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Thursday, 21 March 2013

Naltrexone has TLR-4 activity - is it an "Old Friends" mimic?

(Our first song for today is appropriately Junk by The Puddle, a early post on this topic from 1985.)


Naltrexone is a mu-endorphin antgonist that is used as an opioid antagonist. It will reverse intoxication from opioid analgesics and has a longer duration of action than naloxone. Naloxone is preferred in emergency settings because of its faster onset of action, but naltrexone, unlike naloxone, is effective given by mouth, and its long duration of action (10 hrs) makes it a safer treatment for overdose with long-acting opioids such as methadone (24 hrs, but the half-life of methadone can be significantly decreased by urinary acidifiers such as ascorbic acid or citric acid, and is increased by urinary alkalizers such as sodium bicarbonate as methadone has low solubility at high urinary pH). Naltrexone's most common clinical use is in doses of 50mg daily as a maintenance treatment to blockade the addictive effects of both opiates and alcohol. Recently I spoke to an AOD patient using naltrexone in this way and was surprised that they reported that the drug had an anti-depressant effect. This isn't what would we'd expect under the old endorphin receptor model from a full-time blockade; dysphoria would be the predicted side-effect.


 

A clue lies in the popular off-label use of naltrexone at low doses (3-4.5 mg nocte) as an immune-modulating agent. The Low Dose Naltrexone homepage claims
"FDA-approved naltrexone, in a low dose, can normalize the immune system — helping those with HIV/AIDS, cancer, autoimmune diseases, and central nervous system disorders."
This is not quackery; read the NOLA Hepper blog in the blogroll to your right for a case study of LDN in the treatment of hepatitis C. Naltrexone is some potent juju, but the results have been hard to explain till recently. The mechanism that was originally stated is as followed; endorphin synthesis is highest at night. Blocking receptors for a few hours while the patient sleeps has the effect of elevating endorphin levels; these elevated endorphins have mood and immune-modulating effects. Hormesis in other words.
"The brief blockade of opioid receptors between 2 a.m. and 4 a.m. that is caused by taking LDN at bedtime each night is believed to produce a prolonged up-regulation of vital elements of the immune system by causing an increase in endorphin and enkephalin production. Normal volunteers who have taken LDN in this fashion have been found to have much higher levels of beta-endorphins circulating in their blood in the following days. Animal research by I. Zagon PhD. and his colleagues has shown a marked increase in metenkephalin levels as well."
The endorphin effects of naltrexone are significant of course, but the much higher dose taken by the patient I spoke to could not work in this way; also, both opioid drugs like morphine, and morphine withdrawal states have been found to stimulate HCV replication in vitro.
Recently (c. 2007) studies started to appear showing that opioids and opoid antagonists have TLR-4 activity. TLR-4 is a PAMP (pathogen associated molecular pattern) receptor. TLRs tell  immune cells what type of pathogens they may (or may not) need to get agitated about. In the case of TLR-4 this is gram-negative bacteria, and the ligand is lipopolysaccharide (LPS).
Except that pathogens aren't the only organisms to produce TLR-4 ligands; probiotics, saprophytes and other benign or beneficial bacteria can also have TLR-4 activity, and this exposure is associated with lower rates of auto-immune disease, allergies, depression, cancer and great deal else besides through the "Hygiene hypothesis" or "Old Friends hypothesis" (there are some good articles on this at Emily Deans' Evolutionary Psychiatry blog linked in my blogroll on the right hand side of the page).
The effects we would expect to see from TLR-4 activation by what I might term FAMPs (friend associated molecular patterns) if the Old Friends hypothesis is correct are the same ones attributed to LDN (which increases TLR-4 expression). The illnesses LDN is claimed to treat are all diseases that have come into prominence in the wake of microbiotal extinctions. In many cases they barely seem to have existed prior to the widespread use of antibiotics, disinfectants, and antithelmetics.
2012 Mar;26(3):480-8. doi: 10.1016/j.bbi.2011.12.010. Epub 2012 Jan 5.

Mu opioid receptor activation modulates Toll like receptor 4 in murine macrophages.

Source

Dipartimento di Farmacologia Chemioterapia e Tossicologia medica, Università degli Studi di Milano, Via Vanvitelli 32, 20129 Milano, Italy.

Abstract

Opioids have been shown to affect both innate and adaptive immunity. We previously showed that morphine affects the macrophage production of pro-inflammatory cytokines after LPS in a NFkB dependent manner.
Toll like receptors (TLRs) play a crucial role in the signaling pathways which lead to NFkB activation. TLR4 is considered the Lipopolysaccaride (LPS) receptor.
The data here presented show that, in murine macrophages, morphine impacts on the immune function acting on the early step of pathogen recognition. Morphine, when added to RAW 264.7 cells and when injected into mice (s.c. 20mg/kg) is in fact able to decrease TLR4 both at mRNA and protein level in RAW cells and peritoneal macrophages. In the same cells, the mu opioid receptor (MOR) antagonist Naltrexone increases TLR4 levels, thus suggesting a role of the endogenous opioid system in TLR4 regulation. The effect of the two drugs is moreover lost in case of co-administration.
Experiments with MOR KO mice and with DAMGO (MOR specific agonist) confirm that the effect of morphine on TLR4 mRNA in peritoneal macrophages is due to the MOR activation. Moreover the effect on TLR4 is blocked by PTX thus indicating the involvement of a G(i) protein after MOR binding. This work unveils a clear link between MOR activation and TLR4, suggesting a new possible mechanism at the basis of the peripheral immunosuppressive effect of opioids.

Make no mistake, this is edge-of-your seat scientific stuff that for my money is predictive of rapid medical progress in unexpected directions. For example: Recent evidence implicates toll-like receptor 4 (TLR4) in opioid analgesia, tolerance, conditioned place preference, and self-administration. In other words, some of the anti-addictive effect of naltrexone blockade is due to its TLR-4 agonism, not its mu-endorphin antagonism (though the two are probably indirectly connected). One obvious implication of this research is that the dose range for both LDN and addiction treatment should probably be much wider. Some people will need larger doses or more frequent dosing for LDN to work, or even smaller doses; VLDN, (0.125-0.250mg) treatment has been used to suport methadone withdrawal; while AOD patients who do not tolerate 50mg naltrexone may well benefit from lower doses. The dose thresholds for endorphin and TLR-4 effects are likely to be different and to vary between indviduals.
Exciting - nay, exhilarating. Sometimes you just gotta love science. It's one of those gifts that just keeps on giving.
 

(more music - Steely Dan "My Old School")

23 comments:

Peter said...

Great post. I have long wondered whether an iv dose of naloxone would give 10 minutes of deep depression followed by 24h upregulated happiness. The reverese of a bolus of iv alfentanil.... That's before TLRs put in an appearance.

Peter

George Henderson said...

Unfortunately everyone I know who tried that experiment was overdosing at the time and expressed mostly ingratitude for the results.

Look at the pictures of the healed Crohn's Colitis intestine on the LDN homepage - isn't that what you'd expect after a successful fecal transplant, or a lucky dose of hookworms?

Morphine after cancer surgery significantly reduces the chances of remission (tramadol is safer) according to oncology papers on the LEF site.

Coley's Toxins certainly had TLR-4 activity. Any anti-cancer effects of LDN are more likely to be due to TLR-4. LDN or VLDN could perhaps be used to block immunosuppressive effects of morphine without compromising analgesia...

This discovery pulls together all sorts of scattered findings into a more coherent picture. It's like the Higgs Boson of medicine.

Bill said...

Lol, the boson.
I've been following the LDN story but felt blindsided after reading your article! This is a crazy & unexpected turn of events - kudos to the researchers involved.
best,
Bill

Peter said...

As an anaesthetist I find it depressing how virtually every drug in my armoury is a bummer to the immune system. Maybe I need some naloxone to cheer me up. I have also wondered whether the salvia active ingredient, which looks to be a kappa agonist, mu antagonist (last time I went reading), might do the same. Trip is short, it may well be blocking mu receptors, and it is reported to be antidepressant on long term use....

Peter

George Henderson said...

Bill, it's amazing how often the really good research comes out of Italy.
You might not want them in your army or your government, but if you're staffing a lab, I'd go for Italians every time.

Peter - adjuvants for anaesthesia then?
From what I've read it looks like naltrexone can be co-administered with opioids without losing analgesia; it's only in pre-existing dependence that it becomes tricky.
For example:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2862024/
The development of analgesic tolerance following chronic morphine administration can be a significant clinical problem. Preclinical studies demonstrate that chronic morphine administration induces spinal gliosis and that inhibition of gliosis prevents the development of analgesic tolerance to opioids. Many studies have also demonstrated that ultra-low doses of naltrexone inhibit the development of spinal morphine antinociceptive tolerance and clinical studies demonstrate that it has opioid sparing effects. In this study we demonstrate that ultra-low dose naltrexone attenuates glial activation, which may contribute to its effects on attenuating tolerance.

http://europepmc.org/abstract/MED/9200746
Ultra-low doses of naltrexone or etorphine increase morphine's antinociceptive potency and attenuate tolerance/dependence in mice.

http://www.ncbi.nlm.nih.gov/pubmed/19443938
The findings of the present work suggest that the combination of opioids and NSAIDs has a direct action on spinal nociceptive processing, which may be achieved via mechanisms that are independent of the activation of opioid receptors. The ineffectiveness of naltrexone to reverse the analgesic activity of opioids + NSAIDs combinations indicates that other complex pain regulatory systems are involved in this effect.

http://www.sciencedirect.com/science/article/pii/S0165614709001369
The “Toll” of Opioid-Induced Glial Activation: Improving the Clinical Efficacy of Opioids by Targeting Glia

In farmers' children with low incidence of allergies (hay fever and asthma), there is 3x higher expression of TLR2 (presumed due to "old friends" exposure). Naltrexone increases expression of TLR4.

Galina L. said...

" how virtually every drug in my armoury is a bummer to the immune system"
Even on a LC diet I have from time to time(way less frequently than before) to take ergots or opiates for my migraine. Every med for migraine did it for me. Every time it gets my asthma to resurface, even though LCarbing took away the need to refill the prescription for inhalers.

George Henderson said...

When I used opiates my hay fever disappeared, it came back as I started withdrawing and was really bad for a while. Many of the more trivial opiate withdrawal symptoms resemble allergy symptoms - yawning, sneezing, runny eyes and nose.

It occurs to me there's a duality here: according to wiki naltrexone is the TLR4 antagonist, morphine the agonist, but naltrexone increases expression of TLR4 and morphine suppresses it.
in the asthma epidemiology exposure to "old friends" was associated with increased expression of TLR2.
How is an increased number of receptors linked to a milder response to their agonist ligands?
Or is it only the macrophages that get increased TLRs via exposure to the antagonists?

George Henderson said...

The Circadian Clock Controls Toll-like Receptor 9-Mediated Innate and Adaptive Immunity
http://www.cell.com/immunity/abstract/S1074-7613(12)00047-7

Circadian rhythms refer to biologic processes that oscillate with a period of ∼24 hr. These rhythms are sustained by a molecular clock and provide a temporal matrix that ensures the coordination of homeostatic processes with the periodicity of environmental challenges. We demonstrate the circadian molecular clock controls the expression and function of Toll-like receptor 9 (TLR9). In a vaccination model using TLR9 ligand as adjuvant, mice immunized at the time of enhanced TLR9 responsiveness presented weeks later with an improved adaptive immune response. In a TLR9-dependent mouse model of sepsis, we found that disease severity was dependent on the timing of sepsis induction, coinciding with the daily changes in TLR9 expression and function. These findings unveil a direct molecular link between the circadian and innate immune systems with important implications for immunoprophylaxis and immunotherapy.

Peter said...

George,

Increased receptor numbers gives a lower occupancy % at a given drug concentration. That seems to fit.

BTW, just from lurking on poppy.org I recall a poppy tea user commenting that effective opioid usage, ie with slowly increasing dose over the months, led to the complete absence of any cold or flu like symptoms whatever the challenge. Until the time came to deal with the gorilla of course.

Our standard analgesic combo is virtually always an NSAID along side the opioid, but we are not performing the sort of surgery where opioids beyond 3 days are needed. Of course the pancreatitis cases often need extended opioids but we get them off of pure agonists and on to bupe as early as they seem comfortable enough.

The inclusion of etorphine in the same category as naltrexone is weird, I always thought of it (and used it in my patients) as an ultra potent mu agonist...

And gliosis = free radicals to me. I have also long thought of addiction as possibly being mediated by the balance of energy demand by a cell VS oxidative phosphorylation efficiency in said cell. Big demand for ATP might well spew free radicals, especially if running through complex I AND glycerol-3-dehydrogenase. Dunno. Needs a lot of thinking about.

Peter

George Henderson said...

Very low dose etorphine - surely that would a homeopathic concentration, measured in nanograms.

Morphine down-regulates glutathione synthesis; more ROS to NF-kappa-beta.

http://www.ncbi.nlm.nih.gov/pubmed/2820428

Injection of morphine causes an acute increase in liver enzymes and a decrease in hepatic glutathione (GSH) synthesis.12 Recent studies have shown that the opioid antagonist naltrexone decreases liver injury in rats and mice with acute biliary obstruction.

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1603777/

So not just gliosis but also hepatotoxicity could be reduced by co-administering LDN.
There was a case in N.Z. where a boy died of liver failure after overdosing on methadone. The "experts" testified that "methadone does not cause liver failure" so the coroner ruled natural causes. Just a co-incidence that his previously unexceptionably functioning liver packed in at that particular time apparently.

George Henderson said...

I think of the mitochondria in addiction as running like pressure cookers under the weight of drug interference with the electron chain and membrane potential - when this is lifted the cells boil.
My experience of withdrawal, and the fact it could be mitigated by a high-fat diet, reminded me of William Burrough's idea that addiction is a hunger of every cell in the body.
It's not just felt in the CNS, but every cell has adapted to the formerly cytotoxic dose, till its removal is even more cytotoxic.

George Henderson said...

This 2007 pdf seems to be the earliest paper (from the ever-relevant Hindawi publishing group and a journal I've never heard of called "The Scientific World") to put the full opioid-TLR4-gliosis hypothesis together; it mentions etorphine as an exception to the rule that mu-endorphin receptor agonists cause gliosis (ref 51, but this doesn't seem to be online).
downloads.hindawi.com/journals/tswj/2007/746941.pdf

George Henderson said...

Lovely:

Bacterial peptidoglycan enhances sickness behaviour induced by
bacterial lipopolysaccharide
(PDF)
http://tinyurl.com/d4weoyv

Background: Lipopolysaccharide (LPS) and peptidoglycan are microbial
products recognized by Toll-like receptor-4 (TLR4) and nucleotide-binding
oligomerization domain 1 (NOD1) and NOD2, respectively.

*LPS has been
found to cause behavioural alterations indicative of sickness and
depressed mood.*

The effect of peptidoglycan on exploratory and affective
behaviour has not yet been explored, although it has been reported that it
promotes sleep and anorexia. Since NOD1 and NOD2 are activated by two
different peptidoglycan components, MurNAc-L-Ala-g-D-Glu-mesodiaminopimelic
acid (M-TriDAP) and muramyl dipeptide (MDP),
respectively, the effects of both compounds, alone and in combination
with LPS, on exploratory and affective behaviour were investigated.

I can't wait till someone tests ibogaine for TLR4 activity.

George Henderson said...

I wonder if it's all relevant here that the only non-opioid intervention that's ever relieved my full-blown withdrawal symptoms has been high-dose I.V. ascorbate.

Here's a small connection:
http://www.ncbi.nlm.nih.gov/pubmed/12885590?dopt=Abstract

In conclusion, strenuous short-term aerobic exercise results in significant increases in plasma LPS levels (endotoxemia) together with increases in markers of oxidative stress. Supplementation with ascorbic acid, however, abolished the increase in LPS and nitrite but led to a significant increase in the ascorbate radical in plasma.

This is an interesting view of LPS considered as a kind of hormone: it looks like a good background resource for understanding what TLR4 does in the body (at least in regard to LPS; it has other interesting roles).
http://cid.oxfordjournals.org/content/41/Supplement_7/S470.full

Peter said...

I've been looking on my hard drive for the "high fat meal increases LPS in the bloodstream" paper, part of fat bashing. The one paper I did find suggested that chylomicrons (and LDL) deliver LPS to hepatocytes rather than macrophages in the liver. My assumption was that low fat meals delivered LPS directly to the liver w/o the detox through chylomicrons and might just ramp up inflammation if it was delivered to macrophages. Measuring LPS in the portal vein of human volunteers is not quite as straightforward as looking in peripheral blood... Can't find the paper, argh.

George Henderson said...

It's not the paper you linked to here?
http://high-fat-nutrition.blogspot.co.nz/2009/02/fats-absorbing-endotoxin.html
I'm in MacDonalds with a kids computer so I don't have time to follow the links, but if in doubt about this stuff I go to http://www.lucastafur.com/2012/07/nutritional-immunotherapy-dietary-fatty.html which I can see I need to re-read in light of the naltrexone-TLR4-HCV situation.
Andrew Kim has also discussed these sorts of papers recently, for example http://www.andrewkimblog.com/2012/12/lipopolysaccharide-physical.html
in a breathless but often productive-of-ideas Peatitarian context.

George Henderson said...

From Lucas Tafur's blog:

Hypertriglyceridemia increases endotoxemia

The human body must be able to cope with acute increases in LPS in plasma, attenuating the inflammatory response induced by fat ingestion. Clemente-Postigo, et al. (20) showed that in morbidly obese subjects, endotoxin increases were strongly correlated to the difference between baseline and postprandial triglyceride levels. They also found that baseline triglyceride level was the best variable that predicted basal LPS level in serum. In this regard, very low carbohydrate diets have shown to reduce baseline triglyceride levels and postprandial lipemia (21, 22). In the metabolically healthy, the immune system is capable of attenuating postprandial endotoxemia (as with inflammation induced by any meal). The inflammatory nature of absorption, digestion and metabolism of macronutrients must be coupled with an anti-inflammatory period, such as fasting (depending on the inflammatory load of the diet, an overnight fast might work). By this way, the body's inflammatory balance is maintained in a healthy range.

Lou Thomas said...
This comment has been removed by a blog administrator.
Lou Thomas said...

The cited article suggests that "the mu opioid receptor (MOR) antagonist Naltrexone increases TLR4 levels, thus suggesting a role of the endogenous opioid system in TLR4 regulation." I had been under the impression that the blockade of TLR4 by LDN was a separate effect from its blockade of opioid receptors. For example this study states that "(+)-naltrexone, (+)-naloxone, and (-))-naloxone...we show here to be TLR4 antagonists in vitro on...microglial cell lines."

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2588470/

George Henderson said...

I think you might be right, because this effect is seen consistently with opioids, but I haven't found a reference to endorphins having TLR activity. That is what I was originally looking for when I came across the naltrexone papers.
Presumably endorphins don't cause gliosis, hyperalgesia, tolerance and so on, and these are all properties of small molecules with (usually) both endorphin receptor and TLR activity.

Some interesting stuff you posted at Cort Johnson about gliadin and green tea. I have read there's an opioid antagonist in coffee. It might be the same thing that can cross-react with gliadin.

George Henderson said...

This paper separates TLR4 from endorphins
http://www.pnas.org/content/109/16/6325

Herein, we provide direct evidence that morphine creates neuroinflammation via the activation of an innate immune receptor and not via classic opioid receptors. We demonstrate that morphine binds to an accessory protein of Toll-like receptor 4 (TLR4), myeloid differentiation protein 2 (MD-2), thereby inducing TLR4 oligomerization and triggering proinflammation. Small-molecule inhibitors, RNA interference, and genetic knockout validate the TLR4/MD-2 complex as a feasible target for beneficially modifying morphine actions. Disrupting TLR4/MD-2 protein–protein association potentiated morphine analgesia in vivo and abolished morphine-induced proinflammation in vitro, the latter demonstrating that morphine-induced proinflammation only depends on TLR4, despite the presence of opioid receptors.

George Henderson said...

Finally, someone connects exorphins with TLR4 and microglia

http://search.informit.com.au/documentSummary;dn=734451123042694;res=IELHEA

Pay access, but note the minute fee - $1.80.

Lou Thomas said...

Just noticed your last two comments; that's great info, thanks.