Wednesday, October 07, 2020

Full blown linoleic acid deficiency: speculation

 Tucker Goodrich emailed me the link to this lovely paper from the 1930s


and included it in a blog post

Fat and Weight Gain (a Note to Peter) and the Essentiality of Linoleic Acid

TLDR, grossly linoleic acid (or the arachidonic acid derived from it) deficient mice are small, emaciated and lose a ton of water through their skin. Think superoxide.

It's an interesting situation, with a number of possible explanations and the sort of data you might expect from 1930, good for its time but we're not going to get the massive detail which people struggle to interpret nowadays.

I started off alongside the authors and caloric loss due to the evaporation of water from both the "leaky" skin and possibly from "leaky" lungs. If you have ever tried to keep a mouse warm under anaesthesia while it breathes anhydrous cold gas and is clipped and water/alcohol prepared for surgery you will sympathise. If not, you can take my word for it that without serious attention to heat conservation, their temperature drops like a stone.

The mice lose about 10ml of water per day through their skin. With a latent heat of vaporisation of 0.541kcal/g this means the 10ml of water steals about 5.41kcal per day of energy as lost heat, just to evaporate the leaked water. That's about an extra gram of carbohydrate or protein they would need.

But the mice were ad lib fed. They may have been pretty sick from LA deficiency but they managed to eat exactly as much as the healthy mice did each day. I find it implausible that they were so skinny because they were simply too ill to manage an extra gram of food. There is absolutely no doubt that they were very sick, but I still don't find this plausible for the emaciation.

A simple extrapolation from high PUFA diets facilitating fat loss in to adipocytes to the converse, that very low PUFA diets fail to allow fat storage is nice but seems very unlikely because the curative dose of linoleic acid seems too small to represent a supply of calories for bulk oxidation.

My core question relates to the failure to store lipid in adipocytes, with a spin off question in to the poor health of the fat free rats. This is all guesswork and speculation.

I have to accept the possibility that it might be as simply as that you cannot build a functional cell membrane or mitochondrial membrane without some bendy/fluid molecules. Given an inadequate fluidity to the mitochondrial inner membrane I can see that molecules such as CoQH2 or reduced cytochrome C, which move within/over a fluid inner mitochondrial membrane might lose electrons prematurely to molecular oxygen if they cannot reach their docking site in the next complex down the ETC. That a continuous, pathological loss of electrons from carriers to molecular oxygen, to give inappropriate superoxide and H2O2 generation, would lead to unrestrained insulin resistance and failure to grow, especially failure of adipocytes to grow, seems to be a plausible explanation.

As Burr and Burr comment, the kidney is also a very energy dependent and might well fail given a failure of insulin signalling due to excessive H2O2 generation. While the rats would lack ROS damage to membrane lipids they would not lack ROS damage to cellular proteins or DNA. The cells, if insulin signalling cannot occur due to excess H2O2 generation, would also be calorie starved as well as ROS damaged.

Quite why the tails fell of of the rats is beyond even my speculation ability.

So. The failure of adipocytes to grow suggests severe insulin resistance within those adipocytes. More generalised application of this idea would lead to explaining failure of the whole rat to grow.

Who knows what actually happened?



I've long carried the idea in my head that PUFA depletion might be protective against radiation induced injury. I think I picked this up early in my low carb journey, probably from a Ray Peat follower. But that will have been over 15 years ago and I am unable to locate where this idea came from. So I went looking on Pubmed and opened a whole can of worms. Some of which are interesting.

The first is that rats fed a completely fat free diet are at increased risk of dying from radiation injury. That was this one:




"More recently, it has been observed that the survival time of male rats, as judged by the intervals at which an LD25, an LD50, or an LD75 were reached, or by the average length of survival, was progressively improved when ethyl linoleate was given in doses of 10, 50, or 100mg daily (Cheng et al., '54)."

The same lab has produced at least four studies to support this finding.

Giving a rat 10mg of a source of linoleic acid will have no effect on substrate oxidation. But it might well allow the development of an effective electron transport chain which is less likely to release a random excess of electrons to molecular oxygen.

Of more interest is this study. It's not clear (to me) whether the diets contained 10, 20 or 30% of cotton seed oil by calories or by weight. Low PUFA diets (butter) were ineffective in small amounts but effective at 20% or 30%:

Deleterious effects of high fat diets on survival time of X-irradiated mice

"At levels of 2% or 10% of the diet cottonseed oil and margarine fat increased survival time over that on the fat-free ration. When these fats were fed at higher levels ( ie. ~ 20% or 30% of the diet), however, survival time was decreased below that obtained at the lower levels of supplementation."

My take home is that rats do, absolutely, need a few milligrams of linoleic acid. Notice that the amounts used in all of the studies are in the same ball park as found by Burr and Burr to prevent their fat deficiency syndrome. I think it is an interesting speculation that LA is particularly need to manufacture an effective membrane for the ETC, especially when electrons are going to be knocked around by x-ray irradiation over and above background radiation conditions. As the dietary dose increases then the deleterious effect of the PUFA eventually predominate, certainly in the radiation injury models.

Interesting findings.



Tucker Goodrich said...

Thank you. I really don't understand this post, which was what I was hoping for.

I'll read again a few more times and hopefully it will sink in.


Peter said...

I have no idea if it is correct...


Rich said...

Thanks Peter, mind bending as always!

Since you brought up ray peat I'll ask, what do you make of him and his ideas?

raphi said...

Credit to Masterjohn who in 2008 said "the omega-6 arachidonic acid (AA) and the omega-3 docosahexaenoic acid (DHA) are the only fatty acids that are truly essential" in his paper titled << How Essential Are the Essential Fatty Acids? >>

Lots of useful references in there

Tucker Goodrich said...

Yes, Chris Masterjohn is a great resource.

Passthecream said...

I haven't finished digesting this paper yet (/dadjoke/) but I have seen problems with experimental diets containing casein before, both in those early papers and also more recent ones. Peter has written about some of these irc.

There's a nicely simplified diagram of typical casein as found in milk, here:

--- that is before it is processed into lab rat food.

To summarise technical casein preparation: it is coagulated (precipitated) from milk with acid such as vinegar or lactic acid, then filtered out and washed repeatedly, defatted by boiling with alcohol for some time then redissolved with eg ammonia or lime. That whole process is repeated many times ending up with a rubbery insoluble mass which is dried (more heat) then ground to make powder. One aim of technical casein preparation is to remove the 'ash' content which in the 1930s meant no calcium or phosphorous, so lime was not used. Since the original micelles in milk were rich sources of both calcium and phosphorous this leaves you wondering.

Next thing to mention is that when animals drink fresh milk, stomach pH plus enzymes curdle the casein ie make it completely hydrophobic so it just clumps together, after which it is one of the slowest proteins of all to be digested. Imagine how indigestible the processed industrial version is though?

From my point of view it really is industrial because just as I think of linoleic and alpha-linolenic acids as prime varnish ingredients, I also think of redissolved casein as an astoundingly durable glue (cheese glue holds the Mona Lisa panels together), and also as an ingredient for gesso (primer) for varnished and painted surfaces. Sorry about the digression.

Just as this 1930s rat paper is the one which most others refer back as establishing the essentiality of LA etc, there is another paper, from 1925, in a similar position of authority wrt the essentiality of vitamin A. It now has almost 1,950 referring papers listed on PubMed and there are many more casual articles and review papers which mention it. It is the holey grail paper of vitA research, if you don't mention this one you are nobody.


Check the diet. It also included problematic casein.

Another aspect of a fresh casein micelle is that rather like lipoproteins, these micelles have a hydrophilic outer surface so they are water soluble, but carry other molecules in their hydrophobic cores. Retinol ( vitaminA) is one of them but retinol is quite fragile wrt heat, light etc, and rapidly oxidises to become retinoic acid. Retinol held in a transport protein such as casein is safe as are the usual ester forms retinoyl palmitate, stearate (not so sure about linoleate?) Casein btw has a high affinity for retinol but about 5 times higher affinity for retinoic acid.

Retinoic acid is pretty nasty. It is usually the end product of retinol metabolism in animals and it is hard to get rid of. Retinoic acid in excess can cause most of the terrible side effects that were noted in both these papers. There is a possibility at least that these seminal papers were using baseline diets with toxic amounts of retinoic acid. It doesn't take much RA or retinol to poison a rat. That gives a laughable view of the 1925 paper, considering it was supposed to demonstrate the horrors of vitamin A deficiency. But both papers should be retracted or somehow annotated due to the problematic diet used.

These experiments would have been much more useful if they had used dried (or fresh) skim milk in the diets rather than technical casein, or maybe peanut flour or lamb chops, anything!

Scrivener said...

This reminds me of William Lands’ s presentation to the us Amy. Too little PUFA bad; too much PUFA bad. With a steep hyperbolic curve from one to the other.

Marius said...

Hey Peter,
after listening to the podcast with Paul Saladino I had a thought which I would love to get your comment on. Would it be correct to say that the major pathologic mechanism which presents itself in the form of metabolic dysfunction with all the associated problems (oxidative stress, degeneration, cancer,...) is simply the inability to restrain energy flux through the cell?

Tucker Goodrich said...

I think the aberrant energy production in the cell is a side effect of the root problem, which is basically food poisoning. See here for the cleanest demonstration I've seen of how to induce mitochondrial dysfunction.

"What's Worse—Carbs or Seed Oils? Understanding a High-PUFA Diet."

Oxidative stress, for instance, is the process of n-6 fats oxidizing. The oxidized n-6 fats then go on to cause the rest of the mayhem, such as DNA damage of the p53 anti-cancer gene, etc.

Peter may have a different opinion, of course!

BTW Peter is interviewed on Paul Saladino's latest podcast. 2 hours, I am planning on listening this morning.

Passthecream said...

Tucker " a side effect of the root problem, which is basically food poisoning."

Very neat way of putting it! Interesting blog post also. (I see that casein made it into one of there though. Hmmmm.)

I read this book by David Gillespie years ago

"Toxic Oil: Why Vegetable Oil Will Kill You & How to Save Yourself"

Similarly blunt. It did me a lot of good.

Are you aware of any studies which specifically look at fructose or sucrose plus pufa as opposed to glucose? I always have in mind Peter pointing out that a bolus of fructose is mostly unregulated by the usual processes which deal with glucose.

Peter said...

Hi Marius, yes, that is an idea I keep getting pulled back to. I think it might hold water…

Tucker, I go down a layer. Linoleic acid fails to limit caloric ingress. Excess caloric ingress will still produce an ROS signal but it will be stopping further caloric ingress at a higher level of calories than the cell really wants. Given that the double bonds in linoleic acid are the problem I see the solution best suited to successful evolution as noting the product of ROS interacting with those problematic double bonds as the signal to really, really shut down caloric ingress. So both locally generated lipoxide derivativess or orally consumed derivatives are how a cell limits caloric ingress when physiological high end ROS signals are not enough in their own righ…


Passthecream said...
This comment has been removed by the author.
Tucker Goodrich said...


"Linoleic acid fails to limit caloric ingress."

It's worse than that, it stimulates caloric excess!

Peter said...

Tucker, are you thinking along the AA/endocannabinoid line? Still not had time to pick at that one...


Passthecream said...

Peter, I'd love to see you do a post on the munchies ...


cavenewt said...

Having always been a little curious about Ray Peat, who Peter seems to respect but whose ideas about protein I'm not sure I agree with, I found this 2013 blog post incredibly dense and thought-provoking. It seems to be a summary overview of fats and cellular processes. It touches on a couple of personally relevant health issues (a mystery peripheral motor neuropathy and a brief excursion into hormone-positive breast cancer last year), as well as possible impacts of polyunsaturated fats on COVID-19 symptoms.

I wasn't so aware of the impact of estrogen on some of these metabolic functions. Can't help wondering if my many years of early birth control pill consumption, which in the 70s and 80s were much higher-dose hormones than later, combined with the dutiful substitution of polyunsaturated oils for animal fats, might have contributed to this to neuropathy and cancer.

I'd be really interested to see what the smart people here have to think about it, although being so long it's kind of an investment in time.

Puddleg said...

This suggests a question:

Does it ever appear anywhere in the literature that atherosclerosis is a symptom of linoleic acid deficiency?