Eric Niederhoffer SIU-SOM

1. Making basic science clinically relevant for learners: the biochemistry example Eric Niederhoffer SIU-SOM

2. Wants and needs Curriculum design, objectives, goals; USMLE Biochemistry as a foreign language Web lessons, resource pages, animations Resource sessions Complement self-directed learning Applied to patient case Start simple, discuss difficult Big picture, relevant details Overlap and redundancy Build upon previous knowledge Clinical probes for content and concepts Self-assessment questions, examinations Glucose metabolism as an example Considerations

3. Red Blood CellBiochemistry A 4-year-old African boy presents with a 2-day history of painful extremities. RBC Structure - size, spectrin, channels Metabolism - glycolysis (2,3-BPG), pentose phosphate pathway (G6PDH, NADPH), glutathione Hemoglobin - Genes, heme, Mb/Hb (normal), O2 binding, HbS (defect), fibers (sickling and inflammation)

4. Students’ Notes Devlin, T. M. (ed.). 2006. Textbook of biochemistry with clinical correlations, 6th ed. John Wiley & Sons, Inc., New York. This is very good for most of what you need. Mehta, A. B., and A. V. Hoffbrand. 2000. Haematology at a glance, Blackwell Science, Malden, Mass. Salway, J. G. 2006. Medical biochemistry at a glance, 2nd ed. Blackwell Science, Malden, Mass. This is very good for general principles and topics, and metabolic pathways and regulation. Good focused clinical correlations.

5. H2O2 H2O GSH GP HK Glc G6P NADP+ + H+ GSSG PGI GR F6P Glycolysis PFK NADPH F16BP aldolase G6PDH lactonase 6PGDH G3P DHAP CO2 G3PDH BPG mutase 1,3-BPG 6PG PGK 2,3-BPG PPP 3PG PGM 2,3-BPG phosphatase 2PG 3-7 C metabolites (R5P, F6P, G3P) enolase PEP PK No O2 Lactate Pyr LDH RBCMetabolic Pathways

6. Students’ Notes Glc: glucose HK: hexokinase G6P: glucose-6-phosphate G6PDH: glucose-6-phosphate dehydrogenase PGI: phosphoglucose isomerase PFK: phosphofructokinase DHAP: dihydroxyacetonephosphate BPG: bisphophoglycerate PEP: phosphoenolpyruvate Pyr: pyruvate PK: pyruvate kinase (2 genes, 4 isozymes) NADP+/NADPH: nicotinamide adenine dinucleotide R5P: ribulose-5-phosphate F6P: fructose-6-phosphate G3P: glyceraldehyde-3-phosphate GSH: reduced glutathione (GSH = Glu-Cys-Gly) GSSH: oxidized glutathione LDH: lactate dehydrogenase PPP: pentose phosphate pathway 6PGDH: 6-phosphogluconate dehydrogenase GR: glutathione reductase GP: glutathione peroxidase 3PG: 3-phosphoglycerate 6PG: 6-phosphogluconate Defect in HK, PGI, aldolase, or BPG mutase/2,3-BPG phosphatase  decreased [2,3-BPG]; defect in PK  increased [2,3-BPG] BPG mutase(or synthase)/2,3-BPG phosphatase is a bifunctional enzyme (one protein, two activities), regulated by hypoxia and T3 MIultiple inositol polyphosphate phosphatase acts on 2,3-BPG to give 2-PG Fetal Hb - lower affinity for 2,3-BPG compared with adult Hb; 2,3-BPG binds to and stabilizes deoxyHb; it is easily displaced from oxyHb Common deficiencies: G6PDH - X-linked PGI - autosomal recessive PK - autosomal recessive Sodium fluoride inhibits enolase, used to preserve blood samples for glucose determinations.

7. Hemoglobin Structure Changes http://www.mfi.ku.dk/PPaulev/chapter8/images/8-3.jpg

8. Factors Affecting Binding of O2 Depends on pH ([H+]), CO2, BPG (DPG), Temp pH  BPG or T _; left shift pH _ BPG or T ; right shift

9. What metabolic pathways are used in erythrocytes? What clinical observations would you make concerning patients with SCD? Review Questions

10. Metabolism in skeletal muscle Pathways overview Regulation in skeletal muscle Metabolism in nervous tissue Pathways overview Clinical aspects Clinical aspects Clinical/laboratory findings GSD, PDHCD Glycogen storage disease type VII Pyruvate dehydrogenase complex deficiency Inborn errors of metabolism Metabolism in Skeletal Muscle and Nervous Tissue

11. Glycolysis Glycogenolysis -oxidation (ketone bodies) Krebs (tricarboxylic acid) cycle Branched-chain amino acids Electron transport chain Calcium regulation Key enzyme regulation Metabolism in Skeletal Muscle A 21-year-old woman comes to the physician with pain in her right mid-arm. A 5-year-old boy is brought to the physician to have sutures removed.

12. Ketone bodies Glucose Glycogen Glycolysis Fatty acids Glycogenolysis b-Oxidation Ca2+ PKa Ca2+ PDH No O2 Lactate Pyruvate Acetyl-CoA Electron Transport Chain Krebs cycle BCAA Ile, Leu, Val Ca2+ ISDH, aKGDH Pathways Overview G6P Production of ATP

13. Ep AR ATP Citrate AC ATP PKA Ca2+ PKa PFK-2 PP F26BP Pi IMP AMP NH4+ AMP Pi PDHP Ca2+ PDHP PDHK PDH Regulation in Skeletal Muscle Glc Glycolysis cAMP G6P F6P PFK-1 F16BP Glycogen Glycogenolysis PEP PK PDH Pyr Acetyl-CoA

14. Glycolysis Glycogenolysis (stress) -oxidation (ketone bodies) Krebs (tricarboxylic acid) cycle Branched-chain amino acids Electron transport chain Metabolism in Nervous Tissue A 21-year-old woman comes to the physician with pain in her right mid-arm. A 19-year-old man is brought to the emergency department after a diving accident. A 63-year-old woman is brought to the physician for her “parkinsonism.”

15. Lactate (glial) Krebs cycle Pathways Overview Glucose Ketone bodies Fatty acids Glycogen Glycolysis Glycogenolysis G6P b-oxidation No O2 Lactate Pyruvate Acetyl-CoA Electron Transport Chain BCAA Ile, Leu, Val Production of ATP

16. Toxic accumulation disorders Protein metabolism disorders (amino acidopathies, organic acidopathies, urea cycle defects) Carbohydrate/intolerance disorders Lysosomal storage disorders Energy production/utilization disorders Fatty acid oxidation defects Carbohydrate utilization, production disorders (glycogen storage, gluconeogenesis, and glycogenolysis disorders) Mitochondrial disorders Peroxisomal disorders Metabolic acidosis (elevated anion gap) Hypoglycemia Hyperammonemia Clinical Aspects for Inborn Errors of Metabolism in Muscles

17. Evidence of familial coincidence Progressive decline in nervous functioning Appearance and progression of unmistakable neurologic signs General symptoms State of consciousness, awareness, reaction to stimuli Tone of limbs, trunk (postural mechanisms) Certain motor automatisms Myotatic and cutaneous reflexes Spontaneous ocular movements, fixation, pursuit; visual function Respiration and circulation Appetite Seizures Clinical Aspects for Inborn Errors of Metabolism in Nervous Tissue

18. Clinical/Laboratory Findings

19. Pentose Phosphate Pathway Glucose Glycogen R5P nucleotides G6P Glycogenolysis Glycogenesis Glycolysis Tarui disease Glycogen Storage Disease Type VII F6P PFK F16BP PDH complex deficiency PDH Acetyl-CoA Pyruvate Krebs cycle Glycogen Storage DiseasePyruvate Dehydrogenase Complex Deficiency

20. Classic, infantile onset, Late onset Exercise intolerance, fatigue, myoglobinuria Phosphofructokinase Tetramer of three subunits (M, L, P) Muscle/heart/brain - M4; liver/kidneys - L4; erythrocytes - M4, L4, ML3, M2L2, M3L General symptoms of classic form Muscle weakness, pronounced following exercise Fixed limb weakness Muscle contractures Jaundice Joint pain Laboratory studies Increased serum creatine kinase levels No increase in lactic acid levels after exercise Bilirubin levels may increase Increased reticulocyte count and reticulocyte distribution width Myoglobinuria after exercise Ischemic forearm test - no lactate increase with ammonia increase Glycogen Storage Disease Type VII (Tarui Disease)

21. Neonatal, infantile, childhood onset Abnormal lactate buildup (mitochondrial disease) Pyruvate dehydrogenase complex E1 -  (thiamine dependent) and  subunits, 22 tetramer E2 - monomer (lipoate dependent) E3 - dimer (riboflavin dependent) common to KGDH and BCAKDH X protein - lipoate dependent Pyruvate dehydrogenase phosphatase Nonspecific symptoms (especially with stress, illness, high carbohydrate intake) Severe lethargy, poor feeding, tachypnea Key feature is gray matter degeneration with foci of necrosis and capillary proliferation in the brainstem (Leigh syndrome) Infants with less than 15% PDH activity generally die Developmental nonspecific signs Mental delays Psychomotor delays Growth retardation Laboratory studies High blood and cerebrospinal fluid lactate and pyruvate levels Elevated serum and urine alanine levels If E2 deficient, elevated serum AAs and hyperammonemia If E3 deficient, elevated BCAA in serum, KG in serum and urine Pyruvate Dehydrogenase Complex Deficiency

22. Carbohydrates (Glycogen storage diseases) Amino acids (Maple syrup urine disease) Organic acids (Alkaptonuria) Mitochondrial function (Pyruvate dehydrogenase deficiency) Purines and pyrimidines (Lesch-Nyhan disease) Lipids (Familial hypercholesterolemia) Porphyrins (Crigler-Najjar syndromes) Metals (Hereditary hemochromatosis) Peroxisomes (X-linked adrenoleukodystrophy) Lysosomes (GM2 gangliosidoses - Tay Sachs disease) Hormones (hyperthyroidism) Blood (Sickle cell disease) Connective tissue (Marfan syndrome) Kidney (Alport syndrome) Lung (1-antitrypsin deficiency) Skin (Albinism) Inborn Errors of Metabolism

23. How does muscle produce ATP (carbohydrates, fatty acids, ketone bodies, branched-chain amino acids)? How is skeletal muscle phosphofructokinase-1 regulated? What are the key Ca2+ regulated steps? How does nervous tissue (neurons and glial cells) produce ATP (carbohydrates, fatty acids, ketone bodies, branched-chain amino acids)? How do glial cells (astrocytes) assist neurons? What are some key clinical features (history, physical, laboratory test results) associated with defects in metabolism that affect muscles and nervous tissue? Review Questions

24. For the third example taken from the ERG Unit, what would you choose for the resource session? Carbohydrate Metabolism in Diabetes A 59-year-old man is brought to the emergency department for evaluation of his semiconsciousness and minimal responsiveness

25. Regulation of glycolysis, glycogenesis, glycogenolysis, gluconeogenesis by insulin/glucagon PFK-2 (PKA, AMP-dependent PK) PK (PKA) PDH GS (PKA, PPK, GSK-3, PP-1) GP (PKA, PPK, PP) PEPCK (glucagon) G6Pase (glucagon) Regulatory differences among tissues Liver Muscle Cardiac muscle Key clinical features (history, physical, laboratory test results) associated with carbohydrate metabolism that occur in diabetes Carbohydrate Metabolism in Diabetes

26. Remember curriculum wants and needs Practice new language skills Use resource sessions effectively Complement self-directed learning Applied to patient case Start simple, discuss difficult Big picture, relevant details Overlap and redundancy Build upon previous knowledge Clinical probes for content and concepts Summary