insulin, besides being an indicator of the severity of diabetes, plays some incompletely understood physiological roles. For example it has been hypothesized that it may play a role in the reparation of the atherosclerotic damage of the small arteries. Thus said, I am not aware that it plays a direct role in preventing diabetic polyneuropathy. Diabetic neuropathy has at least two causes: the microvascular damage of the arteries of the nerve (the vasa nervorum), and a direct effect of hyperglycemia and decreased and irregular insulin supply on the nerve metabolism. Diabetic neuropathy is observed in both IDDM and NIDDM, and requires several years to develop. Since the levels of the C peptide differ in IDDM and NIDDM, this would suggest that the role of the C peptide in diabetic neuropathy is not a major one. If you do have better information please share it on this site! In acute intermitted porphyria and congenital erythropoietic porphyria why do the end product of the affected enzymes accumulate instead of their substrate?? Bellelli: First of all, congratulations! This is an excellent question. Remember that a condition is which the heme is not produced is lethal in the foetus; thus the affected enzyme(s) must maintain some functionality for the patient to be born and to come to medical attention. All known genetic defects of heme biosynthesis derange but do not block this metabolic pathway. Congenital Erythropoietc Porphyria (CEP) is a genetic defect of uroporphyrinogen III cosynthase. This protein associates to uroporphyrinogen synthase (which is present and functional in CEP) and guarantees that the appropriate uroporphyrinogen isomer is produced (i.e. uroporphyrinogen III). In the absence of a functional uroporphyrinogen III cosynthase other possible isomers of uroporphyrinogen are produced together with uroporpyrinogen III, mostly uroporphyrinogen I. The isomers of uroporphyrinogen that are produced differ because of the positions of propionate and acetate side chains, and this in turn is due to the pseudo symmetric structure of porphobilinogen. Only isomer III can be further used to produce protoporphyrin IX. Thus in the case of CEP we observe accumulation of abnormal uroporphyrinogen derivatives, which, as you correctly observed are the products of the enzymatic synthesis operated by uroporphyrinogen synthase. The case of Acute Intermittent Porphyria (AIP) is similar, although there may be variants of this disease. What happens is that either the affected enzyme is a variant that does not properly associate with uroporphyrinogen III cosynthase or presents active site mutations that impair the proper alignement of the phoprphobilinogen substrates. In either case abnormal isomers of uroporphyrinogen are produced, as in CEP. Also remark that in both AIP and CEP we observe accumulation of the porphobilinogen precursor: this is because the overall efficiency of the biosynthesis of uroporphyrinogens is reduced. Thus: (i) less uroporphyrinogen is produced, and (ii) only a fraction of the uroporphyrinogen that is produced is the correct isomer (uroporphyrinogen III). is it possible to take gulonolactone oxidase to synthesize vitamin C instead of vitamin C supplement? Bellelli: no, this approach does not work. The main reason is that the biosynthesis of vitamin C, as almost all other metabolic processes, occurs intracellularly. If you administer the enzyme it will at most reach the extracellular fluid but will not be transported inside the cells to any significant extent. Besides, there are other problems in this type of therapy (e.g. the enzyme if administered orally, may be degraded by digestive proteases; if administered parenterally, may cause the immune system to react against a non-self protein). In theory one could think of a genetic modification of the inactive human gene of gulonolactone oxidase, but the risk and cost of this intervention would not be justified. In addition to these considerations, except for cases of shipwreckage or other catastrophes, a proper diet or administration of tablets of vitamin C is effective, risk-free and unexpensive, thus no alternative therapy is reasonable. However, I express my congratulations for your search on the biosynthesis pathway of ascorbic acid. Resorption and not reabsorption would lead to hypercalcemia ie bone matrix being broken down. Bellelli: I am not sure to interpret your question correctly. Resorption indicates destruction of the bone matrix and release of calcium and phosphate in the blood, thus it causes an increase of calcemia. Reabsorption usually means active transport of calcium from the renal tubuli to the blood, thus it prevents calcium loss. It prevents hypocalcemia, and thus complement bone resorption. To avoid confusion it is better use the terms "bone resorption" and " renal reabsorption of calcium". If you have a defect in renal reabsorption, parthyroid hormone will be released to maintain a normal calcium level by means of bone resorption; the drawback is osteoporosis. In Reed and Frost model: I haven't understood what is the relationship between K and R reproductive index. Thank you Professor! Bellelli: in the Reed and Frost model K is the theoretical upper limit of R0. R the reproductive index is the ratio (new cases)/(old cases) measured after one serial generation time. R0 is the value of R one measures at the beginning of the epidemics, when in principle all the population is susceptible. What is the link between nucleotide metabolism and immunodeficiencies and mental retardation? Bellelli: the links may be quite complex, but the principal ones are as follows: 1) the immune response requires a replication burst of granulocytes and lymphocytes, which in turn requires a sudden increase of nucleotide production, necessary for DNA replication. Defects of nucleotide metabolism impair this phase of the immune defense. Notice that the mechanism is similar to the one responsible of anemia which requires a sustained biosynthesis of nucleotides at a constant rate, rather than in a burst. 2) Mental retardation is mainly due to the accumulation of nulceotide precursors in the brain of the newborn, due to the incompletely competent blood-brain barrier. How can ornithine transaminase defects cause hyperammonemia? Is it due to the accumulation of ornithine that blocks the urea cycle or for other reasons? Bellelli: ornithine transaminase is required for the reversible interconversion of ornithine and proline, and thus participates to both the biosynthesis and degradation of ornithine. The enzyme is synthesized in the cytoplasm and imported in the mitochondrion. Depending on the metabolic conditions the deficiency of this enzyme may cause both excess (when degradation would be necessary) or defect (when biosynthesis would be necessary) of ornithine; in the latter case, the urea cycle slows down. Thus there is the paradoxical condition in which alternation may occur between episodes of hyperammonemia and of hyperornithinemia. When we use the Berthelot's reaction to measure BUN do we also have to measure the concentration of free ammonia before adding urease? Bellelli: yes, in principle you should. Berthelot's reaction detects ammonia, thus one should take two identical volumes of serum, use one to measure free ammonia, the other to add urease and measure free ammonia plus ammonia released by urea. BUN is obtained by difference. However, free ammonia in our blood is so much lower than urea that you may omit the first sample, if you only want to measure BUN. Why do we have abnormal electrolytes in hematological neoplasia e.g. leukemia? Bellelli: I do not have a good explanation for this effect, which may have multiple causes. However, you should consider two factors: (i) acute leukemias cause a massive proliferation of leukocytes (or lymphocytes depending on the cell type affected) with a very shortened lifetime; thus you observe an excess death rate of the neoplastic cells. The dying cells release in the bloodstream their content, which has an electrolyte composition different from that of plasma: the cell cytoplasm is rich in K and poor in Na, thus causing hyperkalemia. (ii) the kidney may be affected by the accumulation of neoplastic white cells or their lytic products. Gaussian curve: If it is bimodal is it more likely to be a "certain diagnosis" than if it is unimodal or does it only show the distinguishment from health? Bellelli an obviously bimodal Gaussian curve indicates that the disease is clearly separated from health: usually it is a matter of how precise and clear-cut is the definition of the disease. For example tuberculosis is the disease caused by M. tuberculosis, thus if the culture of the sputum is positive for this bacterium you have a "certain" diagnosis (caution: the patient may suffer of two diseases, e.g. tuberculosis and COPD diagnosis of the first does not exclude the second). However, in order to have a "certain" diagnosis it is not enough that distribution of the parameter is bimodal, it is also required that the patient's parameter is out of the range of the healthy condition: this is because a distribution can be bimodal even though it is composed by two Gaussians that present a large overlap, and the patient's parameter may fall in the overlapping region. Thus, in order to obtain a "certain" diagnosis you need to consider not only the distribution of the parameter(s) but also the patient's values and the extent of the overlapping region. Prof can you please elaborate a bit more on the interhuman variability and its difference with the interpopulation variability please? Bellelli: every individual is a unique combination of different alleles of the same genes; this is the source of interindividual variability. Every population is a group of individuals who intermarry and share the same gene pool (better: allele pool). Every allele in a population has its own frequency. Two population may differ because of the diffferent frequencies of the same alleles; in some cases one population may completely lack some alleles. The number and frequencies of alleles of each gene determine the variance. If you take two populations and calculate the cumulative interindividual variance of the population the number you obtain is the sum of two contributions: the interindividual variance within each population, plus the interpopulation variance between the means of the allele frequencies. For example, there are human population in which the frequency of blood group B is close to 0% and other populati ons in which it is 30% or more. Prof can you please explain again the graph you have showed us in class about thromboplastin? (Y axis=abs X axis= time) Bellelli: the graph that I crudely sketched in class represented the signal of the instrument (an absorbance spectrophotometer) used to record the turbidity of the sample (turbidimetry). The plasma is more or less transparent, before coagulation starts. When calcium and the tissue factor (or collagen) are added. thrombin is activated and begins digesting fibrinogen to fibrin; then fibrin aggregates. The macroscopic fibrin aggregates cause the sample to become turbid, which means it scatters the incident light. The instrument reads this as a decrease of transmitted light (i.re an increase of the apparent absorbance) and the time profile of the signal presents an initial lag phase, which is called the protrombin or thromboplastin time depending on the component which was added to start coagulation (tissue factor or collagen). Prof can you please explain the concept you have described in class about the simultaneous hypercoagulation and hemorrhagic syndrome? How can this occur? Bellelli: The condition you describe is observed only in the Disseminated Intravascular Coagulation syndrome. Suppose that the patient experiences an episode of acute pancreatitis: tripsin and chymotripsin are reabsorbed in the blood and proteolytically activate coagulation causing an extensive consumption of fibrinogen and other coagulation factors. Tripsin and chymotripsin also damage the vessel walls and may cause internal hemorrages, but at that point the consumption of fibrinogen may have been so massive that not enough is left to form the clot where the vessel has been damaged, causing an internal hemorrage. Pancreatitis is a very severe, potentially lethal condition, and DIC is only one of the reasons of its severity. You said that certain drugs (ethanol, cocaine, cannabis, opiates...) cause a necessity of higher and higher dosage, for two reasons: the enzyme in the liver is inducible and the receptors in the brain are expressed less and less. So, first, I am not sure I got it right, and second I did not understand how expressing less receptors leads to a necessity of higher dosage. Bellelli: You got it correctly, but the detailed mechanism of resistance may vary among different substances, and not all drugs cause adaptation. The reason why reducing the number of receptors may require an increased dosage of the drug is as follows: suppose that a certain cell has 10,000 receptors for a drug. When bound to its agonist/effector, each receptor produces an intracellular second messenger. Suppose that in order for the cell to respond 1,000 receptors must be activated. The concentration of the effector required is thus the concentration that produces 10% saturation. You can easily calculate that this concentration is approximately 1/10 of the equilibrium dissociation constant of the receptor-effector complex (its Kd), the law being Fraction bound = [X] / ([X]+Kd) where [X] is the concentration of the free drug. After repeated administration, the subject becomes adapted to the drug, and his/her cells express less receptors, say 5,000. The cell response will in any case require that 1,000 receptors are bound to the effector and activated, but this now represents 20% of the total receptors, instead of 10%. The drug concentration required is now 1/4 of the Kd. Continuing administration of the drug further reduces the cell receptors, but the absolute number of activated receptors required to start the response is constant; thus the fewer receptors on the cell membrane, the higher the fraction of activated receptors required. Why does hyperosmolarity happen in type 2 diabetes and not in type 1? Bellelli: Hyperosmolarity can occur also in type 1 diabetes, albeit infrequently. The approximate formula for plasma osmolarity is reported in the lecture on electrolytes: osmolarity = 2 x (Na+ + K+) + BUN/2.8 + glucose/18 this is expressed in the usual clinical laboratory units (mEq/L for electrolytes, g/dL for non- electrolytes). The normal values are: osmolarity = 2 x (135 + 5) + 15/2.8 + 100/18 = 280 + 5.4 + 5.6 = 291 mOsmol/L Let's imagine a diabetic patient having normal values for electrolytes and BUN, and glycemia=400 mg/dL: osmolarity = 280 + 5.4 + 22.4 = 307.8 mOsmol/L The hyperosmolarity in diabetes is mainly due to hyperglycemia, even though other factors may contribute (e.g. diabetic nefropathy); however the contribution of glucose to osmolarity is relatively small. As a consequence in order to observe hyperosmolarity the hyperglycemia should be extremely high; this is more often observed in type 2 than in type 1 diabetes, for several reasons, the most relevant of which is that in type 1 diabetes all cells are starved of glucose, and the global reserve of glycogen in the body is impoverished: there is too much glucose in the blood and too few everywhere else, thus reducing, but not abolishing, the risk of extreme hyperglycemia. Usually in type 2 diabetes the glycogen reserve in the organism is not impoverished, thus the risk of extreme hyperglycemia is higher. Hemostasis and Thrombosis lecture: I don't understand why is sodium citrate added to the serum solution to measure the prothrombin time. Bellelli: in order to measure PT or PTT you want to be able to start the coagulation process at an arbitrary time zero, and measure the increase in turbidity of the serum sample. To do so you need (i) to prevent spontaneous coagulation with an anticoagulant; and (ii) to be able to overcome the anticoagulant at your will. Citrate (or oxaloacetate; or EDTA) has the required characteristics: it chelates calcium, and in this way it prevents coagulation; but you can revert its effect at your will by adding CaCl2 in excess to the amount of citrate. You cannot obtain the same effect with other anticoagulants (e.g. heparin) whose action cannot be easily overcome. Dear professor I cannot do the self evaluation test because it says the the time has expired It is not possible because I havent even started them Bellelli: this is due to the fact that the program registers your name and matricola number from previous attempts. I shall fix this bug. Meanwhile try to use a fake matricola number. How is nephrotic syndrome associated hypoalbuminemia as you described in methods of analysis of protein because seems counterintuitive Bellelli: nephrotic syndrome is an autoimmune disease in which the glomerulus is damaged and the filtration barrier is disrupted; diuresis is normal but there is loss of proteins (mostly albumin) in the urine. I m sorry i confused polyurea with hypoalbuminemia but my question still stands during glomerulonephritis you mentioned something of polyurea as compensation i could not follow how this compensation mechanism works and collapse after some time in glomerulonephritis Bellelli: the condition you describe is NOT characteristic of acute glomerulonephritis. In glomerulonephritis there is damage of the glomerulus and severely impaired GFR. Thus the diuresis is severely reduced, and due to impaired filtration proteins appear in the urine. The condition you describe corresponds to the initial stage of chronic kidney failure, usually due to atherosclerosis, diabetes, hypertension or other type of damage of the kidney tissue. In this case GFR is impaired, albeit to a lesser extent than in glomerulonephritis, and the excretion of urea is reduced. This leads to increased BUN. However the increased concentration of urea reduces the ability of the tubuli to reabsorb water, because of osmotic reasons, yielding compensatory polyuria. The patient has reduced GFR but normal or increased diuresis (urine volume in 24 hours). To some extent this effect is beneficial, as it favors the elimination of urea; however it cannot completely solve the problem and in any case the progression of the disease leads to kidney insufficiency. In its essence the point is that a moderately reduced GFR can be partially compensated by reduced tubular reabsorption; a severely reduced GFR cannot.       Home of this course Slides for this lecture