MEDICINE AND SURGERY "F" Course of LABORATORY MEDICINE Disturbances of the endocrine system HORMONES 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.       Hormones are produced by endocrine glands and serve to regulate the physiological functions of many organs and tissues. Their concentration in the blood is finely regulated. The blood concentration of some hormones may vary within a limited range, but that of other hormones may experience large changes, in response to external stimuli or because a circadian rhythm. As a consequence of their possible variability, the concentrations of hormones in blood should be interpreted with reference to other clinical and laboratory information. Dysregulation may occur both in the sense of excess secretion (possibly due due to benign, hormone-secreting tumours of the gland, or to overstimulation and hyperplasia of the gland) or in the sense of insufficient secretion (possibly due to damage of the gland due to metabolic defects, malignancies, insufficient stimulation, insufficient blood supply, or infections). Moreover, alterations of the circadian rhythms or the regulation of secretion occur (often in association with hypersecretion), cause symptoms and have diagnostic value. Audio: Hormones LABORATORY METHODS FOR HORMONES: DIRECT DETERMINATION OF HORMONE CONCENTRATION       The concentration of hormones in our blood is usually extremely low and, for some hormones, quite variable during the day or in response to external stimuli. To overcome these problems, hormone concentrations may be tested at time intervals over the whole day, or in the 24 hours urine or under stimulation.The most important method for measuring hormone concentration is the radioimmunoassay. There are several variants of this method, which relies on radioactively labelled probes (the antibody or its target). In the case of hormones, the method developed by Rosalyn Yalow (a Nobel laureate), is based on competition with radioactively labelled hormone. The measurement is carried out as follows: the (unlabelled) antibody against the hormone is immobilized on a substrate and is challenged with a mixture of the patient's serum and a solution of the radioactively labelled hormone at the appropriate concentration. Then the immobilized antibody, with the bound hormones is washed and radioactivity is measured. The bound radioactively labelled hormone reflects the ratio between labelled and unlabelled hormone in the mixture: the higher the hormone concentration in the patient's serum, the lower the radioactively labelled hormone bound to the immobilized antibody. This type of test requires that the total hormone (labelled and unlabelled) is in excess with respect to the antibody binding sites.       The basic principle of the immunoassay allows for some variations: e.g. one can use radiolabelled antibodies (rather than radiolabelled antigens; RIA radiolabelled immuno-assay)), or antibodies labelled with enzymes whose reaction produces an absorbance or fluorescence signal (ELISA: enzyme linked immuno-assay) to detect either antibodies or antigens (hormones in the present case)in the patient serum. In this type of test the immobilized antibody (or antigen) should be in excess with respet to the antigen (or antibody) in the liquid phase. SUPPRESSION AND STIMULATION TESTS       The concentration of hormones is regulated by positive and negative feedback; this allows us to carry out stimulation or suppression tests.       When insufficient hormone production is suspected, stimulation tests may provide more relevant information than simple measurements of the serum hormone concentration. A stimulation test is carried out by administering to the patient a stimulus to secretion of a specific hormone and measuring its concentration increase (e.g. ACTH stimulation test for adrenal insufficiency), or a physiological response that parallels it (e.g. the glucose tolerance test). This type of test detects the level of the hormone regulation cascade that is affected. Consider, as an example, the case of the ACTH stimulation test: if administration of ACTH causes increased prodution of cortisol, the adrenal cortex is functioning correctly but is understimulated by the hypohysis; if administration of ACTH fails to increase the level of cortisol the adrenal cortex is affected and unresponsive.       When excess hormone production is suspected, suppression tests may be used. Suppression tests are carried out by measuring the decrease of hormone concentration (or a physiological response parallel to it) induced by stimuli that provide a negative feedback to secretion (e.g. the dexamethasone suppression test for Cushing's disease, see below). Suppression tests may provide crucial information for the differential diagnosis of hypersecreting conditions. The clinical reasoning is analogous to that in suppression test, but the stimulus is of the opposite type. BIOLOGICAL AND FUNCTIONAL TESTS       Biological and functional tests measure the hormone effect, rather than its actual concentration. Examples are the measurement of glycemia for insulin or that of basal metabolism for thyroid hormones. The interpretation of biological and funcional tests is often not unequivocal because the same biological effect may be controlled by several hormones, thus this type of tests is usually associated to the direct determination of hormone concentration or to suppression/stimulation tests. Audio: Clinical laboratory tests for endocrine diseases tests for endocrine diseases test clinical indication examples direct measurement of hormone concentration in the blood when the hormone concentration is not subject to irregular variations during the day T3 and T4 for thyroid cortisol for the adrenal cortex etc. indirect estimate (measurement of the hormone effects) when the effect is clearly attributed to a single hormone basal metabolism for thyroid glycemia for insulin calcemia for parathyroid hormone etc. stimulation test, coupled to direct or indirect measurement in many cases of insufficient production of the hormone (endocrine insufficiency); establishes the step of the hormone regulation cascade that is affected glucose tolerance test for insulin TRH for thyroid hormones etc. suppression test, coupled to direct or indirect measurement in many cases of excess production of the hormone; establishes the step of the hormone regulation cascade that is affected T3 suppression test for the thyroid - hypophysis - hypothalamus axis cortisol suppression test for the adrenal cortex etc. imaging methods applied to the gland, possibly coupled to biopsy when an anatomical lesion is suspected (e.g. thyroid adenomas) thyroid scintigraphy NMR imaging of the relevant body segment etc.       An important comment to the above Table is that laboratory tests measure the organ's function, whereas imaging methods detect anatomical lesions. Although ultimately function damage must depend on a lesion, the two techniques complement each other and, depending on the specific disease, either may provide optimal sensitivity. E,g, a cleary detectable anatomical lesion may affect only a portion of the organ and may not cause a detectable impairment of the function. Conversely a microscopical or submicroscopical but diffuse lesion may be undetectable by imaging methods, yet it may cause significant functional impairment, reflected by obvious laboratory findings. Thus one should not consider laboratory and imaging evidences as alternative but as complementary.       From a biochemical standpoint, hormones belong to several different families of compounds, and every endocrine gland may secrete one or more hormones, usually belonging to the same biochemical family. An exception is the thyroid which secretes two hormones belonging to different biochemical families: thyroxin is an aminoacid derivative, whereas thyrocalcitonin is a small protein. The adrenals do secrete hormones of two very different chemical types (catecholamines are aminocid derivatives, corticoids are cholesterol derivatives), but from an embriological viewpoint they are two different glands one of which surrounds the other. As a consequence of this variety several analytical methods are required, as summarized in the Table below: HORMONES AND THE CLINICAL BIOCHEMISTRY LABORATORY Endocrine gland Hormone Chemical nature Measurement method Serum conc. Epiphysis (pineal gland) Melatonin Tyrosine derivative Extraction in dichloromethane followed by HPL Chromatography     Neurohypophysis (posterior pituitary) Vasopressin (Anti Diuretic Hormone, ADH) Polypeptide (extraction followed by immunoassay) <12 pg/mL Oxytocin Polypeptide Immunoassay; mass spectrometry     Adenohypophysis (anterior pituitary) Growth Hormone (GH, somatotropin) Protein Immunoassay (biological test) <7 ng/mL Prolactin (PRL) Protein Immunoassay <20 ng/mL (women) <15 ng/mL (men) Gonadotropins (LH and FSH) Glycoproteins Immunoassay (biological test)   Thryoid stimulating hormone (TSH) Glycoprotein Immunoassay 0.5-5 mU/L Adrenal Cortex stimulating hormone (ACTH) Protein Immunoassay     Thyroid Thyroxine (tetraiodothyronine) and triiodothyronine (T4 and T3) Tyrosine derivative RadioImmunoAssay T4=4-12 μg/dL; T3=80-100 ng/dL Thyrocalcitonin Protein Immunoassay     Pancreas Insulin Protein Immunoassay   Glucagon Protein (Immunoassay)     Adrenal cortex Glucocorticoid hormones (e.g. cortisol) Cholesterol derivatives RadioImmunoAssay, usually after extraction with organic solvents 3-24 μg/dL Mineralocorticoid hormones (e.g. aldosterone) Cholesterol derivatives RadioImmunoAssay, usually after extraction with organic solvents; chemiluminescence 1-5 ng/dL Sexual hormones (e.g. estrogens, progesterone, testosterone) Cholesterol derivatives RadioImmunoAssay, usually after extraction with organic solvents     Adrenal medulla catecholamine hormones (e.g. adrenaline) Tyrosine derivatives HPL Chromatography     Measurements methods in parentheses are possible, but not commonly used. Disorders of the anterior pituitary       The anterior pituitary gland produces protein hormones that control several body functions, and notably several other endocrine glands. Hormone secretion is stimulated by releasing factors produced by the hypothalamus and secreted in the special vascular structure of the hypothalamus-hypophysis portal system. The hypothalamus responds to peripheral hormones by reducing the production of the appropriate releasing factor, with a negative feedback mechanism. Each hormone is produced by a specific cell type and no cell produces two hormones. As a consequence of this anatom-functional organization, hyperfunctionality of the hypophysis is usually selective (e.g. because of a secreting adenoma of one cell type), whereas hypofunctionality is often generalized (panhypopituitarism due to ischemic or compresssive destruction of the gland with all its cell types), even though some cell types may be affected to a greater extent than others, due to their different location in the gland. An exception to this rule occurs in the rare case of genetically determined selective hypofunctionality (e.g. isolated GH decrease). An interesting and not uncommon condition is observed when the patient has a pituitary adenoma that produces one hormone and suppresses all other because of compressive lesion of the rest of the gland: in this case a selective hyperfunctionality is associated to a generalized hypofunctionality. In all cases of laboratory-demonstrated hyperpituitarism or hypopituitarism, an evaluation of the hypophysis by imaging methods is recommended, and the possibility of ectopic adenomas that produce pituitary hormones but do not reside in the hypophysis should be considered if the imaging analysis yields a negative result.       Growth hormone (GH, somatotropin) is released in response to hypothalamus' GRH; its secretion is inhibited by somatostatin. At the periphearl level, GH acts by promoting the release of secondary hormones: insulin and somatomedins that stimulate the growth of tissues (notably of bone). Paradoxically, GH is an antagonist of insulin-induced glucose uptake. Release of GRH by the hypothalamus is stimulated by insulin, by hypoglycemia and by some aminoacids (e.g. arginine). Thus GH stimulation tests may be carried out using insulin and glucose, arginine or synthetic GRH. Basal measurements of GH are poorly informative because GH may be below detectable levels for much of the day. Selective hypersecretion of GH may be due to adenomas of the pituitary; more rarely to GRH-secreting adenomas that may develop in some endocrine glands (notably in the Langerhans islest of the pancreas). It causes gigantism in the infant and acromegaly in the adult, and is frequently associated to insulin resistance and subclinical type 2 diabetes mellitus, and to hypercalciuria. Diagnosis of acromegaly is confirmed by the suppression test with glucose: in normal subjects serum GH concentration, if detectable drops to < 5 ng/mL after oral administration of 100g glucose; in acromegalic patients no such drops is observed and GH is often > 10 ng/mL even after glucose administration. Insufficient production of GH or insufficient peripheral response to it causes armonic dwarfism (see Table). Differential diagnosis of dwarfism hypophysary dwarfism (insufficient production of GRH or GH) Normal proportions of the body parts; decreased serum concentration of GH; growth rate < 4 cm/year Laron dwarfism (defective GH receptors; peripheral hypophysary dwarfism) like hypophysary dwarfism Congenital hypothyroidism Non proportionate dwarfism, short extremities with almost normal body and head Achondroplasia and similar osteochondrodysplasias (inherited, usually autosomal dominant) Non proportionate dwarfism, short extremities with almost normal body and head       Thyroid Stimulating Hormone (Thyrotropin, TSH) is a heterodimeric glycoprotein released as a response to hypothalamus' tripeptide TRH, whose secretion is inhibited by thyroid hormones. Adenomas of the TSH-producing cells in the pituitary may cause secondary hyperthyroidism. The suppression test is carried out by measurement of TSH in the serum after oral administration of the thyroid hormone triiodothyronine (see below).       Adrenocorticotrophic Hormone (ACTH) is a 39 polypeptide dervied from cleavage of a higher molecular weight protein precursor. It stimulates the secretion of steroid hormones by the adreanl cortex, most notably cortisol. Secretion of ACTH is stimulated by the hypothalamic protein CRH, whose secretion is inhibited by cortisol and synthetic steroid hormones. Hypersecretion of ACTH, usually due to benign adenomas, causes Cushing's syndrome (see below). Suppression test is carried out by administration of the synthetic steroid dexamethasone.       Gonadotropins (LH and FSH) are protein hormones that control the production of gametes and most other sexual and reproductive functions. They act especially (but not only) on the gonads and regulate the production of (steroid) sex hormones. Excess production of gonadotropins (e.g. because of a pituitary adenoma) deranges the production of gametes, and may induce sterility. It is important to remember that gonadotropins are produced also by the placenta, and that may be ectopically produced by adenomas of other glands or tissues, in the presence of a perfectly normal (i.e. adenoma-free) pituitary.       Prolactin is a protein hormone that stimulates the development of the breast in women and promotes lactation. Release of prolactin is inhibited by dopamine, which provides a convenient suppression test. Hyperprolactinemia may cause galactorrhea and infertility (both in men and women); it may be due to a prolactin producing adenoma, and may be associated to some breast tumors. Normal prolactin levels are < 20 ng/ml in non pregnant women and < 15 ng/mL in men. Serum levels > 300 ng/mL are usually due to adenomas; serum levels > 100 ng/mL in non-pregnant women (or in men) are suspect and warrant further investigation, unless they are explained by therapy with dopamine antagonists.       Panhypopituitarism hypofunctionality of the adenohypohysis can be selective or generalized (panhypopituitarism). Generalized hypoituitarism is more frequent and is the result of compressive or ischemic destruction of the hypophysis, often due to tumours; a less common condition is the Sheehan syndrome (ischemic necrosis of the hypophysis due to post partum hemorrages and shock). Gonadotropins are lost first, followed by GH, and then by TSH and ACTH. Death may ensue, due to the lack of thyroid and adrenal stimulation. Diagnosis is by imaging methods (lesions of the sella turcica) and by laboratory determination of the hormones' concentration in the serum. Infertility and a syndrome indistinguishable from Addison's disease characterize the clinical picture. An important differential diagnosis is with anorexia nervosa. Diagnosis is confirmed by the measurement of the concentration of pituitary hormones, and of the hormones they stimulate (T3, T4, cortisol, etc.) that result low.       Selective hypopituitarism, with isolated pituitary deficiencies, is suspected because of failure to grow (pituitary nanism due to deficiency of GH), or sterility and lack of menstruations (hypogonadotropic hypogonadism,may be due to genetic defects, e.g. Kallmann's disease). Isolated, pituitary induced hypothyrodism or adrenal insufficiency (pituitary Addison's disease) are uncommon, these conditions usually occurring (and some times dominating) a panhypopituitary condition.       The pars intermedia of the hypophysis is active in the fetus but very small or absent in the human adult (it remains functional in lower vertebrates, and may be responsible for skin colour changes linked to camouflage). In the fetus it produces the Melanocyte Stimulating Hormone (melanotropin, MSH), derived fromthe cleavage of the precursor protein proopiomelanocortin. It is not responsible of diseases in humans. Audio: Hormones of the anterior pituitary Disorders of the posterior pituitary       The posterior part of the hypophysis is of neurological origin and is directly connected to the hypothalamus; indeed the secreting cells are neurons whose body resides in the hypothalamus, and whose axon extends in the pposterior lobe of the hypophysis. These cells release two polypeptide hormones, the antidiuretic hormone (ADH) and oxitocin.      Vasopressin (antidiuretic hormone, ADH) is a nonapeptide containing two Cys residues involved in a disulfide bond. It promotes water reabsoprtion in the distal tubuli of the nephrons and contributes to the control of the osmolarity of biological fluids (by which its secretion is regulated). Its serum concentration can be measured by radioimmunoassay.       Diabetes insipidus is due to insufficient or absent production of ADH. Since this hormone causes the kidney to reabsorb water, its deficiency is associated with poliuria and diluted urine (to be distinguished from diabetic mellituria, in which the urine flow is increased because of the osmotic effect of glucose, and from nephrogenic diabetes insipidus, in which the kidney's function is reduced). The causes of diabetes insipidus are variable; often this condition is caused by neoplastic lesions or aneurisms compressing the hypothalamus. Diabetes insipidus is suspected in the presence of dilute urine and serum hyperosmolarity, and confirmed by the laboratory finding of severely reduced serum ADH. This condition is potentially lethal.      Oxitocin is a nonapeptide hormone differing from ADH because of two aminacid residues. Its main function is to promote the contractility of the uterus during parturition and insufficient production of oxitocin may cause a dynamic dystocia (the expulsion of the fetus is slowed down or blocked because of insufficient contractions and independently of a mechanical incompatibility of the pelvis). Audio: Vasopressin and oxitocin       Clinical examples:       1) 40 year old patient admitted to the intensive care unit with hemorrhagic shock due to postpartum hemorrhage. After recovery from the initial acute phase lactational failue and hypglycemic crises. Laboratory findings: Fasting glycemia = 40 mg/dL T4 = 2.8 μg/dL (normal range 4-12 μg/dL) T3 = 48 ng/dL (normal range 80-100 ng/dL) Serum cortisol (morning) 4 μg/dL (reference range, 6-23 μg/dL) TSH = 0.05 mU/L (normal range 0.5-5 mU/L) FSH = 0.2 mU/L LH = 0.1 mU/L ACTH 12 pg/mL (reference range, < 46 pg/mL) prolactin = undetectable       Comment: panhypopituitarism due to post-partum hemorrage and shock (Sheehan syndrome). Patient was treated with cortisol and T4 (for life). Melatonin and the epiphysis       The epiphysis is the second intracranial gland. It produces one hormone, melatonin, which regulates the sleep-wake cycle. Secretion is regulated by light, hence it may be disregulated by artificial illumination or by long plane voyages. Melatonin is a Tyr derivative. No true human diseases are associated with disfunctions of the epyphis but they may cause insomnia or hypersomnia. Disorders of the thyroid       The thyroid gland produces two Tyrosine derived hormones, called tetraiodothyronine (T4, Thyroxine) and triiodothyronine (T3). T4 and T3 regulate the metabolism of most tissues in our body. In addition, thyroid's parafollicular cells produce a completely different protein hormone called calcitonin or thyrocalcitonin that is important for the regulation of calcium metabolism and calcium concentration in the blood. These two types of hormones should be considered separately: they are produced by different cells in the gland, and under different stimuli. The action of thyrocalcitonin will be discussed further on, in conjunction with that of PTH, the hormone produced by the parathyroid glands.       Biochemistry and physiology of T4 and T3       The thyroxine-like hormones are iodinated derivatives of the aminoacid tyrosine. They are synthesized by chemical modification of Tyr residues in the sequence of the storage protein thyroglobulin (represented as P in the figure below); when necessary the protein is digested and the hormones are released in the bloodstream. Audio: Thyroid hormones       The hormones released in the blood are transported by a specific protein carrier called TBG (thyroxine-binding globulin), and by other proteins (e.g. albumin); only a tiny minority of the total T4 and T3 is freely dissolved in the plasma. Secretion of T4 and T3 is stimulated by the pituitary Thyroid-Stimulating Hormone (TSH), whose release is controlled by the hypothalamus' TRH. TRH secretion is suppressed by a negative feedback mechanism by T4 and T3. Some T4 may be deiodinated in the liver and converted to T3; this fraction of T3 is called the reverse T3 (rT3).       The action of T4 and T3 is to stimulate the tissue metabolism and O2 consumption, and is mediated by protein synthesis.       Hypothyroidism is the condition in which the production of thyroid hormones is reduced. It may be due to lack of iodine in the diet, to inflammatory diseases of the gland, or to inherited, congenital defects of the biosynthesis or secretion of T4 and T3. Congenital hypothyroidism may cause mental retardation (cretinism) and disarmonic dwarfism; these cases deserve special attention because the newborn is normal at birth (the hormones having been obtained from maternal blood during pregnancy), thus prompt treatment may prevent the development of symptoms. The thyroid is usually enlarged (goiter) beacuse of overstimulation by the pituitary, and the concentration of T4 and T3 in the blood is decreased. The clinical picture is ususally quite characteristic and diagnosis is confirmed by laboratory findings. If iodine administration does not cure the syndrome a substitutive therapy must be started.       Hyperthyroidism (Basedow's disease; Graves' disease; thyrotoxicosis) may be due to diffuse enlargement of the gland (usually due in turn to the anomoalous overstimulation of the gland by autoantibodies or because of a benign adenoma of the pituitary, which secretes TSH) or to a T3/T4 secreting adenoma. 131I scintigraphy is diagnostic, and the laboratory analyses will confirm increased serum concentration of T4 and T3 (and possibly of TSH). Surgery is indicated.       Euthyroid sick syndrome is a condition in which hypothyroidism-like symptoms are present in spite of normal thyroid tests. It is observed in several systemic diseases (e.g. liver cirrhosis, malnutrition, sepsis), and may be due to alterations in the peripheral metabolism of T4. TSH is normal.       Clinical laboratory analyses of thyroid function       T4 and T3 are the most important iodine-containing compounds in our body and thyroid the most important (and almost only) organ to utilize iodine; this allows the clinical laboratory several easy tests of thyroid function. The radioactive iodine uptake can be measured, and can be coupled to thyroid scintigraphy in order to reveal disomogenous captation by the gland.       The concentration of T4 and T3 in the serum can be measured by radio-immuno assay; the normal ranges are: T4 = 4-12 μg/dL;       T3 = 80-100 ng/dL;       rT3 = 10-40 ng/dL Audio: Analytical methods for thyroid dysfunctions       The concentration of TSH can also be measured by radio-immuno assay; its normal concentration in the serum is < 5 μ U/mL. Comparison of the concentrations of TSH and T4 / T3 is very important to establish whether a case of hyperthyroidism or hypothyroidism due to a disease of the hypophysis or the thyroid, as summarized in the table below:   disease T4 / T3 TSH stimulation test suppression test   hyperthyroidism due to pituitary hyperstimulation increased increased not indicated with T3: usually no effect   hyperthyrodism due to thyroid disfunction increased strongly decreased not indicated with T3: usually no effect   hypothyroidism due to pituitary insufficiency decreased decreased with TSH:strong increase of T3 and T4 with TRH: usually no response with T3: usually no effect   hypothyrodism due to thyroid disfunction or iodine deficiency decreased strongly increased with TSH or TRH: usually no response with T3: strong decrease of TSH       Stimulation and suppression tests to assess thyroid function and the hypothalamus-hypophysis-thyroid axis are available. The TRH stimulation test is useful for the differential diagnosis of hypothyroidism. It is carried out by the intravenous administration of TRH, and measurement of the TSH and T4/T3 responses. In healthy subjects TRH causes an increased release of TSH by the hypophysis (provided that the gland is functional) followed by an increased release of T4/T3 (provided that the thyroid is functional). Absence of a TSH and T4/T3 response is indicative of pituitary disfunction; presence of a TSH response but absence of a T4/T3 response suggests thyroid disfunction.       The thyroid suppression test is useful in cases of hyperthyroidism. It is carried out by oral administration of L-triiodothyronine (100 μg a day for ten days). In healthy subjects this treatment causes a strong reduction of iodine uptake, and a strong decrease of the concentrations of T4 and TSH in the serum. A decrease in TSH, but not in iodine uptake and T4 suggest primary hyperthyroidism; no decrease of TSH, iodine uptake, and T4 suggests that hyperthyroidism is secondary to hyperfunctionality of the pituitary, e.g. because of an adenoma. Audio: Thyroid suppression and stimulation tests       An important additional test is the measurement of basal metabolism. This is carried out by measuring the oxygen consumption (indirect calorimetry). The reference values for this test are as follows: inspired air: 5 L/min inspired O2: 5x0.2=1 L/min espired air: 5 L/min espired O2: 5x0.15=0.75 L/min O2 consumption: 1 - 0.75 = 0,25 L/min (= approx. 10 mMoles) energy equivalent in animal metabolism: 117 kcal/mol O2 basal metabolism: 0.01 Moles/min x 60 x 24 x 117 = 1680 kcal /day According to the above table, the normal reference values for adults are in the order of 1700 kcal/day (for more precise estimates corrected for age, sex and body weigth use appropriate tables) that can be measured from the O2 consumption, and the energy equivalent of O2 (117 kcal /mol). Hyperthyroidism causes a significant increase of basal metabolism / O2 consumption, hypothyrodism a significant decrease.       Clinical examples       1) A patient referring weight loss, restlessness, tachycardia, bulging eyes, anxiety, with gradual onset since approximately 6 months. Laboratory tests: normal hematochemical and hemocytometric parameters; Free T4 = 14,5 μg / dL (normal range: 4 - 12 μg / dL) TSH = 0.03 μU / mL (normal range 0.5 - 5 μU / mL) Comment: increased T4 with decreased TSH points to a primitive thyroid hypersecretion; TSH is reduced because of feedback inhibition. Investigate diffuse hypersecretion (Basedow disease) or, possibly, hypersecreting nodule (benign thyroid tumor).       2) A 47-year-old woman presents to her primary care provider with complaints of fatigue, weight gain, and constipation. Gradual onset since two years. Laboratory findings: Sodium 130 mEq/L Potassium 4.4 mEq/L Chloride 100 mEq/L BUN 12 mg/dL hemoglobin 12.5 g/dL Endocrine cause is suspected, thus a screening of thyroid and adrenals is prescribed: T4 = 3.1 μg/dL (normal range 4-12 μg/dL) T3 =57 ng/dL (normal range 80-100 ng/dL) TSH = 8 mU/L (normal range 0.5-5 mU/L) Serum cortisol (evening) 4.5 μg/dL (reference range, 1.7-16.6 μg/dL) Serum cortisol (morning) 12 μg/dL (reference range, 6-23 μg/dL). ACTH 32 pg/mL (reference range,< 46 pg/mL) Comment: Thyroid hormones are low, TSH is high. Cortisol is normal as is ACTH. Hypothyroidism due to a lesion of the thyroid. Prescribe measurement of serum iodine, thyroid scintigraphy, ecography of the thyroid. Possible diagnostic hypotheses are thyroid cancer, thyroiditis (possibly autoimmune), ischemic damage of the thyroid. Disorders of the parathyroids (and of thyrocalcitonin)       The parathyroid hormone, thyroid calcitonin, and vitamin D are involved in the homeostasis of calcium and control the levels of this ion in the human serum. This subject was hinted to in the lecture on electrolytes, but is dealth with here in greater detail. The parathyroid glands are four in number and adhere to the thyroid. They are quite small and may be accidentally removed in the course of thyroidectomies. Except for this event, diseases of the parathyroids are uncommon.       The parathyroid hormone (PTH) is a small 84-residues protein. It has direct effects, and effects mediated by vitamin D, which it activates. It causes calcium to be mobilized from its reservoir in the bone matrix (as calcium phosphate), and to be released in the blood plasma, inducing hypercalcemia. The normal calcium concentration is 2-2.5 mM (4-5 mEq/L; 8-10 mg/dL). Since calcium is constantly lost in the urine, and constantly absorbed in the gut (in a process that requires vitamin D), PTH is required to maintain its concentration constant. Thyrocalcitonin plays the opposite role of promoting calcium deposition in the bone matrix and lowers its serum concentration. Secretion of both PTH and thyrocalcitonin responds to the serum calcium levels; however, the effectiveness of their action requires that dietary apport of the ion and of vitamin D is sufficient. The serum concentration of all the molecules and ions involved in this system can be measured in the clinical laboratory (calcium by potentiometry or by atomic spectroscopy; vitamin D, PTH and tyrocalcitonin by radioimmunoassay).       Calcium is essential for muscular contraction, and spastic muscular palsies may occur if its concentration falls below the normal range. Paralysis of respiratry muscles may cause death. Hypoparathyroidism causes hypocalcemia, with possible tetany (sustained, painful, involuntary muscular contractions), and, possibly, neurological symptoms (pseudodementia). Pseudohypoparathyroidism (PHP) has similar symptoms but is due to unresponsiveness of the peripheral organs (kidney, bone) to PTH, whose concentration in the serum is normal or increased. Differential diagnosis is confirmed by analysis of the urine under stimulation of PTH (that in hypoparathyroidism causes phosphaturia and excretion of cAMP, whereas in PHP has no effect). We remark that the relationship between true hypoparathyroidis and PHP is reminiscent of that between type 1 and type 2 diabetes mellitus. Vitamin D deficiency is another possible cause of hypocalcemia. Since vitamin D is required for the intestinal absorption of calcium, this condition is associated to insufficient availability of the ion and insufficient deposition in the bone matrix, an effect which is absent in hypoparathyroidism and PHP. In childrem vitamin D deficiency causes rickets, in the adult osteomalacia.       Hyperparathyroidism causes hypercalcemia, excess bone resorption (osteoporosis) and possibly nephrolythiasis. Osteoporosis is frequent in the elderly and may be counteracted by administration of calcitonin and vitamin D. There are several possible causes of hyperparathyroidism, including malignancies of the parathyroids; some malignancies (e.g. hematologic) may produce substances resembling PTH and causing bone resorption, in a paraneoplastic syndrome.       Clinical laboratory analyses of parathyroid function and calcium homeostasis.       The main laboratory datum for parathyroid function is calcium concentration in the serum, measured together with other electrolytes. Normal calcium concentration in the serum is 4-5 mEq/L.       PTH can be measured by radioimmunoassay; its normal concentration range in the serum of healthy adults is 15-60 pg/mL. Calcitonin can also be measured by radioimmunoassay; its normal concentration is ≤19 pg/mL in healthy adult males; significantly less in females (< 10 pg/mL).       Vitamin D can also be measured by radioimmunoassay; normal values are in the range 20-50 ng/mL.       PTH and calcitonin secretion is regulated by calcium concentration in the serum; thus there are no hypophysis hormones to be tested in this case, nor easy to carry out stimulation or suppression tests (caution: never attempt to alter electrolyte concentrations for diagnostic purposes!). Endocrine disorders of the pancreas       Insufficiency of the endocrine functions of the pancreas causes type 1 diabetes mellitus; hyperfunction, most often due to a benign insulin-secreting adenoma causes hypoglycemic crises. Both diseases are potentially lethal if undiagnosed and untreated. These diseases have been discussed in the lecture on glycemia, and will not be discussed again here; however it is important to underline that they conceptually belong to the class of endocrine disfunctions. Disorders of the adrenal cortex       The adrenal cortex produces steroid hormones, derived from cholesterol. They are classified, according to their main action as glycocorticoid hormones (e.g. cortisol), which regulate the cell metabolism; mineralcorticoid hormones (e.g. aldosterone), which regulate the kidney function; and sexual hormones (e.g. testosterone, estrogens, progesterone).       Adrenal insufficiency (Addison's disease). The symptoms of Addison's disease are dominated by the reduced production of glucocorticoid hormones (hypoglycemia, fatigue, reduced basal metabolism, low arterial pressure); hyperkalemia is present because of the reduced production of aldosterone. The disease is fatal if untreated. The major causes of the destruction of adrenal cortex are: tuberculosis, cancer, amiloidosis and inflammatory necrosis. A clinically similar condition may be due to panhypopituitarism and by the (uncommon) selective reduction of hypophisary secretion of ACTH. Addison's disease due to the destruction of the adrenal cortex is characterized by increased serum ACTH; that due to panhypopitutarism by decreased or absent serum ACTH. Due to the numerous physiological effects of cortisol, several laboratory parameters are altered in Addison's diseas: hypoglycemia is common, as is a brownish discoloration of the skin. Serum sodium may be decreased and potassium increased.       Glucocorticoid hypersecretion (Cushing's disease) may be due to a functioning adenoma of the hypophysis (that secretes ACTH) or of the adrenal cortex (that secretes cortisol). Less frequently, tumors of other tissues (e.g. the lung) can produce ACTH. In typical cases of Cushing's disease, secondary to an ACTH-secreting adenoma of the hypophysis, the secretion of both glucocorticoid and mineralocorticoid hormones is increased. Hyperglycemia is common.       Clinical laboratory methods for steroid hormones and ACTH.       Cortisol and cortisone can be measured by a competition radioimmunoassay using 125I-labelled cortisol or cortisone. Cortisol secretion follows a circadian rhythm being higher in the morning (serum concentration 9-24 μg/dL) and lower in the afternoon (3-12 μg/dL); thus the measured concentration should be referred to the time of sampling. Alternatively the concentration can be measured in the urines collected over the 24 hours (cortisol: 20-100 μg/24 hours; cortisol metabolytes - 17 hydroxycorticoid derivatives: 2-10 mg/24 hours).       An important diagnostic test is the dexamethasone suppression test. 0.5 mg of dexamethasone every 6 hours should strongly depress the production of ACTH and hence that of cortisol. Adenomas of the adrenal usually do not respond to dexamethasone suppression.       The measurement of glucocorticoid hormones should always be associated to that of ACTH (also by radioimmunoassay) in order to decide whether the primary lesion affects the hypophysis or the adrenal. Attention should be paid to the following consideration: Addison's disease may be the initial manifestation of panhypopituitarism; thus adrenal insufficiency warrants the investigation not only of ACTH, but also of the other hormones of anterior hypophysis (GH, TSH, gonadotropins). Moreover, ACTH stimulates (to a lesser extent) also the secretion of aldosterone, whereas gonadotropins stimulate the secretion of sexual steorid hormones; thus in the presence of Cushing's and Addison's disease, measuring aldosterone and sexual steroid hormones is mandatory. Audio: Dysfunctions of the adrenal cortex       Hyperaldosteronism. The mineralcorticoid hormone aldosterone regulates the kidney function and promotes Na+ retention and K+ loss; it has an antidiuretic effect. It is regulated mainly by the renin-angiotensin system, and to a lesser extent by ACTH. Aldosterone can be measured in the plasma (normal value 1-5 ng/dL) or in the urine (2-10 μg/24 hours).       Sex hormones are produced primarily by the gonads, but also by the adrenal cortex. As a general rule, their secretion is estimated from the metabolytes (17 ketosteroid derivatives) in the urine (normal values: male 7-25 mg/24 hours; female 4-15 mg/24 hours). > Laboratory findings in adrenal disorders   electrolytes glycemia hormones Adrenal hypofunction (Addison's disease) decreased Na+ increased K+ arterial hypotension decreased decreased cortisol; decreased aldosterone; ACTH stimulation test has no effect in primary Addison's disease, has effect in Addison's disease secondary to panhypopituitarism Adrenal hyperfunction (Cushing's syndrome)   increased increased cortisol; dexamethasone suppression test has no effect in primary Cushing's disease, but may have an effect in Cushing's disease secondary to pituitary adenomas producing ACTH Hyperaldosteronism (Conn's syndrome) increased Na+ decreased K+ arterial hypertension normal increased aldosterone       The adrenal cortex and the thyroid are the most important glands whose function is governed by the anterior pituitary; their dysfunction may cause severe, potentially lethal disease, and may occur isolated or in combination. Thus it is useful to compare, in the Table below, the dysfunctions of these glands, which may occcur in various combinations. Comparison of primary and secondary dysfunctions of the thyroid and the adrenals Disease TSH ACTH T4 and T3 cortisol stimulation test suppression test Hypofunctionalities Panhypopituitarism ↓ ↓ ↓ ↓ increase of T4/T3 or cortisol not indicated Primary hypothyroidism ↑   - ↓   - no effect not indicated Primary Addison's disease   - ↑   - ↓ no effect not indicated   Hyperfunctionalities TSH-producing hypophysary adenoma ↑   - ↑   - not indicated moderate effect ACTH-producing hypophysary adenoma   - ↑   - ↑ not indicated moderate effect Primary hyperthyroidism ↓   - ↑   - not indicated no effect Primary hyperfunction of the adrenal cortex   - ↓   - ↑ not indicated no effect       Clinical examples:       1) Patient refers fatigue, weakness, episodes of dizziness. Presents hyperpigmentation of gradual onset since 1 year. Laboratory tests: Fasting glycemia = 50 mg / dL Serum electrolytes: Na = 125 mEq / L; K = 5.5 mEq / L; Na/K = 22.7 BUN = 30 mg /dL Comment: the laboratory results are typical of Addison's disease; measure cortisol in the 24 h urine; measure ACTH.       2) Patient refers amenorrhea since 14 months (from Medscape). Laboratory findings: Hemocytometric analysis: White blood cell count: 8700 cells/μL, with 62% neutrophils (reference range, 4500-11,000 cells/μL, with 40%-70% neutrophils); Hemoglobin: 13.8 g/dL (reference range, 12.1-15.1 g/dL); Platelet count: 310,000 cells/µL (reference range, 150,000-450,000 cells/µL). Electrolytes: Sodium: 136 mmol/L (reference range, 134-145 mmol/L); Potassium: 3.3 mmol/L (reference range, 3.5-5.1 mmol/L); Chloride: 101 mmol/L (reference range, 96-106 mmol/L). Hematochemical parameters: BUN: 10 mg/dL (reference range, 7-20 mg/dL); Creatinine: 0.7 mg/dL (reference range, 0.6-1.1 mg/dL); Alkaline phosphatase: 42 U/L (reference range, 40-150 U/L); ALT: 15 U/L (reference range, 19-25 U/L); AST: 22 U/L (reference range, 9-32 U/L); Total bilirubin: 0.7 mg/dL (reference range, 0.1-1.2 mg/dL) Endocrine analysis Anti-müllerian hormone: 0.064 ng/mL (reference range, 1.0-3.0 ng/mL) Day 3 follicle-stimulating hormone (FSH): 8.4 mIU/mL (reference range, 1.37-9.9 mIU/mL [follicular phase]) Day 3 estradiol: 48 pg/mL (reference range, 25-75 pg/mL) Thyroid-stimulating hormone: 2.34 µIU/mL (reference range, 2-10 μU/mL) Prolactin: 11.69 ng/mL (reference range, 3-27 ng/mL [adult female]) Total testosterone: < 7 ng/dL (reference range, 15-40 ng/dL [women]) Dehydroepiandrosterone sulfate (DHEA-S): 2 µg/dL (reference range, 45-270 µg/dL [women aged 30-39 years]) Comment: the laboratory vaues are unexpected: the levels of female hormones are normal; those of male hormones are reduced (amenorrhea may be caused by an increase of male hormones in a woman). TSH is also normal (thyroid disfunction may also be a cause of amenorrhea). We remark a mild hypokalemia. Thus at present we have detected an abnormal condition, but we cannot explain the case. However, we have some suspect on a possible adrenal disfunction, because: (i) the adrenal cortex in a woman is the principal source of male hormones; and (ii) adrenal disfunctions may cause amenorrhea. Thus a second run of tests is prescribed, which yields the following results: Adrenocorticotropic hormone (ACTH): 3 pg/mL (reference range, 9-52 pg/mL) Cortisol (PM): 12 μ/dL (reference range, 3-13 μ/dL) Cortisol (AM): 12.1 μg/dL (reference range, 5-23 μ/dL) 24-hour free cortisol in the urine: 74 μg/24 h (reference range, 4-40 μ/24 h) Aldosterone: 8.1 ng/dL (reference range, < 10 ng/dL) Renin activity, plasma: 0.6 ng/mL/h (reference range, 0.6-4.3 ng/mL/h) Metanephrines, plasma: < 10 pg/mL (reference range, 12-60 pg/mL) These tests provide important information: ACTH is reduced and cortisol, though within normal ranges at the time of sampling, is increased as the average of the 24 hours (look at the urine test); thus we may suspect a Cushing's disease of adrenal origin, possibly a secreting adenoma (increased cortisol vs. decreased ACTH). Hypokalemia may be due to a moderate hyperaldosteronism, which parallels the increased cortisol levels. 3) 17 years old with dysmenorrhea and repeated infections (from Medscape). Laboratory findings: sodium 140 mEq/L potassium 5.6 mEq/L urea 60 mg/dL creatinine 1 mg/dL Serum cortisol (evening) 0.11 μg/dL (reference range, 1.7-16.6 μg/dL) Serum cortisol (morning) 4.2 μg/dL (reference range, 6-23 μg/dL). ACTH 75.8 pg/mL (reference range,< 46 pg/mL) TSH 3.5 mIU/L (reference range, 0.5-4.70 mIU/L) triiodothyronine (T3) 4.4 pmol/L (reference range, 3.5-6.5 pmol/L) thyroxine (T4) 18.7 pmol/L (reference range, 10-23 pmol/L) luteinizing hormone (LH) 1.17 mIU/mL (reference range, 0.5-9.0 mIU/mL) follicle-stimulating hormone (FSH) 4.72 mIU/mL (reference range, 3-10 mIU/mL) parathyroid hormone (PTH) level is 58 pg/mL (reference range, 16-87 pg/mL). Comment: alterations in the menstrual cycle always warrant a full endocrine examination. In the present case serum cortisol is decreased, whereas ACTH is increased. TSH and thyroid hormones are in the normal range as are LH and FSH. PTH is also normal. In addition we remark that hypernatremia and hypokalemia are present. These data suggest Addison's disease due to damage of the adrenals; the hypophysis is normal and the increase in ACTH levels is due to the low cortisol levels. A condition of polyglandular failure (probably inherited) may be suspected. Pheochromocytoma       The pheochromocytoma is a benign tumour of the adrenal medulla that secretes catecholamines and causes severe arterial hypertension, either continuous or occurring in acute episodes. The laboratory diagnosis of this conditions rests on the demonstration of catecholamine metabolytes in the urine (metanephrines and vanillyl-mandelic acid) usually by means of High Pressure Liquid Chromatography (HPLC). False negatives are not uncommon unless the urine sample is collected after an hypertensive crysis. Endocrine disorders of the gonads       The gonads produce the gametes and elaborate sexual steroid hormones. Their function is controlled by LH and FSH produced by the pituitary. The development of the testis and the ovary is dependent on the sex chromosomes, and is sensitive to sexual aneuploidies (e.g. Turner syndrome, 45 X0; Klinefelter syndrome, 47 XXY). Both sexes produce male and female steroids, but of course males produce more androgenic hormones and females more estrogens and progesterone.       Male hypogonadism may be primary, due to Leydig cells dysfunction, or secondary to insufficient stimulation by the hypophysis (hypogonadotropic hypogonadism e.g in adiposo-genital and adreno-genital syndromes like Babinsski-Froelich's, or in Kallmann's syndrome). Testosterone production is decreased and sterility (aspermia, azoospermia) is present in both cases. The stimulation test is carried out by adminstration of he hypothalamic factor GnRH.       Female hypogonadism with sterility and anovulation occurs in Turner syndrome, dysfunction of the hypophysis, anorexia nervosa, polycystic ovary, etc. The differential diagnosis is difficult because the causes can be numerous, and the endocrine dysfunction may be subtle (e.g. dysruption of the cyclic hormone changes).       Hyperproduction of sex hormones may cause two very different disturbances, depending on the offending hormone being that of the patint's sex or of the opposite sex. Excess production of testosterone in males is scarcely symptmatic, unless it is due to diseases that cause non-sexual problems (e.g. neoplastic lesions of the testicle). Excess production of estrogens in males (e.g. because of an adenoma of the adrenals or the testicle) causes femininization. In women, excess production of estrogens may derange the gametogenesis resulting in reduced fertility; excess production of androgens causes hirsutism.       Clinical examples:       1) Idiopathic hypogonadotropic hypogonadism. 34 years old male with sterility, impotence, absence of pubic hair, gynecomastia. Laboratory findings: luteinising hormone (LH) 0.8 mlU/mL (normal limit in males: 1.2–7.8) follicular stimulating hormone (FSH) 1.5 mlU/mL (normal limit in males: 1.5–15.4) ante meridiem (AM) testosterone 56 ng/dL (normal limit in males: 249–836) prolactin 5.2 ng/mL (normal limit in males 2–18 ng/mL) thyroid stimulating hormone (TSH) 1.03 ulU/mL (normal limit: 0.4–4.2) free thyroxine 1.13 ng/dL (normal limit: 0.89–1.76). Comment: the only observed endocrine defects are: low LH and low testosterone. Clearly this patient present a primitive idiopathic disfunction of the hypophysis, limited to only one hormone (FSH, TSH and prolactin are within normal limits; ACTH was not measured). The low testosterone level is a consequence of the hypophysary disfunction. The patient responded to replacement testosterone therapy, and became fertile.       2) 14-year-old boy with poorly developed secondary sexual characteristics, low stature, undiscended testes, hypoosmia. Laboratory findings: follicle-stimulating hormone (FSH) 0.16 mIU/mL luteinising hormone (LH) <0.10 mIU/mL testosterone 75 ng/dL Comment: in this case all sex-related hormones are low, and a general growth defect is observed. This may suggest a genetic disease. The association with hypoosmia is consistent with this hypothesis. The boy was subjected to chromosomal and genetic analaysis and mutation of the KAL gene (Xp22.3) on the X chromosome was found consistent with the diagnosis of Kallmann syndrome. Kal gene encodes anosmin-1, a neural cell adhesion molecule necessary for the migration of olfactory neurone axons and gonadotrophin-releasing hormone (GnRH) synthesising neurones toward their final location. Further readings Doubleday AR, and Sippel RS Hyperthyroidism Cherella CE and Wassner AJ Congenital hypothyroidism: insights into pathogenesis and treatment. Stieg MR, Renner U, Stalla GK, and Kopczak A Advances in understanding hypopituitarism. Mihailidis J, Dermesropian R, Taxel P, Luthra P, Grant-Kels JM Endocrine evaluation of hirsutism Questions and exercises: 1) Dwarfism can be: armonic in congenital pituitary insufficiency, disarmonic in congenital thyroid insufficiency disarmonic in congenital pituitary insufficiency, armonic in congenital thyroid insufficiency disarmonic in both congenital pituitary insufficiency, and congenital thyroid insufficiency 2) A patient presents reduced basal metabolism, reduced T4 and T3, and reduced TSH. A likely diagnosis is: Hypothyroidism due to a thyroid disease Hypothyroidism secondary to a disease of the pituitary Hyperthyroidism due to a thyroid disease 3) Insufficiency and hyperfunction of the endocrine functions of the pancreas cause, respectively type 2 diabetes mellitus and type 1 diabetes type 2 diabetes mellitus and hypoglycemic crises type 1 diabetes mellitus and hypoglycemic crises 4) Diabetes insipidus is due to reduced secretion of ADH by the neurohypophysis reduced secretion of insulin by the pancreas reduced sensitivity to insulin of the tissues your score: 10 Attendance not registered because of insufficient score - Retry! You can type in a comment or question below (max. length=160 chars.): 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: