One of the tests I came across to help figure out more of what’s going on with my cholesterol status was sterol testing. What this test does is measure particles in your blood that tell you whether the changes in your cholesterol are related to too much absorption or too much formation.
Here are some of the labs that offer this type of testing.
Unfortunately none of this was taught… or even available when I was in med school, so I didn’t know much about this. I haven’t seen much of any of this written in the paleo-sphere so I’m not sure how much awareness there is out there on this subject.
This lead me to watch hours of lectures put out by Dr. Dayspring on LecturePad.org, a free site anyone can join.
As I normally do, I took a series of notes that I’ll share with you. I plan on writing a post later synthesizing all of this a more digestible format, but until then, here are my raw notes.
Get ready to dive deeper into the cholesterol rabbit hole. Warning… it gets pretty dense.
- Most people don’t have much phyto sterols or stanols in their blood but if they do, then that can indicate they have increased cholesterol absorption.
- Absorption and synthesis are inversely related. People with more of one tend to have less of the other.
- Most cholesterol is synthesized and utilized in the extrahepatic organs.
- Under dietary conditions equivalent to those found in Western humans, the extrahepatic tissues probably account for >80% of whole animal sterol synthesis.
- Acetate and Acetoacetyl CoA is eventually converted to Mevalonate, via the enzyme HMG CoA reductase and then to lanosterol.
- HMG CoA reductase is the enzyme acted on by statins.
- Lanosterol can then be converted into either lathosterol or desmosterol which can both be subsequently converted into cholesterol.
- If lathosterol and desmosterol are high, then this indicates that there is HYPER-synthesis going on.
- Insulin Resistance – Hypersynthesis is the cholesterol abnormality in insulin resistant metabolic abnormality.
- Desmosterol – One of the biomarkers of cholesterol synthesis.
- Now being linked to Alzheimer’s disease, hepatitis C, prostate cancer, and arterial macropage-dependent inflammation.
- Desmosterol levels are decreased in Alzheimers.
- Dr. Dayspring points out that the most common cause of low desmosterol is statin therapy, so there may be a link.
Intestinal Cholesterol Absorption
- Majority of absorbed from the hepatobiliary absorption.
- One factor that affects cholesterol absorption is having working lipases
- In normal systems, ingested phytosterols are absorbed by intestinal enterocytes via the NPC1L1 membrane transporter and then quickly excreted via the ABCG5/G8 membrane transporter to be pooped out.
- Loss of function of the ABCG5/G8 efflux protein on the surface of intestinal enterocytes can cause hyperabsorption and increased phytosterols in the blood.
- Intestinal cells can also make HDL particles, and in fact make a majority of them.
- The #1 way humans get rid of cholesterol as by the liver converting it into bile acids that can be pooped away.
- 90-99% of cholesterol absorbed is endogenous in origin and has nothing to do with what is eaten.
- Genetic expression of NPC1L1 and ABCG5, ABCG8 help regulate cholesterol homeostasis.
- According to the Framingham Offspring Study, people who are cholesterol hyperabsorbers are at increased risk of cardiovascular disease.
- The markers of increased absorption are Campesterol, Sitosterol, and Cholestanol.
- Apo E Genotype – People with E4s have hyperabsorption of cholesterol.
- People with inactivating mutations of NPC1L1 are hypo-absorbers, and have a 13% reduction in total cholesterol and 12% reduction in LDL-C. They have a 53% relative risk of reduction of coronary heart disease.
- Even with frequent metabolic syndrome and diabetes, a low cholesterol absorption was associated with fewer recurrent cardiovascular events, and especially with better survival in elderly cardiovascular patients.
Scandinavian Simvastatin Survival Study (4S) – Simvastatin, drug used to treat cholesterol hyper-synthesizers, was less efficacious in patients with higher cholesterol absorption.
Ezetimibe – Drug used to prevent absorption, significantly lowered plasma concentrations of both sitosterol and campesterol (markers of hyper-absorption) from baseline compared with placebo.
However with atorvastatin monotherapy, this actually caused an INCREASE in sitosterol and campesterol, meaning that while the statin DECREASED cholesterol synthesis, this lead to a compensatory INCREASE in absorption.
A combination of both Statin and Ezetimibe lead to a decrease in both sitosterol and campesterol markers.
Fenofibrate – decreases sterol absorption via inhibitory effect on NPC1L1 expression in the proximal small intestine.
Cholestyramine – Bile acid sequestration
Lactobacillus Reuteri – Can decrease cholesterol absorption.
Phytosterol supplements – This can competitively inhibit absorption of cholesterol, however raising the levels of phytosterols in the blood may be bad.
Why monitor sterols?
- Improve risk assessment
- Hyperabsorption of sterols is a risk factor
- Hypoabsorption is beneficial in reducing CHD events
- Design better therapeutic strategies
- Monitor desmosterol in patients on statins
- Use to monitor phytosterol supplements
Sterols and Cholesterol Homeostasis: Lecture 1
- As a general rule cholesterol absorption is inversely related to cholesterol synthesis: i.e. persons with hyperabsorption suppress synthesis and hypersynthesizers suppress cholesterol absorption
- Increased fat intake is often associated with decreased cholesterol absorption – likely due to fat-induced increased cholesterol synthesis suppressing cholesterol absorption.
- Too much cholesterol build up in a cell can lead to problems because it forms crystals within the cell that can cause damage.
- The storage form of cholesterol is a cholesteryl ester (CE). What this means is that a long chain fatty acid forms an ester bond at hydroxyl group of the free cholesterol. This makes the molecule non-polar.
- Acyl CoA cholesterol acyl transferase (ACAT) is the enzyme that catalyzes this. This is present in all cells, but is mainly in the liver and intestinal.
- This is the key step in the formation of chylomicrons by intestinal cells and VLDL particles by the liver.
- Only free cholesterol can be absorbed by the liver from bile and by the intestine from the gut.
- Cholesterol is a cycloalkane with a double bond at C5. If the double bond is removed (saturation), then it is called a stanol, or cholestanol. Stanols can be used as a functional food because these cannot be absorbed.
- Sitostanol is a commercial product that people can buy to reduce cholesterol absorption.
- In the whole animal, and presumably in humans, most cholesterol is synthesized and utilized in the extrahepatic organs.
- Lathosterol and Desmosterol, markers of cholesterol hyper-synthesis, are typically found at a concentration of 1:1000 to cholesterol, but this is genetically dependent.
- High cholesterol + high lathosterol and high desmosterol = cholesterol hyper-synthesis. Statins work remarkably well in this case.
- High cholesterol + low lathosterol and low desmosterol = cholesterol hyper-absorber. You can also check the phytosterols directly to determine this.
- The body desaturates cholesterol into a stanol when it doesn’t need it anymore. The liver converts cholesterol into cholestanol which then goes down into the bile acid pathway which is then excreted into the gut, so there are trivial amounts of cholestanol within the blood.
- When cholesterol makes it to the gut via the biliary system, intestinal bacteria can convert it into cholestanol or coprostanol. Usually stanols are very poorly absorbed, so if you find cholestanol in the blood, then that tells you that the patient is a hyperabsorber.
- Normal human serum contains small amounts of the cholesterol precursors, squalene, cholestenol, desmosterol, and lathosterol, which reflect cholesterol synthesis, especially in ratios to serum cholesterol.
- Small concentrations of cholestanol, campesterol, and sitosterol are also detectable in serum (these should almost never be absorbed), the ratios of which are related to cholesterol absorption.
- The two groups (synthesis and absorption markers) are negatively (inversely) related to each other in the general population.
Sterols and Cholesterol Homeostasis: Lecture 2
- Dietary cholesterol & plant sterols total around 500 mg/day
- Synthesized cholesterol is 400 mg/day
- Secretion of 24 gm/day of bile salts of which 2 gm is cholesterol
- The intestine absorbs 55% of the cholesterol that is presented to it, but this can vary depending on genetics with a range of 29-81%. This has nothing to do with diet. Vegetarians have the same rate of absorption.
- The average diet consists of 200-600 mg of cholesterol, and consumption has no substantial effect on coronary risk.
- The majority of cholesterol (~85%) absorbed is endogenous biliary cholesterol, NOT ingested cholesterol.
- Intestines convert absorbed cholesterol into chylomicrons and HDL to be brought to the liver.
- Niemann-Pick C1-Like 1 Protein (NPC1L1) – the intestinal sterol influx protein, and is critical to cholesterol absorption. It’s expressed by enterocytes and by hepatocytes.
- NPC1L1 activity is stimulated by low cholesterol and inhibited by high cholesterol.
- The ileum reabsorbs bile salts. 95% of the bile salts that make it to the ileum is reabsorbed, and brought back to the liver.
- In the average person fecal sterol excretion is 400 mg/day.
- The main routes of cholesterol excretion are as bile acids and as free cholesterol.
- Intestinal cells can absorb cholesterol from serum HDL.
- Intestinal cells can also absorb cholesterol from serum LDL.
How do intestinal cells get rid of this absorbed cholesterol?
- Efflux free cholesterol into HDL particles
- Efflux free cholesterol within chylomicrons into lymphatics
- Excrete free cholesterol and non-cholesterol-sterols into the gut lumen.
How does the liver get cholesterol?
- LDL receptors allow influx of cholesterol from serum LDL particles.
- Cholesterol can be absorbed from serum HDL particles
- Cholesterol can be absorbed from bile acids via the NPC1L1 transporter (this is increased in the setting of statin therapy)
How does the liver get rid of cholesterol?
- Excretion of cholesterol as bile acids via the ABCB11 transporter
- Efflux of cholesterol into HDL particles
- Efflux of cholesterol into LDL particles
- Efflux free cholesterol into bile via the ABCG5/G8 transporter (this transporter can also excrete phytosterols)
- Genetic expression of NPC1L1, ABCG5, and ABCG8 help regulate cholesterol homeostasis.
- At the enterocyte, the NPC1L1 absorbs cholesterol and phytosterols. Any unused cholesterol and phytosterol is excreted via ABCG5, and ABCG8.
- Phytosterols are absorbed readily from the intestinal lumen, ARE NOT esterified (unless present in large numbers), and therefore ARE NOT internalized into HDL or chylomicrons, and so are excreted back into the intestinal lumen.
- The easiest way for enterocytes to accumulate LARGE amounts of phytosterols is to have genetic variants of the ABCG5/G8 transporters that are not so effective at evicting the phytosterols, or even absence of the ABCG5/G8 transporters which will not allow eviction of phytosterols from the enterocytes.
- If there are large enough amounts of phytosterols accumulated within the enterocyte, the phytosterols can be excreted into the serum on the surface of HDL particles.
- One enough cholesterol is inside the cell, there is down-regulation of the NPC1L1 transporter, and less cholesterol is absorbed.
- Neither dietary cholesterol nor dietary fat significantly altered % absorption of cholesterol. Regardless of diet type, the individuals within the group differed markedly in the percentage dietary cholesterol absorption.
Sterols and Cholesterol Homeostasis: Lecture 3
- The use of surrogate markers like xenosterols is qualitative in nature, however it is more affordable and convenient.
- Cellular cholesterol precursors are also found in normal human plasma, at concentrations roughly 1:1000 of that of cholesterol.
- Thus the plasma levels of lathosterol and desmosterol are commonly used as measures of the cholesterol biosynthetic activity of the individual.
Do not confuse entry of a sterol into the intestinal enterocyte with sterol absorption which refers to the incorporation of a sterol into a lipoprotein (initially a chylomicron and HDL) and entry into the systemic circulation
- Only an unesterified sterol can enter an enterocyte via sterol influx transporters.
- One internalized the sterol may or may not be esterified by ACAT
- Unesterified sterols are usually exported back to the gut lumen by sterol efflux proteins. Some can attach to chylomicron or HDL surface layers
- Esterified sterols (esters) enter the chylomicron core.
- Cholestanol blood concentration is very well correlated with fractional cholesterol absorption measured by the fecal excretion of isotopic cholesterol and the isotopic nonabsorbed marker (silosterol). Cholestanol is produced by the body.
- Since they cannot be synthesized by the human body and therefore result exclusively from absorption, phyosterols (silosterol and campesterol) are used as more reliable markers of cholesterol absorption.
- Plasma levels of Sitosterol were raised little when intakes were increased greatly, and on fixed intakes they were constant from week to week.
- On diets devoid of plant sterols, the plasma and feces rapidly became free of sitosterol.
- Cholestanol normal dietary intake is < 2 mg/day whereas cholestanol synthesis is > 10 mg/day.
- Coprostanol is formed by the conversion of cholesterol to coprostanol in the gut by intestinal bacteria.
- Phytosterols are trafficked within or on lipoproteins. They do not exist within the body as free molecules. Most often in HDL and LDL particles.
- Because plasma phytosterols are carried in lipoproteins, it’s advocated that they are expressed as a ratio with total cholesterol to adjust for total cholesterol concentration.
- On the other hand, using adjusted values is not correct in statin studies, as the decrease in total cholesterol can increase the phytosterol to cholesterol ratio even though there is no change in the phytosterol concentration.
- The main disadvantage of using ratios is that they are difficult to interpret when used in correlation and regression models.
Plasma sterols being reported as absolute values vs as ratios with cholesterol
- Since both cholesterol and plant sterols are carried by lipoproteins in the plasma, more lipoprotein particles are often present in the plasma of hypercholesterolemic subjects.
- Therefore, it can be assumed that hypercholesterolemic subjects might have a higher absolute plasma concentration of plant sterols, even with the same dietary background or with a similar plant sterol absorption rate.
- To improve the estimation of cholesterol or total sterol absorption, circulating plant sterol levels are often standardized by reporting the plant sterol to cholesterol ratio.
- Dr. Dayspring sees trouble here, lipoprotein concentrations vary between individuals AND vary within the same individual.
In patients with familial hypercholesterolemia, not only are cholesterol concentrations significantly higher compared with normocholesterolemic controls, but those of the plant sterols/stanols and cholesterol precursors are also significantly higher.
- However, after correction for serum concentrations of cholesterol, the ratios of plant sterols and cholesterol precursors no longer differ.
Some say it depends on the question being asked:
- When investigating synthesis and absorption differences among two groups, one would like to be certain that these effects are independent of the cholesterol concentration and therefore values should be expressed relative to cholesterol (as a ratio).
- However, investigating the relationship between absorption markers, synthesis markers, and cholesterol levels, the absolute sterol concentrations should be used due to the fact that normalizing or adjusting for total cholesterol would mean that you are masking the outcome variable you are interested in assessing.
Sterols and Cholesterol Homeostasis: Lecture 4
Dietary intake of plant sterols without plant sterol supplementation played only a minor role in the variations observed.
Sources of plant sterols:
- High – plant oils
- Moderate – nuts and seeds
- Low – fruits and vegetables
Dietary intake of phytosterol supplements:
- When plant STEROLS are administered in supplement form at a dose of 1.8-2.0 g/d for 4-8 weeks, there is a 52-99% increase in campesterol levels and a 23-96% increase in sitosterol levels.
- In contrast, when plant STANOLS are supplemented at a dose of 1.5-3.0 g/day for 4 weeks, there are decreases of 28-113% in campesterol levels and 24-50% in sitosterol levels.
Apo E4 tend to have absorption markers in the hyper ranges and synthesis markers in the hypo ranges.
NPC1L1 – Affinity for the different sterols differs, and these differences in sterol absorption can contribute to the different plasma sterol concentrations
- Cholesterol > campesterol > sitosterol > campestanol > sitostanol
People with insulin resistance and/or diabetes and/or obesity have decreased cholesterol absorption.
Insulin resistant patients have higher levels of cholesterol synthesis markers (desmosterol and lathosterol) and lower levels of absorption markers (cholestanol and sitosterol). Will have better LDL-C response to statins.
Statin treatment may increase plant sterol concentrations in the plasma by increasing intestinal sterol absorption or by lowering the free cholesterol pool in the liver, which may result in decreased secretion of cholesterol and plant sterols into bile.
- Significant correlations between the lathosterol:cholesterol ratio and cholesterol synthesis rates were observed
- Plasma lathosterol concentrations increased by 53% and L:C ratio increased by 72% on ezetimibe
- L:C Ratio is a marker of hepatic HMG-CoA reductase activity and total cholesterol synthesis.
LDL can return cholesterol to the gut, bypassing the liver. This is called TICE.
Approach to the Patient with Xenosterol Disorders Part 1
The pattern associated with the most CV risk is hyperabsorption with hyposynthesis of cholesterol.
When LDL-P (apoB) are elevated in the face of normal sterol values, there is a problem with overproduction (typically related to TG abnormalities) or a decreased clearance of ApoB particles (LDL receptor issues, defective apoB, PSCK9 gain of function)
After lifestyle modification, statins are always the first line LDL-P lowering therapy unless TG are > 500 mg/dL.
- Had the most reduction in coronary events in cholesterol hypersynthesizers (4S study).
- Almost no effect in coronary events in cholesterol hyperabsorbers.
- The major effect of statins is to reduce cellular cholesterol synthesis, resulting in an up-regulation of LDL receptor activity, enhanced fractional clearance of LDL from plasma, and reduction in plasma LDL cholesterol levels.
- These effects may be offset due to compensatory increase in absorption
- Because ezetimibe significantly reduces intestinal cholesterol absorption, but increases synthesis, and because statins have the opposite effect, it would appear that combination therapy would be ideal.
- Patients with high levels of cholesterol synthesis AND absorption displayed the most effective LDL-C lowering by statin monotherapy.
- Patients with a low cholesterol synthesis AND a high level of absorption, or with a high level of synthesis AND a low level of absorption responded poorly to statin monotherapy.
- Lipids in the gut lumen present to the microvilli as a biliary micelles
- Ezetimibe inhibits the NPC1L1 transporter, preventing uptake of cholesterol from the gut lumen
- Thus, the chylomicron the enterocyte excretes will be cholesterol depleted.
- The chylomicron will go to the liver and will deliver it’s decreased amount of cholesterol.
- Ezetemibe also blocks NPC1L1 activity at the hepatobiliary porter too, to decrease backward flux of cholesterol from the biliary system to the hepatocyte.
- Because of the depleted cholesterol pool, this will cause upregulation of LDL Receptors, which will lower Apo B, LDL-C, Non HDL-C, and TG
- There will also be reduced VLDL assembly and secretion in hepatocytes. Which will also reduce LDL-C, TG, ApoB, LDL-P, and Non HDL-C
- Ezetimibe prevents NPC1L1 from entering the AP2-mediated clathrin-coated vesicles, thus inhibiting endocytosis.
Approach to the Patient with Xenosterol Disorders Part 2
Bile Acid Sequestrants (Colesevelam, Colestipol, Cholestyramine)
- When the enterohepatic circulation of bile acids is interrupted with bile acid sequestrants, cholesterol 7-hydroxylase activity increases to maintain bile acid synthesis.
- This increase LDL receptor activity, binding more Apo B lipoproteins, and bringing them into the liver.
- The demand for bile acid synthesis is satisfied through increased catabolism of plasma LDL, with concomitant decreases of plasma cholesterol and plant sterols.
Fibrates (Clofibrate, Fenofibrate, Gemfibrozil)
- Fibrates are PPAR alpha agonists
- This causes reduced expression of the NPC1L1 and decreasing sterol absorption at the enterocyte
- Similar to Ezetemibe, causes the gut to excrete cholesterol depleted chylomicrons, leading to delivery of less enterocytic cholesterol to the liver.
Sterol and Stanol Therapy (Campesterol, Cholestanol, Sitosterol, Sitostanol)
- Therapeutic doses of phyosterols and phytostanols displace cholesterol from the micellar nucleus, so less cholesterol is available in micelles for absorption
- People who supplement with phytosterols expose their intestines to way more phytosterols than someone on a vegetarian diet
- Apo E4 have higher incidence of cholesterol absorption.
- Apo E3/E3 are considered normal
- Regardless of genotype, administration of phytosterols there is significant reduction in total cholesterol and LDL cholesterol.
- Higher plant STEROL intake in the form of supplements increases circulating levels of plant sterols, while plant STANOL supplementation decreases these levels.
- The only phytostanol available is sitostanol, aka “Benecol”
- If you’re not going to measure phytosterols in the blood, then you should recommend patients to take a stanol.
- The observation that hyperabsorption of phytosterols can produce premature CAD and aortic stenosis in individuals with defects in ABCG5/8, raises the possibility that increased serum levels of phytosterols can contribute to CAD in the general population.
- From a review article in 2008, “plant sterol supplementation impairs endothelial function, aggravates ischemic brain injury, effects atherogenesis in mice, and leads to increased tissue sterol concentrations in humans.”
- Lot of data suggesting phytosterols are bad and can induce atherogenesis.
- Currently there are no data available indicating that cholesterol lowering through plant sterol ingestion results in prevention of CVD.
- So it’s important to monitor serum phytosterol concentrations if using sterols as supplementation.
- Lactobacillus reuteri
- It’s hypothesized that increased in deconjugated bile acids may result in reduced FXR activation, increased cholesterol catabolism, upregulation of ABCG5/G8
- Lactobacillus reuteri increase deconjugated bile acids… leading to decrease in LDL-C.
- Results from the L. reuteri study suggest that deconjugation of intraluminal bile acids results in reduced absorption of non-cholesterol sterols and indicate that L. reuteri NCIMB 30242 capsules may be useful as an adjunctive therapy for treating hypercholesterolemia.
*Image found here