Insulin: Are you hungry? Part 2

Well, over the years I have made the occasional serious blooper on Hyperlipid.

Perhaps the worst of these, to my intense shame, is the acceptance of insulin as a satiety hormone. This is complete bollocks and, thankfully, some deleted-expletive person in obesity research has finally opened my eyes to this. The gift was from Dr Guyenet of course. This is how he convinced me that insulin is not a satiety hormone:


Let's feed some rats standard crapinabag and inject one group with nothing much, one with glargine insulin and another with detemir insulin. But here's the trick. Because we know that hypoglycaemia triggers overeating and the overeating causes weight gain, let's limit the insulin dose to one which does not cause hypoglycaemia... No overeating, active satiety hormone, weight loss...

Because we have been (mis)informed that insulin is a satiety hormone we would expect the insulin-injected rats should eat less, weigh less blah blah blah. What really happens? This does:



I've seen this paper cited as showing insulin can reduce weight gain. By Dr Guyenet no less. Who didn't mention the graphs. Which are core to the paper.

Technically this shows that insulin does bugger all to food intake and fat storage. This is hardly surprising as giving a sub hypoglycaemic dose of insulin will simply attempt to lower blood glucose which will be avoided by reduction of endogenously produced insulin. Total insulin will stay the same. There will be subtleties of peripheral administration vs portal secretion but I guess these are a bit too subtle for this study. There are also fascinating differences in duration of binding of detemir insulin to the insulin receptor vs other insulins. Not surprising as it has a socking great fatty acid tagged on one end but that's another set of stories.

Ah, but what about the effect of detemir insulin on limiting fat gain of rats fed toffeefudgecheescake, aka D12492?

We are talking Fig 3 parts b and e here:



Okay, we're now utterly convinced that insulin limits weight gain. Well, detemir insulin does. Of course glargine insulin doesn't, as Dr Guyenet forgot to mention when citing this paper. It produces a non significant increase in weight over vehicle treated controls.

Just for a giggle, consider changing the grams to kgs on graph e and imagine these rats as humans. Given a group size of 6-8 leading to statistically ns changes in fat mass, would you consider 5kg fat mass gain (a ns change) on glargine, without eating any extra, non significant? Biologically? In dress size? Tee hee.

Now let's look at section e in a little more detail.

The study is very, very carefully set up. The insulin and the D12492 were both started on the same day. It is utterly convincing (to me) that detemir insulin limits weight gain IN THE FIRST 7-10 DAYS of D12492 feeding. From day 10 onwards the fat mass does not change in the control, the detemir or the glargine groups. Not even a trend. A bit like the crapinabag groups demonstrated throughout. In fact, identical to the crapinabag groups. Where's your satiety Guyenet?

Now here's a thought experiment. Let's pretend that all rats were fed D12492 from day 0 to day 10 without injected insulin, so became equally obese with a fat mass of 65 grams, same as the controls on day 10. From day 10 onwards all groups then received their respective insulin or vehicle for four weeks.

Would the fat mass have changed from the 65g starting weight? Of course not, look at the last weeks on graph e. These people are not stupid, though they do like to give that impression.

From this study the follow on question has to be: What is the difference between detemir insulin and either endogenous insulin or glargine insulin during the first 7 days of feeding D12492 to rats?

We all recall from the paper from the Schwartz lab featuring the world's greatest mis-citation expert, that the first few days of sucrose/fat feeding produces an acute inflammatory lesion in the hypothalamus of rats which get fat on D12492. If I had to guess I would suggest detemir insulin limits this injury. How and why cannot be guessed at from this paper but needless to say groups working with gold thioglucose injury have considered what factors influenced hypothalamic injuries. That leads to far out speculation, so I'll limit this post to what Guyenet's citations really do show.

They show that physiological insulin does NOT suppress appetite. Are you surprised? Me neither.

Of course an increased dose of insulin might suppress appetite. But this would need a glucose infusion to maintain life, which would promote DNL in adipocytes and inhibit lipolysis. No hunger while you gain fat. You have to wonder what the point of the above study was, excepting it supports a grant maintaining position and is a self justification for a bizarre mindset.

I also notice Guyenet re-cited this crap. Doesn't he read Hyperlipid????? Giggle... That was a rhetorical question!

Less rhetorical is to ask whether he has actually read the Vanderweele paper at all, particularly Fig 4 of the paper and whether he has reverse engineered said Fig 4 to see the problems with the conclusions of the paper!

Finally he has cited a drug study using an insulin mimetic, not insulin. Well, bully for insulin mimetics. With an insulin mimetic you can mimic lethal doses of insulin without all that inconvenient death. The body does not produce lethal doses of insulin under physiological conditions. If you want to know about physiological doses of insulin within the CNS I can just quote this paper. I feel the authors are being just a teensy weensy bit over the top in their deprecatory attitude to the "centralinsulinisasatietyhormone" brigade. But I can understand why! Here's my fav quote:

"To reduce the likelihood of pharmacological effects of the insulin doses administered, we choose a dose of insulin that is more than 15,000–fold lower than those commonly used for ICV [third ventricle, CSF] insulin infusions"

That's about as rude as you get in Cell Metabolism! You can't use the word "pillock". Drug doses (pharmacologic) of insulin produce drug effects. If you give only physiological dose rates you get physiological effects! Now isn't that amazing?

Oh btw, at physiological levels brain insulin increases peripheral lipogenesis and decreases lipolysis. Did you think insulin would do the opposite through the brain compared to what it does in the periphery?

Duh.

A more believable scenario is that ATP generation within the brain using glucose metabolism, facilitated by insulin in those areas responsible for energy sensing, does occur. But this combination of glucose and insulin will also store fat, as it should, when it occurs post prandially. Which is exactly what excess energy sensing should signal. Insulin without the glucose is pharmacology, unless you suffer from reactive hypoglycaemia.

Peter

BTW I notice over on Woo's blog that there has been some discussion as to whether Dr Guyenet is just dumb or being very deliberately misleading, ie conspiring to mislead. I don't do orchestrated conspiracy theories. I don't really do the financial drive thing either, not for some body who is still as wet behind the ears as Dr Guyenet certainly is. No, for a junior post-doc it has to be:

He has the whole of the knowledge base of the Schwartz lab at his beck and call and the above three citations are the best dross that the Good Doctor can come up with... But still he believes! Stupid.

Happy Solstice

Well Happy Solstice, a day late, to all!

Bit of a bummer yesterday with working a late shift and needing a mega shop for the food for over the next week or so. After my bed time before the car was unloaded!

But I don't think we will run out of cream or butter. Phew!

All the best to all

Peter

Some of us eat a high fat diet

Just a brief post while I try to think of a simple descriptor for The Goof Doctor. Oops, typo there, I think I'll leave it. On to happier subjects:

A birthday card from a few weeks ago to my wife from her best friend Karla. They both understand...



(Card from here http://www.corrinarothwell.co.uk)

However, some people don't need help with butter, except getting it on to the spoon if it has just been self-served from the fridge and is rock solid!

Peter

More yawns on insulin and knockout mice

Ok, I've just got 10 minutes to spare. What shall we play?

Last Saturday was the pathology lab's Christmas Party. In my cracker I was graced by a small and extremely nasty quality plastic magnifying glass which, sadly, I discarded. It might have come in useful in searching for any intellectual understanding in the Good Doctor's latest demolition of the role of insulin in obesity. Actually, the loss of the hand lens is no real problem as I doubt whether even an electron microscope would allow us to find something which is not there.

Let's begin with TNFalpha knock out mice. First thing is that these are based on C57BL/6 mice and, as I have discussed on many occasions, these mice develop an hypothalamic injury which decreases sympathetic outflow to adipocytes and so increases their ability to store fat. You need some technique to make fat mice fat and C57BL/6 are your candidate critters. They are really useful in that they lose fat in to adipocytes, just don't imagine they have anything to do with human obesity. But as a tool, they're great.

Here are the diagrams we need, taken straight from the Good Dr's blog:



All very clear cut.

But what does it mean? Fasting insulin goes up when adipocytes become insulin resistant and so leak FFAs, especially palmitic and stearic acids. Adipocytes become insulin resistant when they become over stuffed with lipid. They don't want any more fat, so they refuse to respond to insulin. Remember that the role of insulin is the storage of dietary fat, plus a little DNL if dietary fat is very low. If adipocytes don't want to respond to insulin they will signal to the rest of the body to do the same. That's working on the basis that adipocytes control whole body insulin resistance based on the ratio of palmitic acid to palmitoleic acid they release, which is in turn dependent on their own insulin sensitivity and SCD1 activity.

Now I have yet to delve in to the mechanics of adipocyte distention induced insulin resistance but I can tell you something here and now for free. It involves the TNFalpha. If you knock out TNFalpha your adipocytes (and pretty much the rest of your body) cannot become insulin resistant. No one gets fat due to insulin failing to act. You become fat due to the action of insulin on adipocytes. When adipocytes refuse to listen to insulin you stop getting fatter, but become hyperglycaemic (unless you eat LC of course!).

So the difference between wild type high fat fed mice (plus sucrose of course, the "cookie dough" they "can't get enough of") and TNFa-/- HFD fed mice is that the wild type mice are sending the signal to the rest of the body that they are fat enough and would like to stop accepting any more calories, fat or glucose. A combination of hypothalamic injury, a sucrose rich diet and a pancreas of steel makes these wild type C57BL/6 mice continue to become obese because they need massive levels of insulin to maintain normoglycaemia. All because insulin resistant adipocytes are signalling that insulin resistance should be produced in insulin controlled cells throughout the body.

Summary: Insulin sensitive adipocytes distend in response to insulin. Insulin resistant adipocytes don't. Hyperglycaemia needs to be corrected. The large dose of insulin needed for this continues to drive obesity by force-enlarging insulin resistant adipocytes. With TNFalpha knocked out the adipocytes become "effortlessly" obese. They, and the rest of the body, will stay insulin sensitive and there will be "easily achieved" normoglycaemia. Healthy obese, but still very obese. Obese due to modest insulin acting on very insulin sensitive adipocytes.


Now, on to iNOS knock out mice. These are really interesting. Again, they are based on C57BL/6 freak mice. A useful model for basic physiology, so long as you have some concept of what is wrong with them. These mice have also had inducible nitric oxide synthetase knocked out ONLY from their muscles. Adipocytes are normal, or as normal as any C57BL/6 mouse can be. Here are the weights etc from the results:



Just look at those gluttonous food intakes and fat gains! Wow.

We can get a basic handle of what is going on from this set of graphs:



Graph a is worthy of the Good Doctor.  HFD fed Nos2-/- mice are not only enormously obese, but they manage it on a fasting insulin which is LOWER (admittedly not significantly so) than that of the WT mice fed crapinabag! Blooooodie hell!

I could spend days discussing graph b, but I'm already half way through my 10  minutes so let's leave it, fascinating though it is...

Graph c. This is the pay dirt. Here they injected insulin in to the various WT and KO mice and tracked the fall in blood glucose levels. The ruler-drawn straight line of black squares is the HFD Nos2-/- mice. These massively obese mice are the MOST insulin sensitive, whole body, of all groups, certainly at the 60 minute mark.

Their adipocytes aren't. These are insulin resistant. All that remains sensitive to insulin are the targeted knock out muscles. On insulin injection these mice simply pour glucose in to their Nos2-/- muscles because the muscles ignore the signals from the insulin resistant adipocytes. Their muscle cells are like a black hole in to which glucose pours. Now, at the risk of quoting the Good Dr yet again: Hypoglycaemia is a very, very potent driver of hunger. Eat, or die.

What's on the menu? Ah, a bowl of lard sweetened with sugar. Death is not an option, let's eat the lard to get the sugar. Good idea, stayin' alive. Now, we've used the sugar, what shall we do with the lard? Ah, as a brain injured C57BL/6 mouse we have adipocytes which are rather more willing to accept fat than a genuine wild type mouse might have. Bye bye fat, in to the adipocytes you go! But at least death due to hypoglycaemia is avoided, all be it at the cost of greater obesity!

If you have been following the protons thread you can see that linoleic acid, ie corn oil, is a mild mimic of TNFa-/- mice and of Nos2-/- mice. It's obesogenic while preserving insulin sensitivity. Your cardiologist made you fat.

Now, in my last 30 seconds: Why are adipocyte insulin receptor knock out (FIRKO) mice healthy and slim? Well you could ask the Good Dr for some sort of platitude, but, hey, that would be stupid.

No. Adipocytes control whole body insulin sensitivity. They see no insulin if they have had their insulin receptors knocked out. They sport minimal (zero?) GLUT4s on their surface. What fat they contain has been accumulated without the assistance of insulin. I think it is reasonable to assume they have some GLUT1s on their surface. Any glucose taken up will be available for lipid synthesis but, without insulin's action, there will be no insulin induced SCD1 desaturase activity. So palmitate it is and, in the absence of insulin's action, this will be freely released and should signal whole body insulin resistance. But it doesn't. It does exactly the same as the palmitic acid does in SCD1 knockout mice. Peroxisiomes. FIRKO mice eat more, weigh less and (probably) generate more heat than WT mice do. They are insulin sensitive everywhere except for their adipocytes. They behave exactly as SCD1-/- mice do but get there by a rather indirect route. Excess palmitate is burned in peroxisomes and the C8 end product in mitochondria.

Life is, in the end, logical. Having the correct tools helps. It must be awful to be wallowing in the mire of the Reward hypothesis.

Peter