BIOCHEMISTRY (II) OF GENE EXPRESSION, HUMAN ORGANS AND FUNCTIONS

Teaching her lesson "Biochemistry (II) of Gene Expression.", of Human Organs and Functions" consists of lectures, tutorials and laboratory exercises.

The elections they develop the material described above while attendance at tutorials and exercises is mandatory.

The tutorials summarize the material and by using questions and examples of clinical cases (problem-based learning) highlight the application of biochemical knowledge in the diagnosis and treatment of serious diseases. For each tutorial, students are advised to have studied the material taught so that they can discuss it with the instructors.

The laboratory exercises (in 4 groups of students, 3 teachers per group of 25-30 students) constitute the students' practical training, are a necessary supplement to the lectures and aim to familiarize them with the use of techniques, the operation of instruments and the conduct of tests that are often used in the Clinic Biochemistry, as well as helping to understand concepts that are not easily grasped theoretically (learning based on practical experience). The completion and delivery for control of the booklet of the laboratory exercises is positively included in the evaluation of the students' performance.

Attendance is mandatory in all tutorials and laboratory exercises.

Information and Communication Technologies are used to prepare lecture materials and provide online information and learning aids to students.

In more detail:

• Common software (eg powerpoint) is used to prepare lecture material and display slides and videos.

• The study guide (analytical material & additional bibliography), the theory, the protocols of the laboratory exercises and the slides of the lectures after each lesson as well as videos and scientific articles related to the subject matter are made available to students electronically and online through the e- class.
• Class announcements, information and communication are available online via e-class. Communication is also done via e-mail.
• Common software (eg excel) is used for the statistical processing of the student evaluation.
• Additional general information about the faculty and their research interests and the operation of the Biochemistry Laboratory is available online on the Laboratory's website

http://www.med.uth.gr/biochemistry/index.html

The evaluation of the students' performance in the course is done:

A. In laboratory exercises: Laboratory Work, Report, Written Exams with short answer and problem solving questions.

The students' participation in the exercises as well or written report of the results of the exercises, which is done by the students during the workshops, is mandatory. The report includes the measurements (presented in tables and plots), and the conclusions (eg whether the expected results were obtained, if not why not, sources of possible errors in the experiments) as requested by each exercise. At the end of each exercise, the written report is checked by the teachers and signed if it has been filled in correctly. In the event that particularly demanding calculations or diagrams need to be made, it is possible for the report to be completed after the end of the workshop and delivered to the lecturers at a later predetermined date. Successful participation in the exercise is certified by the teacher's signature on the written report of the results.

At the end of the semester, students are examined in writing on the content of the Workshops. On his exams Laboratory material to be examined is the theory, the methodology and the methods of processing results included in the Laboratory Exercises Guide or developed by the lecturers during the laboratories. Only those who successfully participated in the laboratory exercises have the right to participate in the Laboratory exams. Passing the laboratory exams is a necessary condition for participation in the course exams.

B. In tutorials: face-to-face discussion between students and teachers, deepening with incidents and questions.

In each tutorial, students are prepared for the material of the chapters that will be discussed and also for the answers to the questions that have been posted in the e-class. Clinical cases will also be presented in each tutorial and they will discuss any questions with the teachers regarding the taught material. The participation of students in tutorials is positively included in the evaluation of their performance.

 C. In the theory-material of lectures: Written Exams with multiple-choice and short-answer questions.

The course exams are written, lasting 2 hours, and consist of multiple-choice questions (examples are available online in the e-class and in the course tutorials) and critical or short-answer questions related to the course material. Examinable material is the material of lectures and tutorials as described above. Only those students who have passed the Laboratory exams have the right to participate in the course exams.

 Final grade

The final grade of the course is calculated by 80% from the grade of the written exams of the course and by 20% from the grade of the written exams of the Laboratory.

All of the above can be accessed by the students as they are contained in the course Guide which is distributed in print to all students and is posted electronically in the e-class of the course.

Upon successful completion of the course, the student will be able to:
• Has understood the regulation of the biochemical functions of the human organism at the molecular, cellular and systemic level.
• Has knowledge of the mechanisms of conservation, transmission and expression of genetic information,
• Understands the mechanism of action of hormones and their role in the homeostasis of the organism as well as the biochemical specificity of various tissues and systems such as liver, sensory organs, blood proteins, hemostasis and adipose tissue.
• Has the necessary knowledge to understand and judge the effect of nutrition on health and in special situations.
• Has the necessary knowledge for the participation of muscle metabolism (exercise) in human health and pathogenesis such as cancer, diabetes, aging.
• Has the basic knowledge to distinguish the laboratory pathological picture from the normal one contributing to the diagnosis of basic metabolic diseases.
• Can collaborate with fellow students in a laboratory environment to perform basic biochemical analyzes and process their results.

General Skills
• Search, analysis and synthesis of data and information, using and
necessary technologies
• Adaptation to new situations
• Decision making
• Autonomous work
• Group work
• Work in an interdisciplinary environment
• Exercise criticism and self-criticism
• Promotion of free, creative and inductive thinking

A. Theoretical clinical skills
I. By theory unit

Unit 1: Mechanisms of genetic information flow
• Use of sequence mapping in diagnosis
• Knowledge of the properties and functions of DNA polymerases and topoisomerases for selective drug targeting and use of anticancer drugs
• Examples of chemical mutagens
• Carcinogenic substances that target molecular mechanisms of the genetic material
• Ames test and detection of carcinogenic substances
• Diseases resulting from dysfunction of repair mechanisms such as Xeroderma pigmentosum, Cockayne's syndrome, Hereditary nonpolyposis colorectal cancer-Lynch syndrome (HNPCC), Huntington's disease.
• Examples of antibiotics used as transcription inhibitors
• Splicing process and diseases resulting from alternative splicing dysfunction
• Protein synthesis and antibiotics used as its inhibitors
• The 'disappearing white matter' disease (VWM disease)
• the example of the action of nuclear receptors
• the example of iron homeostasis regulation

Unit 2: Intercellular communication
• Signal transduction of EGF and the ras-MAP kinase pathway as well as the correlation between signal transduction pathways and cancer.
• Examples of signaling pathway defects, oncogenes and toxins that can lead to cancer and other diseases.
• Targeting signaling pathways to treat pathological conditions.
• Clinical implications in Cushing's syndrome, Addison's disease and Graves' disease.
• Disorders of steroid hormone synthesis due to hereditary lack of enzymes.
• 21α-hydroxylase deficiency and clinical implications
• Blindness, nyctalopia and the genetic basis of color vision and color blindness
• Anosmia and the receptors of taste molecules
• Vitamin Deficiencies: Beri-Beri, Wernicke-Korsacoff Syndrome, Megaloblastic Anemia, Pellagra, Scurvy, Rickets and Osteomalacia
• Insulin-dependent and non-insulin-dependent diabetes mellitus
• Leptin and diseases of fuel regulation

Unit 3: Systems Biochemistry
• Assessment of liver function
• Liver proteins and molecules in clinical diagnosis and treatment
• Enzymes naturally present in plasma
• Diagnostic value of enzymes of tissue origin (transaminases, LDH, CPK) and their isozymes
• Coagulation factors and mechanism of action of anticoagulants such as aspirin, heparin.
• Haemophilias.
• Connection of hemostasis mechanisms with cell signaling in drug administration
• Adipose tissue and inflammation, obesity
• Nutrition: Essential fatty acids, amino acids, macronutrients (proteins, carbohydrates, fats).
• Diets, poor nutrition, malnutrition, Special conditions, Pathological conditions
• Muscle contraction and stimulation, control of muscle function
• Energy needs of muscles during exercise
• Effect of nutrition on the choice of energy sources during exercise
• Kama (regional and central)
• Advantages and disadvantages of physical exercise

B. Practical and clinical skills
In the mandatory workshops – learning based on practical experience
Acquisition of general and specific practical skills by medical students during
elaboration of the laboratory exercises:

1. General skills

• observation of phenomena in the laboratory, recording and analysis of data
• drawing conclusions from data
• solving practical problems
• building graphs from data
• extracting information from graphs
• handling basic equipment of Clinical Chemistry and Biochemistry laboratories
• effective team work
• safe work in a laboratory environment
• following and following instructions
• perception of laboratory errors and identification of their sources

2. Special skills

Clinical applications Preparation of recombinant plasmid DNA and digestion with restriction endonucleases
 Preparation of plasmid DNA from bacteria by the alkaline lysis method.
 Quantitative determination of DNA by the spectroscopy method
 Construction of agarose gel and electrophoretic separation of DNA based on its molecular weight.
 Mapping of recombinant plasmid after restriction endonuclease digestion and agarose gel electrophoresis
Clinical application of laboratory analysis of GOT and GPT transaminases
 Diagnostic significance of determination of aspartate aminotransferase transaminase (GOT or AST) and alanine aminotransferase (GPT or ALT).
 Pathological causes of decrease or increase of GOT and GPT in the blood
 Determination of concentration of GOT and GPT in the blood of patients photometrically
 Evaluation of laboratory results
Clinical application of laboratory analysis of urea
 Diagnostic significance of serum urea determination
 Pathological causes of decrease or increase of urea
 Determination of urea concentration in the blood of patients photometrically
 Evaluation of laboratory results and correlation with the results of the concentration of GOT and GPT in the blood of patients
Clinical application of laboratory analysis of bilirubin
 Diagnostic value of serum bilirubin determination
 Pathological causes of decrease or increase of bilirubin
 Determination of bilirubin concentration in the blood of patients photometrically
 Evaluation of laboratory results

C. Knowledge – Lecture material

• Flow of genetic information
• The central tenet of molecular biology
• the double helix model and types of DNA double helix
• The properties of DNA in solution
• The higher structures of double-stranded DNA
• The structure, species and biological role of RNA
• The genetic code: properties, decryption history

• Evolution
Biochemical evolution:
• Oparin/Haldane's 'primordial soup' theory, Stanley/Urey's experiment.
• The 'RNA world' theory
• The evolution of the biochemical processes of life until today, based on observations and simulation experiments.
• Examples of organisms of evolutionary interest (D. discoideum, C. elegans)
Exploring evolution
• The comparison of gene sequences and genomes has provided information over the last half century about the evolutionary relationship of man to his environment:
• LUCA
• Ancient
• Position on the tree of life (evolutionary trees)
• The closest evolutionary relative of man is the chimpanzee
• Neanderthals, Denisovans and Homo sapiens
• Why we care about always keeping an evolutionary eye as doctors

Copying DNA
• The mechanism of replication in prokaryotic and eukaryotic organisms and comparison
• The enzymes involved and their functions:
• DNA polymerase I, II, III, E. coli
• Primase, ligase, major eukaryotic DNA polymerases
• Enzymatic editing of the reading by polymerases
• PCNA as a prognostic marker of cancer
• The etiology of Huntington's disease
• Helicases and syndromes
• Type I and II topoisomerases: functions and drugs
• Telomerase: the ribonucleoprotein with reverse transcriptase activity and its importance

DNA repair and recombination
Creating DNA damage:
• Autogenous or spontaneous
-Identity of bases
-Polymerase errors
-Products of metabolism
• Foreigners
A) Chemical mutagens
a) analogous bases
b) modification of bases
c) interfering factors
B) radiations
a) ionizing
b) ultraviolet
Repair mechanisms
• A) Damage reversal
• B) - Repair of bad connections
– Base excision
– Nucleotide excision
Ames test
Recombination
– Recombinase
– Holliday structures

RNA synthesis and regulation in bacteria
Inception
An RNA polymerase: the binding sites, the p subunit
Elongation
• 8 base DNA/RNA hybrid
• the DNA 'bubble' moves
• topoisomerases are necessary
Termination
• With factor p, independent of factor p
• Transcription inhibitors-drugs
Regulation
• General for all carbohydrate operons in the presence of glucose (catalytic repression via CAP protein)
• Specific to each, the example of the lactose operon (the meaning of the operon, its regulation)

RNA synthesis and regulation in eukaryotic organisms
• The eukaryotic RNA polymerases
• Specializations (what does cis trans elements mean)
• A-amanitin: its effect on each RNA polymerase, liver disease
• Initiation of transcription: Promoters, formation of basal initiation complex and its properties, cis-elements, trans-elements
• Key transcription factors
• Role of chromatin structure: histones (details), nucleosome, regulation
• Regulation: specific transcription factors and their properties: structures
• Regulation at the chromatin level
• Histone modifications (acetylation-deacetylation-drugs)
• Nucleosome rearrangements
• Example of chromatin remodeling

RNA maturation and regulation
• Cover
• Polyadenylation
• Splitting
• Breakpoints between introns/exons are selected with absolute precision
• The mechanism
• The splicing particle
• Alternative splicing

mRNA processing
Protein synthesis (Translation) and post-transcriptional regulation
• Ribosome
secretory pathway of proteins
• Transfer RNA (tRNA)
Wobble
• Aa-tRNA synthetases
their importance for translation accuracy (editing)
• Protein Synthesis Mechanism
-Inception
• Opening complex
• Eukaryotic-prokaryotic differences
-Elongation
Elongation factors and their functions
-Termination
• What is the fidelity (accuracy) of protein synthesis
• Antibiotics, diseases, toxins
• Post-transcriptional regulation. Generally
• the example of iron homeostasis
• miRNAs

Intracellular signaling
• The ternary system of hormone message transduction with G protein-coupled membrane receptors and the mechanism of action of cholera toxin.
• The second messengers: characteristics, who they are, how they are produced, how they act, how they are degraded.
• The action of adenylate cyclase: cyclic AMP, activation of PKA.
• for the action of phospholipase Cβ: IP3 and diacylglycerol, Ca2+ and calmodulin, activation of PKC.
• Examples of hormone receptors / growth factors that have tyrosine kinase activity.
• Receptors acting through JAKs-STATs: growth hormone, α-interferon
• Brief description of the EGF signal transduction and the ras-MAP kinase pathway as well as the correlation between the signal transduction pathways and cancer.
• Examples of signaling pathway defects, oncogenes and toxins that can lead to cancer and other diseases.
• The targeting of signaling pathways to treat pathological conditions.

Biochemical Endocrinology
• Peptido-hormones and Steroid hormones: characteristics of structure, synthesis, maturation and transport of some hypothalamic hormones.
• Hormones of the hypothalamus, pituitary gland, endocrine glands: regulation of the circuit through retrograde feeding.
• The regulation of steroid hormone secretion: ACTH action on the production of cortisol from the adrenal cortex, FSH and LH action on the production of testosterone, estradiol and progesterone from the gonads, multiple regulation of aldosterone secretion from the adrenal cortex.
• Clinical implications in Cushing's syndrome, Addison's disease and Graves' disease.
• Steroid hormone synthesis disorders due to hereditary lack of enzymes. 21α-hydroxylase deficiency. Clinical implications.
• The steroid hormone receptor family: structure and function (transcriptional coactivators).
• The connection of hormone action with signal transduction mechanisms and their targeting to treat pathological conditions.

Biochemistry of sensory organs
 The nicotinic and muscarinic acetylcholine receptors: structure, special structural features and mode of action.
 Biochemical basis of vision. Operation of rods, cones and differentials.
 Relationship between structure and function of rhodopsin, conversion of photons into a nerve signal and the adaptation of rods to light
 Genetic basis of color vision and color blindness
 Biochemical basis of olfaction and osmogen signal transduction
 Biochemical basis of taste and receptors of taste molecules
 Biochemical basis of hearing.
 Biochemical basis of touch.

Reminder of metabolic pathways - Vitamins
• Vitamins: their meaning and importance
• Classification of vitamins and connection with metabolism
• Stimulation test
• Biochemical explanation of potential deficiency and disease for each vitamin
• The characteristic participation of each vitamin in the functioning of the body
• Conditions and possibilities of vitamin deficiencies in Western culture

Hormonal regulation of metabolism
• Blood glucose levels remain stable in circulation
• The use of glucose by the body
• Insulin: role, synthesis, maturation and secretion
• receptor: structure, activation, signal transduction
• actions on the metabolism of sugars, lipids, proteins
• Glucagon: structure, action, receptor
• Production of ketone bodies in chronic starvation (secretion of glucagon)
• The biochemical basis of TDM1 (Diabetes Type 1) and TDM2. Similarities and differences, therapeutic management.
• Hypoglycemia
• About leptin: where is it produced, where does it act, how does it affect the body's fuel and energy balance

Biochemistry of the liver
• The functions of the liver as a central metabolic organ and its communication with other tissues
• The functions of the liver as a metabolic organ of amino acids and proteins
• The functions of the liver as a metabolic organ of nitrogen and urea
• Liver functions in biotransformation, drug metabolism and detoxification
• The functions of the liver in the synthesis and degradation of glycogen as well as in the maintenance of blood glucose levels
• The functions of the liver in heme metabolism
• The functions of the liver in the metabolism of bilirubin
• Assessment of liver function through biochemical tests
• The importance of metabolic mechanisms, their understanding and their applications in clinical diagnosis and practice.

Blood biochemistry
A. Biochemistry of Plasma Proteins
• Composition of plasma and its difference from serum.
• The functions and electrophoretic sorting of plasma proteins, acute phase proteins and C-reactive protein
• The nature and biological role of albumin, α1-antitrypsin, ceruloplasmin, transferrin and γ-globins and clinical implications of disturbances in the levels or activity of these proteins.
• Characteristic changes in electrophoretic separation patterns of plasma proteins – indication of pathological conditions.
• The enzymes normally contained in plasma and the diagnostic value of enzymes of tissue origin (transaminases, LDH, CPK) and their isozymes.
B. Biochemistry of Hemostasis
• The role of platelets in blood clotting and which substances promote their activation and adhesion.
• The role of phospholipase A2, cyclooxygenase and thromboxane synthase in blood coagulation and the mechanism of action of aspirin.
• the nature of blood clotting factors..
• The structure of fibrinogen and the polymerization stages of cross-linked fibrin clot (thrombin, transglutaminase action).
• Limitation of coagulation: antithrombin III and the role of heparin
• The mechanism of fibrinolysis: tissue plasminogen activator and plasmin.
• Coagulation disorders due to genetic deficiency of factors (hemophilia) and for the therapeutic administration of recombinant factors.
• Connecting the mechanisms of hemostasis with cell signaling.

• Functions of adipose tissue
• Adipose tissue in general: distribution in the body, morphology, separation between white and gray.
• Growth and differentiation of adipose tissue cells. Molecular differentiation, PPARγ and coactivators.
• The biochemical function of WAT as a storage tissue.
• The sensitivity and response of adipose tissue to nerve stimuli.
• The function of adipose tissue as an endocrine organ: Leptin, actions, adiponectin.
• Adipose tissue in relation to inflammation
• Adipose tissue in relation to obesity
• Gray adipose tissue.
• The action of PGC-1 and UCP-1.
• The biochemical function of BAT as thermoregulatory tissue

Nutrition and metabolism I
• Calorie content of macromolecules.
• The body's energy balance. The concept of BMP.
• In general about macromolecular food components.
• Especially for carbohydrates
• Content of specific foods in specific carbohydrates
• Reminder of the metabolic pathways of fructose and galactose
• Especially for fats
• Essential fatty acids, foods that contain them, the biochemical basis of their deficiency, their functions, symptoms and frequency of deficiency.
• Trace elements
• Normal diet and nutrition. Food preservatives
• Digestion of carbohydrates, fats and proteins.

Nutrition and metabolism II
• The evolution of human nutrition. Relation to 'diseases of culture'.
• The metabolic state of the organism during feeding, early starvation, prolonged starvation, refeeding.
• Obesity:
o As a result of overnutrition.
o As a genetic disease.
o As an inflammatory disease.
o Leptin resistance.
o Insulin resistance.
o The relationship between obesity and diabetes.
• Lipodystrophy.
• Slimming diets.
• Anorexia nervosa, bulimia.
• Malnutrition

Functions of the muscle
• Muscle contraction and the proteins involved: Actin, myosin, tropomyosin, troponin, sarcoplasmic Ca++ pump, taso-dependent Ca++ channel.
• The muscle's stores of ATP.
• The role of creatine phosphate.
• The types of muscle fibers, their function and their importance.
• Myocyte differentiation, myogenic regulatory factors.
• Satellite cells, their activation factors.
• Muscle tissue also contains intracellular fat droplets. (IMTG- intramuscular triglycerides).
• Muscle tissue is important for movement, thermogenesis and the body's glucose homeostasis.
• Muscle fuel, what factors affect it.

Biochemistry of exercise I & II
Generally for exercise.
• At the start of the exercise. The role of adenylate kinase (myokinase), fuels and their flow. Exercise parameters.
• High intensity exercise.
• Duration exercise:
o Metabolism of free amino acids.
o Metabolic changes in duration exercise, when increasing either duration or intensity or both.
• Muscle hypertrophy: when it happens, molecular mechanism.
• Mitochondrial biogenesis: when it occurs, molecular mechanism.
• Muscle fibers interconvert depending on the exercise state of the muscle.
• The importance of the transcriptional coactivator PGC-1α for metabolism.
• Kama:
o Regional kama (in the muscle): causes
o Central stroke (in the brain): definition, possible causes.
o Disadvantages and advantages of physical exercise