Hypothyroidism Hypothyroidism Nephrotic syndrome Type 2 diabetes Insulin resistance syndromes/obesity Chronic liver disease Insulin resistance syndromes Chronic renal failure Cholestasis + biliary obstruction Cushing \200\231s syndrome Cholestasis + biliary obstruction Cushing \200\231s syndrome Pregnancy Chronic renal failure Peritoneal dialysis Pregnancy 115-Draznin-Chap115.indd 674 4/1/2011 6:47:24 PM CAUSES 675 Diet Excessive consumption of saturated fats and carbohydrate can contribute to this condit ion. Anorexia nervosa can also cause hypercholesterolemia, as a strict low-fat, low-calorie diet results in reduced cholesterol and bile acid turnover, thus increasing circulating le vels. Obesity b See Clinical features (p. 528). Diabetes mellitus b See Lipid abnormalities found in patients with diabetes (p. 629). Hypothyroidism This is estimated to occur in 4% of tho se with hyperlipidemia, and com- pensated (subclinical) hypothyroidism in a further 10%. It usually results in hypercholesterolemia with a TC of 349 \200\223776 mg/dL. It also worsen s gen- etic dyslipidemias through d synthesis of hepatic LDL receptors. Chronic renal disea se Increased creatinine clearance is accompanied by hypertriglyceridemia and a reduction in HDL cholesterol. Proteinuria in the nephrotic syndrome is associated with hypercholesterol emia. Peritoneal dialysis may increase gut glucose load and worsen hypertriglyceridemia. Re member \200\224cyclosporine increases LDL cholesterol. Liver disease Cholestatic liver disease results in hypercholesterolemia. Primary biliary ci rrhosis produces marked increases in LDL and may elevate TC to >462 mg/dL. Severe hepatocel lular damage may lower LDL cholesterol by d production of its component parts and the enzym es that metabolize it. Cushing \200\231s syndrome Glucocorticoids increase VLDL production and thus hypertriglycer idemia. The associated weight gain and glucose intolerance can make this effect more pronou nced. Lipodystrophies This is a very rare group of congenital or acquired disorders whose hall- m ark is regional, partial, or generalized fat loss associated with hyperlipi- demia, especia lly unusually raised triglycerides. They are also associated with severe insulin resistance . Glycogen storage diseases Elevated lipids (triglycerides, cholesterol and mixed hyperlipide mia) are common, but usually only in type I (Von Gierke disease), type III (Forbes disease) , and type IV (Hers disease). Gout Hypertriglyceridemia occurs in about one-third of those with gout. Alcohol excess play s a part but it is not the sole cause. 115-Draznin-Chap115.indd 675 4/1/2011 6:47:24 PM CHAPTER 115 Secondary hyperlipidemias 676 Drugs Medications commonly implicated are as follows: \200 B -blockers, especially the no ncardioselective ones \200 Thiazide diuretics \200 Exogenous estrogens \200 Anabolic steroids \200 Glucocorticoids \200 Isotretinoin \200 Protease inhibitors (said to cau se hyperlipidemia in 50% after 10 months of treatment) Excessive alcohol consumption There is a J-shaped relationship between alcohol consumption and CHD. The American Heart Association (AHA) recommends no more than 1 alcoholic drink per day for women, and 2 drinks per day for men. Pregnancy Cholesterol rises throughout pregnancy, mostly in the second trimester, with LDL- C levels peaking mid-third trimester. Triglyceride levels follow a similar pattern to that of LDL-C. Levels decrease by 6 weeks postpartum but may take up to 1 year to return to norm al. 115-Draznin-Chap115.indd 676 4/1/2011 6:47:24 PM PRIMARY AND SECONDARY PREVENTION 677 Management of dyslipidemia Background The emphasis will depend on the cause of the hyperlipidemia and is aimed at redu cing cardiovascular, peripheral vascular, and cerebrovascular risk. There are now recommend ed target values when treating patients. Primary and secondary prevention It is universally accepted that tackling hypercholesterole mia in secondary prevention works. We now have solid evidence to support this, especially i n the context of CHD, where a 10% fall in TC is estimated to result in a 25% decrease in CH D risk. Fatty lesion regression has also been clearly demonstrated. The gain is within 2 years of l owering the cholesterol, whether by diet or a combina- tion of diet and drugs. Two groups need special mention: patients with type 2 diabetes mellitus and those with acut e coronary syndrome. Both groups show early bene \201 t with statin treatment (high-dose a torvastatin 80 mg). In post-MI patients, omega-3 fatty acids are recommended as they reduce the incidence of sudden death due to arrhythmias. Tackling cholesterol in primary prevention is controversial. There have been primary preven tion trials showing bene \201 t from lowering cholesterol. The \201 nancial burden of th is has to be confronted. Speci \201 c interventions Dietary advice All patients should see a dietitian. Total fat i ntake should constitute <35% of energy consumed (with the majority of fat intake being from polyun- saturated and monounsaturated fats) and saturated fats must be <30% of the total f at content. Vegetable and marine mono- and polyunsaturated fats should be i in the diet. To tal dietary cholesterol should not exceed 200 mg/day. Foods advocated include fresh fruits and vegetables, which are also important sources of an tioxidants. These foods should be tried for 3 months to see if dietary manipulation will wo rk (monitor at 6-week inter- vals). It is unusual to see >15% fall in cholesterol from diet ary measures; a reduction of 5% is more likely in free-living individuals on a diet. Alcohol intake should be no more than 1 drink per day for a female and no more than 2 drink s per day for a male. At this level, alcohol intake con- fers CHD protection in men >40 yea rs old and postmenopausal women. 116-Draznin-Chap116.indd 677 4/1/2011 6:52:41 PM CHAPTER 116 Management of dyslipidemia 678 Plant sterols and stanols, 2 g daily, can reduce blood cholesterol by 10 \200\22315%. They are available commercially in enriched margarine spreads, yogurt, and milky drinks. Weight control All overweight patients must be encouraged to lose weight. Weight reduction is closely attuned to dietary advice and physical exercise. Most weight is lost in the \201 rst 4 months of a regimen. A 10 kg weight loss in an obese subject can reduce LDL chol esterol by 7% and increases HDL cholesterol by 13%. Physical activity Physical activity, especially aerobic exercise, is recommended. This shou ld involve moderate intensity exercise performed for a minimum of 30 min- utes, at least 5 times per week. Acute exercise will transiently change lipoprotein levels and increase lipoprotein lipase a ctivity. These effects become more permanent with regular training. Triglyceride levels fall, HDL cholesterol levels rise, especially the HDL 2 subfraction, wi th more vigorous exercise, and the LDL cholesterol is of the less-dense variety, which is n ot so atherogenic. The changes are dose dependent with i exercise, and a 20% alteration in each variable is achiev- able after 6 weeks. Modi \201 cation of other risk factors Other risk factors such as hypertension, smoking, a nd diabetes mellitus must also be addressed. Drug therapy Numerous agents can be used for both primary and secondary prevention in patie nts in whom diet has been ineffective. HMG CoA reductase inhibitors (statins) Indications These include high LDL or VLDL, i.e., pr imary hypercholesterolemia, heterozygous + homozygous FH, mixed hyperlipidemia. Not all of these medications are licensed for use in children and should be used with cau- tion in wom en of childbearing age because of insuf \201 cient safety data on pregnancy and of potenti al teratogenicity. The medication should be stopped for at least 3 months before pregnancy is planned. Women m ust be warned about pregnancy when on these drugs. Mechanism of action There is competitive inhibition of 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA), which is the rate-limiting enzyme in cholesterol synthesis. Its activation inc reases hepatocyte LDL receptor numbers and reduces VLDL synthesis while enhancing its hepat ic clearance. b See Table 116.1 for a comparative lipid-lowering pro \201 le and Table 116.2 for dosage. 116-Draznin-Chap116.indd 678 4/1/2011 6:52:41 PM DRUG THERAPY 679 Table 116.1 Comparative lipid lowering pro \201 le of statins % Fall in triglycerides % Rise in HDLc Statin % Fall in LDLc Atorvastatin 38 \200\22354 13 \200\22332 3 \200\2237 Fluvastatin 17 \200\22334 8 \200\22312 3 \200\2236 Pravastatin 18 \200\22334 5 \200\22313 5 \200\2238 Rosuvastatin 50 \200\22363 10 \200\22328 3 \200\22314 Simvastatin 26 \200\22348 12 \200\22338 8 \200\22312 Side effects Statins are usually very well tolerated but are known to cause headaches, naus ea, and some abdominal discomfort. The main concerns are a hepati- tis-like picture and myo pathy. LFTs and creatine kinase should be measured before statins are pre- scribed. They should be used with caution in those with excessive alcohol intake. It is recommended that the medic ation be discontinued if liver enzymes aspartate transaminase (AST) and/or alanine transami nase (ALT) show >3-fold elevation above the upper limit of normal. LFTs should be checked every 3 \200\2234 months initially and at least annually in the long term. Statin-associated myopathy is rare and tends to occur when prescribed with other med ications, e.g., cyclosporine, \201 brates, or macrolide antibiotics or when renal impairm ent or untreated hypothy- roidism is present. Clinically, there is a picture of swollen tender muscles and creatine kinase >10 x the upper limit of normal. Rhabdomyolysis is even rarer. Interactions All stat ins can interact with cyclosporine and nicotinic acid and are used with caution with \201 brates. Warfarin interacts with atorvastatin, simvastatin, and rosuvastatin. Erythromycin interacts with all statins. Digoxin interacts with atorvastatin and simvastatin. Rifampicin interacts with \202 uvastatin and pravastatin. Atorvastatin and rosuvastatin may also interact with the oral contracep- tive pill, antacids, and some antifungals. Fibrates Indications These are high VLDL, triglycerides, or IDL, i.e., mixed hyperlipidemia s that have not responded adequately to diet or other therapy. These medica- tions are less effective than the statins at lowering cholesterol but better at i HDL cholesterol and mor e effective in lowering triglycerides. Reduce triglycerides by 20 \200\22360%, increase HDL by 15 \200\22330%, and reduce LDL by 5 \200\22325%. 116-Draznin-Chap116.indd 679 4/1/2011 6:52:41 PM CHAPTER 116 Management of dyslipidemia 680 Mechanism of action Fibrates alter lipoprotein metabolism to reduce VLDL triglyceride synth e- sis by i lipoprotein lipase activity and LDL-receptor-mediated LDL clear- ance while inc reasing HDL synthesis. The effect on LDL cholesterol may vary in isolated hypertriglyceridemia to increase LDL cholesterol. b See Table 116.2 for dosage. Side effects Occasionally, \201 brates cause nausea, anorexia, or diarrhea and precipitate gallstones (mostly clo \201 brate, although this is not approved in the U.S.). Also, pruritus, rashes, hair loss, and impotence can occur. They should not be used in patients with severe liver disease (AST/ALT >2 \200\2233 x upper limit of normal) and renal dysfunction (creatinine >2 mg/dL), as they are conjugated in th e liver prior to excretion by the kidney. Myopathy, although rare, is the main concern, and the risk of this increases if \201 brates are used with statins. Table 116.2 Dosage of lipid-lowering drugs Drug Dose/day Statins Atorvastatin Fluvastatin Pravastatin Rosuvastatin 10 \200\22380 mg 20 \200\22380 mg 10 \200\22340 mg 10 \200\22340 mg (5 \200\22320 mg in pat ient of Asian origin) 10 \200\22380 mg Simvastatin Fibrates Clo \201 brate Feno \201 brate Gem \201 brozil 50 \200\22365 kg, 1.5 g; >65 kg, 2 g 67 \200\223267 mg 0.9 \200\2231.2 g Anion-exchange resins Cholestyramine 12 \200\22336 g (in single dose or up to 4x day) 5 g 1 \200\2232 x day (max 30 g/day) 1.875 g (oral suspension or 3 tablets) twice daily with meals Colestipol hydrochloride Colesevelam Nicotinic acid group Niacin Niaspan 1.5 \200\2236 g in divided doses 500 \200\2232000 mg Omega-3 fatty acids Omacor Lovaza 1 \200\2234 g 1 \200\2234 g Cholesterol absorption blocker Ezetimibe 10 mg 116-Draznin-Chap116.indd 680 4/1/2011 6:52:41 PM DRUG THERAPY 681 Interactions Use with caution in combination with statins. Fibrates can enhance the effects of warfarin and antidiabetic agents and are contraindicated in those on orlistat. Anion exchange resins (bile acid sequestrants) Indications These include high LDL, i.e., hy percholesterolemia. Although largely superseded by statins, these are now best used as adju ncts when LDL has not fallen enough with a statin alone. They are also the only drug licens ed for use during pregnancy. Mechanism of action These agents bind to bile acids in the gut, thus reducing their enteroh e- patic circulation and increasing bile acid excretion. This increases hepato- cyte choles terol requirements, which increases LDL receptor production, thus reducing circulating LDL levels. Under optimum conditions, LDL cholesterol can be reduced by 20 \200\22330%, triglycerides rise by 10 \200\22317%, and HDL increases by 3 \200\2235%. Side effects These agents remain in the gut, so constipation, bloating, nausea, and abdomin al discomfort are not uncommon. Constipation, found in 35 \200\22340% of those on these age nts, can be helped with bulking laxatives. Less often a bleeding tendency due to vitamin K malabsorption can be seen. These agents can also exacerbate hypertriglyceridemia, but HDL cho- lesterol is often i sli ghtly. To reduce the side effects, patients should start with low doses and build up gradua lly over the next 3 \200\2234 weeks while maintaining a good \202 uid intake. Interactions These agents can reduce the absorption of warfarin, digoxin, B -blockers, prav astatin, \202 uvastatin, and hydrochlorothiazide. Many other agents such as simvastatin h ave not been checked, so to avoid any potential interac- tion, advise patients to take all other drugs 1 hour before or 4 hours after the resin. Nicotinic acid Indications These include high LDL, VLDL, IDL, or triglycerides. Nicotinic a cid is the most effective medication for i HDL-cholesterol. In practice, however, their use is limited by the side-effect pro \201 le, espe- cially \202 ushing. Niaspan , a modi \201 ed-release nicotinic acid product, is now available and does seem to cause fewer unwanted side effects and should be the \201 rst choice. Mechanism of action Nicotinic acid works by inhibiting lipolysis in adipocytes, thus alteri ng fatty acid \202 ux and reducing VLDL synthesis and HDL clearance. VLDL levels fall, as do those of triglycerides (by 20 \200\22350%) and LDL (by 5 \200\22325%), whereas HDL leve ls rise (by 10 \200\22350%). 116-Draznin-Chap116.indd 681 4/1/2011 6:52:41 PM CHAPTER 116 Management of dyslipidemia 682 Side effects Side effects are common, with 30% of patients being unable to tolerate these a gents. Vasodilatation giving cutaneous/facial \202 ushing occurs in most patients but ten ds to improve after 2 \200\2233 weeks of therapy or if patients are given prostaglandin inh ibitors such as aspirin in combination with nicotinic acid. Patients should avoid taking it with hot drinks as these increase its absorption and worsen the \202 ushing. Gastritis is also a common problem; the more severe hepatitis occurs in only 3%. Tachycardias, dry skin, and exacerbations of gout and precipitation of acanthosis nigricans and retinal edema are also recognized side effects. Nicotinic acid can adversely affect glucose control in diabetes mellitus. Interactions These agents potentiate the effects of antihypertensive therapies. Omega-3 fatty acids (Omacor , Lovaza ) Indications These include severe hypertriglycer idemia. Mechanism of action They inhibit the secretion of VLDL through i intracellular apo B-100 de struction. Normal patients see both a fall in VLDL and LDL, but LDL may rise in the hypertr iglyceridemic individual. Side effects As high doses are needed, this high-calorie load may increase obesity. More co mmonly it gives nausea and belching. Omacor produces fewer unwanted effects and is less calori \201 c, as it is used in a smaller dose. Interactions There are none that are signi \201 cant. Cholesterol absorption blocker Presently only one preparation, ezetimibe, is available, and is now the second-line of treatment after a statin. Indications This agent is used for LDL cholesterol-lowering in patients who are intol- eran t of statins or in combination with a statin in those whose levels are not adequately contr olled with a statin alone. It is also used in the very rare condition of sitosterolemia, where there is an i absorption of plant sterols. Mechanism of action Dietary and biliary cholesterol ab sorption is selectively inhibited. Ezetimibe (10 mg) monotherapy produces an 18% fall in LD L cholesterol, an increase in HDL cholesterol of 1 \200\2233%, and triglycerides are not af fected. In combina- tion therapy with a statin, ezetimibe reduces levels by an additional 2 2% to that obtained by statin alone. 116-Draznin-Chap116.indd 682 4/1/2011 6:52:42 PM DRUG THERAPY 683 Interactions None are signi \201 cant. Which drug to use when (for diet-resistant dyslipidemias) Elevated LDL cholesterol only Fir st choice \200 Statins Second choice \200 Cholesterol absorption blocker Then: \200 B ile acid sequestrants \200 Nicotinic acid \200 Fibrates Combination therapy if the abov e fail to reduce LDL adequately includes: \200 Statin + bile acid sequestrant (can give 5 0% fall in LDL + 10 \200\22315% rise in HDL) Statin + cholesterol absorption blocker (can give an additional 22% fall in LDL + 3% rise in HDL) Statin + nicotinic acid (can give 50% fall in LDL + 25 \200\22350% rise in HDL) \200 Nico tinic acid + bile acid sequestrants (can give 35% fall in LDL + 25 \200\22350% rise in HDL) Statin + bile acid sequestrant + nicotinic acid (can give 66% fall in LDL + 25 \200\22350% rise in HDL) Elevated triglycerides only First choice \200 Fibrates Second choice \200 Omega-3 fatty acids Third choice \200 Nicotinic acid All three groups can occasionally be used togethe r if one or two of the above groups are insuf \201 cient. Mixed hyperlipidemia First choice \200 Fibrate Second choice \200 Fibrate + statin (but needs close monitoring) \200 Fibrate + nicotinic acid As with elevated LDL alone, combin ation therapy may sometimes be use- ful with emphasis on avoiding side effects and interact ions as above. 116-Draznin-Chap116.indd 683 4/1/2011 6:52:42 PM CHAPTER 116 Management of dyslipidemia 684 Aims of treatment In the United States the decision to treat lipids is based on an assessme nt of any individual \200\231s cardiovascular risk: \200 For individuals with 0 \200\2231 risk factor, the LDL goal is <160 mg/dL. \200 For individuals with multiple (2+) risk fa ctors, LDL goal is <130 mg/dL. For patients with CHD and CHD risk equivalents, the primary treatment goal is as follows: \200 LDL cholesterol <100 mg/dL with a therapeutic option f or <70 mg/dL \200 Although an optimal triglyceride value is <150 and an optimal HDL is >40 (men), the primary goal of treatment is LDL lowering. An alternative to the above is to begin assessment with the Framingham risk scoring (10-yea r risk assessment) (see Tables 113.1, pp. 664 \200\223665 and 113.2, pp. 666 \200\223667) a nd use the associated goals listed in Table 116.3. Table 116.3 10-year risk LDL goal (cid:2)20% <100 mg/dL 10 \200\22320% <130 mg/dL <10% with multiple (2+) risk factors <130 mg/dL <10% with 0 \200\2231 risk factor <160 mg/dL Further reading American Diabetes Association (2004). Position statement, Dyslipidemia mana gement in aduts with diabetes. Diabetes Care Suppl 21:s68 \200\223s71. Bhatnagar D (2006). Diagnosis and screening for familial hypercholesterolaemia: \201 ndin g the patient, \201 nding the genes. Ann Clin Biochem 43:441 \200\223456. Cannon CP, Braunwald E, McCabe CH, et al. (2004). Intensive versus moderate lipid lowering with statins after acute coronary syndromes. N Engl J Med 350:1495 \200\2231504. Colhoun H, Betteridge DJ, Durrington PN, et al. (2005). Rapid emergence of effect of atorva statin on cardiovascular outcomes in the collaborative atorvastatin diabetes study (CARDS). Diabetelogia 48:2482 \200\2232485. De Backer G, Ambrosioni E, Borch-Johnsen K, et al. (2003). European guidelines on cardiovas cular disease prevention in clinical practice. Eur Heart J 24, 1601 \200\22310. Downs JR, Clear \201 eld M, Wies S, et al. (1998). Primary prevention of acute coronary ev ents with Iovastatin in men and women with average cholesterol levels: results of AFCAP/Tex CAPS. Air Force/Texas Coronary Atherosclerosis Prevention Study. JAMA 299:1615 \200\2231621 . Durrington P (2003). Dyslipidaemia. Lancet 362:717 \200\223731. Dyslipidaemia Advisory Group, on behalf of the Scienti \201 c Committee of the National He art Foundation of New Zealand (1996). 1996 National Heart Foundation guidelines for the ass essment and management of dyslipidaemia. N Z Med J 109:224 \200\223232. 116-Draznin-Chap116.indd 684 4/1/2011 6:52:42 PM AIMS OF TREATMENT 685 Hippisley-Cox J, Coupland C, Vinogradova Y, et al. (2007). Derivation and validation of QRI SK, a new cardiovascular dis-ease score for United Kingdom: prospective open cohort study. BMJ 335:136 \200\223141. International Atherosclerosis Society (1996). Clinician \200\231s Manual on Hyperlipidaemia , 4th ed. London: Science Press. Joint British Society 2 (2005). Joint British Societies \200\231 Guidelines on prevention o f cardiovascular disease in clinical practice. Heart 91 (Suppl 5):1 \200\22352. National Institute for Health and Clinical Excellence (2006). Statins for the prevention of cardio- vascular events. Technology Appraisal 94. http://www.nice.org.uk/TA094 Reckless JPD (1996). Economic issues in coronary heart disease prevention. Curr Opin Lipido l 7:356 \200\223362. Royal College of General Practitioners (1992). Guidelines for the management of hyperlipida emia in general practice. Occas Pap R Coll Gen Pract 55:1-15. Sacks FM, Pfeffer MA, Moye LA, et al. (1996). The effect of pravastatin on coronary events after myocardial infarction in patients with average cholesterol levels. N Engl J Med 335:1 001 \200\2231009. Scandinavian Simvastatin Survival Study Group (1994). Randomized trial of cholesterol lower ing in 4444 patients with CHD: the Scandinavian Simvastatin Survival Study. Lancet 334:1383 \200\2231389. Shepherd J, Cobbe SM, Ford I, et al. (1995). West of Scotland Coronary Prevention Study Gro up. Prevention of CHD with pravastatin in men with hypercholesterolemia. N Engl J Med 333:1 301 \200\2231307. 116-Draznin-Chap116.indd 685 4/1/2011 6:52:42 PM This page intentionally left blank Part 13 Laboratory endocrinology E. Chester Ridgway 117 Pitfalls in laboratory endocrinology 118 Reference intervals 688 693 117-Draznin-Chap117.indd 687 3/31/2011 5:40:54 PM CHAPTER 117 Pitfalls in laboratory endocrinology 688 Pitfalls in laboratory endocrinology Introduction The previous issue of this Handbook contained a chapter entitled \200\234Norm al ranges \200\235 for endocrine-related laboratory tests. The purpose of this chap- ter is to explain why this compendium of data needs to be viewed with a degree of caution and why you, as a clinical endocrinologist, should be actively liaising with your laboratory servi ce about the assays that they provide, their reference ranges, and the performance of these assays. Laboratory methods are complex and are affected by a number of factors (Box 117.1) that can often only be considered and explored if information is shared by frequent liaison between clinicians and laboratory scientists. Uncritical use of numbers obtained from the literatu re may lead to errors and bad management decisions. Box 117.1 Factors to be considered for laboratory investigations Pre-analytical factors \200 Sample timing \200 Which tube? \200 Sample transportation factors \200 Biological variation \200 Which stimulation test? Analytical factors \200 Assay speci \201 city \200 Assay standardization \200 Analyt ical performance \200 Hook effects \200 Antibody interference \200 Particularly probl ematic assays Post-analytical factors \200 Reference ranges \200 Units \200 Interpretation 117-Draznin-Chap117.indd 688 3/31/2011 5:40:54 PM PRE-ANALYTICAL FACTORS 689 Pre-analytical factors Half of all errors in the diagnostic process are due to pre-analytic al factors (Box 117.2), and 20% of errors are related to sample collection. Even in hospita ls where there is a heightened awareness of these problems, there is a prevalence of 1% pre -analytical errors. These effects can be of suf \201 cient magnitude to alter the analysis enough to create si tuations for clinical errors. Most problems can be prevented by clear instructions and docu mented policies for sampling. Some issues are relatively straightforward, such as collecting the sample into the correct blood tube and ensuring that samples are transported to the laboratory fast enough at the c orrect temperature. A comprehensive list can be found at http://www.specimencare.com/. In order to be certain whether two or more samples taken from an individual are different, it is important to consider the biological variation that naturally occurs (see Box 117.3 ) . A comprehensive list can be found at http://www.westgard.com/intra-inter.htm. In addition, the clearance t of each hormone should be considered: endogenous hormones an d tumor markers are like pharmaceuticals and 90% of the change occurs after 4.5x t . Box 117.2 Examples of pre-analytical errors \200 Incorrectly or unlabeled samples \200 Unde \201 ned or inappropriate sample timing \200 Wrong tubes \200 Hemolysis \200 Li pemia \200 Delayed transportation \200 Incorrect transportation temperature Box 117.3 Examples of within-person (biological) variation Aldosterone Androstenedione CA125 CA153 CEA DHAS Prolactin SHBG Testosterone (male) TSH 29% 16% 36% 5.7% 10.6% 3.4% 24% 9% 10% 20% Variation expressed as cv(%) [Coef \201 cient of variation = (standard devia- tion/ mean)* 100] 117-Draznin-Chap117.indd 689 3/31/2011 5:40:54 PM CHAPTER 117 Pitfalls in laboratory endocrinology 690 Many endocrine function tests require stimulation tests. What is the evidence for the one y ou plan to use? How much difference exists for dif- ferent stimulation tests? For examples of the effects of different stimulants of GH see Rahim et al. (1996). 1 Analytical factors Most endocrine assays are immunoassays that rely on binding of a hor- mo ne or metabolite by a diagnostic antibody. The binding speci \201 city will depend on the care and attention with which the manufacturer has chosen the reagents. Typical interferences may be due to slightly different molecular forms such as occurs with steroid or thyroid hormones. Peptide hormones are more complex because so many differently glycosylated isoforms of those peptides exist. Standards for pituitary hormones are generally derived from puri \201 ed pituitary extract s that contain a mixture of peptides. This leads to differ- ent assays having quite marked biases between each other due to different binding af \201 nities of the antibodies to the different isoforms. A similar situa- tion exists with hCG, for which many multiple molecul ar forms co-exist. Attempts to \201 nd international consensus for standards that can be used in diagnostic systems are being made, particularly for growth hormone and glycated hem oglobin. Analytical performance or assay reproducibility is important in determin- ing the critical differences between patient samples. A signi \201 cant difference between consecutive samp les at 95% con \201 dence will require a difference of 1.96x the method standard deviation at the appropriate concentration. The hook effect occurs when very high hormone concentrations \202 ood the available antib ody in vitro, leading to artifactually low results. It is less common as assays have larger dynamic ranges, but new cases continue to be reported, and there are published case report s for all hormones and tumor markers. Antibody interference is a widespread problem that affects approxi- mately 1% of immunoassa ys. It is insidious and due to endogenous anti- bodies that interfere with hormone binding in vitro. Most importantly, they cannot be detected by usual-quality control mechanisms. Th ere are a number of laboratory techniques that can be used to clarify whether such interfer ence is present, but it is inherent on the clinician to alert the laboratory to a potential clinical mismatch. Some assays can only be classi \201 ed as problem assays. These include thyroglobulin and low concentrations of estradiol (<300 pmol/L) and testosterone (<5 nmol/L). The former is d ue to the high prevalence of endogenous antithyroglobulin antibodies, which are particularl y prevalent in patients with thyroid disease. The latter are due to antibody speci \201 ci ty; however, it is hoped that this will be resolved with the introduction of mass spectrome try into routine clinical practice. 1 Rahim A, Toogood AA, Shalet SM (1996). The assessment of growth hormone status in normal young adult males using a variety of provocative agents. Clin Endocrinol 45:557 \200\223562 . 117-Draznin-Chap117.indd 690 3/31/2011 5:40:54 PM POST-ANALYTICAL FACTORS 691 Post-analytical factors Interpretation of assay results is made in relation to reference ra nges pro- vided by the laboratory. These usually represent the 95th percentiles of a popula tion of healthy individuals. However, the de \201 nition of normality is subjective, and r eference ranges are affected by such factors as age, gender, and, in some cases, ethnicity. Moreover, for hormones, time of day, month, and season are important. For some hormones, it is dif \201 cult to obtain appropriate samples to con- struct ranges, such as in children , and in circumstances that make it dif \201 cult to obtain samples in health, e.g., follo wing pharmacological stimulation or samples of CSF. If the central 95th percentile reference ranges are used, there is a 5% chance that a resul t will be out of range due to chance. As the number of tests taken are i, so will the risk of a chance abnormality. The increase will be x% (where x = 1 \200\223 0.95 a , and a is t he number of tests performed). Literature from U.S. and European journals may use different units \200\224be- ware! SI units use molar or mass (g) and volumes are reported in liters. Most assays are standardized with international preparations. These are usually the molecul ar forms that are most prevalent when basal samples are taken. However, following stimulati on, nonstandard molecules are released into the circulation that have different clearance r ates and differ- ent binding characteristics to the diagnostic antibodies in vitro. This can lead to marked differences between methods. For example, fol- lowing stimulation b y ACTH, corticosteroid precursors are released that will compete with cortisol for binding in the assay. Similarly, following stimulation of GH release, different isoforms of GH are secreted and the most abundant 20 and 22kDa isoforms clear at different rates, leading to v ariations in recog- nition by the diagnostic antibodies at different times during the test. 117-Draznin-Chap117.indd 691 3/31/2011 5:40:55 PM CHAPTER 117 Pitfalls in laboratory endocrinology 692 Box 117.4 Disastrous outcomes Assay interference A series of patients has been described in whom aggressive therapy for c horiocarcinoma was instituted for diagnoses based on assays affected by in vitro artifacts. Antibody speci \201 city A patient has been described who had prolactin measured by three different assays, giving three different answers. The solution awaited a clinical answer wh en the hyperprolactinemia resolved after the offend- ing medication was stopped. Interference by insulin autoantibodies A patient has been described with recurrent hypoglyc emia due to insulin autoantibodies caused by myeloma. This patient demonstrated in vivo int erference by endogenous antibodies as well as in vitro interference. Hook effect A patient presents with a large pituitary tumor that appears nonfunc- tioning; prolactin is normal and the patient is treated surgically. The next day, the blood sample i s reassayed after dilution, and high prolactin is uncovered when the antibody is no longer \202 ooded by excess prolactin. 117-Draznin-Chap117.indd 692 3/31/2011 5:40:55 PM THYROID FUNCTION 693 Reference intervals Introduction Box 118.1 De \201 nitions \200 Serum A serum sample is collected in a plain tube, left t o permit clotting, then centrifuged and separated. Plasma A plasma specimen is collected in a tube containing EDTA or lithium heparin, centrifuged immediately, and separated. All values are for serum unless speci \201 ed otherwise. Thyroid function Table 118.1 Thyroid Function Tests Traditional units Conversion factor Hormone SI Units TSH 0.35 \200\2234.50 mU/L 0.35 \200\2234.50 mU/L 1 Total T 4 Free T 4 Total T 3 Free T 3 Thyroglobulin 60 \200\223140 nmol/L 4.5 \200\22311.0 mcg/dL 12.9 10.3 \200\22322.7 pmol/L 0.8 \200\2231.8 ng/dL 12.9 1.2 \200\2232.8 nmol/L 80 \200\223180 ng/dL 0.0154 3.5 \200\2236.5 pmol/L 2.3 \200\2234.3 pg/mL 1.54 1.0 \200\22330 mcg/L 1.0 \200\22330 ng/mL 1 Acid-containing container (20 mL 6M HCl). 118-Draznin-Chap118.indd 693 3/31/2011 5:41:54 PM CHAPTER 118 Reference intervals 694 Adrenal and gonadal function Table 118.2 Adrenal and gonadal function tests Traditional units Conversion factor Hormone SI units Cortisol (9 A.M.) 140 \200\223635 nmol/L 5 \200\22323 mcg/dL 27.6 Aldosterone Supine Erect 0.8 \200\2233.0 nmol/L 1.4 \200\2238.0 nmol/L 3 \200\22310 ng/dL 5 \200\22330 ng/dL 0.0277 0.0277 Plasma renin activity Supine Erect 0.6 \200\2233 mcg/L/hr 3 \200\22310.8 mcg/L/hr DHEAS Women Men 6 \200\22310 micromol/L 6 \200\22317 micromol/L 22 \200\223380 mcg/dL 22 \200\223640 mcg/dL 0.0027 0.0027 Androstene- dione Women Men 0.3 \200\2237.4 nmol/L 1.0 \200\2234.6 nmol/L 0.1 \200\2232.1 ng/mL 0.3 \200\2231.3 ng/mL 3.49 3.49 17 \200\223hydroxy- progesterone Women Men 0.3 \200\22310 nmol/L 0.3 \200\2234.6 nmol/L 0.1 \200\2232.9 ng/mL 0.1 \200\2231.4 ng/mL 3.3 3.3 Estradiol Women Pre-M Post-M Men 110 \200\2231468 pmol/L 0 \200\223110 pmol/L 36 \200\223185 pmol/L 30 \200\223400 pg/mL 0 \200\22330 pg/mL 10 \200\22350 pg/mL 3.6 3.6 3.6 Progesterone Follicular Luteal Men <3 nmol/L 6.5 \200\22380 nmol/L <3 nmol/L <1.0 ng/mL 2.0 \200\22325 ng/mL <1.0 ng/mL 3.2 3.2 3.2 Testosterone Men Women 6.9 \200\22338 nmol/L 0.35 \200\2232.45 nmol/L 200 \200\2231080 ng/dL 10 \200\22370 ng/dL 0.035 0.035 Dihydro- testosterone Men Women 1 \200\2232.6 nmol/L 0.3 \200\2230.8 nmol/L 29 \200\22376 ng/dL 8.7 \200\22323 ng/dL 0.034 0.034 Sex hormone binding globulin Women Men 18 \200\223114 nmol/L 13 \200\22371 nmol/L 18 \200\223114 nmol/L 13 \200\22371 nmol/L 1 1 A variety of different units are used by non-UN labs. 118-Draznin-Chap118.indd 694 3/31/2011 5:41:55 PM PITUITARY HORMONES 695 Pituitary hormones Table 118.3 Pituitary hormones Traditional units Conversion factor Hormone SI units FSH Follicular Mid-cycle Luteal Post- menopausal Men 0.5 \200\2235 IU/L 8 \200\22333 IU/L 2 \200\2238 IU/L >30 IU/L 0.5 \200\2235 mU/mL 8 \200\22333 mU/mL 2 \200\2238 mU/mL >30 mU/mL 1 1 1 1 1.0 \200\22318 IU/L 1.0 \200\22318 mU/mL 1 LH Follicular Mid-cycle Luteal Post- menopausal Men 1 \200\22310 IU/L 20 \200\22380 IU/L 3 \200\22316 IU/L >30 IU/L 1 \200\22310 mU/mL 20 \200\22380 mU/mL 3 \200\22316 mU/mL >30 mU/mL 1 1 1 1 3 \200\2238 IU/L 3 \200\2238 mU/mL 1 Prolactin Women Men 0 \200\22325 mcg/L 0 \200\22320 mcg/L 0 \200\22325 ng/mL 0 \200\22320 ng/mL 1 1 Growth Hormone 0 \200\22320 mU/L 0 \200\22310 ng/mL 3 IGF-1 20 years 40 years 60 years >60 years 16 \200\223115 nmol/L 14 \200\22346 nmol/L 10 \200\22335 nmol/L 7 \200\22328 nmol/L 120 \200\223885 ng/mL 105 \200\223353 ng/mL 79 \200\223263 ng/mL 52 \200\223210 ng/mL 0.13 ACTH* 2 \200\22318 pmol/L 10 \200\22380 pg/mL 0.22 Antidiuretic hormone 0 \200\2234.7 pg/mL Lithium heparin tube is cold spun immediately. 118-Draznin-Chap118.indd 695 3/31/2011 5:41:55 PM CHAPTER 118 Reference intervals 696 Bone biochemistry Table 118.4 Bone biochemistry tests Traditional units Conversion factor Hormone SI units Parathyroid hormone* 1.0 \200\2236.0 pmol/L 10 \200\22360 pg/mL 0.1 Total 25-hydroxy- cholecalciferol* 75 \200\223200 nmol/L 30 \200\22380 ng/mL 2.5 1, 25-dihydroxy- cholecalciferol* 35 \200\223180 pmol/L 15 \200\22375 pg/mL 2.4 Calcitonin \200 0 \200\2237.5 pg/mL Serum, cold spun, \202 ash frozen. \200 Lithium heparin tube is cold spun immediately. Plasma gastrointestinal and pancreatic hormones Table 118.5 Plasma GI and pancreatic hormones Traditional units Conversion factor Hormone SI units Insulin (fasting)* 21 \200\223133 pmol/L 3 \200\22319 mU/L 6.9 C-peptide (fasting) 0.25 \200\2231.2 nmol/L 0.8 \200\2233.5 ng/mL 0.33 Gastrin* 0 \200\22345 pmol/L 0 \200\223100 pg/mL 0.45 Glucagon* 11 \200\22336 pmol/L 40 \200\223130 pg/mL 0.28 Vasoactive intestinal polypeptide (VIP)* 0 \200\22325 pmol/L 0 \200\22360 pg/mL 0.42 Pancreatic polypeptide* 0 \200\223104 pmol/L 0 \200\223435 pg/mL 0.24 Somatostatin* 0 \200\223150 pmol/L Chromogranin A* 0 \200\22350 pmol/L Vasopressin 0 \200\2234.7 pg/ml Fasting, Trasylol tube, lithium heparin, cold spun and \202 ash frozen. 118-Draznin-Chap118.indd 696 3/31/2011 5:41:55 PM URINARY COLLECTIONS 697 Tumor markers Table 118.6 Tumor markers Tumor marker Traditional units BhCG 0 Carcinoembryonic antigen (CEA) 0 \200\2233.0 ng/mL Prostate-speci \201 c antigen (PSA) A-Fetoprotein Thyroglobulin 0 \200\2234 ng/mL 0 \200\22315 ng/mL 0-30 ng/ml Urinary collections Table 118.7 Urinary collections: hormones Urine hormone Traditional units (mcg/24 hours) Norepinephrine 0 \200\223100 mcg/24 hours Epinephrine 0 \200\22325 mcg/24 hours Dopamine 60 \200\223440 mcg/24 hours Cortisol, free 0 \200\22360 mcg/24 hours 5-hydroxyindoleacetic acid 2 \200\2238 mg/24 hours Table 118.8 Urinary collections: analytes Urinary analyte Traditional units (mg or mmol/24 hours) Calcium 100 \200\223250 mg/24 hours Phosphate 400 \200\2231300 mg/24 hours Potassium 25 \200\223125 mmol/24 hours Sodium 50 \200\223285 mmol/24 hours 118-Draznin-Chap118.indd 697 3/31/2011 5:41:55 PM This page intentionally left blank 699 Index pheochromocytoma, 364 management of, 364 autoimmune adrenalitis associated with, 197 Abetalipoproteinemia, 673 Abnormal thyroid function Adrenalectomy, 188 clinical features of associated conditions, 198 chronic, 198 laboratory investigations, tests, 542 bilateral, for congenital adrenal hyperplasia, 244 Acanthosis nigricans, 650 Acarbose, 657 \200\2238 ACE inhibitors for Cushing \200\231s disease, 122 laparoscopic bilateral, 198 for hyporeninemic in pregnancy, 362 primary adrenal insuf \201 ciency causes of, 196 \200\2237 investigation of, 200 \200\2231 screening