News in laboratory diagnostics of metabolic syndrome Prof. David Stejskal, MD, PhD, MBA, EurChem ( ) Dept. o f Medical Chemistry and Biochemistry Faculty of Medicine and Dentistry Palacky University, Olomouc Czech Republic

Outline Definition Pathophysiology & dg. markers New markers

Definition(s) 15 December, 20153

Definition(s) - comments (Reaven sy. X, syndrome of insulin resistance) 1. WHO (1998) – used esp. in research 2. NCEP ATP III (2001) (National Cholesterol Education Program expert panel and Adult Treatment Panel) – in clinical settings 3. IDF (2005) (International Diabetes Federation) – in clinics All were different… common factors: IGT (or IR) Obesity (visceral adiposity) Hypertension (endothelial dysfunction) Dyslipidemia (atherogenic) 4.Harmonized definition (2009) 4

Harmonized definition (2009) 5 Metabolic syndrome = 3 and more of the following risk factors: Central obesityWaist circumference (for EU population): males ≥ 94 cm (  risk) ≥ 102 cm (  risk) females ≥ 80 cm (  risk) ≥ 88 cm (  risk) Hypertriacylglycerolemia≥ 1.7 mmol/l (or specific treatment) HDL-cholesterolmales < 1.0 mmol/l (or specific treatment) females < 1.3 mmol/l (or specific treatment) Arterial blood pressure≥ 130 / ≥ 85 mmHg (or specific treatment) Fasting glycemia (IFG)≥ 5.6 mmol/l (type II DM)

Prevalence Positively correlates with population age and obesity In the EU:  15 – 30% adults (depending on the definition)  ca. 40% of senior citizens suggesting that prevalence of type II DM and CV complications will sharply rise in the following decades In the Czech Republic (productive age yrs):  ca. 32% of men and ca. 24% of women In Slovakia: ca. 20% (NCEP) or 40% (IDF) of adults 6

Metabolic syndrome (MS) MS develops in time depending on: genetic predisposition (susceptibility) lifestyle factors (  activity,  E intake) age

Pathophysiology – older concept Insulin resistance – hyperglycemia (and  sympathetic tone) Visceral adiposity –  waist circumference (obesity), neck circumference Endothelial dysfunction – hypertension Atherogenic dislipoproteinemia –  TAG,  HDL 8

Pathophysiology proinflammatory prothrombotic proatherogenic

DM II as a manifestation of MS Type II DM insulin resistance insulin resistance hyperinsulinemia hyperinsulinemia visceral obesity visceral obesity hypertension inflammation fibrinolysis impairment fibrinolysis impairment prothrombotic state prothrombotic state dyslipidemiahormonalchanges

Consequences of MS (adopted from: Pelikánová. T.: Inzulinová rezistence a metabolický syndrome, 2004) Note: patients with MS have: 8x greater DM II risk 2-3x greater CV risk high total morbidity and mortality due to atherosclerosis and cardiovascular events as well as tumors (esp. colorectal Ca, prostate Ca, lung Ca, Ca of the uterus…) Insulin resistance Hyperinsulinemia Central obesity DLP (  HDL,  TAG, sdLDL and postprandial lipemia) IFG, IGT, type II DM Proinflammatory state Hyperuricemia (  CRP, TNF-α, IL-6 Essential hypertension and adhesion molecules) Impaired (  ) hemocoagulation Endothelial dysfunction (  PAI-1, fibrinogen) Microalbuminuria Accelerated atherosclerosis Tumors, non-alcoholic steatohepatitis (NASH), polycystic ovaries syndrome (PCOS)

Insulin resistance (IR) Defined as decreased biological response * to normal conc. of insulin IR leads to endothelial dysfunction and systemic inflammation *Biological response is typically referred to in terms of blood glucose level, but other effects of insulin (lipid and protein metabolism) should also be considered

Dyslipoproteinemia Typical MS-phenotype:  HDL and apo A  LPL activity,  HL and CETP activity  HDL 2 /HDL 3 ratio  TAG (total and VLDL 1) and FFA  TAG (apo CIII)-rich LP remnants  postprandial lipemia  small dense LDL (LDL 3 ) and apoB 100  LP modification (glycation, oxidation) TC and LDL may be normal or 

Superiority of Apo B over LDL source: prevalence of large, buoyant LDL (lbLDL) vs. more atherogenic small, dense LDL (sdLDL) source: Clin Lipidol 2011

Hypertension in MS „High normal pressure“ (i.e. ≥ 130/85 mm Hg) already is a risk factor Treatment: antihypertensives ACEi and AT1B – advantageous due to interference with the RAAS system alphaB and CaB – also preferable because they are metabolically indifferent BB – indispensable in patients with concurrent ischemic CHD (they decrease CV risk) Diuretics – only in small doses, which have little -ve metabolic effects (increase insulin sensitivity when combined with ACEi)

Obesity Quetelet´s „body mass index“ (BMI) BMI = body mass (kg) / body height 2 (m 2 ) BMI does not work in low and high age groups and in athletes with a developed musculature. BMIinterpretationobesity grade 20 – 24.99Normal body mass 25 – 29.99OverweightI 30 – 39.99ObesityII > 40Morbid obesityIII

Obesity Genetic factors – 23 obesity-associated genes have been identified until recently – presumably > 40% of primary obesity cases have a genetic background Lifestyle – genetics is the gun, lifestyle is the trigger – consumption of E-rich foods (polyphagia) inadequate to physical activity

Obesity Endocrine diseases associated with obesity: – Hypothyreosis – Cushing syndrome – Hyperprolactinemia GH deficiency and hypogonadism adiposity but not necessarily obesity – Diabetes mellitus (type II)

Obesity localization of fat tissue accumulation! abdominal adiposity – may be indirectly determined using the waist to hip ratio visceral fat is metabolically active – releases FFA, which: are transported to the liver and peripheral tissues induce insulin resistance (and in concert with  LPL/HL activity result in dislipoproteinemia in obese patients)

Significance of the visceral fat According to J. P. Despres Waist circumference is a good predictor of visceral fat, fat around liver..!!

Laboratory tests - classic Serum (plasma): intact proinsulin best marker of IR C-peptide TAG hsCRP (ultrasensitive) Plasma glucose (or HbA 1c ) Uric acid NEW MARKERS News – Adipokine expression in peripheral leukocytes or adipose tissue

New markers of MetSy Adipovascular axis – adipose tissue hormones (adipokines) have a direct effect on the vascular wall and endothelium control energy metabolism involved in atherosclerosis… – determination of adipokines and selected liver peptides or proteins – extrapolation from animals and use in clinical practice?

Definition of endocrine organ Secretory proteome contains at least 15,000 genes (it is only those which have the mRNA after transcription of the signal sequence and is therefore secreted via the classic pathway). Among them, there are hundreds or thousands of proteins that come from cells of non-classic way without the assistance of a signal sequence There is not a cell or tissue in the human body which would not produce any protein, including regulatory It is likely that each tissue is an endocrinally active and the whole then can be considered an endocrine organ

New concept Binder as an endocrine organ Skin as an endocrine organ Skelet Muscle as an endocrine organ Adipose tissue as an endocrine organ (the subject of our interest ) …

Adipose tissue as an endocrine organ White adipose tissue (WAT) adipocytes, preadipocytes, "non- adipose" macrophages, stromal cells, endothelial cells, fibroblasts, leukocytes. This composition defines the fatty tissue as an important mediator of metabolic and inflammatory processes 1993 Discovery of secretion of TNF-a? 1994 Discovery of first adipokines in the true sence of the word - slimness hormone LEPTIN (deficit led to extreme obesity). Gradually, it was found that no adipokines secreted not only adipocytes or fat tissues. There are many definitions inflammatory/non-inflammatory; adipokines similar to hormones/cytokines/chemokines; the new division at sensu stricto/sensu lato and adipokines helpful to humans and harmful (set in contemporary evolutionary and geo-economic situation

Metabolic syndrome (cluster of cardiovascular risk factors -obesity, DM II, dyslipidemia, hypertension..) If we add to the knowledge of the most well- known effects of adipokines Direct influence of TRG (fats)  steatosis of organs (liver, muscle) + Lipotoxic effect of FA for most tissues in the body (incl. pancreas) The cause of metabolic syndrome (Metsy) is now clearer

Selected adipokines Leptin – disc 1994 (via leptin receptor) regulates (  ) appetite and (  ) E expenditure high leptin in obese individuals does not lead to decreased appetite ( leptin resistance ) Adiponectin  production in obese, IR, endotelial dysfunction, CHD and MS patients decreases endothelial expression of adh. molecules, ROS, gluconeogenesis, Ma activation and foam cell accumulation increases glucose utilization and skeletal muscle FA oxidation, antidiab, antiats ef FGF-21 marker and independent positive risk factor of MS and type II DM produced by the liver and adipose tissue (correlates positively with fat mass) regulates insulin-independent transport of glucose into cells antidiabetic effect (leads to  glucose uptake by adipocytes by  GLUT-1 expression ) admin. of FGF-21 leads to  glycemia and  HDL (comparably to insulin) A-FABP (Adipocyte Fatty Acid Binding Protein) best parameter and predictor of MS, most dangerous adipokin, connection with CV mortality and morbidity regulates LP (  lipolysis) and glucose metabolism modulates proinflammatory cytokine production and CHE accumulation in Ma Macrophage A-FABP expression is  by oxLDL and  by statins

Omentin-1 is another new adipokine with anti-inflammatory effects, which affects the formation of AMI. Individuals with AMI had lower omentin-1 in serum and regular exercise leads to an increase of omentin-1 in serum and have decreased inflammation.

Omentin-1 had 97.7 sensitivity and 100% specificity, cut- off 207 mg/l 15 December, 2015medical biochemistry lecture29

A „new“ syndrome associated with MetSy: Obesity hypoventilation syndrome A number of unknown consequences Excessive production of carbon dioxide CO2 leads to hypercapnia OHS is considered as a disease with multiorgan diurnal hypercapnia and hypoxia, leading to many metabolic disturbances New: this is a cluster of MetSy (part of MetSy)

New risk factors: dermatological patients Psoriasis, systematic LE, PCOS, androgenic alopecia, lichen planus Psoriasis, systematic LE, PCOS, androgenic alopecia, lichen planus

New risk factors: New risk factors: The relationship between MetSy and carcinomas, ie CaP (BPH) Obesity and fat intake is associated with risk of prostate cancer (CaP) and BPH. In a pilot US study ( ) on 199 people with CaP it was found that most were overweight with increased trg and reduced HDL cholesterol.

New risk factor or part or cluster: NAFLD NAFLD has a high prevalence (33%), manifested variously (steatosis, fibrosis, cirrhosis, hepatocellular carcinoma). It is associated with insulin resistance, lipotoxicity and markers of MetSy Insulin resistance leads to the accumulation of fat (trg) in the liver increased with FFA, there is inflammation and stress in the endoplasmatic reticulum. Visceral fat in the liver is considered the most risky for cardiovascular diseases. Men usually have a deficit of free testosterone in blood.

Neck circumference - a new risk factor Circumference of the neck is connected to the CV risk studies in Asia A cut-off was determined for the presence of MetSy – for men: 39 cm – for women: 33 cm

GHDA - a new critical point in MetSy GHDA - growth hormone deficiency in adults is associated with a high risk of MetSy. Ghrelin induces its release (ghrelin also decreases with age as GH). GHDA mean- high waist circumference, low HDL, higher trg, hypertension, it is MetSy And Ghrelin deficiency. Treatment: rhGH (improves the parameters of MetSy and the quality of life, but there is no data in subjects over 80 years old). New form of therapy: Ghrelin administration?

Conclusion Some basic clusters of MetSy are gradually – Expanded – Reduced so we need to take into account the latest scientific information and translational research 15 December, 2015medical biochemistry lecture36

MetSy is a polygenic and multifactorial disease with a dominant and progressive prevalence worldwide. Laboratory medicine is one of the areas which can use a variety of etiopathogenetic factors in experimental designs or complement such designs. From their interlinking and connecting with other clinical symptoms of the underlying disease or its complications arises a somewhat modified picture of obesity, which suggests further possibilities for its treatment and prevention.