Saturday, April 10, 2021

The ginger paradox (5) and some speculation

TLDR: If resisting insulin is rendered impossible due to very, very high PUFA diets then mitochondria resort to uncoupling.

reiterated the well known essentiality of FFAs for the activation of UCP-1 and added in the significant role of reactive alkenyl species. In more common parlance that would be lipid peroxides, primarily those derived from linoleic acid. I thought about it in this post.

This is the next step, which I specifically went looking for because this is how I think life should be organised. It is a perfect example of me having a huge bias, knowing what I want, and going looking for it. You have been warned.

It is utterly clear that UCPs in general suppress ROS generation and that some members of the family are not being used for thermogenesis at all, more likely for prevention of the generation of supra-physiological levels of ROS.

We have an interesting situation where PUFAs, the prime pathology of which is their failure to generate adequate ROS, are also extremely effective at facilitating uncoupling, which needs significant ROS generation to happen. Where does this superoxide come from?

This is the core question.  

Warning: I think step f) is the one I have least evidence for. Life is like that sometimes.

Okay. The story so far:

Caloric ingress in to a cell is controlled to an appropriate level while calories are freely available by limiting insulin signalling.

Insulin signalling is curtailed by driving electrons in reverse through complex I to generate superoxide and hydrogen peroxide which disable/limit said insulin signalling.

This reverse electron transport rises in direct proportion of FADH2 supplied compared to the amount of NADH.

The presence of double bonds in fatty acids limits FADH2 generation, limited FADH2 generation limits the control of insulin facilitated caloric ingress and continued insulin signalling diverts calories in to storage.

I have to accept that not all excess calories will be stored, some will continue to enter the electron transport chain as both NADH and FADH2. If electrons enter the electron transport chain when there is a low (relative) demand for ATP then

a) mitochondria membrane potential will rise, a direct consequence of the inability to limit electrons entering the ETC through complex I and multiple FADH2 entrance points. Protons are pumped but not used

b) the cytochrome C pool will become progressively more reduced, limiting it's ability to accept electrons from complex III

c) electrons will be transported through the cytochrome b centre of complex III back on to the CoQ couple

d) these electrons will replace those absent as a result of double bond containing fatty acid oxidation and will restore superoxide production

e) superoxide production, under these circumstances, is being facilitated only at the cost of high membrane potential secondary to low ATP synthase activity in parallel to excess calorie storage

f) if the double bond content of the lipids providing input to the CoQ couple from beta oxidation is very, very high then the need for negative feedback through complex III will be very, very high as well and will be associated with an excessively high membrane potential

g) the answer to excess membrane potential is uncoupling

h) superoxide positively regulates the activity of UCP-1 as do superoxide derivatives of LA

i) uncoupling allows the flow of electrons down the electron transport chain and lowers membrane potential, it immediately stops reverse electron transport/superoxide generation and converts the excess calories entering the cell in to heat, water and carbon dioxide


Normally regulated calorie ingress to a cell occurs when linoleic acid is low.

Excess ingress from LA is dealt with by diversion to storage, leading to obesity.

Marked excess is dealt with by uncoupling which does not "require" obesity.

There are no hard boundaries between the above conditions, there simply appears to be an inverse "U" shaped curve in response to the content of linoleic acid in the diet. Such curves are common in nature.

A few last thoughts:

Simply looking at the experimental data the situation appears to be that including linoleic acid at levels between somewhere just under 10% up to somewhere in the high 20%s will facilitate excess insulin action and excess lipid storage. As we increase the supply of linoleic acid through the 30%s and in to the 40% region of calories then uncoupling becomes the primary molecular solution to a progressively more intractable problem.

It is worth pointing out that in a given mitochondrion there are lots of CoQ/CoQH2 molecules, lots of beta oxidation sites and, while the calculation of FADH2:NADH is useful for insight as to how RET works for a single fatty acid molecule, the mitochondria are integrating lots of FADH2 production sources, lots of NADH sources and are continuously monitoring the membrane potential (which is an integration of energy input/need) as well as the redox state of the CoQ couple.

What happens on a whole organism basis is also an integration of all of these effects. The mechanism for weight loss in high PUFA diets is both comprehensible and plausible.

The studies are real, the data are real.

Personally I still wouldn't go there, but understanding them is key. 



Tucker Goodrich said...

From 2017:

"Strong indications that UCPs evolved in association with pressure to reduce ROS formation during β-oxidation can be found in regulation of expression and activity. The first important experiments were performed by the Brand group using yeast mitochondria expressing mammalian UCP-1 [137]. Here, palmitate (saturated C-16) and an oxidized (!) FA (4-hydroxy-2-nonenal) enhanced UCP proton transport. Synergistic effects were observed using both [137]. Oxidized FAs like 4-hydroxy-2-nonenal signal encountering ROS. The uncoupling predicted by the kinetic model indeed should be enhanced during β-oxidation, and even more so upon ROS production, as monitored by oxidized FAs."

"The "(!)" is his, so we know how surprising he found this pathway!"

From the comments.

But it makes sense, HNE is a signal to disengage the clutch before damage occurs.

Peter said...


I keep going back to that paper

Brief episode of STZ-induced hyperglycemia produces cardiac abnormalities
in rats fed a diet rich in n-6 PUFA

and it won't leave me alone. If I pull anything useful out of my musings I'll comment. It fascinates me that the PUFA/diabetic mice had FFAs of 1.8mmol/l in addition to the glucose of 25mmol/l. Given the picture painted in these current post I'm wondering about the PUFAs, ROS and uncoupling.

Might well come to nothing. But I'm mulling it over.


cavenewt said...

I was with you up until g), but clearly have lots of thinking still to do, because I kept picturing it as:

"g) Then the mitochondrion explodes."

Tucker Goodrich said...

They implode, actually. 😉

karl said...

Interesting - you might add something about the levels in really bad human diets. I would expect these experimental levels to generate an uncoupling fever?

Levels of LA exposure in diets we evolved with would be MUCH lower - (I remember reading biographies of people captured by primitive people - there was a bit that repeated in their narratives - a hunt - followed by a feast - followed by hunger - often for days at a time. I wonder if hunger periods were the same for people living on the ocean or tropics? )

I wonder if a high enough level of LA could trigger apoptosis?

Trying to understand apoptosis vs autophagy vs insulin levels ..
Insulin appears to suppress both? I'm assming insulin sensitivity is involved - LA modulated?

If it was simple as a switch, it would seem that people with sky high insulin would have seriously compromised immune action - if the immune system can't tell a cell to self destruct. I don't think it is so simple. Must not be total suppression? We need some AI to help us out.

Passthecream said...

Talking about imploding and exploding mitochondria, I sometimes wonder about the dielectric strength of the inner and outer mito membranes. Given a high enough potential difference does it arc over or does the charge just leak away? And what is the nature of whatever it is that occupies the space that the protons are stored in? Strictly speaking that is the dielectric and the membranes are electrodes I suppose. But this is biology and/or chemistry and things aren't usually that straightforward.

Perhaps a membrane with a high level of pufa derived cardiolipin would have a lower dielectric strength?

Uncoupling is a structured and controlled ( you hope) form of leakage as is the usual harnessing of a proton current to regenerate ATP. Are there other controlled leakages, other conduction channels?

Peter, remember there was a Brit. sitcom called "Coupling"? Good to see you working on a treatment for the sequel! :)

Passthecream said...

Hmm no, the conduction channels and other leakages are analogous to wires or electrodes where the currents flow in and out. Diodes, more or less. Internal goop where the protons are kept is another electrode, the insulating properties of the membranes are dielectric.

I think.

With protons on one side and electrons on the other the potential diff is actually across each membrane. Negative charge would be attracted to the outer membrane also so there is a possibility of harnessing current flow outwards also. This is my primitive electrical brain trying to understand biochemistry.

Passthecream said...

Ah, think if it as like a supercapacitor, ultracapacitor or pseudocapacitor, many variants:

" The working mechanisms of pseudocapacitors are redox reactions, intercalation and electrosorption (adsorption onto a surface). "

Peter said...

Pass, proton leakage was an intrinsic feature of the genuinely primitive membranes which were pretty much accidentally generated in LUCA where metabolism and evolution started in the absence of biological membranes. The original membrane energetics actually used Na+ ions because crude, non specific membranes can be formed which were opaque to Na+ but would easily allow H+ to cross. No point in pumping H+ if it just leaks back through. Pumping Na+ works over a membrane which is an insulator to Na+ but not H+.

Nowadays the capacitor effect is absolutely dependent on a proton opaque insulator. Fine unless you are a hyperthermophile in which case it's back to pumping Na+ as hot lipid membranes are H+ transparent...


Gyan said...

Doesn't the mainline keto diet for epileptic children contain PUFA in these >30-40 percentages?
I wonder how long children are supposed to keep themselves on this diet and how they fare with it.
Do they tend to be slim, not feeling cold?

Peter said...

Yes it does but it's also profoundly hypo-insulinaemic so PUFA don't matter, as in F3666 rodent diet. They would probably do even better on a saturated fat diet but hey...


Passthecream said...

That's interesting about sodium vs hydrogen ions.

The mitochondrial 'capacitor' is what it is but isn't the same as an ordinary capacitor as in small electronic circuits because those store electric fields via charge separation and no charged particles enter or leave the insulating layer --- they are Helmholtzian Capacitors. The mito membrane potential capacitor is of a type known as a pseudo-capacitor because charged ions are stored in it by being pumped in. It is a Faradaic capacitor.

How the protons are actually held in the intermembrane space could be interesting to understand. It's unlikely they'll just be drifting around in there freely. They may be intercalated in some protein-polymer structure, or ???

There will be limits to the ability of the membrane to contain potential difference and at some level it will break down although waxy-fatty compounds are very effective insulators. Hopefully uncoupling cuts in and runs fast enough that this never happens.

Passthecream said...

Also you might expect that at a certain value of Delta Psi the electrons running along the etc will simply not have enough energy to be able to power the uphill proton pump.

Peter said...

Absolutely, the protons don't just sit there. The most interesting bacteria on this front live in alkaline lakes. Not a lot of point pumping protons when the external pH is 11 and your proton gradient turns to water! There is thought to be a specialised local environment between the pumped protons and ATP synthase, possibly some sort of channel.

Of course the advantage of Na+ pumping in oceanic circumstances is that the external Na+ concentration is fixed by the ocean and the Na+ gradient is achieved by decreasing the intracellular Na+ concentration. Which is probably why cytoplasm is a low Na+ environment...

Re delta psi and resistance to electron flow, yes, this is exactly how electrons double back through complex three to the CoQ couple and generate ROS.

These people weren't looking at superoxide, rather at lipoxides, but they are just a surrogate for superoxide...



Passthecream said...

That's an interesting paper.
It's another deep rabbit hole.

Fascinating details about ANT and a nice diagram here

But wait, there are ROS in the intermembrane space and ATP is going in, ADP coming out, what the hell is going on in there ?!?

As Hamilton Mattress said "I feel so ANTy"

Peter said...

ANT2 is interesting! Clearly mitochondria require ATP, they do a lot more than generate the stuff... Whatever goes wrong with ox-phos in cancer cells, their mitochondria have ATP requiring processes. Now, it will be interesting to go and look at ANT2 and rho zero cells.......


Tucker Goodrich said...

"ANT possess diverse functions including the electrogenic exchange ADP/ATP across the IM, the participation to the PTPC stimulate by Ca2+ and ROS, a mild uncoupling activity that is stimulated by anion superoxide, fatty acids and peroxydated lipids....

"Interestingly, the ANT crystal analysis confirmed the existence of a threefold repeat of about 100 amino acids and the binding of the IM-specific lipid cardiolipin...

"The ADP/ATP translocase inhibitor CAT is shown in yellow and cardiolipins residues in brown."

Sadly they skirt this topic, despite oxidized cardiolipin having been shown to induce apoptosis. Seems like this may be part of that process...