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PHASE 1 Revision
Akanksha SinhaRegina Sarbaratnam
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• METABOLISM(Carbohydrates, Protein, Lipid, Alcohol, Glycogen)
• ATP(ATP-ADP cycle, Glycolysis, Kreb’s cycle, Oxidative Phosphorylation)
• FATTY ACID OXIDATION/KETONES(Beta oxidation, Ketogenesis)
• ACID/BASE/BUFFER (Henderson-Hasselbach equation)
• OXYGEN TOXICITY (ROS, Haber-Weiss and Fenton reactions)
• HISTOLOGY (Stains, Epithelia)
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Aims - Overview
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Metabolism The chemical processes that occur within a living organism in order to maintain life
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Metabolism 4 main energy sources:
Carbohydrates: Starch, fructose, sucrose, glucose, lactose Mainly come from starchy foods and fruit.
Proteins: Largely come from meat ,dairy ,pulses , fish
Alcohol: typically ethanol
Lipids: triacylglycerols
4kcal/g
4kcal/g
7kcal/g
9kcal/g
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What happens to the excess energy intake? 1. Stored as triglycerides (approximately 15kg)
2. Stored as glycogen (approximately 200g in liver, 150g in muscle)
3. Stored as protein (approximately 6kg)
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BMR
Energy needed to stay alive at rest
Increase BMR:•Increased body weight •Hyperthyroidism•Low ambient temperature•Fever/infection•Caffeine/stimulant intake•Pregnancy
Decrease BMR:•Increased age•Being female •Dieting/Starvation•Hypothyroidism
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ATP
ATP ‘Adenosine Triphosphate’ Structure components: Adenine, Ribose, 3 Phosphate
How does it provide Energy?
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ATP
-The ATP molecule has two phosphoanhydride bonds.
-These are RELATIVELY WEAK bonds, hence require relatively less energy to break
-Overall more energy is released in forming the products that used to break bonds in the reactants
ATP
ADP
Hydrolysis
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ATP-ADP Cycle ATP
ADP + Pi
Respiration:Energy productionCarbohydrateLipidProtein
Energy utilization:Biosynthesis of macromoleculesMuscle contractionActive ion transportThermogenesis
CO2
O2
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ATP- ways to generate ATP? - Glycolysis- Kreb’s cycle- Oxidative phosphorylation- Substrate level phosphorylation- Via electron transport chain- Beta oxidation (Discussed in fatty acid oxidation)
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Glycolysis• GlycolysisPathway of 10 stepsTakes place in cytoplasmIn short – Glucose 2Pyruvate + 2ATP + 2NADH (AEROBIC)2Lactate + 2ATP (ANAEROBIC)
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Glycolysis Energy Investment Phase
- 2 ATP
Energy Generation Phase
+ 4 ATP
Glycolysis generates a NET 2 ATP plus 2 NADH
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• Facilitated diffusion of Glucose into cell mediated by Insulin
• Phosphorylation of Glucose by Hexokinase• -ATP
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Step 1
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• Isomerism• G6PF6P
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Step 2
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• F6P F1,6BP• Via phosphofructokinase (PFK is primary
regulated step of glycolysis; responds to both cellular energy and hormonal regulation)
• -ATP
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Step 3**
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• 6: NADH (x2) produced• 7: ATP (x2) produced• 8:• 9:• 10: 2Pyruvate + ATP (x2) produced• Remember all the enzymes• “Good Gracious, Father Franklin Did Go By Picking
Pumpkins (to) Prepare Pies
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Step 6-10
Pyruvate enters the TCA cycle
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• PFK1 is pH dependent– Inhibited in acidosis
Control of glycolysis via:
• Step 1• Step 3* -key regulatory step in glycolysis
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Acidosis affecting glycolysis
Positive regulators: AMP Negative regulators: H+, ATP, Citrate, hormonal
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Kreb’s/ TCA cycle
In Mitochondrial matrixAcetyl CoA can come from glycolysis/ fatty acid breakdown/ amino acids’ carbon skeletons
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Kreb’s/TCA cycle • Inhibited
by ATP, NADH, succinyl CoA
• Activated by ADP
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Electron Transport Chain• The electron transport chain (ETC) is a
process in which the NADH and [FADH2] produced during glycolysis, β-oxidation, and other catabolic processes are oxidized
thus releasing energy in the form of ATP. The mechanism by which ATP is formed in the ETC is called chemiosmotic phosphorylation
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The chemiosmotic theory Based on the following:-inner mitochondrial membrane is impermeable to protons, hence the mitochondrial matrix is a closed environment-the proton pumping of the ETC leads to the proton motive force which in turn provides the energy for ATP synthesis (Proton flow through the ATP synthase protein drives ATP synthesis)
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The chemiosmotic theory
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Substrate level phosphorylation • some ATP can be made in the cytoplasm
through a process called substrate-level phosphorylation
• With this type of phosphorylation you have an adenosine diphosphate (ADP), which is a unit of adenosine attached to two (di) phosphate groups
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oxidative phosphorylation • Oxidative phosphorylation is the metabolic
pathway in which the mitochondria in cells use their structure, enzymes, and energy released by the oxidation of nutrients to reform ATP
(involves ETC!!!)
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TOTAL ATP • Total ATP made from one molecule of glucose:34 ATP
But this is very debatable- Some sources will say 30 or 32 ATP
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Fatty acid oxidation/ Beta oxidation Examples of fatty acids: •Palmitoleic acid•Linoleic acid•Palmitic acid
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Fatty acid oxidation/ Beta oxidation
Fatty Acid Acetyl CoA (used in TCA/Krebs cycle)
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Fatty acid oxidation/ Beta oxidation
Fatty Acid Acyl adenylate
ATP PPi
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Fatty acid oxidation/ Beta oxidation
Acyl adenylate
Acyl CoA
CoA AMP
Acyl CoA synthetase
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Fatty acid oxidation/ Beta oxidation
The carnitine shuttle !!!
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The Carnitine Shuttle
Acyl CoA Acyl Carnitine
Carnitine CoA
Carnitine Acyl Transferase I
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Inner mitochondrial membrane
Acyl Carnitine
Acyl CoA
CoA Carnitine
Carnitine Acyl Transferase II
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Fatty acid oxidation/ Beta oxidation
Acyl CoA Acyl CoA – 2C
Acetyl CoA
• Sequential removal of 2- carbon units • Each round of β-oxidation produces: 1 mole of NADH 1 mole of FADH2 1 mole of acetyl-CoA
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Ketogenesis The process by which ketone bodies are produced as a result of fatty acid breakdown.
Ketones are used for energy at low fat states.
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Ketogenesis 2 Acetyl-CoAthiolase CoA-SH
Acetoacetyl-CoAHMG-CoA synthase
Acetyl-CoA + H20
CoA-SH
HMG-CoA HMG-CoA lyase Acetyl-CoA
Acetoacetate
Acetone β- hydroxybutyrate
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I know, no pathology … Diabetic ketoacidosis
• Reduced supply of glucose due to a significant decline in circulating insulin and an associated increased in circulating glucagon.
• The increased production of Acetyl-CoA leads to ketone body production
• This lowers the pH of the blood
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Acid/Base/Buffer • Acid = proton (H+) donor
HA H⇔ + + A-
• Base = proton (H+) acceptor
B + H+ BH⇔ +
• Buffer = weak acids or bases that act to maintain H+ concentration
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Acid/base/buffer • Acidosis can either be respiratory or metabolic• Alkalosis can either be respiratory or
metabolic
• pH range is 7.35-7.45. But ideal pH is 7.4
• Compensation: metabolic is slow, respiratory is fast.
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Henderson-Hasselbalch equation • Henderson’s formula:- [H+ ] x [HCO3 - ] = K x pCO2
• Hasselbalch converted it to pH pH = pK + log ([HCO3 - ] / [CO2])
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• Respiratory• Insufficient ventilation retains CO2
so there is more carbonic acid in the blood leading to acidosis
• Causes include COPD (chronic bronchitis + emphysema), asthma
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Acidosis
• Metabolic• Low bicarbonate levels leading to
acidosis• Causes include bicarbonate losses
from the GI tract e.g. diarrhoea, DKA, lactic acidosis (anaerobic respiration), renal failure
Carbonic anhydrase
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Alkalosis
• Respiratory• Blowing off too much CO2 leading
to alkalosis• Causes hyperventilation, pink
puffers COPD (i.e. type 1 respiratory failure), anxiety, fever
• Metabolic• Loss of H+ or too much
bicarbonate leading to alkalosis• gastric secretions contain large
quantities of hydrogen ions.• Causes include excess vomiting
(because you’re losing H+), pyloric stenosis, anorexia nervosa, ingestion of bicarbonate
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Anion Gap The anion gap is calculated by subtracting the serum concentrations of ANIONS (Cl- and HCO3-) from the concentrations of CATIONS (Na+ and K+)
BUT
potassium concentrations, being very low, usually have little effect on the calculated gap. This leaves the following equation:
Anion gap = [Na+] − ([Cl−] + [HCO3−]) =16 meq/lit
The magnitude of this difference (i.e., "gap") in the serum is often calculated when attempting to identify the cause of metabolic acidosis.
If the gap is greater than normal, then high anion gap metabolic acidosis is diagnosed (due to increased cation: H+)
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• Proteins (51%)– Weak acid/base groups
• Bicarbonate (43%)– CO2 removed via lungs, Bicarbonate regenerated
by kidneys
• Haemoglobin (6%)– Deoxy Hb binds H+ HHb + Bicarbonate– Amine group on Deoxy Hb bind CO2
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Types of Buffer systems
Most important
Haemoglobin binds both CO2 and H+ and so is a powerful buffer
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Oxygen Toxicity Toxic reactive oxygen derivatives with:•Free radicals•Unpaired electrons in outer shell•High reactivity
ORDEF: Chemically reactive molecules with unpaired electrons in outer shell & free radicals (unpaired electron) derived from oxygen
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Formation of ROS
O2
Oxygen
O2
Superoxide
H2O2
Hydrogen peroxide
OH• + H2O
Hydroxyl radical
H2O
e- e-, 2H+
e-, H+
e-, H+
Produced by ETC
Generates radicals with transition metals (Fenton or Haber Weiss rx)
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OH• (hydroxyl radical)•Most potent radical•Lipid soluble•Chain reaction forms lipid peroxides & organic radicals
H202 (hydrogen peroxide)•Not actually a radical•Oxidising agent in presence of Fe2+ or other transition metals•Generates hydroxyl radical•Lipid soluble
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ROS
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Formation of hydroxyl radical
H2O2
H2O2Fe2+
Fe3+
OH•
OH-
+
+
+
+
+
+
O2
H2O
OH•
H+
O2
-
Haber-Weiss reaction
Fenton reaction
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• Endogenous – natural by-product of O2 metabolism– Produced mainly inside cell organelles ie.
(mitochondria)
• Exogenous– UV radiation, smoking, inflammation
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Production of ROS
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• Respiratory bursts– ROS released during phagocytosis of bacteria– Damages bacterial cell membrane– Fenton’s reaction
• O2 O2-H2O2+Fe2+Fe3+ + OH- + OH•
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+ve ROS
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• Cellular damage• Damage to membranes of nucleus,
mitochondria, endoplasmic reticulum & cell• Increased permeability leads to influx of
calcium, water & sodium
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-ve ROS
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• Enzymes– Catalase H2O2 2 H20 + 02– Superoxide dismutase O2- H2O2– Glutathione peroxidase
• Antioxidant vitamins/Free radical scavengers (vitC, vitE, carotenoids)
• Cellular Compartmentation• Repair
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Anti-oxidants
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Histology of Epithelia • Roles of epithelia - Protection(skin) - Absorption(gut)- Secretion(pancreas)
Simple Epithelium Stratified Epithelium-Almost always squamous
Pseudo-stratified Epithelium
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Simple squamous epithelia
Found in: -Alveoli -Serosa
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Simple cuboidal epithelia
Found in: -Kidney tubules -Ducts of glandsSweat Salivary Pancreatic
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Simple columnar epithelia
Found in: -Small intestine -Gall bladder -Bronchus
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Protection •Continually being worn down•Cells replaced from below•Found in areas where there is continuous abrasion
Squamous:Keratinised – skinNon keratinised – mouth, oesophagus, vagina
Cuboidalfound in some large ducts
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Stratified epithelium
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Stratified squamous non-keratinising
Found in:-Mouth-Oropharynx-Oesophagus -Vagina
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Found in: -Skin (hairy or non-hairy skin)
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Stratified squamous keratinising
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Found in:•Trachea and large airways (respiratory epithelium) •Urinary tract
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Pseudostratified columnar epithelia
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1) Adherent (tight) junctions
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Cell Junctions
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2) Desmosome
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Cell Junctions
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3) Gap junction
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Cell Junctions
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Staining
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Alcian Blue
GAG-rich structuresMucous goblet cellsMast cell granulesCartilage matrix
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Staining
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Eosin Colloidal proteins (e.g. plasma)Keratin
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Staining
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Haematoxylin
NucleiRNA
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Staining
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PAS Hexose sugars
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Questions?