1. Cholesterol Absorption, Synthesis, & Metabolism I Chapter 34
Nov. 4th 2011

2. Cholesterol Background
Atherosclerotic vascular disease
Stabilizes cell membrane
Precursor to bile salts and steroid hormones
Cholesterol precursors converted to ubiquinone, dolichol, & vitamine D

3. Cholesterol Background Synthesis
Obtained through diet or synthesis
Synthesized in many cells, but mostly in the liver and intestine
Acetyl coenzyme A (acetyl CoA) is the precursor to cholesterol synthesis

4. Cholesterol Background (Transport)
Chylomicrons & VLDL transport cholesterol to other cells through the bloodstream
Chylomicrons package cholesterol in intestine, while VLDL package in liver
Triacylglycerols are also transported by Chylomicrons and VLDL
HDL – reverse cholesterol transport

5. Student Learning Outcomes
Describe the rate-limiting step in cholesterol synthesis and how the HMG-CoA reductase is regulated
Briefly describe the fates of cholesterol
Describe the VLDL to LDL pathway
The role of HDL
RCT, apoprotein & lipid exchange
Explain what occurs during receptor mediated endocytosis
Describe the aspects of Atherosclerosis

6. Cholesterol Synthesis
Perhydrocyclopentanophenanthrene structure consists of four fused rings
Cholesterol contains a hydroxyl group at C3, double bond between C5 & C6, eight-membered hydrocarbon chain at C17, & methyl groups at C10 & C13
Fig.2
Fig. 1
Perhydrocyclopentanophenanthrene
Cholesterol

7. Cholesterol Synthesis Stage I: Acetyl CoA to MevalonateB. C.
Fig.3
Rate limiting step

8. Cholesterol Synthesis Stage I: Transcription Control
Fig. 4A
Feedback regulatory system
Rate of HMG-CoA reductase mRNA synthesis controlled by sterol regulatory element binding protein (SREBP)
Once in the Golgi, SERBP is cleaved twice by S1p & S2P to release the transcription factor

9. Cholesterol Synthesis Stage I: Proteolytic Degradation of HMG-CoA Reductase
Fig. 4B
When sterol present, enzyme undergoes sterol accelerated ERAD (ER associated degradation)
HMG-CoA is ubiquitinated and extracted from membrane where it is then degraded by proteosomes

10. Cholesterol Synthesis Stage I: Regulation by Covalent Modification
Short-term regulation by phosphorylation & dephosphorylation
Adenosine monophosphate (AMP) activated kinase phosphorylates HMG-CoA
Glucagon, sterols, glucocorticoids & low ATP levels inactivate HMG-CoA
Insulin, thyroid hormone, high ATP levels activate enzyme
Fig. 4C

11. Cholesterol Synthesis Stage 2: Mevalonate to 2 Activated Isoprenes
Transfer 3 ATP to Mevalonate in order to activate C5 & OH-group of C3
Phosphate group at C3 & Carboxyl group of C1 leave, which produces a double bound
This allows for two active isoprenes
Fig.5

12. Cholesterol Synthesis Stage 3: Condensation of Isoprenes to for Squalene
1) Head to tail attachment of isoprenes to form Geranyl pyrophosphate
2) Head to tail condensation of Geranyl pyrophosphate and isopentenylpyrophosphate to form Farnesyl pyrophosphate
3) Head to head fusion of two Farnesyl pyrophosphate to form squalene
Fig.6

13. Cholesterol Synthesis Stage 4: Squalene to Four-Ring Steroid Nucleus
Fig. 7
Squalene monooxygenase adds oxygen to form an epoxide
Unsaturated carbons (double bonds) are aligned to allow cyclization and formation of lanosterol
After many reaction get cholesterol

14. Fates of Cholesterol Membranes Cholesterol Ester Biliary Cholesterol
Bile Acids

15. Cholesterol Esters
Acyl-CoA:cholesterol acyl transferase (ACAT) is an ER membrane protein
ACAT transfers fatty acid of CoA to C3 hydroxyl group of cholesterol
Excess cholesterol is stored as cholesterol esters in cytosolic lipid droplets
Fig. 8

16. Bile Salts Bile acids & salts are effective detergents
Synthesized in the liver
Stored & concentrated in the gallbladder
Discharged into gut and aides in absorption of intraluminal lipids, cholesteral, & fat soluble vitamines
Bile acid refers to the protonated form while bile salts refers to the ionized form
The pH of the intestine is 7 and the pKa of bile salts is 6, which means that 50% are protonated
These terms are sometimes used interchangeably

17. Synthesis of Bile Salts
Fig. 9
Fig. 10
Rate-limiting step performed by the 7α-hydroxylase (CYP7A1) and is regulated by bile salt concentration
End product: Cholic acid series & Chenocholic acid series
Bile salts can be conjugated & become better detergents

18. Fate of Bile Salts
Fig. 12

19. Cholesterol Transport by Blood Lipoproteins
Cholesterol, cholesterol esters, triacylglycerols, & phospholipids are insoluble and must travel via lipoproteins

20. VLDL to LDL
Fig. 14
The TG, free & esterified cholesterol, FA, & apoB-100 are packaged into nascent VLDL
Nascent VLDL are secreted to bloodstream and acquire apoCII & apoE from HDL to form a mature VLDL
Hepatic triglyceride lipase (HTGL) hydrolyzes additional triglycerides to produce LDL
40% of LDL transported to extrahepatic tissues
Excess LDL is taken up by macrophages

21. Reverse Cholesterol Transport (RCT)
Oram, JF & Vaughan, AM. (2000) ABCA1-mediated transport of cellular cholesterol & phospholipids to HDL apolipoproteins. Curr Opin Lipidol. June;11(3):253-60
HDL removes cholesterol from cells and returns it to the liver
ABC1 transport protein uses ATP hydrolysis to move cholesterol from inner leaflet to outer leaflet of membrane
HDL receives cholesterol and uses the LCAT enzyme to modify & trap the cholesterol

22. Fate of HDL
HDL binds SR-B1 receptor
Transfers cholesterol & cholesterol ester to cell
Depleted HDL dissociates & re-enters circulation
HDL can bind to specific hepatic receptors, but primary HDL clearance occurs through uptake by scavenger receptor SR-B1
Present on many cells
SR-B1 can be upregulated in cells that require more cholesterol
SR-B1 is not downregulated when cholesterol levels are high

23. HDL Interactions with Other Particles
Fig. 16
Fig. 17
HDL transfers apoE & apoCII to Chylomicrons & VLDL
HDL either transfers cholesterol & cholesterol esters directly to liver or by means of CETP to VLDL (or other TG-rich lipoproteins)
In exchange, HDL receives triacylglyceroles
Prior to CETP mature HDL particles are HDL3, post CETP they become larger and are called HDL2

24. Receptor-Mediated Endocytosis of Lipoproteins
LDL receptor are located at coated pits, which also contain clathrin
Vesicles fuse with lysosome where cholesterol esters are hydrolyzed into cholesterol & re-esterified by ACAT
This avoids damaging effects of high concentrations of free cholesterol on membrane
Unlike cholesterol esters of LDL, these cholesterol esters are monosaturated
Fig. 18

25. Feedback Regulation of Receptors
Regulation by SREBP or its cofactor
Low levels of cholesterol leads to up regulation of receptor genes
Increase amount of cholesterol in cells
High levels suppress expression of receptor genes
Reduces amount of cholesterol that enters cells

26. Lipoprotein Receptors
LDL receptor most well characterized & contains 6 different regions
LDL receptor-related proteins are structurally related but recognize more ligands
Macrophage scavenger receptor : SR-AI & SR-A2
Take up oxidatively modified LDL
When engorged with lipids macrophages become foam cells

27. Anatomical & Biochemical Aspects of Atherosclerosis
Fig 21. Layers of arterial wall
Initial step is formation of fatty streak (foam cells) in subintimal space
Foam cells separate endothelial cells exposing them to blood, which leads to plaques & thrombin at these sites
When plaque content exposed to procoagulant elements in circulation, acute thrombus formation occurs
Further thrombus formation leads to complete occlusion of lumen & eventually AMI or CVA

28. Key Concepts
HMG-CoA conversion to mevalonate is the rate limiting step of cholesterol synthesis
HMG-CoA reductase regulated by feedback, degradation, modification
Cholesterol fate: membranes, esters, biliary cholesterol, bile salts
Bile salts aide in absorption of lipids
Hydrolysis of VLDL leads to LDL, which transport TG & CE to peripheral cells & macrophages
HDL involved in RCT & apoprotein/lipid exchange
LDL enters cells via receptor-mediated endocytosis
Excess LDL taken up by macrophage leads to the formation of foam cells, which is the beginning of atherosclerosis