Ex vivo IL-1β, IL-6, TNF-α, and IL-8 responses 2 years after critical COVID-19 are associated with age and innate immune activation
Abstract
A cytokine storm drives critical COVID-19, yet the determinants and long-term consequences of this exaggerated inflammatory response remain unclear. We investigated whether cytokine responsiveness remains altered 2 years after recovery from critical disease. In a cohort of 117 individuals with mild, severe, or critical COVID-19, acute-phase clinical and laboratory parameters were analyzed in moderate–severe and critical cases. Long-term immune function was assessed by ex vivo stimulation of whole blood with PMA/ionomycin, lipopolysaccharide (LPS), and phytohemagglutinin (PHA), followed by cytokine quantification and correlation with clinical parameters. A biobank of 76 patient-derived lymphoblastoid cell lines (LCL cells) was established to evaluate the stability of cytokine responses. During acute illness, critically ill patients exhibited marked inflammatory dysregulation, including lymphopenia (72%), neutrophilia (51%), elevated neutrophil-to-lymphocyte ratios, and increased C-reactive protein (median 91 mg/L) and ferritin (median 1,153 μg/L), consistent with a hyperinflammatory–hypercoagulable phenotype. Two years post-infection, individuals with prior critical disease showed more than threefold higher secretion of IL-1β, IL-6, TNF-α, and IL-8 following PMA/ionomycin (p < 0.0001) and LPS stimulation (p < 0.05) compared with younger mild cases, whereas PHA-induced responses remained comparable. PBMC counts were also higher in the moderate–severe and critical groups 2 years post-infection (p < 0.001), highlighting differences in cellular composition. LCL cells from critically ill donors secreted 79% more IL-8 and TNF-α than those from mild cases (p < 0.05). Collectively, these findings indicate differences in cytokine responsiveness 2 years after COVID-19 which correspond to prior disease severity and are associated particularly with age and innate immune activation.
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BibTeX
@article{hiti2026vivo,
title = {{Ex vivo IL-1β, IL-6, TNF-α, and IL-8 responses 2 years after critical COVID-19 are associated with age and innate immune activation}},
author = {Luka Hiti and Maša Kandušer and Tilen Burnik and Nataša Karas Kuželički and Jerneja Farkaš Lainščak and Mojca Miholič and Emil Pal and Mitja Lainscak and Irena Mlinarič-Raščan and Tijana Markovič},
journal = {Scientific Reports},
year = {2026},
month = oct,
doi = {10.1038/s41598-026-76481-1},
url = {https://www.nature.com/articles/s41598-026-76481-1},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Ex vivo IL-1β, IL-6, TNF-α, and IL-8 responses 2 years after critical COVID-19 are associated with age and innate immune activation
AU - Luka Hiti
AU - Maša Kandušer
AU - Tilen Burnik
AU - Nataša Karas Kuželički
AU - Jerneja Farkaš Lainščak
AU - Mojca Miholič
AU - Emil Pal
AU - Mitja Lainscak
AU - Irena Mlinarič-Raščan
AU - Tijana Markovič
PY - 2026
DA - 2026/10/24
JO - Scientific Reports
DO - 10.1038/s41598-026-76481-1
UR - https://www.nature.com/articles/s41598-026-76481-1
AB - A cytokine storm drives critical COVID-19, yet the determinants and long-term consequences of this exaggerated inflammatory response remain unclear. We investigated whether cytokine responsiveness remains altered 2 years after recovery from critical disease. In a cohort of 117 individuals with mild, severe, or critical COVID-19, acute-phase clinical and laboratory parameters were analyzed in moderate–severe and critical cases. Long-term immune function was assessed by ex vivo stimulation of whole blood with PMA/ionomycin, lipopolysaccharide (LPS), and phytohemagglutinin (PHA), followed by cytokine quantification and correlation with clinical parameters. A biobank of 76 patient-derived lymphoblastoid cell lines (LCL cells) was established to evaluate the stability of cytokine responses. During acute illness, critically ill patients exhibited marked inflammatory dysregulation, including lymphopenia (72%), neutrophilia (51%), elevated neutrophil-to-lymphocyte ratios, and increased C-reactive protein (median 91 mg/L) and ferritin (median 1,153 μg/L), consistent with a hyperinflammatory–hypercoagulable phenotype. Two years post-infection, individuals with prior critical disease showed more than threefold higher secretion of IL-1β, IL-6, TNF-α, and IL-8 following PMA/ionomycin (p < 0.0001) and LPS stimulation (p < 0.05) compared with younger mild cases, whereas PHA-induced responses remained comparable. PBMC counts were also higher in the moderate–severe and critical groups 2 years post-infection (p < 0.001), highlighting differences in cellular composition. LCL cells from critically ill donors secreted 79% more IL-8 and TNF-α than those from mild cases (p < 0.05). Collectively, these findings indicate differences in cytokine responsiveness 2 years after COVID-19 which correspond to prior disease severity and are associated particularly with age and innate immune activation.
SN - 2045-2322
ER -