Saturday, January 19, 2013

Protons: The linoleic acid fed mice

I've sent a great deal of time trawling through Masseria et al's paper on linoleic acid fed mice. I'm going to try to generate some data which aren't reported and see how much of a transgenerational effect really occurs.

This paper is the one where a group of mice were weaned from chow fed parents on to a moderate fat diet mostly based around linoleic acid. They then were bred for four more generations on this linoleic acid based diet. Replacing starch in your diet with linoleic acid makes you fat. I'm assuming no sucrose was added at the diet switch. Really, they wouldn't have added sucrose without saying? No, no...

The most important variable I'd like to see, which was measured once a week throughout the study, was total body weight. I cannot find ANY body weights for 22 week old mice. All mice were weighed weekly, according to the methods.



We can tell from the photograph of a chow fed mouse along side an HF4 mouse that the HF4 mice weigh more than chow fed mice. Fascinating. This image of these two mice is the total information on weights at 22 weeks of age. Why are no weights reported?

Now, if you had a linear increase in body weights from generations HF1 through HF4, would you report it? Draw a graph perhaps? Bear in mind that you can generate this data with a set of electronic kitchen scales from Argos, no PCR or Western Blot required.

Here are my fabricated data: At 22 weeks of age mice of generations HF1 through to HF4 weighed the same. There is not even an non significant trend to increase. Otherwise it would have beeen reported.

Next, fat pad weights: All mice which made it to 22 weeks of age were euthanased. Now, to weigh their fat pads would need a visit to the local head shop for some slightly dodgy digital scales, but it's still not exactly rocket science. Masseria et al appear to have the scales anyway, because they weighed and reported the weight of plenty of fat pads in selected mice. But, in a research project on adiposity, they appear to have thrown almost all of the fat pads from 22 weeks of age mice in to the clinical waste bin. Of course, as per the body weights, they might have the data. If they do they are not saying. Is there a linear increase in fat mass from HF1 to HF4? I think it is reasonable to assume not.

So this is what is happening:

The "STD" chow fed mice are exposed to a "normal" mouse diet throughout. They stay insulin sensitive and have normal weight.

HF0 mice are unique in the HF generations. In this first omega 6 fed generation there is a total of 19 weeks of exposure to the omega 6 diet, starting from weaning at 3 weeks of age and going through to 22 weeks of age. These mice do become over weight at 22 weeks, as judged by their fat pad weights as given in Fig 1, but not by as much as do the HF2 mice in the same Fig 1. They do not develop insulin resistance.

Then there is a third set of mice, the HF1-HF4 mice, which are all exposed to the same diet for the same duration, ie throughout their gestation (assume 3 weeks for this), breast feeding (assume another three weeks) and weaning until 22 weeks of age, ie for 25 weeks in total. They develop an interesting pattern of insulin resistance.

So to clarify: It looks very much as if we have three groups of mice. Chow mice never get exposed to high omega 6 intake, HF0 mice are exposed for 19 weeks and HF1-4 mice are exposed for 25 weeks. The number of weeks of exposure is what increases the bodyweight and this (exposure time) is what increases from generations Chow through HF0 to HF1-4.

Within generations HF1-4* neither weights nor fat pad weights are reported.

*Except of course the isolated value from Fig 1 part C where we just get HF2 pad weight, but we still can't see the "lack of trend" within these generations.


I might have been tempted to just leave it at that and suggest that eating a high omega 6 fat diet increases you weight in proportion to the percentage of your life you were exposed to it, if it wasn't for Fig 4.

If we pull out Figure 4 (for the key) we see this:



and especially the insulin section:



Which makes it clear that the hormonal milieux is completely different between generations HF1, HF3 and HF4, the right hand three columns. Insulin is up at 4.5 times control in HF1,  drops to 3 times control in HF3 and is down at 2.5 times control in generation HF4. HF2 can be interpolated between HF1 and HF3, I'll fabricate (fabrication warning!) a value of around 3.75 times control.

So the very simple concept that a high PUFA diet, due to a failure to generate superoxide in mitochondria, increases adipocyte insulin senitivity, so facilitates weight gain at a given level of insulin, is not as clear cut as I would like. Damn.


The main hint about what is happening comes from Fig 5. Because of the small group sizes I find it very difficult to accept that these lines mean very much in absolute terms. But there is, certainly, the impression that adipocyte numbers, especially numbers of small adipocytes, really does go up transgenerationally through generations HF1-4. Probably.

So HF0 mice hit weaning with a "chow fed mouse adipocyte count" and get the next 19 weeks on a diet which maintains insulin sensitivity due to the limited production of FADH2 per unit NADH by its dietary fat. These mice get fat but stay insulin sensitive because 19 weeks on an omega 6 diet does not have time to produce enough adipocyte distension to generate enough insulin resistance to over ride the effects of the PUFA diet. Quite fat, but insulin sensitive. Utterly straight forward.

Generations HF1-4 get 25 weeks of omega 6 exposure and all become more obese than HF0. But through the same set of generations they also develop progressively more adipose tissue hyperplasia. ie they have more and more fat cells.

As the total number of fat cells goes up, so does the ease at which they accept fat without becoming so distended as to become insulin resistant. So HF1 have just a little adipose hyperpalsia and each adipocyte manages to hit distension induced insulin resistance by 22 weeks of age. By the time we get to HF4 there are far more small adipocytes available to accept fat, they do so easily and we end up, after 4 generations, with obese but far less insulin resistant mice.

It looks like a simple trade off between the obesogenic effect of omega 6 fats vs the distension induced insulin resistance of the whole population of adipocytes.

Omega 6 fats appear to be great for putting fat in to adipocytes. Adipose hyperplasia appears to be the key to maintaining insulin sensitivity during weight gain. Omega 6 fats are great for generating adipose hyperplasia.

Having adipose hyperplasia means lots of half empty adipocytes are willing to signal that they are not full. Did you notice the leptin levels in Fig 4? Very interesting. Losing weight is not going to be easy if you are a human in this situation.

There are hints from birth weights and pre weaning growth rates that this may be a process which starts in utero. There are also hints from previous publications by this group that the obesogenic effect of omega 6s can be blocked by indomethacin, a cyclo oxygenase inhibitor. That is; there is at least one, possibly several, layers of control in place over the F:N driven process.

At a basic level the F:N ratio concept it is quite clear where the obesogenic effect might come from. That there are additional refinements to this concept is not surprising but, to me, tinkering with the control system when the basic engine of metabolism is broken, is worse than pointless.

Try not to base your diet on corn oil.

Peter

BTW: Most annoying quote from the paper:

"These observations indicate that, despite the fact that glycemia in HF4 mice appeared normal at 22 weeks old (151 ± 30 mg/dl for HF4 versus 170 ± 30 mg/dl for STD mice), continuous exposure to the omega 6HFD led to a sustained increase in plasma insulin levels, which strongly suggests the emergence of insulin resistance of adult animals at later generations."

Technically correct, but here is a repeat of the plot they are describing from Fig5:



Hmmmmmmmmm. Do we heed the voice of Author-ity or read the graph?

46 comments:

Kindke said...

" Omega 6 fats are great for generating adipose hyperplasia"

I can only assume they do that by increasing adipocyte insulin sensitivity. Otherwise they are somehow producing endogenous ligands for PPARgamma?

This study was quite revealing about the interplay between bodyweight, leptin, and the degree of hyperplasia.

For a given bodyweight, your leptin level indicates the amount of hyperplasia you have. Low leptin unfortunately signals high hyperplasia. I like how the subjects in the Troglitazone study reported increased hunger. Troglitazone acts via PPArgamma to increase adipocyte hyperplasia. So the subjects got new adipocytes that were not full.

Not full adipocytes throw baby tantrums for more calories, demanding that the host that it eats more through manipulating the hosts hunger and satiety systems.

The idea that the hyperplasia starts in utero may aswell partially explain the current obesity epidemic, mothers are eating more omega6 and so are giving birth to babies with more fat cells?




IcedCoffee said...

Kindke,
Omega 6 fats do serve as ligands for PPARgamma. As do Omega 3s, and lauric acid (and a few other rare ones).
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC20720/
This, coupled with the fact that these EFAs serve as precursors to eicosanoids which also serve as PPAR ligands suggests that linoleic acid consumption could have a major effect on PPARgamma.

However, CLA has been shown to impact PPARgamma to promote adipocyte cell differentiation to small adipocytes, and seems to have a generally protective effect and results in lower bodyweight. This makes me think that its not just the number of adipocytes, but also the type. The HF1-4 generations have more small AND large adipocytes, and I would speculate its these large adipocytes that are the truly problematic ones. Obviously more adipocytes in general is probably not desirable, but when the differentiation is taking place, the preference for small adipocytes that fill up quickly would make sense.
http://www.ncbi.nlm.nih.gov/pubmed/20838573

It also appears that the different types of adipocytes behave very differently. Large adipocytes tend to promote TNFa production, whereas small adipocytes promote adiponectin secretion.

George Henderson said...

What else might increase FADH2 production per NADH?

"Compared with those in the first quartile, the multivariable-adjusted OR (95% CI) of overweight status from the 2nd to 4th quartiles of BCAA intake were 0.97 (0.80-1.17), 0.91 (0.75-1.11), and 0.70 (0.57-0.86), respectively (P-trend < 0.01). BCAA intake and obesity were also inversely associated (P-trend = 0.03)."

http://www.ncbi.nlm.nih.gov/pubmed/21169225

Gys de Jongh said...

The body weights are in the Supplemental Data of the article :

http://www.jlr.org/content/51/8/2352/suppl/DC1

Have a look at the *.pdf file

DLS PABLO said...


fructose... omega 6.. lol. the fastest (not fattest) guy in the world eats...buckets of chicken nuggets,20 bananas a day, etc... run piggy run?

Peter said...

Dear Gys de Jongh,

Please don't scare me like that! The 22 weeks of age weights are what I want, not those 3 and 8 weeks! At 8 weeks no adipocytes are full enough to become insulin resistant...

Ultra brownie points for anyone who really can find the 22 weeks weights!

Peter

Peter said...

Kindke,

They seem to use a cycloxygenase derived product. Certainly parts of the effect are suppressable with indomethacin. But the insulin sensitivity effect seems core to the problem. And differentiating preadipocytes to adipocytes in cell culture involves lots of insulin and glucose.

Peter

Evangelista Nick said...

I hate studies like this... Annoying indeed!

Aaron Blaisdell said...

Peter, thanks for the leg work. I don't think I would have found this article on my own. As mice are habitual seed eaters in the wild, and many (most?) seeds are relatively high in linoleic acid, I wonder how this lab diet compares to a whole-foods (i.e., intact seeds) in terms of both macro- and mirco-nutrients. It would be interesting to compare the mice on standard and experimental lab diets to mice eating an "ancestral" (i.e., wild-type) diet.

Galina L. said...

I decided to check out what mice eat in a wild habitat, it looks like the answer is "anything". They survive in arid environments eating insects as a source of water, in human dwellings besides regular food they even eat glue and soap.

Peter said...

Galina,

Plasterboard and electrical insulation appear to be standard nutritional features. Not sure how essential they are. Better that D12492.

Peter

Galina L. said...
This comment has been removed by the author.
karl said...



There is some supplimentary data at :

http://www.jlr.org/content/51/8/2352/suppl/DC1

No info on who made the diets or anything about the carbohydrates.

http://www.jlr.org/content/51/8/2352/suppl/DC1

I just wrote to the author to see if he might tell me. ( Sucrose apparently is considered a form of fat by many PHDs these days - at least in many rodent studies - wonder if the O-6 was hydrogenated as well? )

I've seen other data that suggests that if the mother had poor BG control during pregnancy, the offspring tend to get fat. This is of real interest to me as a son of a ObGyn ( only ObGyn docs seem to ever run GTT - gestation diabetes tends to produce large babies and delivery problems - thus C-sections ).

I've wondered if the BG level during pregnancy might set the adipose set point in some way. Could be by the number of adipocytes needed to control BG?


IcedCoffee said...

@karl,
Diabetic mothers definitely seem to produce pre-diabetic babies, although I'm unsure if its the high BG per se, or if it comes from some inflammatory response from the mother. A diet this high in O-6 in an obese mother will do some crazy stuff with eicosanoids, and the baby gets to just sit there and enjoy the repercussions.

My mother had gestational
diabetes (barely) and produced me at over 11 pounds, but I've managed to stave off any serious metabolic damage. Maybe I just caught it early.

Galina L. said...

@Peter,
You liked to joke that some rats involved into research programs were secretly going to a gym to keep their weight down. Who knows, probably for mice chewing plaster and cables is the equivalent of humans exercising on a tread-meal, doing something useless to offset an unhealthy life-style. Who can be healthy living among people eating processed garbage and too much grains? Are there any reports about pests and city pigeons getting too fat? I know, it is the internet, so in the case it was not clear - I was joking.
Speaking of rats, how does your Ratty doing? When I saw him last time on your family photos, he looked healthy but ridiculously unaware of opportunistic cats. Do you plan to perform a proper autopsy after the unavoidable end of his life (if some creature will not manage to consume that N=1 first).

blogblog said...

@galina,
wild mice primarily eat seeds and a few insects. A completely natural mouse diet is high fibre, high starch, low protein and has an extremely low fat content.(2-5%).

engineering-health.org said...

Hey Peter, Perhaps you could make some sense of this article on a future blog post? It says impaired autophagy prevented obesity and improved insulin sensitivity- the opposite of what I'd expect. Any ideas?
http://www.ncbi.nlm.nih.gov/pubmed/23202295

Chip Spitter said...

Random anecdote for today: it seems eating loads of fat has reduced my Lp(a) by a third. I'm taking that as a good thing, even though Lp(a) remains a bit of a mystery. My old man's cardiologist reckons it doesn't move much, no matter what you do. I'm guessing "no matter what you do" only involves pills.

I know, I know, lipidology is for idiots - but I'm not quite at the 'understanding protons' stage. Might get there eventually.

donny said...

If you look at rebound weight gain after starvation, like in the Minnesota starvation study, fat regain tends to come before lean mass--and to overshoot. Regardless of what the actual insulin levels might be, I guess you could see this as evidence of greater relative insulin signalling. In very starved mice, the fat cells, on initial refeeding, tend to first fill up on glycogen--much more glycogen than is usual--before appreciably replenishing fat stores. Sort of like overcompensation of glycogen stores in muscle and liver after carb restriction/loading. Perhaps fat cells aren't subject to this until fat storage gets below a certain point, local fat keeps them from needing too much glucose.

I've wondered for a while whether increased insulin sensitivity is all that good an idea for fat cells.

http://ajcn.nutrition.org/content/51/6/970.full.pdf
---------------------------------
Refeeding after fasting in rats: effects of duration of
starvation and refeeding on food efficiency
in diet-induced obesity
-------------------------------

This study has twinkie-fed rats vs control rats, starved and refed. There's greater food efficiency in the twinkie (crisco and sugar) rats--and considerable hyperplasia of fat cells on refeeding. Maybe it doesn't matter if the omega six is hydrogenated or not?

karl said...

@IcedCoffee
Yes, the mechanisms are all hypothetical right now - we need some researchers that have a clue as to how to design experiments. It is probably a 'bad thing'(tm) to be exposed to elevated BG in the womb.

@blogblog
It depends - most of the 'control rodent diets' have more fat than what they would typically get in the wild.

@Chip Spitter said...
"it seems eating loads of fat has reduced my Lp(a) by a third. I'm taking that as a good thing, even though Lp(a) remains a bit of a mystery."

I've got quite a lot of notes about Lp(a) here.

Niacin also seems to lower Lp(a) (flushing is not the only side effect) - the question that goes unanswered is if the methods of lowering Lp(a) produce better outcomes - or are we just treating a lab number. ( are we lowering circulating Lp(a) by filling up lipid pools or by preventing the creation? - we just don't know ).

That being said, I think that Dr. Davis bit where he has Lp(a) folks take high dose fish oil is probably a good bet (vs science). It appears to take 6mo to 2 years - but it seems to have worked for me.

There is also a hunch that having T3 in the upper half of normal may be important with Lp(a) - but again no quality research to base it on.

@engineering-health.org
I sure wish I had the full text of that paper ..

@donny
Just what I've been thinking - the 'eat PUFA to lower BG' mantra was pushed without any long term studies. (how are people going to lose weight if they have inappropriately elevated insulin sensitivity?) Once again, the overlapping of control loops confounds insulin levels - insulin levels need to be looked at relative to insulin sensitivity, autonomic stimulation, etc..

We need a way to measure the net flux of FA into/out-of adipose tissue short term (long term we look at fat mass) to understand this overlapping/nested control loop.

Jack C said...

Another interesting study regarding linoleic acid and weight gain in mice relates to the fact that linoleic acid is the precursor of arachidonic acid (AA)which is the backbone of two endocannibinoids which activate the same receptors activated by marijuana. (It has been known for years that marijuana gives people the munchies.) Control mice were fed a diet with 1% ALA and 1% ALA with total fat of either 35% or 60% fat. The study shows that increased intake of linoleic acid, from 1% of energy to 8% of energy, increases AA levels which in turn causes increased endocanniniboids in red blood cells, the liver and hypothalmus. Mice fed the 8% LA increased food intake compared to controls fed 1% and also increased food efficiency (weight gain per gram of food) and had a much higher adiposity index

It was concluded that the increased endocannibinoids derived from increased LA intake caused increased food intake and increased fat synthesis in the liver (which can be a cause of fatty liver.)

The study was supposedly structured to represent the change in LA intake that has occurred in the western world during the last century with LA intake increasing from about 1% to 8% of calories, but in fact the ratio ALA to LA of 1:1 that was used is not realistic (a ratio of 0.3% ALA and 2% LA is more representative of past intake as per cent of calories) so the increase in endocannibinoids shown in the study is greatly exaggerated.

It was not mentioned in the study that the standard procedure of inducing obesity in mice is to feed a diet with 60% fat including 8% LA. Mice fed 60% fat with 1% LA did not become obese.

On the low LA (1%) diet, mice on the 35% fat gained more weight than mice on the 60% fat diet due mainly to greater food efficiency. On the 8% LA diet, mice on the 60% fat diet gained much more weight that those on the 35% diet. Most of the fat in the high fat diet was from coconut oil. This makes me wonder about the supposed benefits of coconut oil.

http://www.ncbi.nlm.nih.gov/pubmed/22334255

karl said...

@Jack C

Was the coconut oil hydrogenated?

IcedCoffee said...

@karl,
yes, it was hydrogenated. yay science

IcedCoffee said...

Sorry forgot to mention it says so in the notes of Table 1.

john said...

Well it's a stretch, but that study somewhat supports Karl's and others' ideas about those last 15-20lbs or whatever. The coconut oil would need to be hydrogenated to have such low LA. Does that imply there may be a very narrow window, in terms of pufa, for achieving leanness on high fat?

Jack C said...

@Karl, IcedCoffee,

While the study certainly would have been better had they not used hydrogenated coconut oil, I do not believe that the fact that the coconut oil was hydrogenated negates the conclusions that increased intake of LA results in higher liver and brain levels of AA and consequently higher levels of endocannibinoids (Ecbs)that stimulate increased food intake and increased fat synthesis by the liver which contribute to increased adiposity (and fatty liver.

According to the lands Equation, the base diet of 1% ALA and 1% ALA as per cent of energy intake results in an n-6 AA content of 42% of HUFA in red blood cells for both the medium fat (35% of energy) and the high fat (60% of energy)diets. The observed rbc n-6 AA content on this diet was 45% for the medium fat diet and 42% for the high fat diet. In the liver, for the high fat diet compared to the medium fat diet, AA content was 30% lower and AA content as per cent of HUFA was 17% lower while ecbs were 13% lower. There was not a lot of difference between the medium and high fat diets for food intake, food efficiency or adiposity index, but overall it seems that the high fat diet was slightly better.

The base diet of 1% ALA and 1% LA supposedly approximated human food intakes a century ago, but according to a recent study, in 1909 ALA intake was on the order of 0.35% and LA intake about 2.3% of energy which would result in an n-6 AA content of rbcs of about 66% of HUFA compared to 42% on the mouse diet. On a real world diet, about the only way to get the n-6 HUFA down to 42% is to take fish oil or eat some fish. If you can get LA intake down to 2.3% of energy intake it takes only a small amount of n-3 HUFA (0.3% of energy of EPA and DHA) to reduce n-6 AA to about 42% of HUFA.

The diet with 1% energy as
ALA and 8% energy as LA is a fair approximation of the current western diet. The medium fat 8% LA diet, compared to the 1% LA base diet, resulted in: an increase in liver n-6 AA from 37% to 64% of HUFA, a 63% increase in liver AA, a 73% increase in liver endocannibionids, a 3% increase in food intake, a 12% increase in body weight, and a 26% increase in adiposity index. By comparison, the high fat 8% LA diet resulted in an increase in liver AA from 31% to 63% of HUFAm a 160% increase in liver AA, a 215% increase in liver endocannibinoids, a 17% increase in food intake, a 20% increase in body wieight, and a 41% increase in the adiposity index. It is apparent that when the LA intake is on the order of 8% of energy, a high fat diet results in far greater adiposity than the low fat diet.

It is my conclusion that a high fat diet is no problem if you keep LA intake as low as practical (2 or 3% of energy) and eat a enough fish or fish oil to lower n-6 AA to about 50% of HUFA (by Lands equation).

karl said...

A link for those who want to know about the Lands equation
and here

Using hydrogenated means that they changed more than one variable at a time - and as grade school teachers used to teach - a single variable is required for a real experiment.

How do we know if it negates the conclusion or not? I'm not sure anyone really understands just what transfats do to us.

I have no idea what conclusions one can draw from such an experiment, but most of these hypotheses get a life of their own and the needed experiments don't get worked on unless there is a path to large sums of money.

I've read other bits where people were wondering if the ratio of O-6 to O-3 was as important as the lack of O-3. It is clear that this science is not settled - I have hunches, but I think it is really important here to talk about hunches as such and not to talk like they are known things.

There may be more than one effect from the seed oil that we are eating.

The idea that O-3 blocks the pathway of 18:2 ω-6 linoleic acid => 20:4 ω-6 arachidonic acid or AA is attractive - but I'm not sure that is all that is going on - where food ends and endocrine signals begin appears fuzzy right now - and of counting protons..

The other bit is when they use seed oils we need to remember that there are uncontrolled amounts of phytoestrogens. Polyphenol, etc. Until they start using totally synthetic diets, they may be stirring mud into the water rather than clearing up the science. (Real science is really hard to do - most of these guys just go for the grant money and do something that looks like science.)

karl said...

Interesting rodent study using night vision equipment

hithaeglir said...

@karl - "A link for those who want to know about the Lands equation and here"

The first link is broken and the second is a paper behind a paywall of which and I have tried unsuccessfully in the past to obtain a copy.

Bill Lands is an acknowledged expert on EFAs who is credited with discovering the promoting the need to balance O-6 and O-3 fatty acids to optimize health.

It is well worth reading his many accessible papers and his book, "Fish, Omega-3 And Human Health"

IcedCoffee said...

Although the n-3/n-6 interplay is definitely important, I'm somewhat unconvinced that the high n-6 is necessarily bad. It definitely feeds into the pro-inflammatory eicosanoid pathways, but I don't think it controls them. It seems like the bigger problem is the underlying inflammation that is almost ubiquitous. Get rid of the underlying inflammation, and is the excess n-6 still inflammatory?

Low n-3 is definitely a problem, but there are enough traditional populations with a higher n-6 intake to make me question that it is bad in its naturally occurring forms. And even if we have a higher n-6 intake, does this necessarily mean that our fat cells will be oxidizing them? Or if we force our fat cells to oxidize through mass consumption, will this not also result in a lower insulin secretion from the pancreas for similar reasons?

karl said...

corrected link:

http://digitalcommons.unl.edu/cgi/viewcontent.cgi?article=1713&context=usdaarsfacpub

I'm not saying Land is wrong ( not everyone agrees with his explanations - I think fish oil is important - yet I don't think we understand the details as to why) - I just think that we should all be clearer about what is theory vs fact. There are a lot of bits about O-3 /O-6 that need quality research to ground the abstractions in reality.

Jack C said...

I am sure your grade school teacher would find fault in almost all studies because they inevitably have more than one variable. The study in question really has more variables than just the fact that the coconut oil was hydrogenated. For example, the ratio of monosaturated fats to saturated fats varies in the different diets. The use of coconut oil as the predominant fat can be questioned as it is not a large constituent of the diets of either humans or mice.

Receptors for endocannibinoids (Ecbs)such as marijuana and ecbs derived from AA (anandamide, or AEA and 1-AG and 2-AG) are found in the liver, gut and hypothalmus.
It is well established that activation of Ecb receptors stimulates increased food intake and also results in an increase in synthesis of fat by the liver. Since essential fatty acids such as AA can not be endogenously synthesized by humans, the presence of Ecbs derived from AA in humans only results form intake of either AA or LA, the precursor to AA.

The intent of subject study was not to determine whether increased intake of LA resulted in increased synthesis of AA, or whether increased AA resulted in increased Ecbs, or whether increased Ecbs caused increase food intake and increased fat synthesis. Those facts are fairly well established. The purpose was to estimate how much the increase in LA intake which has occurred during the past century has contributed to increased obesity.

The study compares a diet, in mice, with 1% ALA and 1% LA as percent of energy intake, with a diet of 1% ALA and 8% LA in both a medium fat diet (35% energy from fat) and a high fat diet (60% of energy from fat). In both the medium fat and high fat diets, the increase in LA content was offset by decreased saturated fat from coconut oil. The increased LA content of the diet resulted in increased Ecb in the liver and brain and increased food intake, increased food efficiency (due to increased fat synthesis by the liver) and increased obesity. It seems illogical to me to think that a decrease in intake of hydrogenated coconut oil would result in an increase in synthesis of Ecbs.

The main criticism I have of the study is that the base diet, which is supposed to represent EFA intake by humans a hundred years ago, includes 1% ALA and 1% LA as per cent of energy which results in an AA content of red blood cells, as percent of HUFA of 42%. Increasing LA intake to 8% of energy resulted in n-6 AA of rbcs of 72% of HUFA, an increase of 30% AA over the 1% LA diet.

By comparison, the AA content of rbcs based on the real estimated human diet a century results is 66% AA HUFA, while the current dietary intake of EFAs results in 81% AA HUFA, or an increase of 15%.

Thus it appears to me that the estimated actual increase in AA% HUFA that has occurred during the past century is about 15%, or half of 30% increase that estimated in the subject study. The impact of increased LA intake on obesity is therefor less than suggested in the subject study.

karl said...

@Jack C

The point is there is no shortage of junk-science. I think it has gotten to the point where most people have no idea of the difference ... as long as someone does the required brown nosing, to get a phd, writes a proposal that doesn't disturb any egos, the grant money flows and we get papers where sucrose is considered a key ingredient in a high-fat diet. Junk science, instead of clearing things up simply muddies the water.

These papers, at best suggest where a real experiment might be run. Until then, it is just stamp-collecting.

I would suggest that you start by reading Richard Feynman's Cargo Cult Science and the rest of what he wrote on science philosophy, then perhaps Junk Science Judo, and Black Swan etc.

Yes, it sure looks like long chain O-3's do us good, but most of the evidence is correlative and the hard work to nail down the hows and whys has yet to be accomplished. Those details could easily turn medicine on it's head, yet instead we get garbage papers.

George Henderson said...

@ karl

"Until they start using totally synthetic diets, they may be stirring mud into the water rather than clearing up the science."

But until we start eating totally synthetic diets, that science won't be relevant.

A high-fat mouse diet is much closer to a commercial icecream recipe (or similar sweet milky, fatty pudding - cheesecake perhaps) than it is to a dish of say, steak and chips or nuts and bananas.
The results may only apply to icecream eaters.

Why not do at least a few studies comparing real diets?
Did you ever read Gibbon on the Byzantine chariot races? We should ask diet gurus to support and feed their own teams of mice or rats (pigs would be even better) then race them or fight them in team colours. The races or fights could be televised, and the supporters of the losing team could riot for days wearing the colours of their losing team so everyone would know what losers they backed.

It's the only way, man.

Jack C said...

Karl,

In spite of the hydrogenated coconut oil and the synthetic diet, the observed red blood cell content of n-6 AA, as a percent of HUFA, came out damn close to that predicted by the Lands equation on all six diets. Apparently neither the hydrogenation of the coconut oil or or the synthetic diet distorted accuracy of the Lands equation. I therefor think that it is reasonable to believe the relative accuracy of the study finding that increased LA intake results of increased AA and Ecb content in the liver and brain of LA fed rats which results in increased cholesterol and fat content of the liver. This is consistent with the observation that real live people eating real food can reduce fatty liver by reducing LA intake and by taking omega-3 HUFA.

I recognize the faults of the study but I think I derived some important information from it.

Their is an abundance of information on the adverse effects of excessive intake of LA, both from scientific studies, many of which are poor,and from observation of live people eating common foods. Israel has the highest intake of LA of any major country and the result has been "the Israeli paradox", the observation that LA, rather than being beneficial as claimed by the American Heart Association, Israel has been beset with a paradoxically high prevalence of cardiovascular diseases, hypertension, non-insulin-dependent diabetes mellitus, obesity and cancer.

Peter noted in his post above that a high intake of LA causes cirrhosis.

IcedCoffee said...

George,
I think your comment just won the internet. Although now I'm worried about PEDs skewing the results of the mouse battle royal. Something tells me those myostatin knockout mice will have the upper hand.

Jack C said...

Karl,

In spite of the hydrogenated coconut oil and the synthetic diet, the observed red blood cell content of n-6 AA, as a percent of HUFA, came out damn close to that predicted by the Lands equation on all six diets. Apparently neither the hydrogenation of the coconut oil or or the synthetic diet distorted accuracy of the Lands equation. I therefor think that it is reasonable to believe the relative accuracy of the study finding that increased LA intake results of increased AA and Ecb content in the liver and brain of LA fed rats which results in increased cholesterol and fat content of the liver. This is consistent with the observation that real live people eating real food can reduce fatty liver by reducing LA intake and by taking omega-3 HUFA.

I recognize the faults of the study but I think I derived some important information from it.

Their is an abundance of information on the adverse effects of excessive intake of LA, both from scientific studies, many of which are poor,and from observation of live people eating common foods. Israel has the highest intake of LA of any major country and the result has been "the Israeli paradox", the observation that LA, rather than being beneficial as claimed by the American Heart Association, Israel has been beset with a paradoxically high prevalence of cardiovascular diseases, hypertension, non-insulin-dependent diabetes mellitus, obesity and cancer.

Peter noted in his post above that a high intake of LA causes cirrhosis.

john said...

George,

I think there are a few studies that use real foods. I remember a gerontology article about adrenal lipofuscin and collagen crosslinking in rats. The high fat "meat and milk" group had least, while the cereal-based group had the most (high polyunsaturated fat rats were in the middle).

There is also an Italian group I think that gave mice ground meat and other reasonable ingredients. In their work, high fat (~20% carbs) eating from butter caused an increase in caloric intake, metabolic rate, and t3, while weight remained the same.

Jack C said...

@Icedcoffee: You said:

"Low n-3 is definitely a problem, but there are enough traditional populations with a higher n-6 intake to make me question that it is bad in its naturally occurring forms."

I was unaware that of any traditional populations with a high n-6 intake. I would appreciate it if you would tell me what populations you are referring to and what foods they eat that are high in N-6.

Jack C said...

@ George Henderson:

"Why not do at least a few studies comparing real diets?"

Using "real diets" in such studies introduces a huge number of variables.

Milk fats, for example, contain over four hundred different lipids and many vitamins, enzymes and minerals which vary greatly depending on the diet and breed of the cows. Pasteurization damages or destroy many of the nutrients in milk the content and the degree of damage varies depending on the temperature and time of pasteurization.

For example, milk from grain fed cows contains very little vitamin K2 and beta carotene whereas milk from pastured cows has much higher but varying amounts of these nutrients depending on the quality of the pasture.

Super market eggs, for another example, have an n-6/n3 ratio of 19.9 compared to a ratio of 1.3 for eggs from pastured chickens. (according to Simopoulos)

The number of variables in real food diets could be quite high. I can understand why synthetic diets are usually used.

IcedCoffee said...

Jack C,
The one that immediately comes to mind is the San bushmen and their consumption of Mongongo nuts, but I'm pretty sure there are a couple others. (Basically any group that ate lots of nuts.) There are other people who follow this blog who can probably give you a dozen examples.

It seems like any time we try to isolate a single nutrient as THE problem someone brings out the traditional society that flourished eating primarily that thing.

PUFA in nature tend to be either in very small amounts, or come packaged with a proportionally large quantity of vitamin e. Perhaps the failure to incorporate vitamin e into membranes in conjunction with the PUFA oxidation results in mis-signaling regarding the relative insulin sensitivity of tissues.

karl said...

@George Henderson

Real foods vary too much - are we going to use tomatoes that were gown with fertilizer or not? In sandy soil or not? I partial shade or not? - Which breed? the Roma from 1975 or 1993? What if the control group gets a tomato with a bit of mold on it?

And yes, it could make a big difference.

The idea is to have two diets that are exactly the same - except one variable ( even this is tricky - is it fair to swap equal amount of calories of sucrose for a starch? Or should it be by molar quantity? ) Real science is really hard to do - perhaps only 5% in this field know how to do it.

(For example - What is wheat? The wheat they grow today is quite different to what they grew in the 1950's. )

Here is something else to think of - as they breed plants to be 'pest resistant', often the plant makes some chemical that is toxic to the pest - it is always present, even in years when the pest isn't. Compare that to applying a pesticide - only applied when the pest is present in controlled quantities - of a well tested chemical. The natural pesticides go untested.

The much valued 'organic' foods are breeds that have lots of pest resistance. Is this really a good thing? ( I don't think we know).

'Natural' foods can be quite toxic - there has been chemical warfare going on between plants and animals for a long time - resulting in things like hemlock, poisonous mushrooms, cyanide laced seeds.

Where would we get 'real foods'? The produce in the grocery store is hardly 'natural' - all of it is a result of genetic engineering via selection. It would be quite a process to find wild plants that have not been contaminated with pollen from cultivated crops..

Synthetic diets have been used - even on humans, and the results of these papers become quite interesting.

George Henderson said...

@ Karl,
The last rat diet I pulled up was from a microbiota study.
Surely cellular structures are important when it comes to microbiota.
I can see how pure chemicals are defensible in metabolic studies, but there are number of ways microbiota is influenced by cellular/accellular differences; not just the mechanisms in the Spreadbury paper, but also differences between fermentation of choline (used in diets) and actual phospholipids found in foods.
If nothing else, the more refined diet will feed microbiota higher in the gut, and therefore a different population will be produced. The gist of the paper was that this was down to macronutrients, but clearly the degree of refining would influence it too.
It doesn't matter if the authors aren't trying to generate results that apply to humans. It wouldn't matter much, perhaps, if the results weren't being discussed on the internet, and never made their way into the popular media.
if a mouse gets fat in the forest and no-one is there to weigh it...

George Henderson said...

The Lands equation:
http://jn.nutrition.org/content/132/6/1642S.full

"Analysis of the PL-FA in this study extends findings in humans and rats to dogs and supports the concept of a competitive and “saturable” hyperbolic relationship between dietary PUFA and plasma and tissue LCPUFA accumulation in dogs.

These results support the long-held belief that adequate amounts of dietary essential FA may not be >0.5 en%.

Furthermore, an eightfold relative difference between these constants was seen. This difference is consistent with the observed low metabolic conversion rates of ALA compared to those of LA from numerous other studies (8,9)."

karl said...

@George Henderson

I would agree that gut bacteria play an important role ( we exist merely to keep them alive.. )

I'm not sure that a synthetic diet would interfere with the intestinal flora - in fact figuring out just what role the flora plays again might require a synthetic diet - more research that has not been done...


As far as essential PUFA consumption - there has been an assumption that if a little bit is good - more is better - with out any evidence to back up these claims. I'm not a paleo dieter ( I would find it hard to eat all those bugs and having a chronic worm infection would seem disconcerting ) .. yet I think that unless we have hard evidence otherwise - it is probably a good idea not to change our diet in ways that early man couldn't..

karl said...

@George Henderson

I would agree that gut bacteria play an important role ( we exist merely to keep them alive.. )

I'm not sure that a synthetic diet would interfere with the intestinal flora - in fact figuring out just what role the flora plays again might require a synthetic diet - more research that has not been done...


As far as essential PUFA consumption - there has been an assumption that if a little bit is good - more is better - with out any evidence to back up these claims. I'm not a paleo dieter ( I would find it hard to eat all those bugs and having a chronic worm infection would seem disconcerting ) .. yet I think that unless we have hard evidence otherwise - it is probably a good idea not to change our diet in ways that early man couldn't..