"Hello, Can We Have Your Liver?": Understanding a High-PUFA Diet.

tl;dr: A diet high in omega-6 and omega-3 polyunsaturated fatty acids has some positive effects on the body, but also results in progressive liver failure.

tl;dr: A diet high in Ω-6 and Ω-3 polyunsaturated fatty acids has some positive effects on the body: lower weight gain, better preservation of lean mass, improved blood lipids, and increased brown adipose tissue; but also results in increased oxidative stress, mitochondrial dysfunction, and beginning of progressive liver failure.

This paper is a classic

"Fat Quality Influences the Obesogenic Effect of High Fat Diets [HFD]" (Crescenzo et al., 2015)

Sounds benign enough, right?  We all like quality fats...

"To investigate whether polyunsaturated fats could attenuate the above deleterious effects of high fat diets, energy balance and body composition were assessed after two weeks in rats fed isocaloric amounts of a high-fat diet (58.2% by energy) rich either in lard or safflower/linseed oil. Hepatic functionality, plasma parameters, and oxidative status were also measured. The results show that feeding on safflower/linseed oil diet attenuates the obesogenic effect of high fat diets and ameliorates the blood lipid profile...."

That's terrific!  So we just need to eat more Ω-6 and Ω-3 fats, and we'll be thinner with better blood cholesterol!

A nice example of a rat diet study

So these two sets of rats got two remarkably well-constructed diets, with 58.2% of fat from either lard (L) or safflower and linseed oil (S).  Lard is the classic bogeyman of rat diets. "Eat your carbohydrates, children, or Lard will get you!" Nice to see them adding in the Ω-3 fats from linseed oil—gold star. This accomplishes two things: First, it's well known that sufficient Ω-3 ameliorates the obesity often induced by high Ω-6 diets, so they're assuring the outcome. I don't know if they knew this, but I did and noticed it immediately. Second, they get around the Ω-3/Ω-6 ratio question, as the S diet actually has a better ratio than the lard diet does: the L diet has an Ω-6 to Ω-3 ratio of 12:1, which is pretty bad, and the S diet has a ratio of 4:1, which is pretty good, by lab-rat standards.

(Substack doesn’t support tables. Data at the bottom.)

So the L rats got 8.11% of their daily bread (“% E”—Per cent. of Energy) from linoleic acid (LA), and just a smidge from the Ω-3 fatty acid alpha-linolenic acid (ALA).  The lucky S rats, fed the healthy, anti-obesogenic, cholesterol-lowering high-PUFA diet got 34% (!) of their calories from LA, and a whopping 8.42% from ALA.

The full diet is in the following two images. By lab-diet standards, this is incredibly well done, as they even give the exact breakdown of the individual fatty acids (FA). They get a bit of a demerit for including "chow" as a line-item, and not breaking that out, but at least both arms are getting the same amount of chow, and both are getting the same 20.7%E from carbohydrates. We can infer this is not a high-sugar diet.

I'll say it again.  This is a lovely diet, they're really doing a neat job of controlling their variables here. Lots of heart-healthy PUFAs, and a big dose of plant-based Ω-3 fats to balance the somewhat troublesome Ω-6 fats.

How to make lab rats fat

Now we know that 8% E of LA is plenty to induce obesity in rodents.  See here:

"Dietary linoleic acid elevates endogenous 2-AG and anandamide and induces obesity." (Alvheim et al., 2012)

And sure enough, these rats got fat.  Interestingly, the L rats got fatter than the S rats, despite eating much less LA.

"After two weeks of isocaloric high fat feeding, obesity development was evident both in L and S rats, since their percentage of body lipids about doubled compared to initial value, although the final value was significantly lower in S rats than in L rats (Figure 1A)." (Crescenzo et al., 2015)

Rats don't do very well on HFDs with LA, as the Alvheim paper shows, so it's not surprising that these rats all got fat. It's also been shown that adding Ω-3 fats helps prevent obesity in rats, so that added linseed oil seems to benefit the rats, although it clearly didn't prevent obesity here.

Benefits of a high-PUFA diet, versus lard

What's worse, the L rats got worse fat than the S rats did; visceral (abdominal) and epididymal (the man parts) fat.

"In addition, the percent of epididymal and visceral white adipose tissue (WAT) increased during dietary treatment, reaching a final value that was significantly lower in S rats than in L rats (Figure 1C,D)."

Not only that, but the L rats lost more lean mass than the S rats did, so they're going to have to go to the gym more often.  

"Therefore, it appears clear that L rats exhibit an impaired metabolic flexibility that exacerbates obesity development."

So far, it appears that lard is an unhealthy fat, and that the safflower/linseed combination is far superior. The higher-PUFA diet is also in line with the dietary recommendations, so that's also a benefit.

Brown Adipose Tissue

There were a number of other benefits to the high-PUFA HFD, including better fuel burning, and increased brown adipose tissue (BAT), which is used to turn fuel into heat to keep the animal warm, and also helps with disposing of excess calories. Cholesterol also went down, and for those who like the current dietary guidelines, that's also a plus.

Unfortunately those blood lipids went somewhere...

The Catch

Oh, wait a minute...

"Plasma metabolic characterization evidenced lower cholesterol but higher lipid peroxidation and ALT activity in S rats compared to L rats (Table 3). At variance with plasma lipid profile, livers from S rats had higher lipids, triglycerides, and cholesterol, as well as higher lipid peroxidation..."

And...

"...Conversely, hepatic steatosis and mitochondrial oxidative stress appear to be negatively affected by a diet rich in unsaturated fatty acids."

Oh, darn.  "Negatively affected"? Here's where things go off the rails. One expects a lard-based diet to be bad for rats, who reliably get fat on such diets. But swapping out the MUFA and SFA for PUFA is supposed to make things better, and it has so far in this study.

Until we get to the mitochondria and the liver.

TBARS is a test looking for a marker of lipid peroxidation (LPO), which is the oxidation of PUFAs, essentially; see table 3.

TBARS are notably higher in the S diet. It's not surprising, I suppose, that the oxidation of PUFAs should be much higher on a higher-PUFA diet. But LPO is not a good thing, as the products of LPO are toxic. TBARS is a marker of malondialdehyde (MDA) one of the worst such LPO products. They should have also tested for 4-hydroxynonenal (HNE), in my opinion, but such is life. (We discuss MDA vs HNE testing in episode 10 of the podcast.)

LPO has very negative effects on mitochondria and the liver, and sure enough that's exactly what is seen in these rats. A paler liver indicates a fattier liver, and non-alcoholic liver disease (NAFLD) is a major problem in both rats and humans fed a HFD. Here we see that NAFLD is far worse (the paler colors) in the bottom row of images, from rats fed the supposedly-healthy S diet. What's really notable, and a particularly brilliant part of this paper, is these are rats with a high Ω-6 diet, but a low n-6/n-3 ratio. Which doesn't seem to have helped their livers at all.

They’ve come for your liver.

Liver Ω-6 concentrations change:

"...there was a significant increase in the omega 6 fatty acids linoleic, gamma-linolenic, eicosadienoic, and dihomo-gamma linolenic and in the omega 3 fatty acid docosapentaenoic (Figure 5B), in S rats compared to L rats."

Except the liver mitochondria was altered:

"However, some differences were evident in the content of specific fatty acids, such as the monounsaturated fatty acid oleic acid and the omega 6 fatty acids gamma linolenic, arachidonic, and docosapentaenoic, that were found to be significantly decreased, while the omega 3 fatty acids alpha linolenic, eicosapentaenoic, and docosapantaenoic were found to be significantly higher, in S rats compared to L rats (Figure 5A)."

This might confirm a post of mine: How To Prevent Oxidative Damage In Your Mitochondria

"These reactive oxygen species readily attack the polyunsaturated fatty acids of the fatty acid membrane, initiating a self-propagating chain reaction." (Goodrich, 2016)

In that post I discussed how a high Ω-6 diet affects tissue composition, and how the mitochondria could be damaged by such a concentration. In this study we get to see it in action. Increased TBARS suggests that the Ω-6 membranes are undergoing the reaction described in the post above, and the Ω-6 fats have been replaced by non-peroxidizable SFA and MUFA fats. It's the same process described in this study:

"...patients with ARDS decrease their percentage plasma concentrations of total plasma linoleic acid, but increase their percentage concentrations of oleic and palmitoleic acids. As plasma linoleic acid concentrations decreased, there was usually an increase in plasma 4-hydroxy-2-nonenal [HNE] values, one of its specific peroxidation products, suggestive of severe oxidative stress leading to molecular damage to lipids. (Quinlan et al., 1996)

I won't go into it here, but these findings seem to confound much research into negative effects of omega-6 fats: the simple n-6 fat level alone does not tell you about the pathological process, as less can mean it's further along.

Conclusion

This is one of the neater studies I've seen, and it seems to have been created as a refutation of the premise, stated in the paper:

"In fact, some authors have hypothesized that HFDs rich in unsaturated fatty acids are less deleterious for human health than those rich in saturated fat [12–15]...." (Crescenzo et al., 2015)

They successfully illustrate that higher PUFA in the diet do have the effect they surmise:

"...However, polyunsaturated fatty acids exhibit the highest sensitivity to reactive oxygen species (ROS)-induced damage, their sensitivity to oxidation exponentially increasing as a function of the number of double bonds per fatty acid molecule [16]. As a consequence, if antioxidant defense systems are unchanged, a higher degree of fatty acid unsaturation in cellular membranes may increase their sensitivity to lipid peroxidation and would also expose other molecules to lipoxidation-derived damage."

The effect on the liver alone is enough to demonstrate that this higher PUFA diet is unhealthy. However they also manage to demonstrate, in the process, that some of the often-touted benefits of higher-PUFA diets are in fact parts of the pathological process.

(Title is from one of the more bizarre Monty Python skits.)

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Data and References

L S

Fat 58.20% 58.20%

LA 13.93% 59.13%

% E 8.11% 34.41%

ALA1.12% 14.47%

% E 0.65% 8.42%

Alvheim, A. R., Malde, M. K., Osei‐Hyiaman, D., Hong, Y. H., Pawlosky, R. J., Madsen, L., Kristiansen, K., Frøyland, L., & Hibbeln, J. R. (2012). Dietary Linoleic Acid Elevates Endogenous 2-AG and Anandamide and Induces Obesity. Obesity, 20(10), 1984–1994. https://doi.org/10.1038/oby.2012.38

Crescenzo, R., Bianco, F., Mazzoli, A., Giacco, A., Cancelliere, R., Di Fabio, G., Zarrelli, A., Liverini, G., & Iossa, S. (2015). Fat Quality Influences the Obesogenic Effect of High Fat Diets. Nutrients, 7(11), Article 11. https://doi.org/10.3390/nu7115480

Goodrich, T. (2023, October 26). Ep. 10: Dalton Graham: How to Induce Fatty Liver—With Dr. Brian Kerley [Substack newsletter]. Tucker Goodrich: Yelling Stop. https://tuckergoodrich.substack.com/p/ep-10-dalton-graham-how-to-induce

Goodrich, T. (2016, February 23). How To Prevent Oxidative Damage In Your Mitochondria [Blog]. Yelling Stop. https://tuckergoodrich.substack.com/p/how-to-prevent-oxidative-damage-in

Quinlan, G. J., Lamb, N. J., Evans, T. W., & Gutteridge, J. M. C. (1996). Plasma fatty acid changes and increased lipid peroxidation in patients with adult respiratory distress syndrome. Read Online: Critical Care Medicine | Society of Critical Care Medicine, 24(2), 241–246. https://doi.org/10.1097/00003246-199602000-00010