Insulin resistance (08) UCP2

Another accidental find from my hard drive, probably saved somewhere around 2020.

Differential effects of saturated versus unsaturated dietary fatty acids on weight gain and myocellular lipid profiles in mice

These are the weight gain profiles with the linoleic acid compositions of the diets added.

A couple of comments. Obviously the low LA cocoa butter diet was indistinguishable from the low LA low fat diet, which probably also contained 35% of calories as sucrose (D12450B). I have to say I'm no fan of sucrose but one thing it does not do is to make mice fat. Some people really do think it makes people fat. Shrug.

Second point is that "olive oil" has an LA content varying from 3-21% LA as it comes from the freshly squashed olive, depending on the cultivar and the weather that year. This may increase if cut with rapeseed oil before it was purchased by Research Diet Services in the Netherlands. The study lab had, and used for tissue analysis, a gas chromatography machine, but did not use it on the diets the check their fatty acid composition. So no one knows what the mice ate in the olive oil group. I'd guess just under 4.5% of energy as LA.

All of which is pretty straight forward.

The group which I like best is the one which ate 35% of their calories from linoleic acid. That's the safflower oil group

Now this is nothing new. I've discussed very high intakes of linoleic acid as a weight loss intervention before. The current study merely replicates:

Prevention of diet-induced obesity by safflower oil: insights at the levels of PPARalpha, orexin, and ghrelin gene expression of adipocytes in mice

discussed here (and yes, these folks did measure the dietary fatty acids).

What is new is that we have both hepatic and intramuscular triglycerides measured. This is what we get in the liver:

We have to bear in mind that the low fat group, on 35% of E from fructose, should have some degree of hepatic triglyceride accumulation awaiting export as VLDLs under fasting conditions. The liver samples where obtained at 14.00h when most respectable mice would be sleeping but not hungry.

The two high LA fed mouse groups gave the two high liver triglyceride samples as you would expect and the cocoa butter fed mice resisted insulin's signal to store liver fat. Nothing exceptional.

But 35% linoleic acid in your diet is very protective against triglyceride accumulation within your liver, in addition to be moderately protective against obesity. Before we go on to mechanisms let's also look at muscle triglycerides. They look like this:

with the one problem of the cocoa butter based diet. Despite complete protection from obesity and hepatic lipid accumulation, stearic acid does not protect against lipid build up in the muscle cells. I find that difficult to understand.

I would guess one of two things. Perhaps the data are wrong. I very much doubt this, the group seem quite innocent. Second, running your metabolism on 45% fat vs 10% fat might require some degree of triglyceride storage in muscle cells, facilitated by insulin, which seems more plausible. Though why the various levels of LA did not produce a differential effect is problematic. File under "think about it".

So, on to the real question. Why is a diet containing 35% of energy as linoleic acid completely protective against lipid accumulation in the liver?

We can start with UCP1. The liver does not, as far as I can find out, ever express the gene for UCP1. Given a caloric overload to hepatocytes they just off-load excess caloric substrate, especially fatty acids, to BAT using FGF21 and let the BAT get on with generating a warm, enhanced oxygen consumption environment so the hepatocytes only see a caloric supply they are happy to deal with. That's the job of UCP1. And thermogenesis is nice for mice at 20 degC.

UCP2 is used in many, many tissues, but is not constitutively expressed in the liver. And it's not thought to be used for thermogenesis. When you supply the liver with FFAs for 24h, the gene is expressed whether the FFA supplied is oleate or linoleate, so I'd guess this is a generic property of FFA exposure.

UCP2, a Member of the Mitochondrial Uncoupling Proteins: An Overview from Physiological to Pathological Roles

We know that FFA oxidation will automatically generate ROS without needing high delta psi if the delta psi is above a certain value. It looks like UCP2 has a function (among many) of reducing the generation of these fatty acid oxidation derived ROS, at least in liver tissue. Three quotes from the authors:

"The UCP2 activity is increased in the presence of ROS in a manner dependent on fatty acid oxidation. As a result, the UCP2 acts to revert ROS production by decreasing the membrane potential of mitochondria through a mechanism of H+ leak that could be different from UCP1."

"The role of UCP2 as a regulator of mitochondrial ROS production is corroborated by results in the presence of nucleotide GDP that blocks the UCP2 activity, causing mitochondrial membrane polarization and ROS production [46]."

"We can assert that the H+ leak activity of UCP2 is a “relief valve” for the polarized IMM (Figure 2) to avoid the formation of superoxide anion in conditions in which the mitochondrial electron carriers of the respiratory chain are in a reduced state [4,68]."


The question is: Does UCP2 lower delta psi enough to effectively limit ROS mediated activating insulin signalling?

The normal limitation applied to insulin signalling, from the Protons perspective, is a surfeit of ROS. However it is equally possible to limit insulin signalling by reducing the generation of ROS or by scavenging the normally produced ROS using drugs such as N-acetyl cysteine or Mito-TEMPO, as in this study:

Insulin-stimulated lipid accumulation is inhibited by ROS-scavenging chemicals, but not by the Drp1 inhibitor Mdivi-1

Major caveats are that is is in vitro and uses those highly malleable 3T3-L1 adipocyte-like cells with which you can show just about anything. There is an in-vivo rodent study using NAC which probably deserves its own post some time.

Anyway, the un-answerable question is whether activating UCP2 within hepatocytes drops the mitochondrial delta psi to limit the generation of ROS to the point that insulin signalling fails and the drive from insulin to store intra hepatocyte triglyceride is limited.

We know that BAM15 can do this, it is largely studied as a management for what used to be called NAFLD. However BAM15 is a pharmacological uncoupler and has no limit to how significantly it can drop delta psi. The beauty of UCP2 is that it is inhibitable by guanosine diphosphate, used pharmacologically as an analogue of ADP. Increasing ADP above a certain level shuts down the uncoupling activity of UCP2. Both BAM15 and UCP2 can increase the ADP:ATP ratio, and so activate AMPK, but there are strict limits placed on this for UCP2 in normal physiology. 

I would posit that linoleate is an excellent agent for the induction of UCP2 gene expression in the liver and, once the protein is in place, it will be a significantly better activator of the uncoupling process than either oleate, palmitate or stearate. As in

Polyunsaturated fatty acids activate human uncoupling proteins 1 and 2 in planar lipid bilayers

probably limited by rising ADP levels, or falling ATP levels if you prefer.

Bear in mimd I am not considering a degree of uncoupling which would measurably increase whole body oxygen consumption, merely a small, targeted lowering of delta psi to reduce ROS generation and so reduce insulin signalling. It's not a "calories out" scenario.

Reduced insulin signalling results in reduced hepatic triglyceride storage, reduced intramuscular triglyceride storage and reduced adipose triglyceride storage. All resulting from LA exposure if the intake is high enough, ie with LA at 35% of daily energy intake. By 45% of calories there is evidence of uncoupling through UCP1 in BAT. As in:

Voluntary Corn Oil Ingestion Increases Energy Expenditure and Interscapular UCP1 Expression Through the Sympathetic Nerve in C57BL/6 Mice

Nobody should doubt that linoleic acid, at obesogenic levels (6-20% E), drives both adipocyte and hepatocyte lipid storage and mediates hypometabolism by accentuating insulin signalling. At around 35% E it activates UCP2 in hepatocytes (and probably in adipocytes) so limits ROS mediated insulin signalling which reverses hepatic lipid accumulation (and obesity) with a minimally detectable increase in metabolic rate. At 45% of E it activates UCP1 in BAT via FGF21 giving a detectable rise in metabolic rate in addition to avoiding pathological lipid accumulation.

It would be nice if linoleic acid metabolism effects were simpler.

They're not.

Peter