L9 Metabolism Lipogenesis, lipolysis and ketogenesis

Overview

This lecture covers the role of fat in energy homeostasis through three linked processes: lipogenesis (fat storage in the fed state), lipolysis (mobilisation of stored fat in fasting and stress) and ketogenesis (conversion of excess acetyl CoA into ketone bodies). The organising principle is insulin: insulin drives lipogenesis and suppresses lipolysis, and the switch between the two is executed at the level of gene transcription (SREBP-1c, ChREBP), allosteric and covalent enzyme control (acetyl CoA carboxylase, AMPK) and mitochondrial fatty acid entry (malonyl CoA inhibition of CPT1). The clinical payoff is understanding why insulin resistance produces fatty liver and hyperlipidaemia, and why ketoacidosis is a feature of type 1 rather than type 2 diabetes.

Definitions

• Lipogenesis: term for both fatty acid synthesis and TAG (triacylglycerol) formation.
• De novo lipogenesis: lipogenesis when the process starts from acetyl CoA.
• Lipolysis: cleavage of TAGs with mobilisation of fatty acids. It is a fuel mobilisation step, occurring when fat needs to be metabolised for energy or glycerol is needed for gluconeogenesis, i.e. the fasted state, starved state and trauma. It yields glycerol and free fatty acids (FFAs).
• $\beta$-oxidation: fatty acid breakdown to acetyl CoA units, NADH and FADH${}_2$, with the acetyl CoA entering the TCA cycle.
• Ketogenesis: generation of the ketone bodies acetoacetate and $\beta$-hydroxybutyrate.

Why it matters: excess fat in organs and within the bloodstream is disadvantageous, and excess fat stores are a risk factor for type 2 diabetes and cardiovascular disease.

Lipogenesis in the fed state

Lipogenesis has two components: TAG synthesis (glycerol-3-phosphate combining with fatty acids) and fatty acid synthesis (acetyl CoA as the precursor to fatty acids). The overall direction is acetyl CoA $\to$ fatty acids $\to$ TAGs.

Liver, fed state (Lippincott’s Fig. 24.3): glucose from the gut enters the hepatocyte and is stored as glycogen, shunted into the HMP (pentose phosphate) pathway, or taken through glucose-6-phosphate to pyruvate. Pyruvate feeds the TCA cycle (which also exchanges with amino acids from the gut, with NH${}_3$ release and protein interconversion) and is converted to acetyl CoA, which is used for fatty acid synthesis. Fatty acids also arrive as chylomicron remnants from the blood. Fatty acids are esterified to triacylglycerol, packaged into VLDL and exported to the blood for delivery to adipose tissue.

Two points about hepatic glucose handling:

• Because of an abundance of GLUT-2 glucose transporters, glucose uptake by the hepatocyte is not rate limiting.
• The liver responds to high blood glucose by increasing phosphorylation of glucose by glucokinase, which has a high $K_m$ for glucose.

When does de novo lipogenesis actually run?

• Lipogenesis from excess dietary carbohydrate occurs mainly in the liver, but also in adipose tissue.
• On most Western diets, dietary fat delivery to liver and adipose tissue suppresses de novo lipogenesis.
• De novo lipogenesis operates significantly when carbohydrate intake exceeds energy requirements and glycogen stores are full.

Adipose tissue, fed state: insulin stimulates GLUT4-mediated glucose uptake; glucose $\to$ glucose-6-phosphate $\to$ DHAP $\to$ glycerol-3-phosphate, which combines with fatty acids (via fatty acyl CoA) to form stored TAG. TAG arriving in chylomicrons (from gut) and VLDL (from liver) is hydrolysed at the cell surface by lipoprotein lipase (LPL), releasing fatty acids into the adipocyte, with lipoprotein remnants and glycerol returning to the liver. The two hormonal controls are:

• Insulin stimulates LPL (fat uptake into adipose).
• Insulin inhibits HSL (hormone sensitive lipase), so TAG breakdown and FFA/glycerol release are suppressed in the fed state.

The de novo lipogenesis pathway in liver

Ordered steps (critical enzymes in bold):

1. Glucose $\to$ glucokinase $\to$ glycolysis.
2. Glycolysis $\to$ DHAP (which branches off to glycerol-3-phosphate) and onward to phosphoenolpyruvate $\to$ pyruvate via pyruvate kinase.
3. Pyruvate enters the mitochondrion and is converted by pyruvate dehydrogenase (PDH) to acetyl CoA.
4. Acetyl CoA condenses with OAA to form citrate. Acetyl CoA itself is trapped in the mitochondrion, so citrate is exported instead.
5. In the cytoplasm, citrate lyase cleaves citrate back to OAA + acetyl CoA.
6. Acetyl CoA carboxylase (ACC) carboxylates acetyl CoA to malonyl CoA. This is the first step of fatty acid synthesis: acetyl CoA + CO${}_2$ + ATP, with biotin as cofactor, gives malonyl CoA + ADP + P${}_i$.
7. Fatty acid synthase, using NADPH, produces palmitate, which is activated to fatty acyl CoA (FACoA).
8. FACoA combines with glycerol-3-phosphate (derived from DHAP) to form TG, which with apoproteins and other lipids is packaged as VLDL and exported to the blood.

TAGs are made in the liver and exported as VLDL for storage in adipose tissue.

Regulation of lipogenesis

Transcriptional (the dominant control). Transcription of all the enzymes critical to lipogenesis is increased by glucose and insulin, which are elevated by a high carbohydrate diet. Specifically:

• Glucokinase: synthesis stimulated by insulin, and activity additionally stimulated allosterically by elevated glucose (two separate mechanisms).
• Pyruvate kinase and pyruvate dehydrogenase: synthesis increased by insulin.
• Citrate lyase, acetyl CoA carboxylase and fatty acid synthetase: synthesis increased by insulin.

The transcription factors. SREBP-1c and ChREBP are activated by insulin and glucose respectively, and both bind response elements in lipogenic genes:

• Insulin acts on the insulin receptor, raising glucokinase and increasing SREBP-1c, which binds the SRE on lipogenic enzyme genes, increasing lipogenesis. Glucagon and PUFAs act negatively at the receptor.
• Glucose enters via its transporter and is converted to G6P, which activates ChREBP, which binds the ChoRE on the same lipogenic enzyme genes.

Allosteric and covalent control of ACC.

• Stimulated by citrate and by insulin (dephosphorylation).
• Inhibited by long chain fatty acyl CoA and by glucagon (phosphorylation).

Blocking $\beta$-oxidation so that lipogenesis is productive. For lipogenesis to be effective it is necessary to inhibit $\beta$-oxidation and to inhibit AMPK. In the fed state:

1. Activation of ACC elevates malonyl CoA.
2. Malonyl CoA potently inhibits CPT1 (carnitine palmitoyl transferase).
3. Entry of fatty acyl groups into the mitochondrion is thereby blocked.
4. $\beta$-oxidation of fat is inhibited.

AMPK, the cellular fuel gauge. When AMPK is activated, lipogenesis is blocked; when AMPK is inactive, lipogenesis is no longer blocked, so AMPK is downregulated during lipogenesis. When AMP levels rise, AMPK switches off energy-expensive pathways (lipogenesis) and switches on energy-producing pathways ($\beta$-oxidation). Rising AMP inhibits fat synthesis; rising ATP promotes fat synthesis. Active (phosphorylated) AMPK:

• Turns ON glycolysis, glucose uptake, HK, GLUT4 and UCP3 in skeletal muscle.
• Decreases SREBP, turning OFF gluconeogenic enzyme expression (PEPCK, G6Pase) and lipogenic gene expression.
• Phosphorylates HMG-CoA reductase, turning OFF cholesterol synthesis.
• Phosphorylates ACC, turning OFF fatty acid synthesis.
• Phosphorylates MCD, turning ON fat oxidation in skeletal muscle.

Fatty liver and its clinical significance

Obesity has overtaken alcohol as the number one cause of liver disease in Canada. Over 1.4 million Canadians have non-alcoholic fatty liver disease (NAFLD), and 15% of obese children have NAFLD. The progression is NAFLD $\to$ non-alcoholic steatohepatitis (NASH) $\to$ cirrhosis $\to$ death. Macroscopically a fatty liver is pale/yellowish against the darker red-brown of normal liver.

Case study. A lawyer presented with tiredness and feeling unwell for some time, initially attributed to work pressure, and was overweight with BMI 31.8.

• U&E and LFTs: sodium 141 (134 to 144 mmol/L), potassium 3.7 (3.5 to 5.0), urea 4.5 (2.6 to 6.8), creatinine 101 (50 to 110), albumin 44 (30 to 50 g/L), protein 82 (60 to 80 g/L). Abnormal and highlighted: ALP 175 (40 to 130 U/L), ALT 110 (8 to 40 U/L), GGTP 105 (6 to 58 U/L). Amylase 103 (25 to 125 U/L) was normal. Total and direct bilirubin were not reported.
• Lipids and glucose, all abnormal: cholesterol 7.8 (target <4.0 mmol/L), LDL 5.3 (<2.0), HDL 0.9 (>1.0), triglyceride 4.1 (1.7), glucose 6.6 (3.5 to 5.4 mmol/L).
• Ultrasound: appearance of the liver typical for steatosis (echogenic/bright), otherwise normal abdominal ultrasound.
• Likely diagnosis: NASH, likely progressed from NAFLD.
• Potential complications: progression to fibrosis and cirrhosis.
• Treatment: diet (weight loss, cholesterol reduction), exercise, alcohol moderation, consideration of medications and further studies.

Dysregulation of lipogenesis in diabetes and insulin resistance

Hyperglycaemia and insulin resistance elevate insulin. Insulin’s inhibitory effect on lipolysis in adipose tissue is impaired, but its stimulatory effect on lipogenesis in liver is impaired less. Therefore the diabetic liver makes increased TAGs.

Sources of hepatic triglyceride, by flux: FFA arriving from subcutaneous fat via the arteriovenous route contributes about 60%, de novo lipogenesis about 25% (driven by lipogenic mRNA via SREBP-1 and ChREBP, activated by glucose and insulin), and dietary fat/chylomicron remnants plus FFAs about 15%. These converge on TG, exported as VLDL to peripheral TG stores.

Three mechanisms by which insulin resistance loads the liver with fat (Choi and Ginsberg):

1. Hepatic TG from plasma fatty acids: more FFA is freed from adipose by HSL and goes to the liver, and more VLDL is made.
2. Hepatic TG from remnant TG-fatty acid: reduced lipoprotein lipase activity means TAGs in VLDL are not properly off-loaded to adipose tissue, so more VLDL and chylomicron remnant fatty acid returns to the liver.
3. Hepatic TG from de novo lipogenesis: elevated insulin, SREBP-1c, ChREBP and PPAR$\gamma$ activity drive more lipogenesis and more VLDL output.

Fat can accumulate in the liver in this situation.

Lipolysis and its regulation

• Stimulated by: glucagon, adrenaline, TNF-$\alpha$, cortisol.
• Inhibited by: insulin.
• Dysregulation of lipolysis can be an important contributor to lipid related disorders.

Hormone sensitive lipase (HSL). In the adipocyte, stored TAG is broken down by activated HSL, releasing fatty acids and glycerol out of the cell into the blood (glycerol going to the liver). In the fasting state, glucagon stimulates lipolysis by activating HSL. In stress/trauma, adrenaline, cortisol and TNF-$\alpha$ stimulate lipolysis.

Glucagon signalling cascade in the adipocyte. Glucagon binds glucagon receptors on adipocytes, which are G-protein coupled receptors (GPCR family), and stimulates mobilisation of TAGs to provide fatty acids. Steps:

1. Glucagon binds the GPCR.
2. The G-protein ($\alpha$, $\beta$, $\gamma$ subunits) is activated.
3. Adenylate cyclase is activated, converting ATP to cAMP + PP${}_i$.
4. cAMP activates inactive A-kinase (PKA).
5. Active PKA phosphorylates inactive HSL, using ATP $\to$ ADP.
6. Phosphorylated, active HSL catalyses TAG $\to$ FFA.

Fatty acid oxidation between meals

Between meals, fatty acids are the main fuel of all aerobic tissues except brain.

1. Fatty acid utilisation is related to FFA concentration in plasma.
2. In the cytoplasm, fatty acids are activated to fatty acid acyl-CoA.
3. They are transferred into mitochondria via the carnitine palmitoyl transferase system.
4. They are oxidised by $\beta$-oxidation to acetyl CoA.
5. In the liver, excess acetyl CoA is diverted to synthesise ketone bodies, which is ketogenesis.

Fasted liver cell, the switch in sequence: glucagon inhibits acetyl CoA carboxylase $\to$ malonyl CoA levels decline $\to$ inhibition of CPT1 is relieved (the acyl-carnitine shuttle opens) $\to$ fatty acid oxidation increases $\to$ acetyl CoA levels rise $\to$ some is channelled to ketogenesis. Citrate is directed toward gluconeogenesis, and the TAG/VLDL and glycerol pathways are reduced relative to the fed state.

Carnitine.

• Made by liver and kidney, supplying heart and muscle; made from lysine and methionine; also obtained from the diet.
• Deficiency leads to muscle fatigue, hypoglycaemia and fatty liver.
• Causes of deficiency: liver disease; vegetarian diet with insufficient sources of carnitine; haemodialysis; genetic defects such as enzyme deficiency; post trauma, illness or surgery through increased loss.

Ketogenesis

Ketone synthesis occurs in the fasted or starved state, where there is increased $\beta$-oxidation and elevated acetyl CoA, some of which goes into ketogenesis, especially if the TCA cycle cannot keep up.

Synthesis in the liver. Amino acid catabolism, fatty acid oxidation and glycolysis all converge on acetyl CoA. Then:

1. 2 acetyl CoA $\to$ acetoacetyl CoA (releasing CoA).
2. Acetoacetyl CoA $\to$ 3-hydroxy-3-methylglutaryl CoA (HMG-CoA).
3. HMG-CoA $\to$ acetyl CoA + acetoacetate.
4. Acetoacetate $\to$ 3-hydroxybutyrate (using NADH + H${}^{+}$ $\to$ NAD${}^{+}$), or by a minor route $\to$ CO${}_2$ + acetone.

Use in peripheral tissues (e.g. muscle, and brain). Acetoacetate and 3-hydroxybutyrate travel in the blood to peripheral tissues, where:

1. 3-hydroxybutyrate $\to$ acetoacetate (NADH + H${}^{+}$ / NAD${}^{+}$).
2. Acetoacetate $\to$ acetoacetyl CoA via thiophorase, using succinyl CoA $\to$ succinate from the TCA cycle.
3. Acetoacetyl CoA $\to$ 2 acetyl CoA, which enters the TCA cycle for energy.

Ketone bodies as fuel. They are an important alternative fuel to glucose in starvation for brain and other aerobic tissues during fasting, thereby helping maintain glucose homeostasis. Over a 40-day fast: $\beta$-hydroxybutyrate and acetoacetate rise steeply from near zero to a plateau around 7 mM by roughly day 20; blood glucose stays flat at about 3.5 to 4 mM throughout; free fatty acids rise gradually to about 1.5; urinary ammonia rises; total urinary nitrogen starts high at about 4.7 g/day, falls sharply and levels off around 1 g/day. The brain can obtain 60% of its energy from oxidation of ketone bodies during fasting and in the neonatal period: fed brain is 100% glucose, whereas in starvation roughly one third is glucose, about 55% $\beta$-hydroxybutyrate and about 12% acetoacetate.

Plasma ketone body concentrations.

State	Concentration
Fed	< 0.1 mmol/L
Fasted (days)	0.3 mmol/L
Fasted (6 weeks)	10 mmol/L
Newborn baby	2 to 3 mmol/L
Type 1 diabetic ketoacidosis	> 30 mmol/L

Important

Fasting levels of ketone bodies in non-diabetics are much lower than those found in diabetic ketoacidosis (DKA), and DKA will not occur from fasting. In diabetics, if insufficient insulin is present, ketone bodies can be made far in excess of the body’s ability to oxidise them, and ketoacidosis may develop.

Ketogenesis in type 1 versus type 2 diabetes

Type 1 diabetes (high ketogenesis):

1. Insulin is not inhibiting lipolysis, so there is a greater supply of FFA to the liver.
2. Insulin is no longer stimulating ACC, so there is less malonyl CoA.
3. This relieves inhibition of CPT1.
4. FFA oxidation to acetyl CoA is increased.
5. Ketogenesis is increased.

Type 2 diabetes (less ketogenesis):

1. Insulin is not inhibiting lipolysis, so there is a supply of fatty acids to the liver.
2. BUT insulin is also stimulating ACC (lipogenesis), so malonyl CoA levels are elevated.
3. Elevated malonyl CoA inhibits CPT1, so $\beta$-oxidation is inhibited.
4. Insulin also stimulates re-esterification of FFA to TAG, so less acetyl CoA comes from $\beta$-oxidation.
5. Less acetyl CoA is available for ketogenesis, so there is less ketogenesis than in type 1 diabetes.

Self-test

1. Define lipogenesis and de novo lipogenesis, and state how they differ.
2. Define lipolysis and ketogenesis, naming the products of each.
3. Describe the ordered steps of hepatic de novo lipogenesis from glucose through to export into the blood.
4. Explain why citrate rather than acetyl CoA is exported from the mitochondrion during lipogenesis.
5. List the enzymes identified as critical to lipogenesis and state how insulin affects each.
6. Glucokinase is regulated by two distinct mechanisms. Describe both, and explain why glucose uptake itself is not the rate limiting step in the hepatocyte.
7. Name the two transcription factors that mediate the lipogenic response, state what activates each, and name the DNA element each binds.
8. Describe the regulation of acetyl CoA carboxylase, distinguishing the stimulatory from the inhibitory inputs and identifying which act by phosphorylation state.
9. Explain the sequence by which activation of lipogenesis in the fed state blocks $\beta$-oxidation.
10. Describe AMPK’s role as a fuel gauge, and list the pathways it switches on and off when activated.
11. Predict what happens to hepatic $\beta$-oxidation if acetyl CoA carboxylase is pharmacologically inhibited, and explain the mechanism.
12. Describe what insulin does in adipose tissue in the fed state, naming the transporter and the two lipases involved.
13. Under what dietary conditions does de novo lipogenesis operate significantly, and what normally suppresses it on a Western diet?
14. List the hormones and cytokines that stimulate lipolysis and the hormone that inhibits it, and state which stimulators act in fasting versus stress.
15. Describe the signalling steps from glucagon binding its adipocyte receptor to the breakdown of TAG.
16. Describe the steps of fatty acid utilisation between meals, from plasma FFA to acetyl CoA, and name the tissue that is the exception to using fatty acids as main fuel.
17. Where is carnitine made, from which amino acids, and what are the consequences and causes of carnitine deficiency?
18. Describe the steps of ketone body synthesis in the liver and their reconversion to acetyl CoA in peripheral tissue, naming the key enzyme of the peripheral step.
19. State the plasma ketone body concentrations in the fed state, after days of fasting, after 6 weeks of fasting, in a newborn, and in type 1 diabetic ketoacidosis.
20. Describe how blood glucose, ketone bodies, free fatty acids and total urinary nitrogen change over 40 days of fasting, and state how much of the brain’s energy ketone bodies can supply.
21. Explain why ketogenesis is greater in type 1 than in type 2 diabetes, giving the mechanism for each.
22. A 45 year old lawyer presents with several months of tiredness and a BMI of 31.8. ALP is 175 U/L, ALT 110 U/L, GGTP 105 U/L, cholesterol 7.8 mmol/L, LDL 5.3 mmol/L, HDL 0.9 mmol/L, triglyceride 4.1 mmol/L and glucose 6.6 mmol/L; abdominal ultrasound shows a bright liver. Give the likely diagnosis, the condition it progressed from, the potential complications and the treatment.
23. Integrative: explain how insulin resistance produces both a fatty liver and hyperlipidaemia, identifying the three sources of hepatic triglyceride and their relative contributions.

Answers

Reveal answers

1. Lipogenesis is the term for both fatty acid synthesis and TAG formation. De novo lipogenesis is the case where the process starts from acetyl CoA.
2. Lipolysis is cleavage of TAGs with mobilisation of fatty acids, yielding glycerol and free fatty acids. Ketogenesis is the generation of the ketone bodies acetoacetate and $\beta$-hydroxybutyrate.
3. Glucose $\to$ glucokinase $\to$ glycolysis $\to$ DHAP and on to pyruvate via pyruvate kinase; pyruvate enters the mitochondrion and PDH makes acetyl CoA; acetyl CoA + OAA $\to$ citrate; citrate is exported and cleaved by citrate lyase to OAA + cytoplasmic acetyl CoA; ACC makes malonyl CoA; fatty acid synthase (using NADPH) makes palmitate; palmitate $\to$ fatty acyl CoA, which combines with glycerol-3-phosphate (from DHAP) to form TG; TG plus apoproteins and other lipids forms VLDL, exported to blood.
4. Acetyl CoA is trapped in the mitochondrion and cannot cross to the cytoplasm, so citrate is exported instead and used to regenerate acetyl CoA via citrate lyase.
5. Glucokinase (synthesis stimulated by insulin), pyruvate kinase and pyruvate dehydrogenase (synthesis increased by insulin), citrate lyase, acetyl CoA carboxylase and fatty acid synthetase (synthesis increased by insulin). Transcription of all lipogenic enzymes is increased by glucose and insulin.
6. Increased synthesis of the enzyme is stimulated by insulin, and increased activity of the enzyme is stimulated allosterically by elevated glucose. Glucose uptake is not rate limiting because of an abundance of GLUT-2 transporters; glucokinase, which has a high $K_m$ for glucose, is the point of control.
7. SREBP-1c, activated by insulin acting at the insulin receptor (and raised via glucokinase), binds the SRE. ChREBP, activated by glucose via G6P after glucose entry through its transporter, binds the ChoRE. Both drive lipogenic enzyme genes. Glucagon and PUFAs act negatively at the insulin receptor.
8. Stimulated by citrate and by insulin, insulin acting through dephosphorylation. Inhibited by long chain fatty acyl CoA and by glucagon, glucagon acting through phosphorylation. So the dephosphorylated enzyme is the active form.
9. Activation of ACC elevates malonyl CoA; malonyl CoA potently inhibits CPT1 (carnitine palmitoyl transferase); entry of fatty acyl groups into the mitochondrion is blocked; $\beta$-oxidation is therefore inhibited. For lipogenesis to be effective it is also necessary to inhibit AMPK.
10. AMPK is a cellular fuel gauge: when AMP rises it switches off energy-expensive pathways (lipogenesis) and switches on energy-producing pathways ($\beta$-oxidation); it is downregulated during lipogenesis, since active AMPK blocks lipogenesis. Active AMPK turns ON glycolysis, glucose uptake, HK, GLUT4 and UCP3 in skeletal muscle, and MCD-mediated fat oxidation; it turns OFF gluconeogenic enzyme expression (PEPCK, G6Pase) and lipogenic gene expression by decreasing SREBP, cholesterol synthesis via HMG-CoA phosphorylation, and fatty acid synthesis via ACC phosphorylation.
11. Malonyl CoA levels fall, so the inhibition of CPT1 is relieved, fatty acyl groups enter the mitochondrion via the carnitine shuttle, and $\beta$-oxidation increases. This is the same sequence glucagon produces in the fasted liver.
12. Insulin stimulates GLUT4-mediated glucose uptake, giving glucose-6-P $\to$ DHAP $\to$ glycerol-3-P for TAG synthesis; insulin stimulates lipoprotein lipase (LPL), which hydrolyses TAG in chylomicrons and VLDL at the cell surface so fatty acids enter the adipocyte; and insulin inhibits hormone sensitive lipase (HSL), suppressing TAG breakdown and FFA/glycerol release.
13. De novo lipogenesis operates significantly when carbohydrate intake exceeds energy requirements and glycogen stores are full. On most Western diets, dietary fat delivery to liver and adipose tissue suppresses it.
14. Stimulated by glucagon, adrenaline, TNF-$\alpha$ and cortisol; inhibited by insulin. In the fasting state glucagon stimulates lipolysis by activating HSL; in stress and trauma adrenaline, cortisol and TNF-$\alpha$ do so.
15. Glucagon binds a GPCR on the adipocyte $\to$ activates the G-protein ($\alpha$, $\beta$, $\gamma$) $\to$ activates adenylate cyclase, converting ATP to cAMP + PP${}_i$ $\to$ cAMP activates inactive A-kinase (PKA) $\to$ active PKA phosphorylates inactive HSL using ATP $\to$ ADP $\to$ active phosphorylated HSL catalyses TAG $\to$ FFA.
16. Utilisation is related to plasma FFA concentration; in the cytoplasm fatty acids are activated to fatty acid acyl-CoA; they are transferred into mitochondria via the carnitine palmitoyl transferase system; they are oxidised by $\beta$-oxidation to acetyl CoA; in liver, excess acetyl CoA is diverted to ketone body synthesis. The brain is the exception: fatty acids are the main fuel of all aerobic tissues except brain.
17. Made by liver and kidney (supplying heart and muscle) from lysine and methionine, and also obtained from the diet. Deficiency leads to muscle fatigue, hypoglycaemia and fatty liver. Causes: liver disease, vegetarian diet with insufficient carnitine sources, haemodialysis, genetic defects such as enzyme deficiency, and increased loss post trauma, illness or surgery.
18. In liver: 2 acetyl CoA $\to$ acetoacetyl CoA $\to$ HMG-CoA $\to$ acetyl CoA + acetoacetate; acetoacetate $\to$ 3-hydroxybutyrate using NADH + H${}^{+}$, or by a minor route to CO${}_2$ + acetone. In peripheral tissue: 3-hydroxybutyrate $\to$ acetoacetate $\to$ acetoacetyl CoA via thiophorase (using succinyl CoA $\to$ succinate) $\to$ 2 acetyl CoA $\to$ TCA cycle.
19. Fed < 0.1 mmol/L; fasted for days 0.3 mmol/L; fasted 6 weeks 10 mmol/L; newborn 2 to 3 mmol/L; type 1 diabetic ketoacidosis > 30 mmol/L.
20. $\beta$-hydroxybutyrate and acetoacetate rise steeply from near zero to a plateau around 7 mM by about day 20; glucose stays flat at roughly 3.5 to 4 mM throughout; free fatty acids rise gradually to about 1.5; total urinary nitrogen starts around 4.7 g/day, falls sharply and levels off near 1 g/day (while urinary ammonia rises). The brain can obtain 60% of its energy from ketone body oxidation during fasting and in the neonatal period.
21. Type 1: no insulin, so lipolysis is uninhibited and FFA supply to liver rises; ACC is not stimulated, so malonyl CoA falls, CPT1 inhibition is relieved, FFA oxidation to acetyl CoA increases and ketogenesis increases. Type 2: lipolysis is again uninhibited so fatty acids reach the liver, but insulin is still stimulating ACC, so malonyl CoA is elevated and inhibits CPT1, blocking $\beta$-oxidation; insulin also stimulates re-esterification of FFA to TAG. Less acetyl CoA is generated, so less is available for ketogenesis.
22. Likely diagnosis NASH (non-alcoholic steatohepatitis), likely progressed from NAFLD; the ultrasound appearance is typical for steatosis. Potential complications: progression to fibrosis and cirrhosis. Treatment: diet with weight loss and cholesterol reduction, exercise, alcohol moderation, and consideration of medications and further studies.
23. Insulin resistance with hyperglycaemia elevates insulin; insulin’s inhibitory effect on adipose lipolysis is impaired more than its stimulatory effect on hepatic lipogenesis, so the liver makes more TAG. Hepatic triglyceride comes from plasma FFA released from subcutaneous fat by HSL (about 60% of flux), de novo lipogenesis driven by SREBP-1c and ChREBP under glucose and insulin (about 25%), and dietary fat/chylomicron remnants with FFAs (about 15%). Reduced lipoprotein lipase activity means VLDL-TAG is not properly off-loaded to adipose, so VLDL and remnant fatty acid return to the liver. The result is both accumulation of fat in the liver and increased VLDL output into the circulation, i.e. hyperlipidaemia.