Preprint Persistent Activation of Chronic Inflammatory Pathways in Long Covid, 2024, Aid et al.

SNT Gatchaman

SNT Gatchaman

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Persistent Activation of Chronic Inflammatory Pathways in Long Covid
Malika Aid; Katherine McMahan; Nicole Hachmann; Jessica Miller; Erica Borducchi; David Hope; Marjorie Rowe; Eleanor Schonberg; Siline Thai; Ai-ris Collier; Janet Mullington; Dan Barouch

Long Covid, or Post-Acute Sequelae of COVID-19 (PASC), involves a spectrum of chronic symptoms following resolution of acute SARS-CoV-2 infection. Current hypotheses for the pathogenesis of Long Covid include persistent SARS-CoV-2, activation of other viruses, tissue damage, autoimmunity, endocrine insufficiency, immune dysfunction, and complement activation.

We evaluated 142 participants, including uninfected controls (N=35), acutely infected individuals (N=54), convalescent controls (N=25), and Long Covid patients (N=28), by comprehensive immunologic, virologic, transcriptomic, and proteomic analyses.

Long Covid was characterized by persistent inflammatory pathways compared with convalescent controls and uninfected controls, including upregulation of IL-6 and JAK-STAT pathways as well as activation of coagulation, complement, metabolism, and T cell exhaustion pathways. Moreover, robust activation of these pathways during acute COVID-19 infection correlated with the subsequent development of Long Covid. In an independent validation cohort (N=47), Long Covid patients had higher levels of plasma IL-6R compared with convalescent controls and uninfected controls.

These data demonstrate that Long Covid is characterized by persistent activation of chronic inflammatory pathways, suggesting novel therapeutic targets and biomarkers of disease.


Link | PDF (Preprint: BioRxiv) [Open Access]
 
This preprint was from May so I'm expecting it to be published soon. Summary quotes —

Methodology
  • In this study, we evaluate the immunologic and inflammatory responses in people with Long Covid compared with convalescent controls at 3-6 months and >6 months after initial COVID-19 infection, using immunologic assays, virologic assays, transcriptomics, and proteomics.

  • samples included uninfected controls (Uninfected; N=35), acutely infected individuals <1 month following COVID-19 infection (Acute; N=54), convalescent controls (CC; N=24), and Long Covid patients (LC; N=28)

  • Clinical symptoms in the Long Covid (LC) cohort included persistent fatigue, shortness of breath, exercise intolerance, brain fog, and abnormal smell and taste

  • Peripheral blood mononuclear cells (PBMC) were collected at 3-6 months (LC: N=25; CC: N=25) and >6 months (LC: N=19) following COVID-19 infection. Plasma samples were similarly collected at 3-6 months (LC: N=23; CC: N=8) and >6 months (LC: N=16; CC: N=6) following COVID-19 infection.
Results: Virology
  • first assessed SARS-CoV-2 neutralizing antibody (NAb) responses using pseudovirus neutralization assays and T cell responses by pooled peptide interferon-g ELISPOT assays against SARS-CoV-2 WA1/2020, Delta, and Omicron BA.1 in the CC and LC groups. We did not detect differences in NAb titers, but we observed higher Spike-specific ELISPOT responses to all three variants in the LC individuals compared with the CC individuals (P=0.015, P=0.006, P=0.002, respectively)

  • may reflect either more severe acute infection or could represent potentially persistent virus […] did not detect plasma SARS-CoV-2 in any CC or LC individuals by RT-PCR genomic or subgenomic viral load assays
Results: Transcriptomics
Viral reactivation
  • did not detect differences in viral reads for multiple viruses in the LC compared with CC groups, including human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV)
Whole transcriptome
  • Unsupervised cluster analysis using the whole transcriptome revealed a separation between the LC and other groups, while the CC group clustered with the uninfected controls
Inflammation, complement, coagulation
  • upregulation of multiple inflammatory markers in the LC compared with the CC groups and uninfected controls, including the signaling molecules LIFR and JAK2; chemokines CXCL2, CXCL3, and CCL3; cytokines IL10, NLRP3, IFNG, IL6, TNF, IL1B, IL1A; and complement and coagulation proteins C5, F3, and THBS1

  • compare the expression profiles in PBMC of the LC and CC groups at 3-6 months following acute COVID-19 infection. The LC group was characterized by higher levels of innate immune cell signatures for monocytes, macrophages, neutrophils and dendritic cells; complement and coagulation cascade signatures; and cytokine and chemokine signaling pathways, including IL6, IL8, IL10, IL12, IL17, JAK_STAT, and type I interferon pathways
NK cells
  • Genes associated with natural killer function such as KLRC4, KLRC1, KLRC2 and transcription factors such LEF1, GATA6, BACH2 were decreased in the LC group compared with the CC and uninfected control groups
T cells
  • We observed downregulation of certain T cell pathways in the LC group compared with the CC group (Fig. 4c), including T cell differentiation and activation pathways and T regulatory cell signatures (Fig. 5a-b). However, signatures of T cell exhaustion and PD-1 signaling pathways were significantly increased in the LC group compared with the CC group (Fig. 5b), suggesting that Long Covid is associated with T cell dysfunction. T cell signaling correlated inversely with IL6, JAK-STAT, and interferon signaling (Fig. 5c).
Metabolism
  • also observed increased metabolic activity in the LC group compared with the CC group involving amino acid and lipid metabolism
Most significant pathways
  • pathways that showed the most significant enhancement in the LC group compared with the CC group were the IL6, JAK_STAT, IL1R, mast cell, coagulation, complement, bile acid metabolism, ascorbate/aldarate, and leptin signaling. No differences in these pathways were observed between the CC and uninfected controls.
Results: Proteomics
  • similar differences between the LC and CC groups, consistent with the transcriptomic data

  • increased levels of immune cell signatures, such as neutrophils, monocytes and epithelial cells; cytokine signaling, including IL6, IL12, IL8, NFKPB, JAK-STAT, and corticotropin releasing hormone; complement and coagulation cascades; and certain metabolic pathways

  • pathways of cytotoxic T cell and NK_DCs cross talk signaling were downregulated

  • IL6 signaling in plasma inversely correlated with cytotoxic T cell, amino acid metabolism, and DNA damage pathways and directly correlated with metabolic pathways

  • During acute COVID-19 infection, increased IL6, complement, and cortisol signaling correlated with the subsequent development of Long Covid at 3-6 and >6 months

  • Feature importance analyses revealed that plasma level of IL6R, IL6ST, STAT1, STAT3, PTPN11 and MAPK1,3,4 during acute COVID-19 infection (< 1month) was associated with the subsequent development of LC at 3-6 and >6 months

  • To validate our findings in an independent cohort, we evaluated plasma levels of proinflammatory cytokines and chemokines and found increased levels of IL6R in individuals with LC (n=19) compared with CC (n=13) and uninfected controls (n=13), using both and ELISA and a meso-scale discovery (MSD) assay at 3-6 months and >6 months after COVID-19 infection
 
The pathways that showed the most significant enhancement in the LC group compared with the CC group were the IL6, JAK_STAT, IL1R, mast cell, coagulation, complement, bile acid metabolism, ascorbate/aldarate, and leptin signaling. No differences in these pathways were observed between the CC and uninfected controls.
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Just picking out those two items Replicated blood-based biomarkers for Myalgic Encephalomyelitis not explicable by inactivity (2024, Preprint: MedRxiv) said —

Considering all cases combined, 54 proteins are significant (FDR < 0.05). Among these are 7 complement proteins (C1RL, C2, CFB, CFH, CFI, CFP and CR2) of the innate immune system, whose levels are all elevated in ME/CFS cases, including CR2 (complement C3d receptor 2)
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and

ME/CFS cases also show increase in levels of leptin (LEP), which has a role in energy homeostasis
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A few previous references to leptin in ME/CFS (there are older ones) —

Distinct plasma immune signatures in ME/CFS are present early in the course of illness (2015, Science Advances)

Cytokine signature associated with disease severity in chronic fatigue syndrome patients (2017, PNAS)

with S4ME threads already for —

Association of circulating biomarkers with illness severity measures differentiates myalgic encephalomyelitis/chronic fatigue syndrome and post-COVID-19 condition: a prospective pilot cohort study (2024, Journal of Translational Medicine)

Proteomics and cytokine analyses distinguish myalgic encephalomyelitis/chronic fatigue syndrome cases from controls (2023, Journal of Translational Medicine)
 
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