Digestive Nutrition

Cardiovascular diseases:

 Atherosclerosis: the narrowing or blockage of arterial blood vessels by the formation of plagues
- building up of cholesterol particles in blood leads to deposition on the lining of the artery
- macrophages in vascular wall take up cholesterol by endocytosis and become cholesterol laden foam cells and initiate formation of atheromatous plaques
- high blood cholesterol content will drive this process
 Risk factors of atherosclerosis: usually factors which cause damage to the vascular endothelium allowing macrophages to infiltrate
- family history
- smoking
- hypertension
- lipid profile
- diabetes
- age
 Triglyeride: three fatty acids bonded to a glyerol center. The macromolecule is nonpolar hence will not dissolve in plasma but is carried around by amphipathic cholesterol and phospholipids packages called lipoprotein.
 Lipoproteins:
- surface: monolayer of phospholipids, cholesterol, apoproteins
- core: triglyeride stabilized by cholesterol-esters (cholesterol with its polar group removed)
- apoproteins on the lipoprotein have specific functions such as enzyme activation, structural maintenance and receptor binding.
 Types of lipoproteins:
- Chylomicrons: largest least dense lipoprotein consisting of 90% TG and 4% cholesterol. Has apoproteins Apo B-48,Apo CII, and Apo E
- VLDL: very low density lipoprotein consisting of 60% TG and 20% cholesterol. Has apoproteins Apo B-100, Apo C-II and Apo E
- LDL: low density lipoprotein consisting of 5% TG and 50% cholesterol. Has apoproteins Apo B-100
- HDL: high density lipoprotein consisting of 5% TG and 20% cholesterol. Has apoproteins Apo-A, Apo C-I and Apo E
 Functions of apoproteins:
- Apo A: structural roles and activates lecithin-cholesterol acyl transferase (LCAT)
- Apo B: structural roles (Apo B-100 can also mediate binding to LDL receptors)
- Apo C: activates lipoprotein lipases (Apo C-I activates LCAT)
- Apo E: binds to receptors and facilitate uptake
 Exogenous pathway: lipid absorption and transport pathway from GI tract
- lipids from the GI is absorbed through small intestine into blood streams and packaged as chylomicrons
- lipoprotein lipase on adipose tissue capillary endothelium rapidly hydrolyze and absorb the TG
- the cholesterol remnants is cleared from the plasma into the liver overnight
 Endogenous pathway:
- TG synthesized in the liver by carbohydrates and exported as VLDL to tissues in blood
- Hydrolysis by lipoprotein lipases leaves VLDL remnants called IDL, and is rapidly reabsorpted by the liver receptors
- LDL, the final stage of “used” VLDL carrying mostly cholesterol, has no Apo E hence is uptaken into the liver slowly mediated by LDL receptors only.
- Accumulation of LDL in blood leads to atheroma plaque formation
 Reverse cholesterol transport pathway:
- HDL mediates reverse cholesterol transport to remove cholesterol from peripheral tissues
- Apo C-I on HDL activates LCAT and converts cholesterol to cholesterol-ester
- The cholesterol-ester are carried and donated by HDL to CM remnants or IDL to be taken into the liver and excreted (as bile)
- HDL is anti-atherogenic and beneficial
 Friedwald formulae: LDL cholesterol = total chol – HDL chol – TG/2.2 (TG/2.2 gives a rough estimation of VLDL chol and formulae is no valid if TG>4.5 mmol/L)
 Healthy plasma levels:
- total chol <5 mmol/L
- total chol and HDL ratio <5 mmol/L
- TG <3 mmol/L
- LDL chol <2 mmol/L
- HDL chol >1 mmol/L
 Primary lipid disorder: genetic defects in lipoprotein metabolism
 Secondary lipid disorder: genetic predisposition plus triggering primary disorder factors
- Obesity, type 2 diabetes and metabolic syndrome: total chol no change; TG increase; HDL decrease
- Chronic renal failure: total chol no change or increase, TG increase, HDL decrease
- Uncontrolled diabetes mellitus: total chol increase, TG increase, HDL decrease
- Nephritic syndrome: total chol increase, TG increase, HDL decrease
- Obstructive jaundice: total chol increase, TG no change, HDL no change
- Hypothyroidism: total chol increase, TG no change, HDL no change
- Heavy alcohol intake: total chol no change, TG increase, HDL increase
 Familial hypercholesterolemia: mutation of the LDL receptor on liver prevents absorption of LDL and cause accumulation of cholesterol in blood. Occurs from early age with 9-12 mmol/L of blood cholesterol in heterozygotes, leading to myocardial infraction at age 30-50
- symptoms: tendon xanthomata resulting from collection of cholesterol plaques (atherosclerosis) in blood vessels around tendon and hence inflammation
- treatment: aggressive treatment with mainly drugs and diet from early on
 Familial binding-defective ApoB: mutation of the Apo B-100 also prevents binding to LDL receptors hence produce a very similar phenotype to familial hypercholesterolemia
 Common hypercholesterolemia: polygenic genetic defect causing elevated levels of cholesterol (up to 10 mmol/L).
- symptoms: no tendon xanthomata but increase CV disease risk
- treatment: diet mainly and optional drug therapy
 Familial dysbetalipoproteinemia: mutation of Apo E protein in homozygous patients impairs the clearance of IDL by Apo E receptor and cause elevation in both TG and cholesterol. The disease is also termed type III mixed hyperlipidemia and is highly atherogenic
- symptoms: palmar crease xanthomata and tuberous xanthomata
- treatment: very diet-responsive
- on a protein electrophoresis, the bands of LDL and VLDL appear merged due to IDL existing as an intermediate
 Familial chylomicronemia: rare autosomal recessive disorders where either lipoprotein lipase or Apo C-II is mutated, which leads to failure of CM clearance from the blood and elevating TG levels.
- symptoms: eruptive xanthomata and pancreatitis
- treatment: extreme low-fat diet (drugs are ineffective)
 Familial hypertriglyceridemia: gene defect in addition to a lifestyle factor (e.g. obesity, alcohol) which causes increased VLDL in the blood. Symptoms are similar to that of chylomicronemia.
 Lipoprotein little A: an apoprotein homologous to plasminogen that in abnormal cases exists on LDL particles and may enhance the cloth formation on atheroma plaque. It’s a possible inhibitor of plasminogen in breaking down clots and cause CVD.
- the plasma level of Apo (a) are genetically determined and not affected by diet or drugs
 Treatment of lipoprotein disorders:
- treat exacerbating conditions such as diabetes, hypertension etc
- lifestyle changes
- diet
- drug therapy
 Drug therapy
- statins: HMG-CoA reductase inhibitor that prevents synthesis of cholesterol
- fibrate: decrease TG and increase HDL through complex mechanism
- cholestyramine: non-absorbable resin that binds to cholesterol and bile salts in gut and prevent absorption
- niacin: high does lower cholesterol
- ezetimibe: cholesterol absorption inhibitor
 Homocysteine: amino acids associated with increase CVD risk. It forms disulphide with protein and damage endothelium of vessels.

Carbohydrate metabolism and diabetes:

 Glycogenolysis: break down of glycogen during fasting in liver muscle and kidney and stimulated by glucagon and adrenalin
 Gluconeogenesis: production of glucose from non-sugar precursors such as fat, protein. Occurs during fast in liver and kidney and stimulated by low level of insulin.
 Lipolysis: breaking down triglyceride into free fatty acid and stimulated by low level of insulin. Free fatty acid. The fatty acid can later on participate in beta oxidation in muscle, brain etc or ketogenesis in liver and partially kidney.
 Insulin: hormone of anabolic reaction in metabolism, i.e. switches off all pathways of the fasting state. Stimulated release by glucose or drugs sulphonylurea.
- features: formed from the beta cells of pancreatic Islets of Langerhans as pro-insulin, consisting of c-peptide chain and insulin. It is later cleaved to activate insulin. Because C-peptide half life is longer, its level is blood is higher and useful to make measurements.
 Regulatory hormone of catabolic pathway:
- glucagons: stimulated in low glucose and acts to increase glycogenolysis, gluconeogenesis, ketogenesis, lipolysis
- adrenalin: produced in the adrenal medulla and increase glycogenolysis and lipolysis
- growth hormone: made in the pituitary and increases glycogenolysis and lipolysis
- cortisol: made in the adrenal cortex and increase gluconeogenesis and proteolysis
 Type 1 diabetes: autoimmune destruction of pancreatic beta cells and produce insulin deficiency (low levels of insulin and c-peptide).
- autoantibodies involves to beta-cell protein are anti-GAD (glutamic acid decarboxylase), anti-IA2 (islet antigen 2)
- associated with human leukocyte antigen DR3 and DR4 (gene coding for cell surface antigen presenting protein)
- Family history of diabetes in only a minority of cases and usually presents in childhood or young adulthood
 Type 2 diabetes: non-insulin dependent disorder relating to insulin-resistance and often due to factors such as obesity linked with strong genetic components and family history.
- insulin resistance: fault in the signal transduction pathway of insulin i.e. tyrosine kinase receptor. Consequently though insulin is present, its effect can not be produced (e.g. stimulate glucose transporters - hyperglycemia)
- insulin resistance occurs in early stage causes high fasting insulin levels but eventually beta cells burns out and fails
 Symptoms of diabetes: ineffectiveness of insulin due to whether low levels or block in transduction pathway will send a wrong signal to liver, adipose tissue and muscles etc and respond as if the body were in fasting state
- catabolic pathways are activated
- uncontrolled lipolysis (severe in type 1) producing weight loss and ketoacidosis
- excess proteolysis and gluconeogenesis produce weight loss and hyperglycemia
- hyperglycemia leads on to glucosuria, osmotic diuresis (increase urine production), thirst and eventually hypotension and dehydration
- ultimately drowsiness, coma and death
 Method of testing:
- hyponatremia: lowering of Na+ concentration due to osmotic shift of water out of cells
- hyperkalemia: increasing K+ concentration due to displacement of intracellular K+ by H+ of acidosis
- low bicarbonate
 Long term complication of diabetes: complication directly related to glycemic control
- diabetic microangiopathy: nephropathy (renal failure), retinopathy (blindness), neuropathy
- atherosclerosis: strongest known risk factor for atherosclerosis
- coronary peripheral vascular and cerebrovascular disease constitutional
 Laboratory tests:
- blood glucose: normal below 6.1, impaired 6.1 – 6.9 and diabetic above 7.0
- urine glucose: plasma glucose above 10 and glucose appears in urine. However this is not an accurate indication of diabetes because individuals may have lower renal threshold, e.g. during pregnancy or kidney problems
- plasma ketone
- glycated haemoglobin: glucose can attach spontaneous to haemoglobin covalently and degrades the RBC. This test provides an index of long term glycemic control
- glucose tolerance test: giving an oral glucose load around 75g and measure plasma glucose for 2 hours after an overnight fast. Normal is below 7.8, impaired is 7.8 – 11.1 and diabetic is above 11.1.
 Treatment of diabetes:
- diet and lifestyle: obesity causes insulin resistance so exercise and avoid large amount of sugar
- oral and antidiabetic drug: stimulate insulin secretion (sulphonylureas) and improve insulin sensitivity (metformin, thiazolidinediones)
- insulin: achieve balance between good control and risk of hypoglycemia

Protein and Nitrogen Balance

 Nitrogen sources: mainly amino acids from dietary proteins
 Nitrogen balance: rate of nitrogen intake (diet) minus the rate of nitrogen expenditure (excretion). Positive balance leads to net protein synthesis and deposition while negative means overall protein breakdown.
- minimum intake to maintain nitrogen equilibrium is 30-50g of high quality protein
 Factors affecting nitrogen balance:
- positive: increase protein intake, growth and pregnancy, recovery from illness
- negative: decrease protein intake, starvation or reduce GI function, injury (trauma), sickness (infections), cancer, lactation
 Adaptation: when amount of ingested protein falls below normal limits, there is an initially period of negative nitrogen balance, but later the liver and other organs adapt to restore nitrogen balance (e.g. lower enzyme production) Due to this short-term balance studies are very unreliable.
 Turn over: about 1-3% of total body protein is normally degraded each day (300g in the endogenous amino acid pool – figure is higher due to sickness). In addition 100g of amino acid from diet producing a total of 400g to be incorporated into protein.
 Excess amino acid: amino acids can not be stored by are deaminated to give ammonia and residual carbon skeletons. Urea detoxify ammonia keeping its concentration low (<20 umol/l)
- main forms of nitrogen excretion: urea (4.2-6.5mmol/l), uric acid (0.1-0.3mmol/l), and creatinine (80-150umol/l)
- residual carbon skeleton can be used to oxidize to produce energy, or used in gluconeogenesis
 Non-essential amino acids:
- alanine
- asparagines
- aspartic acid
- glutamine
- glutamic acid
- glycine
- proline
- serine
 Essential amino acids: any other amino acids with few special exceptions
- Arginine and histidine are semi-essential as the body have synthetic pathway but too low to support optimal growth
- Hydrolysine found as a post-translational modification in collagen and elastin
- Methionine can be provided by alternative sulfur and methyl group sources (Cysteine and choline respectively)
- Tyrosine can be made from phenylalanine
 Biological value of protein: the fraction of the absorbed nitrogen which is converted to tissue protein. Amino acids are only incorporated into tissue protein when sufficient of each amino acid is present. The limiting amino acid govern the extent to which the other amino acids in dietary protein can be incorporated as it’s present in the smallest amount
- absorption and subsequent incorporation of amino acids into protein is essentially complete within a few hours of a meal. Excessive amino acid can provide a source of glucose
 Net protein utilization: fraction of retained nitrogen in tissue to that of the ingested nitrogen. This index takes into account of the possible partial digestibility of the protein.
 Protein complementary: “low quality” proteins each with a different limiting amino acid can be ingested together (within a very short space of time) to compensate for each other’s deficit. The result comparable to that of high quality protein ingestion.
 Activities of amino acid catabolism enzyme fall during protein depletion but increase during starvation where glucose is needed for brain.
 Change in protein requirement: protein requirement declines throughout life, i.e. newborn to elderly. The rate of turnover primary governs the nutritional requirement. During pregnancy and lactation, protein requirement is increase 20 to 25%
 Protein-calorie malnutrition:
- kwashiorkor: disorder that results due to energy intake from solely the carbohydrate of starchy food with no protein
- marasmus: disorder of frank starvation and grossly inadequate diet
 Kwashiorkor: excessive glucose supply causes over-secretion of insulin and impairs starvation response. Consequently lipolysis is retarded and adipose tissue preserved, amino acid redistribution from muscle to liver is prevented and plasma amino acids are broken down and protein synthesis rate falls (e.g. hypoalbuminaemia)
- symptoms: early symptoms involve fatigue, irritability and growth failure and loss of muscle mass as deprivation continues. Dermatitis and pigmentation change (vitiligo) and edema is also seen. Later on, a swollen abdomen and reddish discoloration of hair and skin results and affect mental development and growth.
 Marasmus: utter starvation brings about stunted growth, loss of adipose tissue and generalized wasting f lean body mass without edema
 Treatments of PCM:
- reintroduction of food slowly with carbohydrate followed by protein foods. Vitamin and supplements are essential
- early treatment is important as later stages of kwashiorkor leaves the child permanently mentally and physically disabled

Water soluble vitamins

 Vitamins: organic molecules that acts as co-factors for enzymatic reactions in the body. These are generally not synthesized by mammalian cells and must be supplied in the diet
 Water soluble vitamins:
- ascorbic acid (C): antioxidant and hydroxylation reaction of collagen. Deficiency causes scurvy
- cobalamin (B12): converts homocysteine to methionine and methylmalonyl CoA to succinyl CoA. Deficiency causes pernicious anemia
- Thiamin (B1): oxidative decarboxylations and transketolases. Deficiency causes Wernicke’s encephalopathy and Beri Beri
- Riboflavin (B2): redox reactions (oxidative phosphorylation)
- Niacin (B3): redox reactions (oxidative phosphorylation). Deficiency causes pellagra
- Panthothenic acid (B5): part of coenzyme A for acyl activation and transfer.
- Pyridoxine: amino acid metabolism and phosphorylase modification
- Folate: one carbon transfers. Deficiency causes megaloblastic anemia and homocysteinuria
 Dietary requirement: as the bodies have pathways for degrading them or mechanism of excreting them, therefore the required intake of a vitamin is determined by its rate of degradation or excretion.
 Vitamin storage: vitamins such as thiamin and vitamin C are hardly stored while vitamins such as retinol and ester are stored in substantial amounts. Time until onset of deficiency symptoms is hence body stores/ (daily rate of degradation and excretion).
 Enzymatic malfunctioning: vitamin deficiency will obviously impair enzymes in which they are the co-factors. However sensitivity to decline in the vitamin differ for each enzymes because:
- presence of uptake mechanism for the vitamin of the enzyme in the sub-cellular compartment, i.e. uptake can maintain concentration
- different localization of enzyme and hence different concentration of the vitamin, i.e. some compartment have higher concentration
- differences in Km of enzyme for the co-factor, i.e. enzyme of low Km can still function under low concentration
 Absorption and transport of vitamins:
- water soluble: digested by mucosal hydrolases and absorbed into mucosal cells, subjected to modification and transported to hepatic portal vein to reach liver.
- Lipid soluble: similar process to triglyceride where initially they form minor components of micelles. Their fate after uptake into mucosal cells is to be exported in the lymphatic system and ultimately the plasma as chylomicrons. Stored as body lipids, e.g. fatty acid esters.
 Thiamin deficiency: illness Beri Beri that affects alcoholics and polished grains eaters of 3rd world countries where the grain lack the germ that is rich in thiamine. Severe reduction in cells’ capacity to generate energy
- characterized by neurological symptoms and disordered muscle function and accumulation of lactate and pyruvate in plasma, leading to vasodilation and edema.
- Malabsorption, malnutrition, alcohol and low folate levels, reduced stomach acidity (by antacids etc) all contribute to thiamin deficiency
- Symptoms: earliest symptoms include constipation, appetite suppression, nausea and mental depression. Chronic thiamin deficiency leads to more severe neurological problems such as ataxia, mental confusion and loss of eye co-ordination.
 Dry beri-beri: anorexia, anxiety states and weakness of muscles
 Wet beri-beri: more severe progressed form with cardiac failure, edema and at times sudden death.
- edema: build up of fluid in body tissue due to dilation of peripheral blood vessels caused by accumulated lactate and fluid leaks through the capillaries
 Dietary requirement of thiamin: requirement for thiamine is proportional to the caloric intake of the diet and increased with fever, heavy exercise or high carbohydrate intake
- range from 1.0 – 1.5 mg per day for normal adults and excessive thiamine is excreted in urine
- dietary sources: red meat, whole grains, potatoes, nuts
 Ruldoph Peter’s experiment: pigeons fed with polished grain caused convulsions and acute lesion in the brain, which was reversed by administration of thiamin. The plasma of the animal was high with pyruvate and lactate. The brains of the pigeon used oxygen at a much slower rate than that of normal.
 Thiamin pyrophosphate: active form of thiamin, converted in the brain by thiamin diphosphotransferase
- essential for the activity of enzymes in aerobic oxidation of glucose including pyruvate dehydrogenase, ketoglutarate dehydrogenase and transketolase
- aerobic oxidation: the role of thiamin is to provide a reaction carbon on the thiazole ring that forms a carbonion with alpha keto acids (e.g. pyruvate) when the COOH is liberated as CO2. Carbonyl group such as SCOA can then be added on.
- Transketolase: a carrier of the glycoaldehyde and catalyzing its addition to an aldehyde recipient
- Amino acid decarboxylation: keto aci d such as leucine, isoleucine and valine
 Testing for thiamin deficiency:
- plasma pyruvic acid measurement after exercise (where respiration becomes obvious and important) or oral glucose load
- measurement of transketolase activity in RBC with and without TPP in thiamin deficiency (activity is increase by 25% with TPP)
 Wernicke-Korsakoff syndrome: neuronal loss, gliosis (proliferation of the glial connective tissue cells of the CNS) and vascular damage in regions surrounding the third and fourth ventricles and cerebral aqueduct.
- Cause: increased Km¬ ¬of transketolase for the cofactor thiamine pyrophosphate and/or changed isoelectric forms that impair reactions of the pentose phosphate pathway.
- It is commonly seen in alcoholic patients and also patient with impaired nutrition

Iron uptake transport and storage:

 Iron storage: 60% haemoglobin, 9% myoglobin,, 0.4% cytochrome and remainder 30% are non-haem iron is mostly stored in liver, bone marrow and muscles.
 Iron deficiency: microcytic anaemia where concentration of RBC does not change but they become smaller containing less hemoglobin
 Dietary intake: of a dietary intake of 14mg per day of Fe, only 0.9 mg is absorbed in males and 1.7mg in female which the rest appears in faeces (shedding of GI epithelium cells containing absorbed iron is a major cause)
 Iron loss:
- desquamation: skin, alimentary tract, urinary tract (0.9mg per day in both males and females)
- menstruation: average of 20mL blood cells per menstrual cycle hence 0.8mg per day
- blood loss: ulceration, hemorrhoids and fighting
 Iron requirement: females have almost twice the iron requirement of males due to menstruation, pregnancy where iron is transported to fetus and lactation where iron is given to the baby in milk
 Iron absorption:
- Iron in the GI lumen exists in the ferric form. Fe3+ reducing agents, such as ascorbic acid reduces it to ferrous Fe2+ state to allow absorption into the villi.
- Heme Fe can also be absorbed directly
- In the intestinal cell, the iron is attached with an amino acid and then can be either stored as ferritin (in the Fe3+ state) or it is transported transcellularly to blood.
 Iron transport:
- as iron enters blood circulation it is oxidized by caeruloplasmin, a copper-containing protein of the plasma, to ferric iron
- the iron is carried in the circulation very tightly bound to transferrin
- transferrin: a large molecular weight of 77000 prevents filtration in the kidney. Can bind to 2 Fe3+ molecules and usually 30% saturated. In acidity conditions, affinity of transferrin for iron decreases (bicarbonate being a counter-iron for binding)
 Cellular iron uptake:
- specific transferrin receptor facilitate iron uptake bounded to transferrin. These are present in large amounts on reticulocytes and placental trophoblast cells
- binding of iron-bound transferrin to the receptor initiate a temperature and energy dependent endocytosis of the plasma membrane around and the receptor-complex is taken into the cell
- endosomal acidification lowers pH causing the iron to detach from the transferrin-receptor complex. The iron proceeds intracellularly to either mitochondrion for heme production or ferritin as storage.
- The iron free apotransferrin that remains on the receptor at pH 5.5 is returned to the cell surface where at pH 7.4 the apotransferrin is released. Receptor and transferring are recycled
 Ferritin: large protein consisting of 24 identical units that forms a cage and capable of carrying up to 4500 Fe atoms.
 Mechanism of ferritin control:
- induction and repression of ferritin is affected at the level of mRNA stability
- iron-response element binding protein (IRE-BP) in the cell binds to iron but binds instead to an iron responsive element (IRE) on the mRNA when iron is absent. IRE-BP is a closely related to aconitase
- Low intracellular Fe: IRE-BP binds to IRE on the mRNA of both ferritin (5` head end) and transferrin receptor (3` tail end). As a result translation of ferritin is repressed while transferrin receptor is enhanced


 Prevalence of obesity: 50% of population in 2003 is overweight
 Postulated causes: the key cause of obesity is disregulation of energy intake to expenditure, i.e. energy balance.
- low metabolic rate
- brow fat hypothesis
- monogenetic defect
- appetite regulation
- inactivity
 Low metabolic rate: measured in a whole body calorimetry for the oxygen and carbon dioxide exchange. Metabolic rate is dependent on body size and composition (fat:lean tissue ratio) and is rarely the primary cause of obesity because energy expense for obese people is actually higher
 Brown-fat hypothesis: brown fat in the body is metabolically active (contains many mitochondria) but only comprises of less than 2% of total body fat in adults and is physiologically unimportant
 Monogenetic effect: an example is Ob gene which produced the protein product Leptin. In mice deficiency of leptin (recessive ob gene) leads to obesity produced obesity. Leptin functions to regulate (decrease) appetite and metabolism. In obese humans, excess leptin are produced (hyperleptin) to counteract rise in body energy intake. Humans are generally heterozygous for ob and recessive homozygous are very rare.
 Appetite regulation: the regulation of appetite is under polygenic control. A number of environmental factors such as macronutrient composition of food, family history, exercise.
- a high fat diet results in a much higher energy intake than a high CHO diet. Study where subjects are provided with fat composition-altered but otherwise identical meal and allowed to eat to appetite, those that eat the high fat meals have greater energy intake (e.g. hyperphagia)
- Fat and CHO oxidation rates are quite different. Carbohydrates are rapidly oxidized after it is eaten while fat is preferentially stored in adipose tissue contributing to overweightness

B12 and Folic acid

 Symptoms of deficiency of folic acid:
- megaloblastic anaemia where the bone marrow cells have large reticulated nuclei and the presence in plasma of macrocytic red blood cells.
- This is due to precursor bone marrow progenitor cells unable to divide properly during maturation to generate nuclei of new erythrocytes with inadequacy of DNA synthesis
- Folic acid is an antianaemia factor and cures megaloblastic anaemias of pregnancy, malnutrition and malabsorption
 Mechanism of deficiency: lack of folate lead to impairment in the dTMP synthesis which leads to cell cycle arrest in S-phase of rapidly proliferating cells, especially hematopoietic cells
 Primary folate deficiency: deficiency of folate due to inadequate intake from food. This is rare as folate is normally enough in food.
- causes: chronic alcoholism, impaired absorption or metabolism, increase demand for the vitamin, pregnancy due to an increased number of rapidly proliferating cells in the fetus.
 Spina bifida: increased requirement in early pregnancy causes folic acid deficiency and inadequate intake at time of conception predisposes neural tube defects in fetal development. Taking supplement can reduce the incidence to half of the present rate.
 Folic acid sources: liver, spinach, fresh vegetables, exists mainly as polyglu folic acid. In the intestine, free folic acid is converted to N5 methyl THF.
 Tetrahydrofolic acid: the active form of folic acid that functions as a 1-C carrier unit at various oxidation levels.
- N5-N10-methylene-THF: the most significant form of THF as it’s the prescursor for the synthesis of dTMP from dUMP required for DNA production.
- N5-methyl-THF: the main form of folate in the body but also the “sink” for folic acid as reactions producing it is irreversible. Requires the reaction of homocysteine to methionine to escape sink.
 The folic acid cycle in DNA synthesis: conversion of 2-deoxyuridine monophosphate to 2-deoxythymidine monophosphate for DNA synthesis.
- The reaction involves adding a methyl group on to the uridine’s pyrimidine ring while N5-N10-methylene THF is catalyzed by thymidylate synthetase to DHF.
- DHF is then recycled to tetrahydrofolate by NADPH dependent reduction catalyzed by dihydrofolate reductase.
- THF then acquires a CH2 from serine to regenerates N5-N10-methylene THF while serine is converted to glycine and water as a result. Reaction catalyzed by serine hydroxymethyl transferase
 Trimethoprim: selective inhibitor folate reductase in gram-negative bacteria and has little effect on mammalian enzymes.
 Methotrexate: antagonist of the folate reductase converting DHF to THF as it has a similar structure to THF and can be used as an anticancer drug.
 Vitamin B12 deficiency (pernicious anaemia): secondary folate deficiency with remarkably similar symptoms to folate deficiency. Chief effect is megaloblastic anemia due to failure to synthesize dTMP but RNA synthesis is normal. However in addition there are neurological problems to B12 deficiency as it is required for myelin synthesis in nervous tissue
- cause: destruction (usually autoimmune) of the parietal cells of gastric mucosa that secrete both acid and intrinsic factor for uptake of vitamin B12. Hence absorption of B¬12 is impaired.
 Vitamin B¬12 sources: liver, meat, eggs, diary products.
 Function of B12:
- homocysteine methyl transferase: B12 is a co-factor for an important enzyme that transfers methyl group from N5-methyl-THF on to homocysteine to produce methionine and THF. As most folic acid is trapped in the N5-methyl-THF form, this reaction is the only route by which THF can be restored to continue DNA synthesis.
 Treatment of pernicious anaemia: the megaloblastic anemia that results is in effect also due to a deficiency of THF that is tied up as the useless N5-methyl-THF (in the absence of the transferase).
- Temporary treatment providing folate can relieve the deficiency but eventually, all of it will be converted to N5-methyl-THF (reduced from N5-N10-methylene-THF) and become unavailable.
- Sulphonamides: analogues of para-amino benzoic acid that inhibit DNA synthesis selectively in bacteria and lower folic acid intake from bacterial sources.

Pernicious Anaemia

 Pernicious anaemia: autoimmune destruction of parietal cells and the loss of gastric acid and intrinsic factor secretion produce vitamin B12 deficiency.
 Problems with serum B12 test:
- Assay not reliable
- No valid normal range
- Result may not be clinically relevant, i.e. many patient have low serum level that has no functional meaning
 Requirement for normal B12 absorption
- acid to release food-bound cobalamin
- secretion of intrinsic factor from healthy parietal cells for absorption by specialized receptors in terminal ileum
- normal pancreatic secretion of enzymes to help release B12
- normal ileal function
 Autoimmune gastritis:
- Evidence: antibodies to parietal cells, antibodies to intrinsic factor, inflammation in the stomach (biopsy), evidence of low acid output, raised plasma gastrin, other autoimmune disease
- Clinical manifestation: tiredness, difficulty concentration, weight loss and diarrhea, hypochromic and macrocytic blood cells, but iron and folate levels are normal
- Treatment: B12 replacement by parenteral injections. Monitor response to B12 replacement.
 Surgically removing ileum: loss of specialized receptor on terminal ileum, leads to failure to absorb B12 and bile salts. There is irritant effect of bile salts on colon and impaired fat absorption due to decreased bile salt pool
- Evidence: past medical surgery where 85 cm of ileum was removed due to cancer
- Clinical manifestation: diarrhea, bowel motions 5 times per day, tiredness, hypochromic and low B12 levels, high faecal fat
- Treatment: parenteral replacement
 Gastrectomy: loss of gastric acid and intrinsic acid secretion due to atrophic gastritis. Failure of normal stimulation of pancreatic secretion and mixing of pancreatic juice with food
- Evidence: past medical surgery
- Clinical manifestation: tiredness, mild anaemia, low iron stores, low B12 and vitamin D
 Coeliac disease: loss of small bowel villi along with the endocrine cells that secrete secretin and CCK which stimulate pancreatic secretion. Sever cases involve damage of ileal villi and less ileal receptors
- Evidence: results from duodenal biopsy
- Clinical manifestation: tiredness, abdominal bloating and low B12, folate and iron levels

Vitamin K

 Sources of vitamin K: vitamin K exist naturally as K1 (phytylmenaquinone) in green vegetable and K2 in intestinal bacteria, which have different chemical structures. Half of our requirement comes from the bacteria.
 Absorption of vitamin K: naturally occurring vitamin K is absorbed from the intestines only in the presence of bile salts and other lipids through interaction with chylomicrons. Hence fat malabsorptive disease leads to vitamin K deficiency.
 Vitamin K deficiency: Hemorrhagic syndrome is the primary result.
- malnutrition can deplete body reserve in 2 days
- long term antibiotic treatment can kill intestinal bacteria
 Vitamin K function: co-factor for the enzyme that catalyze the carboxylation of specific Glu residues to gamma-carboxy glutamate (Gla). These occur in microsomes of most cells but high activity in liver hepatocytes. Proteins that must have carboxylated Gla residues for activity include blood clotting factors and osteocalcin.
 Clotting factors:
- factor IX (Christmas factor) and factor X (Stewart factor) require glutamate carboxylation for activation. Factor X, when activated, converts prothrombin to thrombin by cleaving it in two locations.
- Factor II (prothrombin) is formed from the carboxylation of specific glu in the first 30 residues of the amino-terminus of preprothrombin to gla. The thrombin segment on the prothrombin is called prethrombin.
- Gla residues on thrombin are able to bind to Ca2+ and this facilitates binding to the negatively charged surface of phospholipid layer (Ca2+ bridge). Thrombin converts fibrinogen to an active form that assembles fibrin and also activates factor VIII (a transglutaminase which catalyze formation of covalent bond between lys and arg to stabilize fibrin clot), protein C (negative feedback protein that deactivates factor VIII and V),
 Vitamin K cycle:
- Carboxylation reaction of the glutamate residue using O2 and CO2 converts the reduced vitamin K hydroquinone to vitamin K 2,3-epoxide, catalyzed by vitamin K gamma-glutamyl carboxylase/epoxidase.
- Vitamin K 2,3-epoxide is reduced by dithiol-dependent epoxide reductase to vitamin K quinone
- Vitamin K quinone is finally reduced to regenerate hydroquinone by quinone reductase using NAD(P)H.
- The three enzymes that catalyze this cycle are all found on the rough ER
 Poison: warfarin or dicoumarol are vita K antagonists that inhibit the two reduction reaction for Vitamin K hydroquinone regeneration and thus inhibit carboxylation, clotting, leading to uncontrollable haemorrhage.
 Osteocalcin: protein secreted by the osteoblasts that functions to collect calcium using the Gla residues and deposit them to form the hydroxyapatite crystal matrix of the bone.
- fetal warfarin syndrome: treatment with anticoagulants during pregnancy can lead to abnormalities of fetal bone
 Role of Ca2+: clotting factors requires Ca2+ in order to anchor phospholipid and carry out their function. Hence sodium citrate, substance that removes Ca2+ from the blood, is able to prevent clotting.
 Vitamin K and neonates:
- lack of vitamin K diet for a newborn whose intestines is sterile
- vitamin K does not cross placenta
- low prothrombin production in immature liver
- low vitamin K content for human milk
- healing time of umbilicus is used to diagnose vitamin K deficiency
- Treatment: A prophylactic dose of 0.5 – 1.0 mg of phylloquinone is usually given orally and shortly after birth to prevent vitamin K deficiency

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