Week 7 - Ectopic lipids and liver disease

Question 1

What is ectopic fat?

Answer 1

Fat (lipids) stored in places not designed for mass storage (i.e., not adipose tissue)

Question 2

Sites of fat storage:

Answer 2

1) Subcutaneous adipose tissue (ScAT) - fat stores located superficially just under the layer of skin (helps with insulation) - this the lowest cardiometabolic risk for fat storage.
2) Visceral adipose tissue (VAT) - ectopic. Located deeper in abdominal region around internal organs. has a greater cardiometabolic risk than ScAT.
3) Intra-organ - ectopic. lipids stored within internal organs, which causes stress in the organ. has the greater cardiometabolic risk

Question 3

How does ectopic fat accumulate?
Adipose tissue expandability/ fat spillover hypothesis:

Answer 3

• Modern day lifestyles are characterised by a state of chronic positive energy balance – high fat and simple sugar diet coupled with low energy expenditure
• Our body buffers excess energy availability by storing the energy as lipids within adipose tissue or adipocytes. With an increase of lipids, these expand overtime
• Lipid storage has a finite capacity (in the adipocytes). When full the adipocytes become stressed/ inflamed and deprived of oxygen – hypoxia. This results in the production of pro-inflammatory cytokines
• Overtime this promotes insulin resistance within adipose tissue cells
• Within the adipose tissue, insulin promotes the uptake of different substrates such as glucose and lipids, out of the circulation and into the tissues of the body. In the adipose tissue, insulin promotes the uptake and storage of lipids. If adipose tissue is resistant to insulin, the lipids stored within the adipocytes start to get broken down into FA building blocks that spill over into circulation
• These FA get delivered to unwanted (ectopic) sites e.g., liver, skeletal muscle, pancreas (T2DM) and the heart (contributes to CVD).

Question 4

Non-alcoholic fatty liver disease

Answer 4

• NAFL is characterised by the ectopic fat accumulation within the liver
• In the general population around 32% of people have a fatty liver. 10-30% of these individuals will then progress to a more severe stage known as NASH (inflammation of the liver)
• Absence of excessive alcohol intake or other sedentary causes
• Defining feature = hepatic steatosis (liver fat accumulation)

Question 5

NASH - inflammation of the liver:

Answer 5

• This inflammation over time causes liver cells to be replaced by fibrous tissue – has no function in terms of normal function = fibrosis
• Fibrosis = liver loses its normal function
• Cirrhosis = excessive sacring of the liver (build up of fibrosis) – can result in liver failure (end stage liver disease)
• HCC = a type of liver cancer
• All these conditions are lifestyle or obesity related (not caused by other secondary causes such as alcohol/smoking, autoimmune disease, viral causes e.g., hep A,B & C)

Question 6

Change in nomenclature (naming of the disease):

Answer 6

MASLD chosen to replace NAFLD and MASH chosen to replace NASH
MASLD = metabolic dysfunction-associated steatotic liver disease
MASH = metabolic dysfunction-associated steatohepatitis

Question 7

Adult cardiometabolic criteria: At least 1 out of the 5:

Answer 7

• BMI ≥ 25kg/m2 (23 Asia) OR WC > 94cm (M) 80cm (F) OR ethnicity adjusted equivalent
• Fasting serum glucose ≥5.6mmol/L (100mg/d) OR 2 hour post-load glucose levels ≥7.8mmol/L (≥140mg/d) OR HbA1c ≥5.7% (39mmol/L) OR T2DM OR treatment for T2DM
• Blood pressure ≥30/85 mmHg OR specific antihypertensive drug treatment (hypertension)
• Plasma triglycerides ≥1.70 mmol/L (150mg/d) OR lipid lowering treatment (dyslipidaemia)
• Plasma HDL cholesterol ≤1.0mmol/L (40mg/d) (M) and ≤1.3 mmol/L (50mg/d) (F) or lipid lowering treatment

Question 8

MASLD diagnosis - Liver biopsy:

Answer 8

• Take a small slice of liver tissue, stain it and put it under a microscope to see the visible white lipid droplets
• Visible lipid droplets in more than 5% of hepatocytes = MASLD
• Gold standard method
• Key benefit = can look at the different stages of MASLD can be identified (can see fat, inflamed liver cells and any fibrosis occurring)
• Negatives = invasive and specialist e.g., has to cut through the layer of abdominal muscles. Carries risks and complications

Question 9

MASLD diagnosis- MR spectoscropy:

Answer 9

• Liver fat percentage greater than 5.56%
• Non-invasive ‘gold standard’ method
• Allows repeated measurements
• Negatives = MRI scanners are Expensive and specialist radiographers are required

*Other methods: MRI, CT, ultrasound, blood biochemistry (blood based biomarkers to indirectly tell us what’s going on)

Question 10

Prevalence of MASLD:

Answer 10

• Most common form of liver disease worldwide
• Currently affects ~32% of adults (increased dramatically over recent years): T2DM: ~60%, Obesity (BMI ≥30kg/m2) ~70%, Severe obesity (BMI ≥ 40kg/m2): > 90% (9 in 10 people with severe obesity also have excessive amounts of fat in the liver)
• These relationships are bidirectional: T2DM is a risk factor for this obesity related liver disease, and individuals with MASLD also have a 2-fold greater risk of developing T2DM.

Question 11

Impact of MASLD:

Answer 11

• The liver plays an important role in modulating glucose/ lipid metabolism and energy homeostasis – produces lipids and glucose in times of needs and can uptake them (for energy production or storage (small amount)).
• MASLD is closely associated with other metabolic comorbidities: T2DM, metabolic syndrome, CVD, insulin resistance, dyslipidaemia, hypertension
• When compared with ScAT and VAT, liver fat is the strongest predictor of insulin resistance in multiple tissues throughout the body

Question 12

Pathogenesis of hepatic steatosis:

Answer 12

• Hepatic steatosis (excess liver fat) develops when lipid supply to the liver exceeds the ability to dispose of it
  Supply routes: adipose tissue lipolysis, dietary fat and de novo lipogenesis
  disposal routes: fat oxidation and VLDL-TAG export

Question 13

Pathogenesis of hepatic steatosis explained: supply routes

Answer 13

People with MASLD have a high dietary fat intake and high simple sugar intake (glucose and fructose).
Dietary fat can be transported in circulation bound to large lipid transporters called chylomicrons. These chylomicrons can deliver the lipids directly to the adipose tissue
Once adipose tissue reaches the limit, it becomes dysfunctional and insulin resistant. It is then no longer able to supress the breakdown of lipids. We therefore see an Increase in lipolysis (breakdown of lipids into FAs)
Increase of FAs into the circulation – get taken up by the liver – converted back to lipids and stored in the liver.
Dietary fat can go directly to the liver via chylomicron remnants – these can be directly taken up by the liver and also increases triglyceride/ lipid synthesis in the liver.
Increase glucose and fructose in circulation. Take up by the liver and undergo de novo lipogenesis (creating new lipids from non-lipid precursors)
De novo lipogenesis = creating new lipids from non-lipid precursors
- Glucose and fructose are potent stimulators of de novo lipogenesis
- This creates new FAs from glucose and fructose, again contributing to high amounts of fat within the liver
Skeletal muscle – insulin promotes uptake of glucose from circulation into muscle to fuel muscular work or for storage as glycogen. If skeletal muscle becomes resistant to the effects of insulin it won’t effectively promote the uptake of glucose out of the circulation. Contributes to greater glucose accumulation in the circulation and consequent glucose delivery to the liver which supplies greater de novo lipogenesis.

Question 14

Lipid disposal routes:

Answer 14

• Lipid oxidation
• Beta oxidation – occurs in mitochondria in liver cells (lipids undergo oxidation to generate ATP to use as a fuel source)
• In people with MASLD – levels of oxidation increase (to try and buffer the increased lipid availability to oxidise as much as possible to maintain normal levels). In people with MASLD it is increased to an inadequate extent and net gain of lipids occurs
• 2 key reasons for this:
  1) Reactive oxygen species that are generated as part of the beta-oxidation process. These can cause oxidative stress within the liver meaning that oxidation is not as efficient.
  2) By-product of de novo lipogenesis pathway called Malonyl-CoA, which is a direct inhibitor of lipid oxidation.
• Export of lipids bound to VLDL cholesterol – the liver can take the stored triglyceride and package it up bound to VLDL cholesterol molecules and export it from the liver. Harmful side effect = accumulation of VLDL cholesterol in our circulation which is a key contributor to the build of atherosclerotic plaques.

Question 15

Lifestyle modification guidelines for management of MASLD:

Answer 15

• Best Practice Advice 2: Among patients with MASH, weight loss of 5% or greater of total body weight can reduce hepatic steatosis, weight loss of 7% or greater can lead to NASH resolution and weight loss of 10% or greater can result in fibrosis regression or stability. Take home message = at least 5% body weight loss is required for a reduction in liver fat. The more is better…
• Best Practice Advice 7: Regular physical activity should be considered for patients with MASLD, with a target of 150-300 minutes of moderate-intensity or 75-150 minutes of vigorous-intensity aerobic exercise per week. Need to meet or exceed current PA guidelines in the UK
• Resistance training exercise can be complementary to aerobic exercise and can have independent effects on MASLD.

Question 16

Intensive lifestyle interventions:
- Prospective cohort study of 293 patients with MASH.

Answer 16

• In people that achieved 5% weight loss or greater, it was associated with a high likelihood of steatosis improvement (reduction in liver fat)
• In terms of MASH resolution (improvement in inflammation) – a weight loss of 7% or greater was required for a high likelihood of improvement
• A weight loss of 10% or greater was required for a high likelihood of fibrosis regression
• Weight loss is key – greatest improvements seen in those with greatest weight loss. But, small percentage achieved the > 5% weight loss target.
• Key caveat = 70% of people in the study did not reach the 5% threshold in the study – reaching these amounts of weight loss are difficult

Question 17

The effect of exercise training on intrahepatic triglyceride and hepatic insulin sensitivity: a systematic review and analysis (Sargeant et al 2018)

Answer 17

• In the studies that didn’t show weight loss - exercise was associated with 2.1% absolute reduction in liver fat
• In the studies that did show weight loss - exercise was associated with an almost 5% reduction in liver fat
• All studies together = 3.3% reduction in liver fat with exercise
• Meta-regression: Each 1% reduction in body weight was associated with a 0.9% (abs) reduction in liver fat.
• Greater the weight loss with exercise = greater reduction in liver fat
• Exercise is able to reduce liver fat independant of significant weight loss

Question 18

Exercise – HIIT (Seargent et al 2018):

Answer 18

• 3 supervised HIIT session per week for 6 weeks
  No changes in body weight. Unsurprising due to short exercise training period
• Significant reduction in liver fat vs. pre-training (2.1% reduction in liver fat).
• Significant reductions also seen in VAT and ScAT.
• The greater the reduction in liver fat was associated with a greater improvement in peripheral insulin sensitivity.

Question 19

Effects of exercise training on intrahepatic lipid content in humans (Brouwers et al 2016)

Answer 19

• Simplest view: exercise-related EE induces negative energy balance, resulting in an increased mobilisation of hepatic lipids to fuel an energy deficit.

Question 20

Exercise effect on Adipose tissue:

Answer 20

Exercise improves adipose tissue insulin sensitivity (improved ability of insulin to suppress lipolysis)
- If adipose tissue becomes more sensitive to insulin, insulin can more effectively promote the uptake of lipids and suppress the breakdown of lipids (causes lower breakdown of FAs and less FA spillover into circulation, and les delivered to the liver)

Question 21

Exercise effect on skeletal muscle (Brouwers et al 2016):

Answer 21

• Exercise training can increase in uptake and oxidation of circulating FFA – prolonged aerobic exercise switches to fat as a fuel source – less fat going to the liver
• Increase in lipid uptake from diet into the muscle (done through an increase LPL activity – the key enzyme involved in the uptake of lipids out of the circulation)
• Increase glucose uptake from enhanced insulin sensitivity
• Reduced substrate for De Novo Lipogenesis in the liver

Question 22

Exercise effect on the Liver – supply:

Answer 22

• Decrease uptake of lipids from the circulation (decrease hepatic lipase activity – less lipids taken up into liver)
• Exercise also decrease De novo lipogenesis in the liver: No evidence in humans; reduced enzyme activity in rodents in exercise training

Question 23

Exercise training: Liver – disposal:

Answer 23

• Increase lipid oxidation. No evidence in humans: increase markers of b-oxidation and mitochondrial biogenesis in rodents – increase mitochondria in liver cells to enable more lipids to be oxidised as a fuel source
• VLDL-TAG export? decrease rather than increase in export from the liver. Could be a consequence of decreased liver fat levels – if less liver fat is available less needs to be exported – contribute to decrease risk of CVD as less VLDL into the circulation

Question 24

Resistance exercise - Systematic review of 23 exercise training studies in patients with MASLD:

Answer 24

• Similar improvements in BMI and liver fat between exercise modes despite lower intensity (vo2 and METs) and energy consumption with resistance training.
• Resistance exercise worked at lower intensity of vo2 max and expended less energy during the protocol. For clinical populations this may be beneficial (limited exercise capacity/ less mobile)
• Mechanisms: resistance exercise may complement aerobic exercise by modulating liver fat through different mechanisms.

Question 25

Aerobic exercise vs resisistance exercise:

Answer 25

Aerobic exercise:
1) activation of lipolysis
2) upregulation of UCP-1 and PPARy
3) Alteration in adipocytokine

Resistance exercise:
1) hypertrophy of type 2 muscle fibres
2) activation of GLUT4, AMPK, and caveolins
3) alteration in myokines;

Question 26

Resistance exercise – mechanisms:

Answer 26

1) Hypertrophy of type II muscle fibres
- Type II fibres are characterised by anaerobic glycolytic metabolism.
- Hypertrophy = greater glycolytic demand (greater demand of glucose as fuel source) from muscle.
- The greater demand on glucose as fuel source results in decreased circulating glucose meaning less substrate for De Novo Lipogenesis.
2) Activation of GLUT4 and AMPK
- GLUT4 = insulin-regulated transporter protein responsible for glucose uptake into muscle.
- AMPK = energy-sensing kinase which promotes insulin sensitivity.
- Resistance exercise results in an energy expenditure. AMPK senses this energy deficit and improves insulin sensitivity. Enhanced insulin sensitivity increases GLUT4-mediated glucose uptake into muscle which decreases circulating glucose and insulin (decreased de novo lipogenesis).
3) Alteration in myokines:
- Myokine = cytokine which is produced and secreted by skeletal muscle
- Irisin is a myokine which is increased by resistance exercise
- Irisin is suggested to modulate fat metabolism in liver by inhibiting key regulators and enzymes involved in DNL.

Question 27

Is all ectopic fat bad?
The athletes Paradox

Answer 27

• Skeletal muscle lipids are elevated (myosteatosis) in both endurance-trained athletes and individuals with T2DM compared to lean, sedentary counterparts.
• BUT, athletes remain highly insulin-sensitive whilst individuals with T2D exhibit insulin resistance and are less metabolically healthy.
• This may be beneficial in athletes where the fuel demand for muscular work is greater – greater turnover as lipids for a use of fuel.
• Why the difference? The form in which lipids are stored in the muscle appears to be important (e.g. PUFA vs. SFA and their intermediates DAG and ceramide). People with T2DM may store lipids as saturated FAs (converted into harmful lipid intermediates e.g., DAG and ceramide – these may induce insulin resistance). Whilst athletes may store lipids as PUFA (may not induce insulin resistance).