Persistent complement dysregulation with signs of thromboinflammation in active Long Covid, 2024, Boyman et al

Persistent complement dysregulation with signs of thromboinflammation in active Long Covid
Open access: https://www.science.org/doi/10.1126/science.adg7942


Editor’s summary
Some individuals can endure persistent, debilitating symptoms for many months after an initial severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. However, the factors underpinning these health issues, called Long Covid, are poorly understood. Comparing the blood of patients with confirmed SARS-CoV-2 infection with that of uninfected controls, Cervia-Hasler et al. found that patients experiencing Long COVID exhibited changes to blood serum proteins indicating activation of the immune system’s complement cascade, altered coagulation, and tissue injury (see the Perspective by Ruf). At the cellular level, Long Covid was linked to aggregates comprising monocytes and platelets. These findings provide a resource of potential biomarkers for diagnosis and may inform directions for treatments. —Sarah H. Ross

RESULTS
Long Covid patients exhibited increased complement activation during acute disease, which also persisted at 6-month follow-up. The complement system is part of the innate immune system and contributes to immunity and homeostasis by targeting pathogens and damaged cells, among other functions. Interestingly, blood complement levels normalized in Long Covid patients recovering before their 6-month follow-up. The complement system can be activated by various triggers, resulting in formation of the terminal complement complex (TCC), made of the complement components C5b-9. These complexes can integrate into cell membranes and induce cell activation or lysis. Long Covid patients showed imbalanced TCC formation, marked by increased soluble C5bC6 complexes and decreased levels of C7-containing TCC formations that can incorporate into cell membranes. This suggested increased membrane insertion of TCCs in Long Covid patients, contributing to tissue damage. Accordingly, Long Covid patients showed elevated tissue injury markers in blood and a thromboinflammatory signature, characterized by markers of endothelial activation, such as von Willebrand factor (vWF), and red blood cell lysis. Low antithrombin III levels in Long Covid patients were accompanied by signs of increased cleavage by thrombin, a driver of TCC formation. Furthermore, Long Covid patients had elevated platelet activation markers and monocyte–platelet aggregates at 6-month follow-up, particularly in cases where Long Covid persisted for 12 months or more. These patients also showed signs of antibody-mediated activation of the classical complement pathway, which was associated with increased anti-CMV (cytomegalovirus, also known as human herpesvirus 5) and anti-EBV (Epstein-Barr virus) immunoglobulin G (IgG) antibody levels.

CONCLUSION
Our data suggest that active Long Covid is accompanied by a blood protein signature marked by increased complement activation and thromboinflammation, including activated platelets and markers of red blood cell lysis. Tissue injury may also be complement-mediated and, in turn, activate the complement system. Moreover, complement activation may be driven by antigen–antibody complexes, involving autoantibodies and antibodies against herpesviruses, as well as cross-talk with a dysregulated coagulation system. In addition to offering a basis for new diagnostic solutions, our work provides support for clinical research on complement modulators for patients suffering from Long Covid.

 

Looks interesting. A previous preprint on LC also pointed to problems with the complement system:
https://www.medrxiv.org/content/10.1101/2023.10.26.23297597v1

And it was also highlighted in some ME/CFS studies for example here:
https://www.mdpi.com/2077-0383/10/18/4171

 

Abstract

Long Covid is a debilitating condition of unknown etiology. We performed multimodal proteomics analyses of blood serum from COVID-19 patients followed up to 12 months after confirmed severe acute respiratory syndrome coronavirus 2 infection. Analysis of >6500 proteins in 268 longitudinal samples revealed dysregulated activation of the complement system, an innate immune protection and homeostasis mechanism, in individuals experiencing Long Covid. Thus, active Long Covid was characterized by terminal complement system dysregulation and ongoing activation of the alternative and classical complement pathways, the latter associated with increased antibody titers against several herpesviruses possibly stimulating this pathway. Moreover, markers of hemolysis, tissue injury, platelet activation, and monocyte–platelet aggregates were increased in Long Covid. Machine learning confirmed complement and thromboinflammatory proteins as top biomarkers, warranting diagnostic and therapeutic interrogation of these systems.

 

Nature News: Long-COVID signatures identified in huge analysis of blood proteins

 

Cohort

Follow-up of 113 hospitalised Covid-19 patients for 1 year (measurements taken at 0, 6 and 12 months). Thus according to most studies one would expect roughly 5-15 of these to develop LC with maybe 0-2 having severely disabling LC. However, one third of this cohort also had a severe acute infection (this means they had severe pneumonia which required oxygen support) which automatically further complicate observations (many people would also consider an infection that causes hospitalization to automatically be at the more severe end of the spectrum and if not that those people who require hospitalization for a mild infection might generally be more likely to report health adverse outcomes). Furthermore they also included 39 healthy controls (which are a lot younger than the hospitalised cohort).

Of those 113 acute Covid-19 cases they classified 40 patients to be suffering from LC (i.e. more than 1/3 of previously infected people) using the rather useless for research "one or more symptoms" definition (but excluding people that only had anosmia). At the 12 month mark 22 were still considered to be suffering from LC (8 were lost to follow-up). Looking at the supplementary material it appears most people are suffering from 1 or 2 symptoms at both time points and far fewer report more than 2 symptoms. Fatigue, Dyspnea and Dysosmia are the most common symptoms. Nobody appears to have PEM (but I also wouldn't expect anybody in this cohort to actually have PEM).

Often the looseness of the LC definition and typically the lack of attention to essential details (often failing to record the number of symptoms, severity of symptoms or limitations to daily life) have been a hindrance for similar studies, that will naturally always end up with a very mild spectrum of usually self-resolvent symptoms (many people encounter some health issues for whatever reasons after an arbitrary event), the 1 year follow-up can circumvent some of the problems other studies have.

Given they are studying hospitalized patients the focus will naturally be on somewhat overweight and elderly males with comborbidities and the LC group then naturally shifts towards more females.

The authors did a lot of analysis to address confounding factors and also acknowledge that

It seems extremely inefficient to conduct studies where you start of with mostly noise until you try to get somewhere, however this is also one of the very few studies (if not the only study) of hospitalised patients which I would consider of any value for the understaning of the development of LC as their 1 year follow-up adds a lot of value.

 

The Mercury News Long COVID creates changes in the blood, aiding detection, reports new study

quote:

The new findings are important because “they demonstrate dysfunction, which is important to patients,” said Jaime Seltzer, scientific director at the nonprofit MEAction, which advocates for patients with long COVID and myalgic encephalomyelitis/chronic fatigue syndrome, or ME/CFS.

“Secondly, they point the way to potential treatments, and even possibly mechanisms” of disease, she said.

 

NBC News has just run a story on this: Long Covid study reveals major insights on a potential cause (nbcnews.com)

 

Thread from one of the researchers explaining the study:

https://twitter.com/user/status/1748064835982553230

 

Trying to get an overview of all the tests they did in this study.

They started with analysing proteins using the ‘SomaScan platform’. This uses aptamers: DNA or RNA-molecules that bind to specific protein targets a bit like antibodies do. With this method they could screen more than 6000 different human proteins.

The one that showed the starkest difference between LC patients and recovered covid patients and healthy controls was C7, a protein that is part of the complement system. Unbinded C7 was normal in Long Covid patients but C7 complexes, the form where it binds to other proteins was reduced.




That suggested that the complement system was aberrant and so the researchers looked closer at other proteins that are involved in this immune pathway. C7 can bind to the C5bC6 complex, and the researchers found that C5bC6 was increased in LC patients and negatively correlated with C7.



There were even more proteins in the complement increased such as C2, Ba and C3d. These increases suggest that both the classical and the alternative complement pathway were abnormal in Long Covid patients. However, Mannose-Binding-Lectin (MBL) was normal so there was no proof of involvement of the lectin complement pathway.

The researchers were able to confirm abnormal complement proteins using another technique (mass spectrometry) and by testing it in an independent Long Covid cohort from Mount Sinai.

Then they started looking what an abnormal complement pathway might mean. Potential downstream effects of complement activation include destruction of red blood cells (hemolysis), blood clotting (thromboinflammatory responses), and changes in innate immune cells, such as neutrophils.

I think they found most evidence pointing toward blood clotting: they found for example lowered antithrombin III in the serum, a natural anticoagulant that inhibits clotting factors. They also found increased Von Willebrand Factor (vWF) which acts as a bridge between platelets and the blood vessel wall during clot formation. In a separate smaller experiment they used flow cytometry on monocytes cells of 10 Long Covid patients and found that CD41, a platelet marker that is usually absent in these white blood cells, was increased in long covid patients.

Other proteins pointed toward tissue injury and destruction or red blood cells so the authors suspect that this might be what the complement system is responding to. Another possibility is viral cause because the authors also found increased antibody levels against herpesviruses.

 

That other LC study on the complement system (the preprint by Paul Morgan et al.) also found activation of the classical and alternative but not the lectin pathway although the intermediary proteins they measured differed somewhat. I don't see for example c7 or the the C5bC6 complex mentioned.

Here are the main results from that study, which was larger with 166 LC patients and 79 controls.



Source: https://www.medrxiv.org/content/10.1101/2023.10.26.23297597v1

 

Found this simplified overview in a YouTube video that gives an introduction to the complement system.

 

The Spanish ME/CFS study found increased levels of circulating complement factor C1q but the LC preprint mentioned above found these C1q to be decreased.

An Austrian ME/CFS study found reduced Mannose-Binding Lectin (MBL) but this Science paper on LC did not find it to be abnormal.

So I don't think there is a good match between these new LC findings and older complement abnormalities in the ME/CFS field.

 

Everything in this study seems to be 'signs of' or 'tending towards' or 'suggestive of'... maybe 'thromboinflammation'.

But what is thromboinflammation, and more to the point was there any?

If I had thromboinflammation I would expect it to be red, if not purplish, and sore enough to keep me awake at night. I haven't had any bits like that.

As far as I can see they have found suggestions of processes that might have produced thrombosis or inflammation if given the chance - but didn't.

In the old days, as I have said before, papers gave you data. Nowadays all you get is an interpretation. I can make my own mind up thank you.

If C7 is off in LC that might be interesting but to be honest C7 isn't the most exciting of things to find off.

 

And there's this too!

 

Could the so-called "Covid-toes" be a sign of that or would it be more likely that, that would hav something to do with blood pooling or something else?

 

Perhaps it was just coincidence that C7 was the protein that had the starkest difference. They did find other proteins involved in the classical and alternative complement pathway to be abnormal in LC patients with some confirmation in their independent Mount Sinai cohort. The preprint by the Cardiff team also found this.

 

There is still a lot of doubt as to whether Covid toes have much to do with Covid. I wouldn't be surprised if they did but they are reported as part of the acute illness and as far as I know usually as a one off. People with LC are not getting new purple patches on toes every day of the week. It just doesn't fit.

 

I actually note now that they say what was low was C7 containing terminal complexes. I have no idea what that might mean. Circulating activation of membrane attack complex is not normally invoked as a cause of damage. The circulation uses complement as an anti-inflammatory strategy by and large - using CR1 on red cells to clear complement-bound material silently. Membrane attack complex is mostly seen as causing cell lysis outside the circulation. As to what free complexes containing C7 would mean goodness knows. These things may be relevant in septic shock syndrome or lupus vasculitis but LC simply doesn't have any clinical features like that.

 

Eric Topol wrote about this study in his newsletter: https://open.substack.com/pub/erict...id?r=2gi8q&utm_campaign=post&utm_medium=email

He writes:

 

A Swedish (paywalled) news article about this study, including comments by Petter Brodin.

Forskarnas nya fynd: Postcovid kan upptäckas med enkelt test
https://www.aftonbladet.se/nyheter/a/69bX68/ny-studie-postcovid-kan-upptackas-med-enkelt-test

ETA: Some of you might remember this news interview from March 2023 when Brodin, who used to do ME research, said that they will find a cure for long covid before the end of 2023...