Insulin resistance (18) The red line

Before I get on to the absolute treasure trove which is this paper from Tucker I would like to continue a little with this one:

α‐Tocopherol suppresses hepatic steatosis by increasing CPT‐1 expression in a mouse model of diet‐induced nonalcoholic fatty liver disease

In the last post I considered Fig 4 to show the role of α‐tocopherol in reducing the insulin resistance of high levels of ROS to allow a more effective insulin-like signal from those ROS in an insulin-free cell model system.

It's time to think about what is happening in intact mice. Which is different.

Here's the supplementary image again:

which we can simplify to this basic scheme with the measured levels of α‐tocopherol added in:

The red line is easy. I start from the premise that lipid storage in hepatocytes is mediated by insulin. Exactly as per adipocytes, resisting insulin resists hepatic lipid storage. We know that long chain saturated fats protect against fatty liver *because* their oxidation resists insulin's signal better than does the oxidation of linoleic acid. Going back to my old scheme of what I think is happening we have this effect from LA on insulin signalling. First is the normal resistance to insulin's signal limiting adipocyte size. Hepatocytes are not adipocytes but I consider this aspect still holds:

and if we allow extra insulin signalling by oxidising linoleic acid, a fat which fails to adequately resist insulin, we get this:

in which the peak ROS signal is the same, here an hypothetical equivalent to 0.3mM H2O2, but it needs extra caloric intake to achieve this "satiety" level of ROS. If we add in a couple of rising dose rates of α‐tocopherol we get hepatocytes which store even more fat than is the case for plain LA. We still have that peak 0.3mM equivalent of ROS but we need even more caloric ingress to achieve it. Some gets stored as fat in the liver:

This is pure Protons. ROS reduction means more signalling which means more intracellular lipid storage. Note that there is no suggestion of any increase in peak ROS hypothesised, the increase signalling action is mediated by α‐tocopherol limiting the "stop" signal, it's just that more signalling is allowed before that putative 0.3mM H2O2 is reached. So the ALT is not coming from ROS mediated direct damage.

If we push this process in adipocytes we end up with rising basal lipolysis, a process which is protective to individual adipocytes and cannot be suppressed by insulin.

If we push this process in hepatocytes there is no basal lipolysis. FFAs are absorbed by hepatocytes as metabolically active free acids and rendered inert by conversion to triglycerides by combining them with glycerol. These inert triglycerides are exported as VLDL under low insulin conditions. That's normal.

There is, undoubtedly, a "basal" VLDL secretion rate. The problem for hepatocytes is that VLDL secretion is not free from the control of insulin. Excess delivery of FFAs to hepatocytes in the presence of elevated insulin will trap triglycerides in hepatocytes.

Elevated basal lipolysis from adipocytes delivers excess FFAs and is fundamental to hyperinsulinaemia. Hyperinsulinaemia is fundamental to NAFLD.

Aside: The most effective management for NAFLD is caloric restriction. This drops adipocyte size which drops basal lipolysis which drops insulin which drops hepatic lipid storage. This simple management is complicated, in the presence of linoleic acid, by unremitting hunger. So it always fails. End aside.

So my view is that liver cells under normal physiology are insulin sensitive within the limits set by Protons, that this insulin sensitivity is still under the control of ROS and that linoleic acid, or α‐tocopherol, allows too much insulin signalling before the normal storage limiting signal of high ROS occurs. So why the damage?

I hope everyone recognises this image:

These are adipose tissue crown-like structures stained for macrophages in this study. If you want to see the same structures in liver tissue you need to go to a different study to find them. The black arrows are placed by the authors and are specifically denoting crown-like structures. They don't look as neat as in adipocytes because hepatocytes have lots of messy cytoplasm which gets in the way:

I discussed crown-like structures in a previous post or two but we can summarise by saying that, while triglycerides enclosed in perilipin proteins are inert, above a certain size the perilipin storage breaks down and all hell breaks lose on an inflammatory basis. In the soup of TNF-α and IL-6 surrounding the remains of a dead hepatocyte the still viable hepatocytes will, undoubtedly, become insulin resistant and share this disaster signal with the rest of the body.

Okay, okay. If you insist, here are crown-like structures in adipocytes stained for TNF-α and IL-6, because the images are so pretty. Given enough funding for these very expensive antibody stains you could show exactly the same in hepatocytes:

While the macrophages are what release the cytokines it is the dead hepatocytes which release the ALT. That is where the red line is coming from:

This red line process is pure Protons plus "pyroptosis", unlike the blue line.

I'll take a brief pause here because I have a ton of other stuff to do before I get to the blue line, which is much more interesting.

Peter

Insulin Resistance (17) ROS NOX2 RET and alpha-tocopherol in cell culture

I'd like to just state, as I start this post, that I have absolutely no doubt that adding a modest dose of α-tocopherol to a lard based obesogenic high fat diet is protective against the associated fatty liver and liver damage. The mechanism is not clear to me as yet, but while pondering it a perfectly reasonable explanation for the problems caused by higher doses of α-tocopherol became apparent. That's what this post is about.

I want to discuss the supplementary figure S2:

taken from

α‐Tocopherol suppresses hepatic steatosis by increasing CPT‐1 expression in a mouse model of diet‐induced nonalcoholic fatty liver disease

ALT is a routine indicator of liver damage, more specifically, of hepatocellular injury.

I'm interested in how progressively increasing levels of dietary α-tocopherol produce worsening liver damage on an high linoleic acid background. I've added in, from Fig 1 panel C, the measured plasma levels of vitamin E involved for the two oral dose rates which we are given, plus I've removed those parts of the chart which are not relevant to the discussion:

From other parts of the paper the group realised that CPT1, the main mitochondrial fatty acid uptake protein, is down regulated in α-tocopherol liver damage and that down regulating CPT1 function using an inhibitor restores the toxicity eliminated by α-tocopherol added at 50mg/kg to the high fat diet. I agree that the role of CPT1 down regulation might be important to the hepatotoxicity of high dose α-tocopherol.

So they went to a cell culture model to see if α-tocopherol reduced CPT1 activity in HepG2 cells.

This is their bar chart:

and this is the line I wish to discuss:

People may have noticed that I have ignored the value for 1μM of added α-tocopherol so here's an aside to attempt to justify this fudge. I have several reasons for this. Primarily it doesn't fit my hypothesis, you have been warned. Added to this is that it has a higher standard deviation than all other bars on the chart, especially those containing α-tocopherol. It's very difficult to interpret this because the methods do not specify if the cell cultures were replicated, if repeated aliquots of tissue protein from the cell culture were analysed and averaged or whether Western Blots were repeated and the data presented are the averages of several densiometry measurements. We could tell a fairy tale which looks like this, with tightly grouped results in four of the bars but with one outlier at 1μM in culture:

the elimination of which would allow us to redraw the bar chart to look more like this:

I have to emphasise that this is a COMPLETELY hypothetical situation. The only problem is that it makes sense once we think about mechanisms.

So let's think.

It takes thirty seconds on any search engine to ascertain that insulin controls the transcription and expression of the gene for CPT1, negatively.

The more we facilitate insulin signalling, the lower CPT1 will be. These are the terms in which we need to be looking at the cell culture results.

The cells were cultured, for 24h, in "10% fetal calf serum/Roswell park memorial institute (FCS/RPMI)‐1640 (Sigma)" which not only provides 25mM of glucose and everything else a cell might want, it also provides significant amounts of insulin and IGF1.

They were then transferred to 1% FCS/RPMI‐1640, which also routinely provides 25mM glucose but with no insulin, IGF1 or any appreciable fatty acids. These cells are quiescent and give a stable platform for assessing CPT1 production without the complication of rapid cell growth over 24h.

Anyone who has followed this blog for any number of years will be very aware that glucose, acting via an NADPH oxidase, in this case NOX2, is an insulin mimetic in its own right. A jump from 5mmol/l to 10mmol/l in humans with pharmacologically fixed insulin levels will demonstrate this insulin-like signal.

This is the paper and this is my doodle drawn on somebody else's doodle:

Equally, we can go to this paper and see that glucose at 30mmol/l produces a seriously damaging level of ROS and insulin *resistance*. Here's my doodle:

The full discussion is in this blog post.

In cell culture at 25mM glucose I would posit that there is a serious ROS signal being produced in the complete absence of insulin which looks, from an HepG2 cell's perspective, a lot like supra-peak insulin signalling. And it's stable. The cells are not like mice, they don't do meals. There is a constant ROS signal equivalent to supra-physiological insulin. This "insulin-like" ROS signal, by imitating insulin, sets our CPT1 baseline level in this column of the bar chart:

Our next move is to look what happens when we add 1μM (a small amount) of oleate to that base line ROS signal. Oleate does nothing to ROS production via NOX2 but does generate its own ROS signal via RET, as per Protons. The increase is enough to go from insulin resistance levels of ROS to even more insulin resisting ROS, more insulin resistance and this allows more CPT1 production, as in the blue oval:

Bear in mind that there is no insulin. At all. These extra ROS will suppress insulin-like signalling but negative feedback limiting insulin signalling is unable to reduce the ROS because the ROS are not, in this case, from insulin.

But ultimately we can reduce the ROS signal by adding α-tocopherol.

Adding 0.1μM of α-tocopherol is just too little to do anything, we can imagine the blue oval moved one column to the right.

The addition of 1μM of α-tocopherol *should* reduce the ROS signal and allow sightly better insulin signalling which might reinstate a little of the suppression of CPT1 by lowering the ROS signal back closer toward to peak insulin-mimesis. In my imagination like this:

As more and more α-tocopherol is added the levels of ROS are stabilised at lower and lower levels until, somewhere between 25μM and 50μM of α-tocopherol, the added ROS from the oleic acid are neutralised and suppression of ROS now equates the control levels of insulin resisting ROS seen in the control cells.

But these control cells are still looking at 25mM of glucose and, at the high levels of ROS this will be generating, there is still more scope to improve the insulin-like signal towards more effective CPT1 suppression by reducing ROS further out of insulin resistance and toward insulin signalling levels. This happens with 20-50μM of α-tocopherol.

That's what I think is happening.

It's cell culture. It's as close to steady state as we can imagine. It almost certainly tells us something interesting about high levels of ROS at or above peak insulin levels and how manipulating these levels downwards has some effect on insulin signalling downstream of ROS generation.

Just before I take a break we can look at this in terms that I've viewed ROS mediated insulin mimesis before.

That's using the concept from

Evidence for Electron Transfer Reactions Involved in the Cu2+-dependent Thiol Activation of Fat Cell Glucose Utilization

and this well worn graph:

Looking at intact organisms I modified this to show an (arbitrary) upper limit to insulin derived ROS set by the negative feedback cells use to resist the action of real, actual insulin. It looked like this:

But in cell culture with glucose fixed at 25M there is no negative feedback and, though insulin signalling does still become obtunded, ROS generation can exceed that mediated by peak insulin and we end up with a graph very much like the one in Czech's seminal paper where ROS were added from a bottle of hydrogen peroxide:

In the current case we are achieving supra-peak ROS generation using NOX2 and 25mM glucose and then even more by adding to this level of ROS that from oleic acid a 1μM like this:

The in vitro data points are capable of reducing insulin-like signalling by raising ROS. We can then add in the likely ROS reductions mediated by α-tocopherol and see the progressive improvement in insulin signalling.

Which, of course, suppresses CPT1 protein production.

Just as a final "dig" at α-tocopherol 1μM, the two places where it could have acted to markedly reduce insulin signalling, via either high or low ROS, to facilitate CPT1 suppression are these:

I don't buy it.

To summarise: In cells generating stable insulin resisting levels of ROS α-tocopherol moves the ROS signal closer towards imitating an effective insulin-like signal. We can put the same information directly on to Czech's graph like this:

Of course, insulin stores fat.

If you store fat in liver cells you get fatty liver. Does this happen in vivo? When plasma levels are 163μmol/l, rather than the 50μM used in cell culture? Under dynamic insulin signalling conditions?

This not as simple as it seems. A live mouse is not a cell in culture with 4.0mM of added hydrogen peroxide out of a bottle or an added ROS signal from metabolic manipulation via culture medium, both of which can be reduced by a simple antioxidant. In-vivo there are limits to ROS generation and it's the movement of these limits which matters.

Peter

Insulin resistance (16) Yes. Vitamin E can cause weight loss

Part 3

It's time to set out a logical explanation for this graph:

taken from 

Ability of high fat diet to induce liver pathology correlates with the level of linoleic acid and Vitamin E in the diet

The first thing we have to do is to ask the correct question:

To me the correct question which has to be answered is why an high fat diet (35% of calories) with only 1% of calories from LA is obesogenic, at all. If you come from the point of view that the mechanism of obesity is down to LA alone, irrespective of macros, you need to be teasing this paper apart in great detail.

We know from

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

that a mouse eating an high fat diet based on D14521, if you choose your fat correctly, will not become obese at all provided LA is limited to 1.4% of energy intake. We can simplify Figure 1 from the paper:

to this:

This makes it abundantly clear that if you hold one variable constant, that is the percentage of energy supplied by linoleic acid, the nature of the rest of the fat in the diet is not insignificant.

Okay.

Here is the fat used in Graham's study, which is obesogenic with fat at 35% of calories:

and here is the non obesogenic diet with 45% of calories from fat in the D12451 study. I've combined the saturated fats together and expressed all as percent of total energy to allow easy comparison with the obesogenic diet from Graham's study:

They are identical in composition, the only difference is between 35% and 45% of energy from fat causing an absolute difference in overall calories from fat. A complete coincidence, but still very neat. Both studies look like they could have used the same jar of fat to make up the two diets. Except...

Those 27% of calories from saturated fat in Graham's study were largely from hydrogenated coconut oil. In total, hydrogenated coconut oil supplied 89.1g/kg of saturated fat to the diet.

We can ignore all fatty acids of C16 or greater as they are, as always, completely absorbed as chylomicrons through the thoracic duct and in to the systemic circulation, bypassing the liver completely.

For the shorter chain fatty acids I think we can ignore the myristic acid (~17% of the coconut oil) as it too is mostly carried in chylomicrons and treated much like palmitic acid. Only around 5% of it goes directly to the liver as FFAs.

The lauric acid which makes around 45% of coconut oil is partly packaged in to chylomicrons but something around 30% is delivered directly to the liver as FFAs.

Capric acid (C10) and caprylic (C8) (~15% combined in coconut oil) are completely transported directly to the liver as FFAs and do not enter the systemic circulation.

This medium chain triglyceride inclusion is the major difference between the 35% fat obesogenic diet used by Graham et al and the non obesogenic diet based on the stearate/palmitate/oleate mix in the cocoa butter D12451 study.

This matters.

To get some idea of what is happening we have to go somewhat in to reductio as absurdum and use pure MCT oil with a splash of soybean oil giving 50% of calories as fat in this study

A rich medium-chain triacylglycerol diet benefits adiposity but has adverse effects on the markers of hepatic lipogenesis and beta-oxidation

and look at this, Figure 1:

from which we can extract the relevant lines like this:

Here we can see that, on 4% of linoleic acid, MCT oil at 45% (no lard in this group) of energy intake is obesogenic when compared to 5% LA on a low fat background ie without the MCTs.

MCTs, in any significant amount, are obesogenic in their own right.

In this last study the obesity is made much worse by adding lard containing significant LA, that's what the faded-out lines on the unaltered graph show. I'll just add in the all-lard fed group as we need it when we look at pAKT levels:

So now we can look at the degree of insulin signalling present in hepatocytes at the time of euthanasia after a six hour fast. The full panel C from Figure two looks like this:

which we can simplify down to the interesting bits like this:

Clearly there is a marked reduction of insulin signalling in the lard fed mice. They are the most obese and are providing long chain fatty acids to the fasting liver, some LA but also palmitate and oleate. This is the normal physiological insulin resistance of fasting augmented by elevated basal lipolysis.

The lard-free highest MCT oil fed mice also demonstrate hepatic insulin resistance but far less than the lard fed mice. Which looks like a paradox.

The main problem with interpreting the pAKT signal is timing. The levels were measured in liver tissue after a six hour fast, which is quite a long time for a mouse. If you feed an MCT rich meal you would expect to flood hepatocytes with FFAs via the portal vein during the peak absorptive phase, probably around 1-2h post intake.

It is this flood of MCTs, which enter hepatocyte mitochondria with minimal restriction, which necessitates the resistance to insulin. Never forget that insulin resistance, at it's core, protects against the damaging levels of ROS generated by unrestrained elevation of delta psi. Insulin resistance automatically reduces insulin catabolism.

MCTs are not stored, so the excess lipid within the liver or exogenous lipid from the systemic circulation by six hours post intake will reflect the highly controlled oxidation of longer chain FFAs, not the un-restrained catabolism of MCTs at peak nutrient absorption. As the mice consuming MCTs are less obese than those on lard, it is reasonable for the level of insulin resistance to be lower because basal lipolysis is also lower.

In fact there are various influences on the high MCT fed hepatocytes pushing insulin sensitivity in conflicting directions, especially under fasting. But we can tease out what matters, and when, from a few extra studies. 

The Protons prediction from hepatic insulin resistance with associated reduced hepatic insulin extraction is the facilitation of elevated systemic plasma insulin, primarily at the time of peak delivery of MCTs to the liver via the portal vein circulation.

We can get an idea of whether this genuinely happens from this human intervention study here:

Thermic effect of medium-chain and long-chain triglycerides in man

which gives us this graph:

There is a small but significant rise in systemic insulin after MCT ingestion. It was triggered by 400kcal of MCT oil. You can enhance the effect in rats, who can't object to the GI distress caused, by giving them close to half a full day's calorie intake as a gavage. That's in this paper

Relation of ketosis to metabolic changes induced by acute medium-chain triglyceride feeding in rats

including this effect on systemic insulin

which we can tidy up like this:

It's also quite simple to ask how much insulin is extracted by the liver from the portal vein before being passed out through the hepatic vein and in to the general circulation. All you have to do is compare systemic insulin levels after an MCT bolus in a normal person with those generated by people with severe cirrhosis and multiple porto-systemic shunts, which by-pass whatever dysfunctional liver tissue they have remaining in-situ. That will be this study:

Stimulation of Insulin Secretion in Manby Medium Chain Triglycerides

After just 30ml (270kcal) of MCT oil we get this doubling in systemic insulin in cirrhotic patients during the time of peak MCT absorption, but the effect is gone by three hours post ingestion:

This effect is only present under MCT ingestion. Neither corn oil nor other LCFA containing fat sources do the same. My expectation is that, at peak MCT absorption times, flooding hepatocytes with MCTs will generate an high ROS signal, confined to those hepatocytes. This ROS generation can be dealt with by storing the acetyl-CoA as intra-hepatocyte LCFAs derived from acetyl-CoA (to be later exported as VLDLs), by off-loading acetyl-CoA indirectly as ketone bodies and, finally, by simply resisting insulin.

Which allows systemic hyperinsulinaemia.

The action of insulin is the inhibition of lipolysis.

Which is obesogenic.

The Surwit diet does this.

Here we have a fundamentally different form of obesity compared to linoleic acid induced obesity. In LA obesity the fundamental problem is at adipocyte level and here *inadequate* FADH2 driven ROS generation allows excess insulin signalling to distend those adipocytes.

In Surwit's hydrogenated coconut oil diet derived obesity, the fundamental problem is an *excess* of ROS from the MCTs in coconut oil, delivered in high levels to the liver only, allowing passage of insulin through the liver to give systemic hyperinsulinaemia which acts directly to cause simple obesity.

I suppose that the addition to this is that the lauric acid which reaches the systemic circulation might be a factor in the obesity, the shorter chain length gives poorer ROS generation compared to palmitate or stearate. 

MCT obesity is the only form of obesity which should be *reduced* by limiting ROS signalling.

Drop the ROS, restore hepatic insulin sensitivity, allow the liver to extract insulin, so shrink peripheral adipocytes via normalisation of peripheral insulin levels. *Hepatic* ROS scavenging does this:

and the converse is ROS scavenging does this:

Or, more relevantly

Or, with a minor change of one arrow's colour:

I'll leave it at that for the time being. There are, as far as I can find, no studies looking at the effect of vitamin E on Surwit-like diets. That's understandable. Why should vitamin E, considered to stabilise PUFA, have any effect on saturated fat induced obesity? So why bother looking at this?

But, given the ROS hypothesis of obesity, a plausible mechanism for the action of Surwit like diets is clear.

Peter

Insulin resistance (15) Vitamin E for weight loss? No

Part 2

Here is the next study.

α‐Tocopherol suppresses hepatic steatosis by increasing CPT‐1 expression in a mouse model of diet‐induced nonalcoholic fatty liver disease

From the methods:

"Experiment 1: Mice were divided into seven groups (n = 10 in each) and given the following diets for 8 weeks: standard diet (control group; 30% protein, 68% carbohydrate, and 12% fat including vitamin E acetate [500 IU/g]; Research Diet); HF diet (HF group; 20% protein, 20% carbohydrate, and 60% fat including vitamin E acetate [500 IU/g]; Research Diet) and HF diet with α‐tocopherol (α‐Toc) which is one of the natural vitamin E forms supplementation (20, 50, 100, 150, and 200 mg/kg)."

Okay, a ghastly typo.

We have no idea which of the Research Diets these mice were fed on. I am going to assume that the chow resembles PicoLab Rodent Diet 20/LabDiet 5053 and contains, as per my last post, ~100iu/kg of synthetic vitamin E acetate yielding, also as per last post, 44.5mg/kg of active vitamin E in the food.

The typo is to state that this chow contains 500iu/g. That's quite a lot of vitamin E. The correct amount is (almost certainly) 100iu/kg, not 500,000iu/kg.

I feel it is reasonable to assume the high fat diet was something similar to, or in fact was, D12942 which also probably contains around 100iu/kg vitamin E. Obviously the mice would eat less weight of D12942 than chow because they eat to caloric need. This will be met by a lower weight of D12942 so their intake of d-α-tocopherol would also be a little lower than if they ate the chow. More like 40mg/kg if you taken in to account the reduced weight of food eaten.

I am also going to assume that they added their supplementary vitamin E to this standard high fat diet so we're looking at intakes based on diets containing totals of 60mg/kg to 240mg/kg of d-α

-tocopherol per kilogram.

Anyhoo. For the time being I'm going to ignore the changes in everything other than total body weight.

There are no data presented for the effects of most of the supplement levels used, though these were recorded as per the methods. I think it's safe to assume that the effect on weight was consistent across all vitamin E intakes used, otherwise they would have mentioned it. This is what they actually presented:

There are, undoubtedly, effects from vitamin E supplementation on parameters other than total body weight. People may find the liver damage induced by high dose vitamin E fascinating. I do. But that's another story.

So I think we can say that, in a poorly described study, vitamin E supplementation has absolutely no effect on the body weight of mice over eight weeks of feeding an high fat diet. Over a wide range of dose rates.

To continue the catalogue of appalling vitamin E focused studies, it's now time to look at this one:

Effects of d-α-tocopherol supplements on lipid metabolism in a high-fat diet-fed animal model

How bad is it? It's this bad:

"After the adaptation period, the mice were randomly divided into three groups. Nine mice were placed in the control group [CON, regular diet (10% of calories derived from corn oil) and distilled water as a vehicle (0.1 ml, p.o.)]. Another set of nine mice were placed in the high-fat group [HF, high-fat diet (45% of calories derived from lard) with distilled water as a vehicle (0.1 ml, p.o.)], while the rest of the mice were placed in the high-fat diet with daily oral administration of 100 IU/kg B.W. of d-α-tocopherol group [HF-E, high-fat diet (45% of calories derived from lard)]."

So we know nothing about anything. We have no idea of the vitamin E levels of the control chow or of the high fat diet. We don't even know if the high fat diet was manufactured specifically or whether they just added lard to chow to make 45% of calories from the lard which diluted the chow's vitamin E. We don't know what the lard was composed of in terms of LA either. Or even if it was Japanese or from the USA. We *do* know it was high enough in LA to make the mice fat.

None of this matters too much because the vitamin E supplementation was given by oral gavage of 100iu/kg once daily. This was pure d-α-tocopherol so the arithmetic is easy. The 100iu gavage provided 67mg of active d-α-tocopherol, not the racaemic mix and not the acetate ester. The mice weighed 32g so each got ~2mg/d by the end of the study.

If we reverse engineer to translate this in to how much vitamin E would need to be added to food to deliver that same dose we can do this. We can say that a mouse eats ~2.8g/d of high fat diet. So there would need to have been 2mg in 2g of food or 1000mg/kg of food. Though if you used the synthetic acetate ester then around twice that. This is a massive dose of vitamin E and guess what effect it had on body weight? You're waaay ahead of me:

We might also take note of the caloric intake per day which comes out as exactly what anyone would expect for mice on an high fat diet, in contrast to the last post. The current study calorie intake is picked out in blue:

I hope you're not getting too bored with this. I suffered for weeks with these studies. Now it's your turn. 

I guess I'm not selling you vitamin E as a weight loss hack. That's good.

So how do we square these studies (and many others, it's been a rough three weeks of reading) with the results from

Ability of high fat diet to induce liver pathology correlates with the level of linoleic acid and Vitamin E in the diet

where there is a marked decrease in weight gain on an high fat diet with modest vitamin E supplementation?

I absolutely accept that these data are correct as reported.

These are the sorts of findings which test your hypothesis of obesity. It's what makes slogging through the typos and brain farts and shifting definitions of high vs low vs unspecified levels of vitamin E in diet trials worthwhile.

Ultimately we are looking for circumstances where reducing ROS with vitamin E allows weight loss. Some weight loss anyway.

I think there might be an explanation.

Peter