Discussions between members

Spring-Summer-Autumn 2005 - Sterols and atherosclerosis

Jacqueline Walker
Does anyone know anything about a cholesterol lowering drug called 'Ezetrol' (ezitimibe is, I think, the name of the active ingredient)? My father has just been put on it and asked me to find out about it for him.

Barry Groves
Hi Jacqueline.  Further to my last e-mail, . . . .Cholesterol-Lowering Drug Increases Hepatitis Risk

The Australian drugs watchdog is investigating claims that a cholesterol-lowering medication increases the risk of catching hepatitis. Ezetrol (Ezetimibe, also known as Zetia in the U.S.) has been available in Australia since October 2003 and is made by US company Merck Sharp and Dohme.

Therapeutic Goods Administration (TGA) spokeswoman Kay McNiece said the drug increased the risk of catching clinical hepatitis.

"The adverse drug reaction unit has received 115 reports of suspected adverse reactions where Ezetrol is suspected of contributing to the event," Ms McNiece said. "We have 10 cases where abnormal liver enzymes or hepatitis were reported."

Clinical hepatitis has been confirmed in three cases.

"As Ezetrol is a relatively new medication all the risks associated with its use, particularly the less common ones, may not be fully identified," she said.

Ferntree Gully woman Loreto Kelly was diagnosed with clinical hepatitis after taking Ezetrol. "I think it would have killed me, I really didn't think I was going to survive," she said.

Click below to read the full story:

http://www.heraldsun.news.com.au/common/story_page/0,5478,15203139%255E662,00.html  

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Eddie Vos
How long can one flog a dead horse and hope it will spring to life and walk again?  The LRC trial of ~1984 using a bile acid sequestrant in the top 0.8% for cholesterol, average 49 year old, American males had a difference in deaths of 3, between 2 groups of n=1900 after 7.4 years, i.e.:  http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=6361299

Note that the [LDL lowering] 'efficacy' of that drug, the language used until today*) is beyond question. 

What's the rationale thinking stanols, sterols or ezetimibe [combo pill with statin:

http://www.vytorin.com/ezetimibe_simvastatin/vytorin/hcp/index.jsp ]

will save any more lives? 

*) Current AJCN has a study by VWY Lau et al from a Montreal group re sterols [-15 to -27% LDL] with the same statement .. "plant sterol consumption decreases the risk of CVD in this population [29 'hypercholesterolemic' fellow Quebeckers-]."  We knew 20 years ago reducing cholesterol uptake does not save lives.  Talking about wasting lives: what wasted research careers flogging a horse arguably first buried in 1977 by G.V. Mann in NEJM.    

On a positive note: stanol/sterol and ezetimibe are a culinary quantum leap over resin sequestrant, a possible explanation for the: " greater number of violent and accidental deaths in the cholestyramine group. " of the LRC trial.  

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Nicolai Worm
This is something to think about:

J Am Coll Cardiol. 2005 Jun 7;45(11):1794-801.  
Miettinen TA, Railo M, Lepantalo M, Gylling H. Plant sterols in serum and in atherosclerotic plaques of patients undergoing carotid endarterectomy. Department of Medicine, Division of Internal Medicine, University of Helsinki, Helsinki, Finland. tatu.a.miettinen@helsinki.fi
OBJECTIVES: The purpose of this research was to determine whether serum plant sterol levels are associated with those in atheromatous plaque. BACKGROUND: Cholesterol of low-density lipoprotein (LDL) particles contributes to atheromatous plaque formation; LDL also contains most serum non-cholesterol sterols, including plant sterols. The role of plant sterols in atheromatous plaque formation is open. METHODS: Free, ester, and total cholesterol and the respective non-cholesterol sterols were measured by gas-liquid chromatography in serum and arterial tissue of 25 consecutive patients undergoing carotid endarterectomy. The population was ranked to triads according to tissue cholesterol concentration. RESULTS: Cholesterol concentration increased markedly in tissues but not in serum with triads. The ester percentage was lower in the third than in the first triad (47% vs. 56%; p < 0.01) and lower than in serum triads (70%; p < 0.001). Ratios to cholesterol of non-cholesterol sterols decreased in increasing tissue triads, but were unchanged in serum. A major new observation was that the higher the ratio to cholesterol of the surrogate absorption sterols (cholestanol, campesterol, sitosterol, and avenasterol) in serum, the higher was their ratio also in the carotid artery wall (e.g., r = 0.683 for campesterol). Despite undetectable differences in serum and tissue cholesterol concentrations off and on statins, an additional important novel finding was that statin treatment was associated with increased ratios of the absorption sterols in serum and also in the arterial plaque. CONCLUSIONS: The higher the absorption of cholesterol, the higher are the plant sterol contents in serum resulting also in their higher contents in atherosclerotic plaque. However, the role of dietary plant sterols in the development of atherosclerotic plaque is not known  

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Leib Krut
It is amazing how things are rediscovered. I do not have references at my fingertips, but I do recall that several decades (~4) ago there was interest in feeding plant sterols with the idea that these would compete with cholesterol for absorption. This did indeed prove to be the case. Unfortunately the hopes inherent in the concept that lowering cholesterol in plasma is beneficial were dashed when it was found that feeding these plant sterols increased the plant sterol content in arteries. It was not possible to know what the relevance of this was, but it did not seem to be a plus, and, so far as I know, there was no more exploration of this topic, until now. I think Miettinen should be aware of that work done decades ago, although he might need to dig deep into his memory bank to recall those studies. ( I have not seen their paper, and they might well quote the earlier work. I shall look it up). What this work does indicate is that sterols, (and possibly other compounds) contained in LDL do become trapped in the arterial wall when LDL is trapped there. The crucial issue is: How is this relevant to our problem?

Now that I am scratching my own memory bank, I recall that Merck marketed a compound named Mer 29 about 1960 (I think), which blocked cholesterol synthesis at or near its penultimate stage. There was an accumulation of that precursor(s) in the plasma, with lowering of cholesterol, but that precursor also found its (their) way into arteries. I do not remember whether these precursors were also shown to be transported in LDL, but it seems likely they were so transported. There were a number of other deleterious effects of Mer 29 thought to be alarming at the time and it was dropped (it might have survived in todays climate!). This experience would seem to have been the reason for the search for a compound that would block cholesterol synthesis early in its synthetic pathway. And so we acquired statins! Like many of you, I expect that there will ultimately be a high price to pay for blocking cholesterol synthesis early in its synthetic pathway. 

It would seem to me that those who believe that cholesterol from plasma has relevance in atherogenesis need to establish what it is that converts it into a pathogenic moiety in the artery, how that might be prevented, and the relevance of such prevention in the epidemiology of CHD. Surely we are by now beyond the stage of putting all our focus on plasma cholesterol concentration and how to lower that concentration by every conceivable means.    

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Morley Sutter
You are absoulutely right about Mer 29 except that it was not Merck, but a company called Merrell that marketed it.  Mer 29 was taken off the market because it caused cataracts.  The suits almost broke Merck. Merrell subsequently disappeared in some merger or other.  

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Barry Groves 
And you don't have to go back 40 years. See Plat J, Brzezinka H, Lutjohann D, Mensink RP, von Bergmann K. Oxidized plant sterols in human serum and lipid infusions as measured by combined gas-liquid chromatography-mass spectrometry. J Lipid Res 2001 Dec;42(12):2030-2038.

Dr Plat and colleagues at Maastricht University’s Department of Human Biology in the Netherlands, say that plant sterols may actually be more important in heart disease than cholesterol. 

Cholesterol is only thought to be harmful if it is oxidised. Because plant sterols are structurally related to cholesterol, Plat and colleagues examined whether oxidized plant sterols (oxyphytosterols) could be identified in human blood and soya-based fat emulsions. They found that they could. Approximately 1.4% of the plant sterol, Sitosterol, in blood was oxidised. That's 140 times as much as the 0.01% oxidatively modified cholesterol normally seen in human blood. They found the same with two soya emulsions.  

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Leib Krut
Thanks for your input. It did come back to me. The compound that accumulated with Mer 29 was Desmosterol and it was found in plaques and the drug did cause cataracts.

The work of Miettinen et al shows that a number of cholesterol precursors are found in plasma and plaques. I suspect they may be implying too much. It seems likely that these precursors (and plant sterols derived from the diet) are transported in LDL and trapped with LDL in the subendothelial space of arteries. I would suggest that their findings in plaque do not reflect recent depositions. The lipid in plaque is pretty well encapsulated by fibrous tissue. There is usually no lipid in the fibrous capsule. The lipid in plaque is contained in "pultaceous necrotic tissue", according to pathologists. It is not part of a metabolic pool. This lipid must for the most part, if not entirely, be laid down early in life. It is most unlikely that its content or composition could be changed by more recent alterations in plasma, either by reductions in plasma cholesterol concentration, eg with statins, or by induced alterations in the concentration of other sterols in plasma.

The concentrations of cholesterol precursors in plasma and of plant sterols that Miettinen et al measured are an exceedingly small fraction of the total sterols in plasma, the great bulk is cholesterol. We have no idea what the presence of these other sterols means; probably nothing of note, tho' there are a number of other aspects one may infer from that study which are of more general interest.

Dear Barry: Thank you for your input and for the reference to Plat et al on plant sterols. My point really was that the possible role of plant sterols in atherogenesis is a very old idea which had not lead to any contribution to our understanding of what that disease process is about. This does not mean that it is not deserving of another look. However, just considering concentrations of compounds of interest does not seem to have lead us anywhere except into what most THINCS members agree is a conceptual morass.

I am interested in the points you make about oxidized phytosterols. I think expressing their concentration the way you have done does put a slant on things. To say that the amount of oxidized phytosterol is 140 times the amount of oxidized cholesterol will seem alarming to those who believe that oxidized cholesterol, perhaps other sterols, is/are the lethal compound/s. However, the fact that 1.4% of phytosterol is oxidized compared with 0.01% of cholesterol that is oxidized should not be alarming. It can be noted in the Miettinen paper that the amount of phytosterol in plasma or tissue relative to the amount of cholesterol is minute. 1.4% of oxidized phytosterol leaves a minuscule amount of these compounds in plasma. There is no possible way these negligible quantities of oxidized sterols could have relevance in a process such as atherogenesis. Such minute quantities of sterols might conceivably have relevance if they had hormonal implications. We all know that a huge number of factors have been implicated in atherogenesis and related issues. On reflection it is perhaps surprising that no one has as yet (at least to my knowledge) implicated a steroid hormone, conceivably derived from ingested sterols (!), in its genesis. Perhaps that will still be done down the road; anything goes in this field.

I might add that a possible reason for such minute quantities of oxysterols in plasma is attributable to the extraordinarily rapid rate at which oxysterols are cleared from plasma relative to cholesterol. (Krut et al. Arterioscler Thromb Vasc Biol. 1997;17:778-785, Correction 1979;17:1481) The actual amount traversing plasma might be more substantial.

Raising the spectre of oxidized cholesterol does give me the opportunity to plug my views. Oxysterols, as distinct from cholesterol, were first conceptually implicated in experimental atherogenesis by Altschul around 1946. Since then it has over the years been reported by several groups of workers, including work by Altschul, that oxysterols do the reverse. They in fact attenuate lesion formation in cholesterol fed animals. The work implicating oxysterols in atherogenesis would seem to be questionable.

I have postulated that cholesterol from plasma becomes a pathogenic moiety in the arterial wall when it takes on its native character, which is that of a crystalline solid. In this state cholesterol cannot be cleared from tissue and it is sclerogenic. Thus factors that promote or prevent the crystallization of cholesterol determine its role in atherogenesis. This view is based on observations originally made by chance, namely, that oxysterols prevent the crystallization of cholesterol in in vitro systems, and that glucose promotes the crystallization of cholesterol. In addition, oxysterols implanted subcutaneously in rats together with cholesterol results in the solubilization and clearance of a large mass of cholesterol, leaving little residual fibrosis. Implanted pure cholesterol is rapidly sequestrated by fibrous tissue, no cholesterol is cleared and it remains sequestrated in a seemingly permanent granuloma.

The possible relevance of oxysterols to atherogenesis in humans was attributed to their progressive elimination from the human diet since the advent of refrigeration, which prevents the spontaneous oxidation of cholesterol in foods of animal origin. Oxysterols must have been invariably generated in such foods by the techniques practiced for preservation of these foods prior to the development of refrigeration. Oxysterols would have been progressively eliminated from the human diet with progressive application of modern technology in preservation and handling of foods.

You can find references, should you want to follow up on these matters, in: Krut L H. Med. Hypotheses 1979;5:533-548, Atherosclerosis 1982:43;95-104 and 1982:43; 105-118. Recent Res. Devel. in Lipids Res 2, 1998:299-318. Amer J Cardiol 1998:81;1045-1046. Atherosclerosis supplements 2004;5/1:36. Wilkens and Krut. J. Atheroscler Res 1963:3:15-23 and 1965:5;516-523.  

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Bogdan Sikorski  
Leib - When some months ago, during your introduction to the group, you described your research hypothesis about the positive role of oxchol in atherosclerosis, I could "feel" the unease it created - no one, including me, dared to question you on that. Perhaps "dared" is a wrong term, but certainly the typical skepticism of the group was surprisingly dormant.
Now, what you have just again described really makes sense to me (I hope to stir some unease here) considering that I am of the firm belief, or rather conviction, that circulating chol can not have a major influence, if at all, on the pathogenesis of atherosclerosis or formation of the plaque. Whatever the precise mechanism is, if there is just one, the in situ production appears logically to be the major source of chol with a major influence of glucose and insulin, and possibly other factors, all in response to an injury or anoxic assault of some form. This fits in with your notion about the positive role of oxchol in mopping up the injury site in an attempt to restore the functional integrity of the blood vessel. The putative role of glucose and insulin on chol formation in situ has been first documented by Stout in early 1970s using radioligands.

I might add that another experiment which showed that chol can be made in situ, in media, before the arrival of macrophages, subject to anoxic assault, has been done on aortae of living rabbits (this time I am confident also without feeding chol) fitted with inflatable cuffs, after separation from vasae vasorum.  I do not know if these results were ever published, since I heard them reported at a scientific meeting some year ago. I forgot how they differentiated between the in situ mechanism and infiltration of chol from the lumen, but I remember that I asked the experimenter if they used statins to block the in situ synthesis - they did not at the time, but planned to do so.

Anyway, contrary to popular modern dietary habits, I and my family must be just about the only human beings, apart from the remnants of  traditional Northern Indians, who are happy to consume buckets of ghee, which as a rule must be relatively high in oxchol. We (well, my wife) make our own, to which we also add some coconut oil, which makes it a perfect frying, non smoking, fat.
Oh well, this week for a change we will have a bucket of freshly melted lard, after procuring some non-smelly pork fat from a Chinese source. Hopefully, that should also be oxchol rich due to the high-temp melting process.
Here is hoping for clear arteries!  

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Alena Langsjoen
Dear Jacqueline, Zetia (Ezetrol or Ezetimibe) is a pretty strange looking molecule which somehow is supposed to block absorption of dietary cholesterol along the brush border of the small bowel. Here in the US it is marketed by itself and also in combination with Zocor (this combination drug is called Vytorin). Peter has been worried about Zetia and has not prescribed it to any of his patients. But some of his patients are advised by their primary doctors to take it, or worse yet, to take Vytorin which obviously is a double whammy.

If Zetia blocks the absorption of cholesterol, does it also block absorption of dietary fat-soluble vitamins, like vitamins D, E, betacarotene and lycopene, essential fatty acids and also how about our favorite molecule, the fat-soluble CoQ10-some of which we get from our diet? The PDR states that (this is not an exact quote) after 14 days supplementation with Zetia, there is no clinically significant depletion of fat soluble vitamins. Peter and I are now able to analyze patient plasma in our laboratory with our new HPLC system equipped with electrochemical detector (ECD) and UV detector (UVD). Our ECD produces a graph with peaks for vitamin E, lycopene, beta carotene, reduced CoQ10 (H2CoQ10 or ubiquinol) and oxidized CoQ10 (ubiquinone). Our UVD produces graph with peaks for free cholesterol and several cholesteryl esters (linolenate, arachidonate, linoleate and oleate).

Even though I tried objecting to Peter doing this (I like for Peter to stay healthy) towards the end of last year Peter decided to do an experiment on himself by taking Zetia for 2 weeks. We ran his baseline plasma and again after 2 weeks on Zetia. FYI, after I post this I will upload pdf of the resulting analysis of his plasma. First page of the pdf is his plasma while he was taking 600mg CoQ10/day for at least a month (no other supplements). The second page is his plasma after 2 weeks on Zetia in addition to the 600mg CoQ10/day. Two weeks was all he could stand taking the Zetia. Note that his total Q10 (oxidized + reduced Q) level dropped by 43%. And his vitamin E level dropped by 33%.

I cannot quantify the lycopene or beta carotene peaks because I don't have standards for them yet, but after 2 weeks on Zetia Peter's area under the peak for lycopene dropped by 26% and for beta carotene by 48%. It is noteworthy that the cholesteryl esters also dropped, along with the free cholesterol. We do not know how this relates to the status of plasma levels of free fatty acids. Note the total Q10/total cholesterol ratio dropped (by 18%).

Next we thought it would be very interesting to find out what effect Zetia has on people who are not supplemented with CoQ10 so we are recruiting some volunteer doctors and we are in the process of
collecting data. We plan to publish the results.  

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Bogdan Sikorski
Alena - This is very interesting. I hope I can show these HPLC results to few people, but a published study would of course be very powerful indeed. In AUS, the dietary use of phytosterols has been thus far restricted (because of effects on fat-sol Vits) to "healthy" margarine, but the "good" industry has been pushing to have these timber milling byproducts (i.e. normally environmental pollutants) to be included in a wide range of products, including kids "health" bars and milk-drinks for some time. I think, the US FDA being more "progressive" has already allowed a relatively wide use of phytosterols in foods such as milk and soft drinks. Treatment with these could be another arm of your study after a 2-week washout period.   

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Alena Langsjoen
We hope to publish our Zetia findings, but it may be a while before we get to that.  I'll see if I can talk Peter into eating a bunch of phytosterols for an experiment.  Actually right now we really want to concentrate on measuring tissue levels of CoQ10 and correlating that to plasma levels.  We hope to start doing this sometime this fall or winter.  A local cardiovascular surgeon is willing to cooperate with us, giving us a snip of the heart along with a concurrent blood sample.  

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Eddie Vos
Alena, It is wonderful that after blowing the kids' college fund to a chromatograph it is finally spitting out reproducible results!  You did not say what dose zetia Peter was on, I presume 10 mg and where the drop in total cholesterol reported in the P.I.., below, is more like -14%, your finding was -30%  .. http://www.zetia.com/zetia/shared/documents/zetia_pi.pdf  ; other links: 

http://www.vytorin.com/ezetimibe_simvastatin/vytorin/hcp/price_dosing/dosing.jsp

http://www.zetia.com/ezetimibe/zetia/hcp/index.jsp  

It is also amazing to me that cholesterol in serum marries so massively to linoleic acid [excess n-6 in the diet?] and also that the on ezetimibe test showed n-3 linolenate cholesterol ester dropped from 0.04 to 0.01 mMol, but I guess that could be limit of detection, or less alpha-linolenic acid in the diet the day before since it does not store for long.

Final comment: I would not dream about going a day without a high dose multivitamin to keep my homocysteine at a low level; Peter being on no other supplement than Q10 sounds like he's missing something, but that's the subject of a next email.  Best to you both and exiting stuff that new toy!   

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Alena Langsjoen
Peter's Zetia dose was 10mg/day.   

Yes, we're having lots of fun with our new expensive HPLC toy after some stressful times with it.  I agree that the n-3 linolenate chol. ester is very close to detection limit, not only because its concentration is so low but also the peak is not very well separated from cholesteryl arachidonate which comes off right after the chol.linolenate. I got a big bottle of B-50 complex from Sam's on Peter's side next to his sink in our bathroom now and I try to remind him to take one every day.   

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Melchior Meijer
What a creative and courageous experiment. I have some questions. Zetia (Ezetrol) blocks (re-)absorption of cholesterol from the gut and has (unlike statins) no influence on endogenous Co Q10 synthesis. The fall in Q10 in Peter's plasma is thus a reflection of Zetia blocking the uptake of the Q10 supplement and Q10 is his food, right? 

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Alena Langsjoen 
Yes, we think that Zetia blocked the absorption of Peter's large doses of supplemental CoQ10 and it also blocked the ability to carry Q10 (Q10 gets carried by LDL & VLDL).  At 600mg/day you could say that CoQ10 could almost be Peter's food but he still has a good appetite for eggs, butter, steak and all of my Czech cooking!  

Some of our current observations are making us believe that dietary CoQ10 may be much more important than previously thought. 

Nakamura has performed experiment with radioactively labelled supplemental CoQ10 in animals and there was no decrease in endogenous biosynthesis of CoQ10.  I can find the reference if you need It.  

Peter's side effects were weakness, fatigue and some GI distress. Peter said that he would think about experimenting with Becel Pro Aktiv.     

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Leib Krut
Dear Bogdan: Thank you for your note. I must say I was quite tickled to read of the reaction you felt occurred in the THINCS group in response to my concepts. I am not unaware of the general reaction to my "heterodoxy". I must say that I had hoped that in a group of "heterodox" thinkers there might have been more interchange with another view that was very different from the mainstream.

I moved to the USA about a decade ago and my reprints of articles got into a mess, beyond recovery. I mention this because I should have liked to give you a reference to a paper published in the British Heart Journal many years ago by someone named Malhotra (I think). He wrote on the difference in the incidence of Coronary Heart Disease between Northern and Southern Indians in India. He reported

that the southerners were vegetarian and lean and had a CHD incidence 7+ times that of the northerners who were much heavier and who ate a considerable amount of fat (10-20 times that in the south), that this fat is mainly from animal sources, including ghee, and therefore largely saturated. In the south the small amount of fats are mainly from seed oils and therefore largely polyunsaturated.He described the way ghee was made, something your wife clearly knows well, and I have no doubt that oxysterols are generated in the process since they are found in butter made by traditional methods in which cream is allowed to "ripen" simply by holding it at ambient temperature for 3-4 days. This cream is then churned to make butter. I suspect that in making ghee even more oxysterols are generated. I might add that the northerners smoke 8 times as many cigarettes as do their southern neighbours. They would seem to be doing all the wrong things in the north, according to the standard dogma, and yet are protected from CHD, save for the fact that must have a considerable intake of oxysterols, or at least must have done so when Malhotra wrote that paper. It goes without saying that I found that paper very appealing and that clearly is why I recall it so many years after its publication. It may be that I cited it in one of my publications. If I do come across it I shall let you have the reference. As a potential ally, I shall dearly love to provide you with as much information as possible to encourage your support.

I did in fact cite that paper. The reference is: Malhotra SL. Brit. Heart J. 1967;29:895-905.

I think you will find it of interest.  

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Barry Groves
I have a paper of Malhotra's on this subject, but published in Am J Clin Nutr (Malhotra SL. Serum lipids, dietary factors and ischemic heart disease. Am J Clin Nutr  1967; 20: 462-474). In it Malhotra states " . . .occurrence rates of acute myocardial infarction were seven times higher in the South Indians as compared to the North Indians, even though the North Indians consumed nine times more fat, most of which was animal fat derived from milk and ghee, with a preponderance of saturated fatty acids" He gives as a reference for this statement his own paper, Geographical aspects of acute myocardial infarction in India, with special reference to the pattern of diet and eating. Brit. Heart J. 1967;29:777.

Malhotra notes that in the comparison between the two groups, serum cholesterol levels were similar and normal values. Free fatty acids were non-significantly higher in the Northern Indians, but mean values for esterified fatty acids, cholesterol esters and total triglycerides were similar in both groups.

What this study shows is that serum lipid levels can be the same and normal in peoples with very big differences in both their intakes of fats, including animal fats, and incidences of IHD. It also demonstrates that serum lipid levels are not dependent on totals or even proportions between different fats eaten.

Exercise and physical activity are also considered by Malhotra, but he found no significant difference between the two groups -- they both were "habituated to an identical amount of a high degree of physical exercise at work . . ."

After measuring fecal and urine urobilinogen excretions, he puts the blame for the South's higher CHD rates on excess bile entering the intestinal lumen. "These results . . . show unequivocally that out South Indian group on a carbohydrate-rich, lipid-poor regime of boiled rice and lentil soups had significantly larger amounts of bile in their intestinal lumen, as compared with our North Indian group on fat-roughage and cellulose-rich wheat diet; and that these differences are dependent upon the pattern of diet and eating."

Malhotra also talks about other studies that buck the trend -- Bantu, Masai and Samburu in Africa.  

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Bogdan Sikorski
I recall, that we have discussed this paper, or rather these findings, in the past, on more than one occasion.
I think, I looked for the journal in the departmental (work) library and could not find it.  It would be nice to get a copy (scan?) of both.
Anyway, Leib, I can hardly be your ally, considering that I am "running on empty" - I have done no research (except for a dietary trial we are running at home) in the area, but I am very interested in your findings and the "fringe" or heterodox hypothesis you have constructed based on them.  Ultimately, it would be nice to know that our dietary preferences have some scientific support.
As far as I can tell, many members of this group are on a range of fringes, having adopted one or more heterodox positions, with one uniting us all. But, as with any group of people, one has to be aware of not pushing too hard!  In such cases, the polite response is typically no response or no comment.
Being brought up Central European, I don't have much respect for such politeness  - thus my previous comment.  Lets "drill" this hypothesis of yours.  Perhaps Uffe or Malcolm should start?  Ooops - maybe someone less "polite" first? Did I offend anyone?  

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Uffe Ravnskov
First a comment to Bogdan. Hopefully no one in this group who disagrees with thoughts presented in our correspondence would remain quiet because of politeness. I think not, because previous discussions bear witness to many divergent thoughts about this and that and hopefully we shall proceed that way.

As for Leib´s idea about oxidized cholesterol I have speculated much about it without coming up with any wise thoughts, mainly because my biochemical knowledge is too rudimentary. Personally I think that the idea about oxchol being the villain is a typical ad hoc hypothesis. As far as I know there is no evidence of that besides the finding that high oxchol is a better predictor than high cholesterol, but there are probably more than hundred other risk factors that are better predictors than high cholesterol so why just oxchol.

I recently read The Dynamics of Atherosclerosis (Aberdeen University Press, Aberdeen 1976), a most interesting book by British pathologist Jack Duguid.  His hypothesis is that atherosclerosis is the sequels of microthrombi formed at the intimal surface. Very quickly such thrombi are covered with endothelial cells. Their repeated incorporation into the intima throughout life beginning in early childhood eventually results in irregular fibrous thickenings that are too stiff to comply wih pulse movements and so cause disruption and haemorrhage. The extravasated blood disintegrates leaving fatty deposits which accumulate progressively. Thus, plaques are simply scars and he has illustrated his idea with many compelling microphotographs.  

The crucial question is of course, what is causing these microthrombi? Here is room for many suggestions, for instance too much homocysteine, free radicals, microorganisms, etc and none of them exclude any of the others; a complex interaction is likely. 

The finding of plant sterols in atherosclerotic plaques is amusing but didn’t surprise me at all. It is amusing because of the authors´ conclusion. If they think that high cholesterol leads to atherosclerosis, which they obviously do, then why don’t they follow the line and warn against plant sterols? Or have they realised that the finding of a certain substance in a scar doesn’t mean that the same substance is causing the scar?   

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Paul de Groot
Reading the discussions on atherosclerosis (2002-2005), my main interest, some statements are still relevant. I like to comment on two of them. Firstly: intimal enlargement is the first sign of atherosclerosis and secondly: atherosclerotic plaques seem randomly distributed over the arterial wall apart from the branching sites.

My studies of human coronary arteries contradict the first statement while I endorse the second one.

I tell you my story and will try to formulate the way of thinking.

1984 I planned my thesis study on human coronary artery vasa vasorum. The heart foundation  was sceptical about even the existence of vasa vasorum in the coronary arterial wall. The intention was to study the normal coronary artery and the artery circumferentially enlarged, then called hypertrophied. Mind you, in literature, no definition of “normal” could be found and so also not of concentrically enlarged vessels. Of course some histological indications could be found but no hard figures at all. We came up with the following histological classification:

Coronary artery                   N                    AN                 A                    E
INTIMA
Endothelium                          undamaged     undamaged     undamaged     damaged
Sub-endothelial
Layers                                  some              several            many              many
Thickness
Intima/media                         <1/2 media      in between      >1/2 media      >>1/2media
Conc.c.q. ecc.                      -                     conc.              Conc.             Ecc.
Smooth

Internal Elastic Lamina
Single/double                        single             single/double   double            double
Continuous/Fragmented         cont.               cont.               fragmented      fragmented
Undulated/Stretched/
Destroyed                             undulated        stretched        stretched        destroyed

Extenal Elastic Lamina
Changed/Unchanged              unchanged      unchanged      unchanged      unchanged 

VASA VASORUM
Intima/Media/adventitia           adventitia        adventitia        adventitia        intima + adventitia
Outer/inner Border adventitia   outer               inner/outer       inner               inner 

N=normal, AN= between N and A, A=concentrically enlarged, E=eccentrically changed coronary artery 

Morphometrically intimal-, medial and adventitial-width was assessed and to make different sized vessels comparable the relative figures were used: thickness intima/r-lumen, thickness media/r-lumen and thickness adventitia/r-lumen. This resulted in the following mean figures of Th-int, Th-med and Th-adv in micrometers:

Coronary artery                   N                 AN                 A
Th-int                                    23                  136                 199
Th-med                                 77                  136                 141
Th-adv                                  120                 185                 211 

In this way coronary arteries N, AN and A can be identified in numbers. 

The aim of the follow up study was to investigate coronary artery remodelling not only in vessels N, AN and A but also E. Arteries eccentrically enlarged can be divided into vessels containing a part N opposite of the plaque (EN), a part A opposite the plaque (EA) or the plaque is circumferentially (EE). Serial radial measurements were performed and Th-int/r-IEL was used in stead of Th-int/r-lumen  as a measure of the arterial pathology. The following figure shows the correlation between the histological- and measured classification.

It appears the correlation is rather good. The data of Th-int/r-IEL resembles the atherosclerotic process: normal arteries <5, concentrically changed arteries 5-19,9. 20-49,9 the arteries change from concentrically enlarged into eccentrically and the last phase >50 destruction of the arterial wall and closure of the lumen (TH-int/r-IEL 100). In this way 4 stages have been defined. The next figure shows the mean values of r-lumen, Th-int, Th-med and Th-adv for each stage.

?

This figure shows the increase of intimal width (Th-int/r-IEL 5-19,9) accompanied by growth of the media and adventitia and also r-lumen (strong positive remodelling). Or is the increasing media-adventitia width accompanied by intimal growth? In any case the first stage of atherosclerosis (pre-atherosclerosis?) concerns the entire coronary arterial wall which finding I can not relate to “atherosclerosis starts in the intima”.

The focal nature of the atherosclerotic process becomes clear when the segments Th-int/r-IEL <5 – 19,9  from coronary arteries E are compared to the vessels. The next table show the result.

The vessels and segments seem to be similar and so atherosclerosis affect only a part of the vessel wall which proves the focal nature not only lengthwise but even circumferential.  

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Eddie Vos
I just finished first reading of Paul's entire dissertation [thank you for the copy], a handsome histology text indeed. My initial take home impression was that intima enlargement was always found, and I thought early, but that outer changes happen also.

I explained this as possibly caused by a decline of the extra-cellularmatrix, i.e. the elastins, collagens and associated cartilage [proteo-glycans], but the dissertation did not deal with the chemistry of these structural components.   If this comes first [and as per Kilmer] through the myriad ways by which homocysteine attacks these structural layers, such would explain I think the observations of Paul.  Since there is MORE structure in the intima than in the media [cells and elastin structure] and adventitia [structure, cells, fat, vasa vasorum et al, it makes sense that the health of intima structure is vital and an early factor.

Things crystallize in one's mind when realizing that both collagen and elastin are kept together by lysine based linkages, and that homocysteine messes with the lysine AND deactivates the enzyme needed for the linking [lysyl oxidase], and that nothing much happens in linking without sufficient vitamin C and copper.

I was not clear if the vasa vasorum [the vessels of the vessels .. feeding cells, not 'dead' structure like collagen] that Paul looked at was mainly the 20-50 micron diameter type that does not 'invade' the media leave alone the intima [only after elastic barrier fibers or net- and sheet-like elastins are destroyed] and if the vasa vasorum looked at included the 5 micron diameter network of capillaries [marginally smaller than a  flat red cell but >150x larger than LDL]. 

Paul does not say what is in an intima at age 12 but it seems that more cells appear after age 20, his 2 earliest samples of autopsy hearts. Others have suggested that the intima is initially essentially a cell free ayer -only structure, and thus not in need of a permanent blood supply, and not needing an inflammatory defense mechanism.

I would imagine that the dozens of concentric muscle cell layers of the media of larger arteries -separated by a layered network of elastins- does need some capillary blood supply at some early stage.  Paul finds that vasa vasorum infiltrates towards the inside only when structural layers -internal elastic membrane for example- are destroyed.

May I refer you to the first paragraph of http://www.health-heart.org/comments.htm#11 and solicit comments and to the last image on my home page http://www.health-heart.org where I attempt to put things together as to cause.  What should I change?    

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Paul de Groot
The vascularity of the media has been debated fiercely in early days. The arterial media of several animal species contain vessels. However, the anatomy of the media is also different: laminated. Vessels are localized between the lamina. The human coronary arterial media is avascular that is to say I have never seen any vessels using light microscopy. So the structure of the "organ" vesselwall is peculiar consisting of two avascular layers (intima, media) and one vascular layer (adventitia). With that the nutrition of the intima and media depend entirely on diffusion. Although peculiar it is not uncommon for example the eye's cornea and cartilage. It is said the intima plus 1/3 media are dependent on diffusion from the lumen
and the other part from the vasa vasorum in the adventitia. The nutritian of Eddies poor myocyte in the middle of the media is in fact subject to diffusion distance and diffusion gradient, a process easily disturbed. To keep vessel wall homeostasis with an increase of vessel wall width vasa vasorum increase in number and size and grow towards the lamina elastica externa minimalising diffusion distance. This repair mechanism is of use for as long as EEL and media are intact (Th-int/r-IEL 5-49,9). Further increase
of intimal thickness and with that Th-in/r-IEL is accompanied by media destruction and vasa vasorum growth into the plaque. By now I will not comment on the cause of this process (homocysteine?). At
first I want to sow doubt about the primary role played by the intima in the process of atherosclerosis.

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Uffe Ravnskov
As far as I have understood from your dissertation, your mathematical data gave no support to the idea that the primary event in atherosclerosis is damage to the adventitial vessels. But then, why do you ask: Does it really start in the intima??? Your finding that all vessel walls increase, not only the intima, does not contradict that plaque formation starts from the inside, because in your calculations you have only used data from normal vessels and from vessels with concentric stenosis and excluded those from vessels with eccentric ones. As you suggest yourself, eccentric and concentric stenoses most probably have a different pathology. Furthermore, it has to be proven that concentric stenosis has any clinical importance. Isn´t it the eccentric stenosis that matters, the plaque, or better the vulnerable plaque, or the
acute thrombosis? And whatever term you use, these are situated just beneath the endothelium, a strong support to the view that atherosclerosis starts from the inside, at the endothelial surface.

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Paul de Groot
The vascularity of the media has been debated fiercely in early days. The arterial media of several animal species contain vessels. However, the anatomy of the media is also different: laminated. Vessels are localized between the lamina. The human coronary arterial media is avascular that is to say I have never seen any vessels using light microscopy. So the structure of the "organ" vesselwall is peculiar consisting of two avascular layers (intima, media) and one vascular layer (adventitia). With that the nutrition of the intima and media depend entirely on diffusion. Although peculiar it is not uncommon for example the eye's cornea and cartilage. It is said the intima plus 1/3 media are dependent on diffusion from the lumen and the other part from the vasa vasorum in the adventitia. The nutritian of Eddies poor myocyte in the middle of the media is in fact subject to diffusion distance and diffusion gradient, a process easily disturbed. To keep vessel wall homeostasis with an increase of vessel wall  width vasa vasorum increase in number and size and grow towards the lamina elastica externa minimalising diffusion distance. This repair mechanism is of use for as long as EEL and media are intact (Th-int/r-IEL 5-49,9).Further increase of intimal thickness and with that Th-in/r-IEL is accompanied by media destruction and vasa vasorum growth into the plaque. By now I will not comment on the cause of this process (homocysteine?). At first I want to sow doubt about the primary role played by the intima in the process of atherosclerosis.

Eddie Vos
So, intima and 1/3rd of the media is nutrient and signal supplied by diffusion from the inside and the balance of the artery wall from the outside. So, are you saying that IF there would have been 5 micron diameter capillaries in the media, you would have seen them -and that BEFORE the EEL [external elastic lamina/layer] deteriorates around the media, there is no infiltration of capillaries into the media? 

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Paul de Groot
I owe several of you answers to questions posed on my, perhaps, provocative saying “does atherosclerosis really start in the intima???”  I tried to sow some doubt on the subject and now will try to be more specific. Of course Uffe the fact that all three layers of the coronary vessel wall thicken at the same time does not imply the cause is not in the intima. My supposition is “the start of atherosclerosis is in the adventitia”.  Now you all frown and accuse me of fantasy. Here me out! In literature much evidence is available on the subject. Nakata (1) showed experimentally (1967) obstruction of vasa vasorum cause intimal lesion similar to early atherosclerotic lesions. Gutterman (2) induced intimal and medial hyperplasia by dissecting the adventitia from arteries while in the rabbit (3) removal of the adventitia intitiates intimal proliferation and regression of the lesions on regrowth of the adventitia. The same phenomenon is seen with positioning a hollow silastic inflatable collar around the artery (4). At the other hand an external collar inhibited balloon induced intimal hyperplasia (5) while in a pig model external stenting reduced medial and intimal thickening and growth factor expression (6). All right you say all these experimental findings have nothing to do with the “normal” process of atherosclerosis and manipulating endothelium can also induce atherosclerotic like lesions. How about adventitial fibroblasts migrating to the intima and playing a role with neo-intima formation (7). I will not tire you with a full survey of literature much more can be found.  

In my opinion arterial wall hypoxia, disturbance of vessel wall homeostasis, could explain the findings. Hypoxia inducible factor can cause arterial wall hyperplasia. So if I am right the arteriosclerotic process to start in the adventitia the first question you ask is what changes the adventitia? My answer would be inflammation. If you can digest the foregoing I can state my case on inflammation an other time. 

The normal coronary artery can change in different ways: concentrically enlarged and later on eccentric grow or directly from normal into eccentric plaque. I did measure eccentric coronary arteries Uffe as was shown in one of my attachments. I did not emphasize then that coronary arterial remodelling is Th-int/r-IEL dependent and in this way positive- and negative- remodelling can take place in the same artery. This could provoke plaque instability. With these plaques inflammatory infiltrates are found at the shoulders of the plaque but situated in the adventitia. As a matter of course I do not negate the role played by the endothelium but I see that as secondary to the adventitial process. Perhaps that also is the answer to Leslie: adventitia induced hypoxia as a result of inflammation give rise to HIF (hypoxia induced factor) after which the entire process starts.  

1. Nakata Y e.a. Vascular lesions due to obstruction of the vasa vasorum. Nature 1966;212, no.5097
2. Gutternan DD e.a. Adventitia-dependent influences on vascular function. Am J Physiol Haert Circ Physiol 1999;277(4):1265-1272
3. Barker SG e.a. The adventitia and atherogenesis: removal initiates intimal proliferation in the rabbit which regresses on generation of nea adventitia. Atherosclerosis 1994;105(2):131-144
4. Loo van der B e.a. The adventitia, endothelium and atherosclerosis. Int J Microcirc Clin Exp 1997;17(5):280-288
5. Fogelstrand P e.a. External collar inhibits balloon-induced intimal hyperplasia in rabbits. J Vasc Res 2002;39(4)
6. Dheeraf M e.a. External stenting reduces long-term medial and nea intimal thickening and platelet derived growth factor expresson in a pig model os arteriovenous bypass grafting. Nature Medicine 1998;4(2)
7. Saverio S e.a. Contribution of adventitial fibroblasts to neaintima formation and vascular remodelling. Circ Res 2004;89:1111  
   

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Bogdan Sikorski
Dear Paul - Even though I got somewhat lost in your initial paper (too many abbreviations and tables for my liking), I am firmly with you on that hypothesis, and as I have indicated to the group on occasion, I have strong reservations about the source of the cholesterol in the arterial wall being blood from its lumen - I propose it is made right there - in situ. As has been shown, BBB is virtually impervious to cholesterol (brain has to make it on the spot), and BBB closely resembles endothelial lining, if I correctly remember.
As I have mentioned recently, I have seen presentation(s) on external arterial cuffs causing atherosclerotic-like changes in rabbits, including accumulation of cholesterol and various cell infiltrates, all because adventitia and/or vasa vasorum were disrupted/cut.
Again, as shown by Stout in 70s, glucose and acetate showed up in aortal wall as cholesterol.  Glucose is an excellent oxygen donor, and cholesterol is a "healer" in the oxidative stress, as in inflammation. All fits nicely!
I my previous life, I was vivisection pharmacologist and cannulation and isolation of small arteries (carotid; renal) and aorta was a breeze! Oh, I almost feel like doing it again to those poor ratties and piggies.
Have you tried in your experiments on animals, assuming you have done some, to find out if when a cuff is applied to the artery, a statin given IV makes a difference in the cross section picture and cholesterol accumulation of the artierial wall?

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Paul de Groot
Dear Bogdan, that makes two believers. I agree fully the puzzle pieces fit nicely if the atherosclerosis story is told starring the adventitia. Unfortunatally I did no animal experiments, in a hospital animals are not desirable. Formerly tissue samples could be obtaind easily from obductions and so all my investigations concern humans. Perhaps a next time I will try to describe my hypothesis as to the cause(s) of adventitial changes.

Bogdan Sikorski

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Björn Hammarskjöld
Why use cuffs to cause atherosclerotic-like changes in rabbits?  Isn't it enough just to have a high level of glucose in the vessel? 

See Josephine M. Forbes et al in Diabetes 51:3274-3282, 2002 
and other papers like 
S. Kooptiwut et al.High glucose-induced impairment in insulin secretion is associated with reduction in islet glucokinase in a mouse model of susceptibility to islet dysfunction. J. Mol. Endocrinol., August 1, 2005; 35(1): 39 - 48[Abstract] [Full Text] [PDF]. 
J. M. Forbes et al. Advanced Glycation End Product Interventions Reduce Diabetes-Accelerated Atherosclerosis. Diabetes, July 1, 2004; 53(7): 1813 - 1823. 

Glucose binds nonenzymatically to proteins and destroys the thee dimensional structure as well as changing water solubility.This can cause cell damage which in turn cause vessel wall damage and atherosclerosis. THINCs about it!

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Uffe Ravnskov
I doubt that anyone in our group believe that plaque cholesterol comes from blood cholesterol sieving or transported through the arterial wall. My problem with the idea that the primary damage happens in the adventitia is purely based on common sense. Isn´t the main reasons why arteries are atherosclerotic but not veins the intraluminal pressure? And I assume that the pressure in the adventitial capillaries is only a little higher than the venous pressure. What is wrong with Duguid´s hypothesis (see my previous letter)? To avoid misunderstandings – I do not think that the high pressure is the main cause or that high pressure by itself causes atherosclerosis; rather that it predisposes to damage from whatever cause.

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Paul Rosch
Uffe:  I agree.  Sustained hypertension causes stroke but is an associated "risk marker" rather than a causative "risk factor" for coronary atherosclerosis and the same is true for cigarette consumption, which can cause cancer of the lung and emphysema.  The MRFIT study clearly showed that lowering elevated blood pressure, cholesterol and reducing smoking alone or in combination did not lower the rate of heart attacks.  On the other hand, heart attacks were higher in men with certain Type A traits and the WCGS clearly showed that Type A behavior as assessed by the structured personal interview was as significant a "risk factor" for CHD as hypertension, cholesterol and smoking and was also completely independent of these.  What is important about this observation is that although hypertension is not a hallmark of Type A, such individuals do show hyperactive and exaggerated blood pressure responses to stressors and it is these repeated surges that probably damage the intima and predispose to the development of plaque.  As Jim Lynch and I have shown, everybody's blood pressure spikes as soon as we start to speak and the magnitude of this is affected by speed and volume of speech, the perceived relative social status of the audience, the content of the conversation, presence of a pet and other factors.  Although these elevations can be alarming at times, patients have no perception of this and are not aware whether their blood pressure is high, normal or low.  The higher the resting blood pressure, the greater the rise when you start to talk.  No antihypertensive medications are capable of blunting these surges and beta blockers actually accentuate them.  Conversely, blood pressures fall below basal levels when one is listening to someone else or is silent and attending to something in the environment, such as watching tropical fish in a tank.  Note also that the customary rise in blood pressure with age is not seen in secluded orders of nuns who rarely speak and occupy themselves mostly by tending to their plants or crops.

Type A's are poor listeners because they tend to think about what they are going to say next and when to interrupt others in order to emphasize their own points.  One of the most defining Type A traits are vocal stylistics and speech patterns that include rapid, forceful and "plosive" speech that results from inhaling large amounts of air and expelling it while talking to provide further emphasis.  These "plosive" and other speech characteristics seen in Type A's not only lead to proportionately greater increases in blood pressure while talking but blood pressure also fails to fall back to basal levels when they stop because instead of listening they are thinking of what they are going to say next.  Thus, they are caught in an upward spiral of increasing blood pressure surges the longer they continue to talk or try to communicate with others. Most physicians are unaware of this since silence is built into the auscultatory measurement of blood pressure with a stethoscope but is readily demonstrated with the Dinamap we use or other automated computerized devices.  Similarly, ambulatory monitoring studies confirm that the highest blood pressures are seen while talking to someone and especially during phone conversations discussing something that is controversial or stressful.
Deaf mutes show the typical blood pressure surge as soon as they start to communicate with someone by sign language but not when they move their hands vigorously in a meaningless fashion.  The only exception is seen in schizophrenics, who also tend to be hypotensive.  The explanation for this is too complex to discuss here but there is a detailed discussion in The Language of the Heart that also reviews our research results noted above.

Jim and I are quite confident that the link between Type A and coronary heart disease will prove to be these repetitive spikes in blood pressure that damage the inner surface or adventitia of coronary vessels that result from Type A vocal stylistics and poor listening habits.  These and some of our other research findings have led to the development of a very successful non pharmacologic treatment for hypertension by teaching patients to get in touch with their feelings and how to reduce blood pressure surges while talking.

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Leib Krut
The idea that cholesterol from plasma could contribute to cholesterol in the atherosclerotic plaque via the adventitial capillaries is not tenable. It has long been known that the inner half to 2/3 of the media is devoid of capillaries ( and probably lymphatics) and at least one of our contributors has recently pointed this out. This is unlike the situation in veins, even including the pulmonary artery, which do indeed contain capillaries, and probably lymphatics, across the whole of the media. When there is pulmonary hypertension (among other things probably causing compression of the low pressure capillaries and lymphatics in the inner wall) the pulmonary artery becomes susceptible to atherosclerotic lesions.

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Paul de Groot
Uffe, of course it is unbelievable cholesterol is transported from the adventitia to the intima and is not what I mean. What I really believe is (primary) adventitial inflammation resulting in (secundary) permeability of the endothelium after which (perhaps) ox-LDL enters the intima etcetc. Soon I will try to compose my hypothesis.

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Eddie Vos
I hate to be a 'stick in the intima" and while swellings anywhere can obstruct and cause pain, angina is just a warning and does ot kill. It is important to know where decline and blockages start but more important WHY, and it is only when the STRUCTURE of the intima breaks open, an engineering problem, that one initiates a heart attack. From page 2 of PWF Wilson's 'Atlas of Atherosclerosis' re the intima of the 'normal' artery: "1.) the endothelium resting on a thin basal lamina; 2.) the subendothelial space containing thin elastic and collagenous fibers and proteoglycans and; 3. longitudinally oriented smooth muscle cells in large muscular arteries." [the latter would be the 'musco-elastic' layer of the intima as per another 'atlas']. My point and I believe Kilmer's would be that as long as you don't damage the 3 components of item 2 in the 'subendothelial space' with homocysteine [thiolactone], your collagen and elastin should never fragment and weaken and cells and repair mechanisms have no reason to move in and try to remedy. I think the issue is: what is the precise composition of the intima of a micronutrient replete (representing homocysteine <~6 µmol/L) twelve [12] year old. The trick then is to maintain that precise structural engineering masterpiece through out life and to keep what is meant to be acellular just so. What happens in the adventitia would not be able to cause structural failure (infarct) if the structural layers to the inside remain in tact. That is also true re the collagen fiber and elastin (layers & fiber) that position the cells in the media, the structure that is similarly attacked by the slow protein degrading homocysteine [thiolactone]. Then, logically, cells get triggered to move or replicate and then require more capillaries and (re)generate the extra cellular matrix upon which they depend. This is impossible to do well when high homocysteine prevents the integral lysine linkages [directly when the molecule below opens at X or by destroying lysyl oxidase -and m.m. for proline in collagen]. My final point would be that homocysteine but nothing else explains everything historic, i.e the rise-and fall of CHD with heart attacks first described in dull detail in JAMA 1912, i.e. after the first several decades of micro-nutrient removal to generate the long-shelf life products we have to day, and the very year the first cookbook for hydrogenated fat was published [Crisco by Procter and Gamble]. 

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Bogdan Sikorski
OK Eddie - Again, nicely put! and convincing.
Just a simple question - has anyone stuck a radioligand on homocysteine and found it where you and Kilmer say it should go when present in a relative abundance (>6 µmol/L )?

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Paul de Groot
Uffe, Duguid published a lot of micrographs of microthrombi covered by endothelium but AFTER a heart attack. That is essential. A heart attack (myocardial infarction) is a result of blockage of the coronary artery by plaque bleeding c.q. plaque rupture. In both cases thrombi are formed on the bulging mass in the coronary lumen: flow stagnation. The thrombi are incorporated in the plaque and the endothelial lining is restored. The thrombi fibrose and often recanalization takes place. Sometimes such a plaque is laminated as a proove of more than one thrombus. This picture is definitely different from the "normal" atherosclerotic plaques which show a fibrous cap on a mass of undifferentiated tissue (pultaceous mass) in which blood vessels (from the adventitia), spiked spaces seen as cholesterol crystals??, fibrous and collagen connective tissue etc. It shows more or less the composition of scar tissue. This is the reason atherosclerosis is now described as a chronic inflammatory disease. So thrombosis and atherosclerosis are different processes, however, can go together.

With that your reasoning thrombosis, fatty streak, fibrous plaque, complicated plaque is highly unlikely and in comflict with facts.  

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Uffe Ravnskov
Paul, the crucial question is, what causes thrombi and what causes plaque bleeding? Here is Duguid´s interpretation of his findings: 

“Microthrombi occur at as early as three years of age and continue to occur throughout life. By forty years their repeated incorporation into the intima results in irregular fibrous thickening which are too stiff to comply in the normal way with pulse movements and so cause disruption and haemorrhage. The extravasated blood disintegrates leaving fatty deposits, which accumulate progressively, and when haemorrhages are frequent and profuse the deposits predominate whilst the fibrous tissue is relatively sparse, with the result that the thickenings are soft and friable. In such circumstances the surface layers are liable to be torn away leaving ulcers, which promote gross mural thrombosis. The thrombi become organised and form fibrous thickenings which narrow the arteries and destroy their elasticity.” 

As I see it Duguid´s hypothesis leaves room for many factors that may cause the formation of microthrombi, and also for factors that may lead to incomplete healing and thus predispose to the production of soft and friable vascular scars (eg. plaques). 

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Herbert Nehrlich
This seems to beg the question of the apparent absence of these processes in certain populations. Could the disturbed (reversed) ratio of n-6 to n-3 in "modern man" be the key? I am always reminded of Dr. Paul Dudley White's (White House Physician under Eisenhower) statement that he could not find patients to try his new gadget on, it was the EKG machine. Surely this isn't all just one big fault in the design of the human being.

Eddie Vos
Friends, sure, it's a fault of being human and having supermarkets and grocery stores around and there seems little evidence that before about 100 years ago we got such mirco-coagualtion problems leading to infarct and decline.

When animals eating non-processed and non heated foods with say 4-5x the homocysteine lowering B vitamins and none of the refined starch revolution foods don't seem to have heart attacks/atheroma, this alone is a message.

There are clearly 2 methods by which we damage long-living structural proteins: glycosylation from excess carbs [O-bonding glucose to proteins] and thiolation [sulfur bonds/insertion in proteins].  Both methods are nutrition related. Nobody would do animal or culture dish experiments with such clearly malnourished subjects as in current H. Sapiens.   We're overwhelmed with nutritional confounders.

With Uffe: clearly the 'coagulopathy' elements [all micro-nutrients], and via glucose or insulin PAI-1 for example, is a fascinating subject and evidently micro-and macro clots play roles -but are they primary?  Also, in the coagulopathy department, Herbert's n-3 helps while n-6 clearly does the reverse. 

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Paul de Groot
Uffe, To day I am a bit philosophical because we, al together, seem to discuss different things. My good friend Roel put it this way. If you look to our globe from very far off you can see a town. If looking closer you see a town with clogged streets and a still closer look reveals cars in the street for example most cars are Renaulds and Peugeots. My assumption is the town is somewhere in France. A look at a street sign says "champs elisee" and so the town I saw from far of could be Paris although perhaps there are more towns with a Champ Elisee. What clogged the street in Paris? possibly road work, however, in London the street is also blogged but possibly by non working street lights. Although we see in both towns the same thing, street congestion, the cause is quite different. This came to my mind reading your e-mail. Atherosclerosis is the name we use for arterial wall changes showing excentrically plaques and we assume the process is the same in all arterial vessels although we know many a time there is predilection, cerebral vessels, peripheral arteries, renal arteries, coronary arteries and last but not least aorta. So ..."the surface layers are liable to be torn away leaving ulcers..." is a statement clearly concerning the aorta; I have never seen ulcers in a coronary artery. I have studied only (human) coronary arteries and so I can only discuss these vessels. With this in mind I still can not understand why after many years of life all of a sudden the endothelium become permeable for say cholesterol, ox-LDL and/or thrombi. Let's have a look at the so called (coronary) complicated plaque. The big difference between the complicated plaque and the fibrous plaque is its vascularity, the fibrous plaque contain no vessels this in contrast to the complicated plaque. The plaque vessels grow from the adventitial vasa vasorum via a destructed media into the plaque. The vessels are thinwalled, form no pattern and are irregular of size and are prone to disruption according to the many forces executed. The result is plaque hemorrhage and subsequently forming of a cholesterol pool which we later on can see as cholesterol chrystals. With this process external cholesterol does not play a role.
My proposition is to abandon "atherosclerosis" and make up new names describing the process at hand in the various sites and with that make discussions more to the point. Any suggestions?? Perhaps my argument sound a bit pedantic, if so I am sorry, Eddie I rode my "stokpaard" again.

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Eddie Vos
Hi Paul, just a note since you mentioned my name in your response .. how about athero-arteriosclerosis .. of simply 'artery tire failure', artery puncture or re the aorta 'ply delamination' since you made that nice vehicle traffic jam analogy? Let me suggest that it is slow decline of structure that allows your vasa vasorum in, but that the decline of structure comes first ... 
Let's realize the importance of the structural components like elastin and how it degrades and allows calcium and lipid deposits in that broken structure [in later type V, VI lesions]. This attachment of exceptional length [sorry] is a 2005 article from Atherosclerosis that wonderfully describes the process of decline in the inner half or so of the artery: great electron microscope work indeed. This article, read together with the 2000 J. of Nutr. Carlos L. Krumdieck article I sent around about 2 weeks ago, is required reading when one wants to understand the more fundamental processes at play. Re Krumdieck: Melchior commented to me: I finally understand the protein decline theory. The Yuri V. Bobryshev article uses terms as 'vacuolization', holes or 'cavities' in elastin, 'crumbling' elastin, 'splitting of elastin fiber fragments' ... guess what, homocysteine (thiolactone) will do that! Sorry about riding my my own 'stokpaard' [hobby horse].

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Paul de Groot
Hi Eddie, I agree to the importance of the structural components of the arterial wall. However, why degrades elastin it has to have a cause. We all know that sclerotic changes of the arterial wall is not per se age dependent and so is not a fact of life. There ought to be one or more causes and possibly the causes are not the same for different arteries. Why would elastic arteries change in the same way as muscular arteries from the same agens? and why would a different "organ" such as the aorta react in the same way. We just don't understand the differences and still we think that there is just one disease called atherosclerosis which is responsible for the changes of all arteries. This is inconceivable, we still can only see clogged streets in a town unknown and so we must try to acquire the knowledge which town in which state and what caused the jam there. Still your attachment is very interesting but the conclusions are only valid for carotic arteries. I still try to read and understand your papers on homocysteine, however, my chemical knowledge is rusty and from a long time ago but "nooit te oud om te leren".

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Eddie Vos
Right, we're evidently as rusty [crumbly] as our arteries but as you said never too old to learn. Maybe it's more the elastin that comes apart since it ENTIRELY depends on the mirconutrient dependent [Cu, vit. C] and Hcy-lactone degraded X-linking system, while collagen will still function somewhat in the light of the same factors. I can see that collagen 'rope' still pulls well yet in pushing mode a rope may come undone and it's there that the OH-lysine / OH-proline keep the strands together systems gets stressed, at least that is my simplified way of looking at it. What a great term in that study: crumbling elastin. Now, let's jump back to Kilmer's Classic of 1969: 'splitting, irregularity and focal discontinuity of the internal ELASTIC membrane' ..

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Uffe Ravnskov
I have a few questions and objections.
To Paul: I am still sceptical to the idea that the primary event is inflammatory reactions of the adventitial vessels. If intima and inner media receive their nutrients and oxygen by diffusion from the lumen, what role does these inflammatory processes play for the creation of endothelial lesions? Couldn´t the experimental studies, where the authors hurt the adventitia or its vessels be explained otherwise? Saul and Gerard presented an interesting hypothesis a few years ago (Med Hypotheses 1991;36:228-37; ibid. 1999;52:349-51). They considered extra-arterial pressure as a crucial factor. If it is low, the artery wall increases its tension or resistance to prevent its lumen to expand, either by fibrosis or by deposition of other rigid material such as cholesterol. They claimed for instance, that arteries located in bone channels never become atherosclerotic, neither do the small coronary branches that go perpendicular through the myocardial wall because there is no need for strength here. Their hypothesis means that at least some part of the arterial changes, in particular the fibrosis and the growth of the muscular wall, are normal features to prevent dilatation of the arteries and unnecessary pumping of the heart.  

Therefore, if you damage the adventitia the outer muscular wall may not be able to maintain a sufficient tension and thus give rise to endothelial damage.  

Still, as I see it, this mechanism seems possible, but unlikely, biologically seen. Why should adventitial vessels become inflamed? Isn´t it much more likely that the endothelium does, exposed as it is to much higher pressure gradients and dynamic forces and therefore predisposed to become damaged by all the toxic chemical factors we have mentioned so often? 

By the way, Saul and Gerards hypothesis also explain why the coronaries are more prone to atherosclerosis than other arteries because they are exposed to an extravascular negative pressure (as is the aorta). 

I have a question to Leib also. In a previous letter you claimed that LDL “as we all know” is extensively degraded when it enters a cell. As far as I have understood, LDL is not able to pass any cell wall, neither is it degraded when it comes into contact with one. Please explain.

You also wrote that every cell must have access to LDL from plasma if it is to meet its own cholesterol needs. First, do you mean LDL-cholesterol? If so, why call it LDL-cholesterol instead of just cholesterol. Second, I have learnt from more knowledgeable colleagues that any cell is able to synthesise cholesterol. Isn´t that true any longer?  

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