So many possibilities…Is it the mitochondria? Or the blood vessels? Or the liver? Or plasmalogens? Or viral reactivation? How about viral-induced inflammation? Brainstem dysfunction… It’s both exciting and overwhelming to see so many possibilities show up for long COVID, ME/CFS, and related diseases.
We want the answer and we don’t seem to be getting it. Instead, we seem to getting more in the weeds. Maybe, though, we need to step away a bit. It’s possible that many of these theories are, in fact, correct and that we simply have to find a way to link them together. It’s probably going to take a multi-systemic problem, after all, to produce – in ME/CFS’s case – one of the most functionally disabling diseases known to man.
So maybe the best thing to do is just sit back and watch things reveal themselves. Something interesting certainly revealed itself recently in a long-COVID study which actually may be able to tie things up a bit.
The New York Times offered up its review “Scientists Offer a New Explanation for Long Covid” from Pulitzer Prize winner Pam Belluck. Check out what Belluck’s intro says about the state of long-COVID research.
“This is one of several new studies documenting distinct biological changes in the bodies of people with long Covid — offering important discoveries for a condition that takes many forms and often does not register on standard diagnostic tools like X-rays.”
Things, in other words, are happening…
The study, “Serotonin reduction in post-acute sequelae of viral infection“, which included about 50 mostly University of Pennsylvania co-authors (plus some heavyweights (Henrich, Deeks, Peluso) from the UCSF LIINC team), made up for in intensity which it lacked in size. As in the recent NIH WASF3 study, once this group uncovered a bone, they really gnawed on it.
That’s new – at least for ME/CFS. We’re seeing research groups with the resources to really chase down their findings take on long COVID and it shows. Both the Hwang group, with its WASF3 finding, and this group with its serotonin finding, for instance, immediately turned to and expanded their findings using mouse studies – something that has almost never been done in ME/CFS. You’ll see this group take a finding, validate it in several ways, and then expand on it. This will happen multiple times. In the end, you get a preliminary but satisfying and, in this case, possibly quite expansive result.
While the study was not large (n=110), it was certainly not small by ME/CFS stands. It compared the metabolites (using metabolomics) in the blood of long COVID patients between three months and 22 months after their infection to recovered COVID-19 patients and people who were in the midst of a coronavirus infection.
The main pathway that popped up – the serotonin pathway – was a bit of a surprise. Serotonin was the only significant metabolite that did not recover to pre-infection levels in the long-COVID patients.
Serotonin seems like a new entrée in the “discover what’s causing ME/CFS and long-COVID sweepstakes”, but it’s not really. While serotonin itself has never been a major focus, ME/CFS researchers have been looking into a key part of the serotonin pathway – tryptophan – for quite some time. Plus, low serotonin levels have been found in ME/CFS.
One thing for people with ME/CFS and other post-infectious diseases to note is that because these researchers used a number of pathogens to test their hypotheses, this finding is not necessarily specific to the coronavirus. For instance, they found that infecting mice with the vesicular stomatitis virus (VSV) also decreased their plasma serotonin levels, and that after creating a chronic infection with the lymphocytic choriomeningitis virus (LCMV), the same thing happened.
When they introduced a compound (synthetic double-stranded RNA polyinosinic: polycytidylic acid (poly(I: C)), that mimics a viral infection, serotonin levels dropped as well. Once they stopped providing that compound to the mice, their serotonin levels rebounded.
The Interferon Connection
Since the problem is not necessarily the virus – but the immune response to the virus – next they assessed the levels of the main antiviral cytokine – type 1 interferons (IFN-a/IFN-b) and found that a coronavirus, or other infection, as well as a simulated infection, all “strongly upregulated” interferon-stimulated genes.
Interferon alpha (IFN-a) – an antiviral factor produced by the body – was used for many years to treat hepatitis C. Its production of severe fatigue in a significant subset of patients helped to open the door for distinguishing “sickness behavior” and the realization that most of the symptoms experienced during an infection are produced by the immune system.
The IFN-a finding also produced a nice template for understanding how sickness behavior is produced and how to manage it. Subsequent studies that examined what’s happening in IFN-a-treated patients, again and again, seemed to have relevance for ME/CFS. That made sense – from the very beginning, it’s been conjectured that people with ME/CFS are simply caught in a sickness behavior loop.
IFN-a increases serotonin turnover in the prefrontal cortex – an area of the brain impacted in ME/CFS. IFN-a administration targets the reward and movement centers of the brain in the basal ganglia – a brain region of interest in ME/CFS – and induces motor slowing – which has been found in ME/CFS. IFN-a administration has also been associated with reductions in another feel-good brain chemical – dopamine. (Serotonin production in the brain occurs in the brainstem – another area of interest in ME/CFS).
Given the carnitine issues that have shown up in ME/CFS recently, it was interesting that IFN plus L-carnitine reduced the fatigue levels. Another study found that Modafinil – a stimulant – was helpful in reducing IFN-a-induced (i.e. inflammation-induced) depression. A gene expression study noted that similar genes were activated in IFN-a-treated patients and people with ME/CFS. Three years ago, London researchers proposed that IFN-a constituted a “novel, inflammation-based, proxy model of chronic fatigue syndrome“.
A nice, robust increase in interferon-associated genes then, makes sense in both long COVID and ME/CFS.
An Interferon-Serotonin Connection
Next came the big question – could the interferon activation have had anything to do with the reduced levels of serotonin found? Again they turned to the mice and found that serotonin levels stayed at normal levels in mice that were genetically created not to respond via the interferon pathway to the viral mimic.
Since most of the circulating serotonin in the body is synthesized from tryptophan in the gut, the next question was whether the coronavirus (or another viral infection) might be limiting tryptophan production in the gut. (Tryptophan is the precursor to serotonin.)
- We’re starting to see some really in-depth studies; studies that are able to go beyond initial findings and expand greatly on them. That’s obviously the result of research groups with the resources and time to really track down findings. That’s something we haven’t really had with ME/CFS but do, at times, have with long COVID – and that means things can go much more quickly.
- This is a long and complicated study – so much the better! The 50+ research group used metabolomics to assess what was going on in long COVID and then a bunch of mouse studies to expand on the results.
- Finding that serotonin was the only significant metabolite to be downregulated in long-COVID patients, they infected mice with several different viruses and exposed them to a viral mimic – and found (lo and behold) reduced serotonin levels in them as well.
- That prompted them to assess the main antiviral response in cells – the interferon system – and found evidence that it was highly activated. (See the blog for evidence of interferon upregulation in ME/CFS.) Next, they asked if the activated interferon system might be interfering with the production of the precursor to serotonin – tryptophan. Turning back to the mice, they asked if an infection might be interfering with the production of tryptophan – and it was.
- So far so good…But how was tryptophan being depleted? Turning to the main source of tryptophan in the body – the gut. A gene expression analysis of the intestinal tissues revealed a strong upregulation in genes associated with inflammation and viral infections. “Remarkably”, they stated the gene functions “most significantly diminished” by the viral mimic were involved in nutrient metabolism, including amino acid (tryptophan is an amino acid) absorption; i.e. the infection appeared to have affected the ability of their cells to take up tryptophan, in particular.
- Things were really heating up now. Wondering whether tryptophan supplementation could help, they found that both a special diet (containing a glycine-tryptophan dipeptide) and/or supplementation with the serotonin precursor 5-hydroxytryptophan (5-HTP) returned serotonin levels to normal.
- The authors concluded that “collectively, these data demonstrate that viral-RNA-induced inflammation impairs intestinal tryptophan uptake, which causes systemic serotonin depletion.” Notice that they’re not just talking about the coronavirus…this finding could pertain to all infectious events – which, of course, means it could apply to ME/CFS.
- Next, the manufactured small intestinal “organoids” – miniaturized organs that are derived from stem or tissue cells to study what a simulated viral attack might be doing to the intestinal tissues. Rather remarkably, the organoids responded with a downregulation of the ACE2 receptor – which has been implicated in both long COVID and ME/CFS.
- The ACE2 receptor is associated with the “renin-angiotensin-aldosterone” paradox, which makes it impossible to raise the blood volume in ME/CFS to normal levels but until recently has been mostly ignored. ACE2 dysregulation could, though, also be producing inflammation, whacking the mitochondria, causing fibrosis, inhibiting muscle repair, damaging the endothelial cells lining the blood vessels, producing vasoconstriction (narrowing) in the blood vessels, jacking up oxidative stress levels, reducing the levels of nitric oxide – an important vasodilator, and impacting the gut flora.
- Importantly, this study suggests that any infection might be able to dysregulate the ACE2 receptor – thereby potentially explaining why it’s gotten messed up in ME/CFS.
- The authors also showed how low serotonin levels might be impacting the vagus nerve. The authors turned conservative in the end, proposing that supplementation (5-HTP) and selective serotonin reuptake inhibitors might help increase serotonin levels. They are starting to trial to test the effectiveness of fluoxetine (Prozac) and possibly tryptophan.
- Compare that, though, to a 2021 paper focusing on ACE2 dysfunction which proposed using escitalopram, coenzyme Q10, and nicotinamide adenine dinucleotide to restore endothelial functioning, suggested trying angiotensin receptor blockers (ARBs), fat globule membranes (MFGM), b-glucan and metformin to restore gut health, and drugs called senotherapeutics (dasatinib, hyperoside, quercetin, fistein, Navitoclax) to impact cell death and aging.)
- (Although the authors did not mention it, the Epstein-Barr virus – which is commonly reactivated in ME/CFS – has been shown to impact serotonin levels as well.)
- Finally, the authors proposed that serotonin depletion links the four horsemen of the long-COVID apocalypse (viral persistence, chronic inflammation, hypercoagulability, and autonomic dysfunction) together.
- The study garnered a lot of media attention and was well-received by major long-COVID researchers. As many of the results were done in mouse studies, they need to be verified in humans (when possible) and larger studies need to be done. For now, though, the “serotonin surprise” is making waves.
Whence the Tryptophan Deficiency?
Now it was time to chase down the cause of the tryptophan deficiency. Noting reduced levels of tryptophan can be caused by gut problems (reduced absorption) or by having too much tryptophan being converted in kynurenine, they next examined the kynurenine levels in their long-COVID patients and found – to their evident surprise – that they were not elevated.
Once again going the extra mile, they found that even inhibiting kynurenine-producing enzyme levels did not restore serotonin levels; i.e. the tryptophan deficiency was not caused by problems in the kynurenine pathway (as has been proposed in ME/CFS).
They surmised that the origin of the tryptophan and serotonin deficiency, then, must lie in the gut. Thinking that tryptophan/serotonin was simply not being absorbed, they put the mice through a series of dietary tests. Neither putting the mice on a diet, nor feeding them extra supplies of tryptophan (while exposing them to the viral mimic) altered their tryptophan or serotonin levels, indicating that tryptophan absorption in the gut was not the problem. Nor did they find a reduction in serotonin-producing cells in the gut.
Apparently at somewhat of a loss as to what was going on, they turned to the tissues of the small intestine of the mice to try to understand what was going on. A gene expression analysis of those tissues revealed that the mice exposed to the infection mimic demonstrated strong upregulations in genes associated with inflammation and viral infections.
“Remarkably” they stated the gene functions “most significantly diminished” by the viral mimic were involved in nutrient metabolism, including amino acid (tryptophan is an amino acid) absorption; i.e. the infection appeared to have affected the ability of their cells to take up tryptophan in particular. (The gene expression pathway that converts tryptophan into serotonin was not affected: the problem appeared to entirely lie in the inability of the gut tissues to take up tryptophan.)
Next came a fascinating and possibly helpful test. Wondering whether tryptophan supplementation could compensate for the impaired tryptophan uptake, they found that both a special diet (containing a glycine-tryptophan dipeptide) and supplementation with the serotonin precursor 5-hydroxytryptophan (5-HTP) returned serotonin levels to normal.
The authors concluded that “collectively, these data demonstrate that viral-RNA-induced inflammation impairs intestinal tryptophan uptake, which causes systemic serotonin depletion.”
They weren’t nearly done yet. This intrepid group’s next step was to use something we’ve never seen before in ME/CFS research – small intestinal “organoids” – to study what a simulated viral attack did to the intestinal tissues.
Organoids are miniaturized and simplified organs that are derived from stem or tissue cells. The organoids, interestingly enough, responded with a downregulation of the ACE2 receptor – which has been implicated in both long COVID and ME/CFS. The analysis implicated the NF-kB transcription factor and the TLR3 receptor it turns on in the interferon response, and found that type 1 interferons reduced the expression of genes involved in tryptophan absorption in the gut.
The ACE2 Tryptophan Link
ACE2 is the receptor the coronavirus uses to enter cells. A downregulation of the receptor had been found when the mice were subjected to a simulated viral attack. Several small studies have implicated the ACE2 receptor and the renin-angiotensin-aldosterone system (RAAS) in ME/CFS and POTS as well.
Next, the researchers used genetically altered mice to show that downregulation of the ACE2 receptor was associated with a reduction in tryptophan levels. That finding potentially tied together long COVID, ME/CFS and POTS in a very specific way.
From the beginning, it seemed clear the virus would impact the ACE2 receptor in long COVID – it was, after all, how it entered the cell. But what about ME/CFS? What might have caused the receptor to go bonkers in ME/CFS has been a mystery. Now we have a possible cause. It appears that an infection – perhaps any infection – can impair the activity of ACE2 genes.
They also noted that because tryptophan is the precursor for niacin, NAD, and melatonin, those compounds could be impacted as well. Note that each of these has been used in ME/CFS and associated diseases.
The Weird ACE2 Long COVID – ME/CFS Connection
It’s seemed beyond weird that the coronavirus would enter the cells using the same receptor that studies have linked to the renin-aldosterone-paradox in ME/CFS and POTS. ACE2 dysregulation could not only account for the low blood volume in ME/CFS but could also impact many other things (lowered white blood cell counts, impaired oxygen uptake by hemoglobin, increased oxidative stress, leaky gut, problems with lung perfusion, reduced vasodilation of the blood vessels, inflammation, and heart issues).
I’ve been puzzled that the ACE2 situation in ME/CFS has not been addressed more. Until Wirth and Scheibenbogen addressed the renin-angiotensin-aldosterone situation in their hypothesis, it was virtually ignored in ME/CFS circles. Things began to pick up a bit with the coronavirus pandemic, however. A 2021 study failed to find any difference in ACE2 gene expression in ME/CFS and healthy controls, while another found reduced ACE2 expression.
We knew that the coronavirus’s use of the ACE2 receptor could affect it. Except for Miwa’s proposal that the HPA axis, and Wirth and Scheibenbogen’s suggestion of activation of the kallikrein-kinin-system (KKS), we had no explanations for one of the strangest manifestations in ME/CFS – the renin-angiotensin-aldosterone paradox, where the presence of low blood volume fails to kick in the RAAS system to boost it up again.
Now we have a possible answer – any infection may be able to impair ACE2 functioning.
That could be a big deal. In 2021, researchers proposed that the renin-angiotensin-aldosterone system, that ACE2 is a part of, was producing inflammation, whacking the mitochondria, causing fibrosis, inhibiting muscle repair, damaging the endothelial cells lining the blood vessels, producing vasoconstriction (narrowing) in the blood vessels, jacking up oxidative stress levels, reducing the levels of nitric oxide – an important vasodilator, and causing the gut flora to be thrown off in both ME/CFS and long COVID.
Ignoring the really intriguing connection between ACE2 receptor dysregulation in long COVID, ME/CFS, and POTS, the authors did note that “none of the mechanisms described in this study are unique to SARS-CoV-2 infection“.
Finally, they used mouse studies to show that serotonin depletion could also be responsible for a reduction in vagus nerve functioning and cognitive impairment, and noted that vagus nerve functioning is impaired in chronic fatigue syndrome (yah!).
It was quite a tour de force. Given all the experiments carried out, it wasn’t surprising that this study had 50+ co-authors.
The authors acknowledged that not all long-COVID patients have low serotonin levels, and turned conservative at the end, proposing that supplementation (5-HTP) and selective serotonin reuptake inhibitors might help increase serotonin levels. They are starting to trial to test the effectiveness of fluoxetine (Prozac) and possibly tryptophan. While this is going to be a rather crude trial the researchers will be able to see if Prozac increases serotonin metabolite levels and if doing so helps.
(Compare that to the 2021 paper which proposed using escitalopram, coenzyme Q10, and nicotinamide adenine dinucleotide to restore endothelial functioning, suggested trying angiotensin receptor blockers (ARBs), fat globule membranes (MFGM), b-glucan and metformin to restore gut health, and drugs called senotherapeutics (dasatinib, hyperoside, quercetin, fistein, Navitoclax) to impact cell death and aging.)
Again, note that because these researchers used a variety of pathogens as well as simulating a pathogen attack, we know this model is not unique to the coronavirus and could apply to other post-infectious diseases. In fact, the authors proposed it might apply to non-viral conditions such as multiple sclerosis.
Epstein-Barr Virus (EBV) Connection?
One wonders, given the recent association found between multiple sclerosis and Epstein-Barr virus (EBV), how long MS will be considered “non-viral”. The authors also missed a magnificent opportunity (in my mind, anyway) to connect EBV with a similar scenario.
EBV can be found in the mucosal lining of the gut and can affect and be affected by the gut microbiome. In 2021, Ohio State University researchers found that Epstein-Barr Virus dUTPase enzyme altered the expression of tryptophan, dopamine, and serotonin metabolism in ME/CFS, and chronic EBV has been associated with increased tryptophan degradation. Interestingly, EBV interacts with the ACE2 receptor as well.
This group got far enough to propose a mechanism that linked the four horsemen of the long-COVID apocalypse (viral persistence, chronic inflammation, hypercoagulability, and autonomic dysfunction) together in a single devastating pathway, and suggested some treatments.
“All these different hypotheses might be connected through the serotonin pathway,” said Christoph Thaiss, a lead author of the study and an assistant professor of microbiology at the Perelman School of Medicine at the University of Pennsylvania.
The study made quite a media splash showing up New York Times, Yahoo, Medical News Today, NPR, MSN News, ABC News, etc.
Dr. Michelle Monje, a professor of neurology at Stanford University, said “I’m impressed by the study. I think they did a beautiful job showing the causality of these changes.” Akiko Iwasaki, an immunologist at Yale University, said the story shows a “very nice linear story. Everyone who’s engaged in this research should now be thinking about this serotonin pathway,”
Since a major part of the study involved mouse studies, the results need to be verified in humans and in larger studies. It should also be noted that both Cortene’s work and the Metabolic Trap hypothesis predicted that high not low serotonin levels in the brain were the problem. For now, though, the “serotonin surprise” is making waves.
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