PHM142 Lecture 2: Modulation of Oxygen Transport PDF

PHM142 Lecture 2: Modulation of Oxygen Transport, Keeping RBC Oxidation in Check **Isohydric Transport** - Major CO2 transport pathway to remove CO2 from actively respiring tissues to the lungs for excretion. - In actively respiring tissues, CO2 is produced and diffuses out down its...

PHM142 Lecture 2: Modulation of Oxygen Transport, Keeping RBC Oxidation in Check **Isohydric Transport** - Major CO2 transport pathway to remove CO2 from actively respiring tissues to the lungs for excretion. - In actively respiring tissues, CO2 is produced and diffuses out down its concentration gradient into the blood where it enters RBCs. - CO2 reacts with H2O to form H2CO3 which decomposes to HCO3- and H+. - HCO3- is transported out of the RBC in exchange for Cl- via a bicarbonate-chloride transporter down their respective concentration gradients. - Therefore, in venous blood, there is a higher concentration of bicarbonate ions as it is effluxed out of RBCs and a reduced concentration of Cl- as it being taken in. - The purpose of the exchange of HCO3- and Cl- is to maintain electric neutrality so as to not depolarize cell membranes. - The H+ binds with Hb via His residues which will stabilize the deoxy (T) state of Hb and decreases the binding affinity to O2 to help offload more O2 in the active tissue. - When blood enters the lungs where pO2 is high, HCO3- in the blood enters RBCs in exchange for Cl- (in the opposite direction now) where it acted on by CA to form CO2 and H2O which diffuses out into the lung tissue to be respired out. The H+ required to make H2CO3 is provided by Hb-H and Hb reverts to its unprotonated state so as to increase affinity for O2 (good when in lungs). Why important? Opioid overdose, respiratory acidosis, anaesthetics. **2,3 Bisphosphoglycerate (2,3BPG or 2,3DPG)** - 2,3BPG is made as a side reaction of glycolysis. One of the intermediate products of glycolysis (1,3BPG) is shunted to the side via bisphosphoglycerate mutase which shifts the position of one of the phosphates to create 2,3BPG. Note that 2,3BPG can still be converted back into 1,3BPG to feed into glycolysis if needed by the action of 2,3 bisphosphoglycerate phosphatase. - 2,3BPG binds to deoxy Hb (thus stabilizing deoxy form) through ionic cross-linking of beta chains to form salt bridges which decreases O2 affinity. This increases O2 release. - Why the need to convert from 1,3 to 2,3? For differentiating different molecules for different purposes. Body wants to use 2,3BPG for O2 off-loading and not take away 1,3BPG from energy-producing glycolytic pathway. - In fetal-Hb, there is a lower affinity for 2,3BPG to ensure that O2 affinity in fetal Hb is always higher and they are priority for O2 supply. - Defects in glycolytic enzymes? May alter production of BPG such that the rightward shift of binding curve for Hb does not occur and tissue oxygenation is compromised. Think about hexokinase and pyruvate kinase. - 5 negative charges on 2,3BPG form ionic bonds with +ve charged amino acid residues of Hb like K and H. Coordination stoichiometry is 1:1 meaning 1 BPG molecule to 1 Hb. - Binds tightly to deoxy Hb and weakly to oxyHb. - In high altitudes, the body compromises for the lower pO2 level of the environment by doubling the amount of 2,3BPG (in a process called acclimation). This increases the P50 (lower affinity) to offload O2 more efficiently and prevent hypoxia. - Why? Function of 2,3BPG is to give body greater control of tissue O2 demand and supply. Allows for adaptation to environment. **Energy Metabolism in Erythrocytes** - Our tissues love to use carbohydrates as main source of ATP production. It's easy to react on, is a reduced carbon source and thus have many avenues of oxidation pathways that body can utilize to generate ATP. - In the fed state, our body can store glucose molecules by forming a long polymer chain stored predominantly in muscle, liver and adipose. As the body shifts to the fasted state (24h), our body begins to breakdown these glycogen storages to provide glucose for the body. - The liver can create new glucose molecules from non-carbohydrate sources like proteins and glycerol through gluconeogenesis (liver). However, during this phase, some organs still use glucose as main energy source like the brain, RBCs and bone marrow, WBCs and renal medulla. - In the prolonged starvation phase, ketone bodies form from beta oxidation of fatty acids. The brain is able to use ketone bodies for energy. Finally, when stores are depleted, proteins are the last source to provide energy and muscles give up their protein storage (muscle wasting) to prevent death. **Glycolysis** - 10-step anaerobic metabolic pathway that takes a molecule of glucose and oxidize it into two 3C pyruvate molecules, forming 2 ATP and 2NADH. - Dominant form of ATP production in RBCs. - The pathway is ensured to be forward moving by steps 1,3 and 9 because these reactions cost ATP and are irreversible steps. - In RBCs, ATP has to be used sparingly because it does not have an efficient way to generate lots of it. Therefore, it uses ATP for its most important cellular processes like: 1. Na+/K+ ATP-ase: Pumps 3 Na+ out for 2K+ in to maintain proper electrochemical gradient of the cell membrane. If RBCs cannot maintain proper electrochemical gradient, the membrane will become compromised and cell cannot function. RBCs will swell when the Na+ going in is greater than K+ going out and shrink when Na+ going in is less than K+ going out. Digitalis and ouabain are inhibitors of this protein pump. Digoxin is a drug used to increase contractility by greater influx of Na+ in heart. 2. Ca2+ ATP-ase: Maintains Ca2+ gradient (Extremely low inside compared to outside) because Ca2+ accumulation will compromise membrane integrity, and cell volume and cell rheology. Ca2+ accumulation changes the cell shape and ridity occurs to form echinocytes (ages the RBC). RBCs then need to be removed. - Glycolysis also forms NADH which is not used for ox-phos but instead used for an enzyme called methemoglobin reductase to convert the oxidized methemoglobin Fe3+ back into oxyhemoglobin Fe2+. First the oxidation of NADH to NAD+ allows the reduction of FMN to FMNH2. This is done by the nicotinamide adenine dinuc and flavin mononuc. Then, FMNH2 can then be used to reduce Fe3+ to Fe2+ by being oxidized back into FMN. - The oxidation of hemoglobin Fe occurs when it is in high O2 environments or exposed to oxidizing agents. **Pentose Monophosphate Shunt** - Supplies cells with a source of NADPH. NADPH is a source of reducing power for synthesis biochemical reactions (like making fatty acids for example). Not used in energy metabolic pathways. - NADPH is also a source of antioxidation by providing a source of protons to antioxidizing molecules. - Glucose-6-Phosphate dehydrogenase (G6PD) converts G6P from glycolysis to 6-Phosphogluconate by removing a hydrogen (oxidation) and in turn reduces a molecule of NADP+ to NADPH. - 6-Phosphogluconate can get oxidized even further by 6-Phosphogluconate dehydrogenase which generates another molecule of NADPH and is converted to ribulose-5-phosphate. - NADPH is used as a reducing agent to reduce GSSG (oxidized form of GSH) into GSH. This is catalyzed by glutathione reductase. - GSH is our body's source of antioxidants. GSH gets used up and oxidized to GSSG when it reacts with oxidizing agents (ROS) like H2O2, superoxide anion (O2rad-), hydroxide ion (OH-). - H2O2 can get reduced to H2O by glutathione peroxidase using up GSH. Or by catalase using a molecule of NADPH to form O2 and H2O. - Oxidation of Hb forms MetHb and also the superoxide anion which can get antioxidized by superoxide dismutase to form H2O2 and then through the actions of GSH or NADPH (enzymes above) to convert to water. - Selenium is an important inorganic metal required for proper glutathione peroxidase activity. - Ribulose-5-P from the conversion of 6-phosphogluconate by 6-phosphpgluconate dehydrogenase can be converted into ribose-5P or xylulose-5P. - Xylulose and Ribose combine together through action of transketolase to form glyceraldehyde (3C) and sedoheptulose (7C) and then transaldolase converted it to fructose (6C) and erythrose (4C). - Separately, xylulose and erythrose can combine through transketolase to form glyceraldehyde (3C) and fructose (6C) which can feed back into glycolysis. - Stoichiometry: 3R5Ps (15) form 2F6P (12) and 1G3P (3). **Drugs and Toxins Affecting Erythrocyte Function** 1. Form complexes with Hb 2. Oxidize Hb Fe 3. Hemolysis 4. Genetic Diseases on Enzymes - O2 cannot bind to Hb whose Fe have been oxidized to Fe3+. - During cyanide poisoning, the cytochrome oxidase Fe3+ centre binds CN and will disrupt electron transport in oxidative phosphorylation leading to death. An acute antidote uses amyl+Na nitrite to convert the Fe2+ centres of Hb to Fe3+ and since we have so much Hb molecules in our blood, CN will bind to them instead of cytochrome oxidase. - MetHb can be induced by: aniline drugs (dapsone), nitro aromatic drugs, hydrazine, oxidants, chlorates, nitirites, quinones, naphthalene, benzene, arsine. - MetHb patients -- what can you give? Leucomethylene blue is a source of oxidizing power that can oxidize NADPH to NADP+ to reduce Fe3+ to Fe2+. Note: That this is different from the Methemoglobin reductase using NADH from glycolysis. But need to see if patient G6PD status is OK, if mutated, patient can't make NADPH to begin with. - ROS like H2O2 will attack cell membrane of cells including RBCs. This ultimately destroys the RBC and leads to anemia. - People with G6PD deficiency canot make NADPH and therefore the antioxidant processes of their body is compromised. This will lead to an increased concentration of ROS which damage RBCs. - Therefore, drugs that may induce hemolysis like aromatic amines, nitro compounds, hydrazines, antimalarial drugs should not be given to people with G6PD deficiencies due to lack of antioxidant activity. **Muscle Fibres and Oxidative Metabolism** - Fast-twitch muscle fibres use primarily anaerobic respiration to fuel their ATP demand. They fatigue quickly as glycolysis in an anaerobic environment will produce lactic acid and until the lactic acid can get oxidized back into pyruvate. Glycogen content higher (better for anaerobic respiration) and mitochondria content lower (less need for ox.phos.) Little myoglobin (anaerobic). - Slow-twitch muscle fibres are used for sustained activity and do not fatigue easily. They derive energy from oxidative phosphorylation thus have high perfusion of blood vessels, high mitochondria content and a large amount of myoglobin. **Malignant Hyperthermia** - Ryanodine receptor in muscle cells are responsible for initiating the release of Ca2+ ions from the SR for muscle contraction after an AP is sent from CNS. - Mutations of the Ryr receptor is a contraindication for some anaesthetics. Anesthetic will cause excessive Ca2+ to be released from SR in skeletal muscle muscle use. - Active muscles will generate heat and lactic acid which will cause acidosis and death. - Reversible in the first few mins. Start cooling and administer dantrolene. Dantrolene is a ryanodine receptor antagonist which blocks further Ca2+ from being released. Thus it relaxes muscles. - Anesthetics also can cause hepatic damage because 20% of the metabolism is toxic. Can also cause spontaneous abortion in those pregnant.