Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, presents a spectrum of severity, ranging from asymptomatic infection to severe pneumonia requiring mechanical ventilation. While most patients experience mild symptoms, a significant subset develops acute respiratory distress syndrome (ARDS) and faces high mortality risk. Several risk factors for severe COVID-19 have been identified, including age, sex, obesity, and pre-existing comorbidities. Hyperinflammation and microthrombosis are also associated with ARDS and fatal outcomes. The innate immune system's initial response to SARS-CoV-2 appears insufficient to clear the infection, necessitating adaptive immune responses involving T and B cells for viral clearance. Virus-specific T cell responses and neutralizing antibodies are crucial for resolving the infection. However, the role of polyclonal T cell reactivity in COVID-19's pathogenesis remains unclear, with studies reporting both insufficient and excessive T cell responses in severe cases. Existing methods for measuring polyclonal T cell reactivity, such as intracellular cytokine staining or cytokine quantification, have limitations in sensitivity and consistency. This study aimed to develop a more sensitive and reliable method to evaluate polyclonal T cell reactivity in COVID-19 patients and investigate its association with disease severity and clinical outcomes.

Prior research on T cell responses in COVID-19 has yielded inconsistent results. Some studies reported decreased interferon-gamma expression in severe COVID-19, while others found no change or even increased responses. The majority of research focused on SARS-CoV-2-specific T cell responses which are frequently reduced in critically ill patients. These studies highlighted the need for a more comprehensive and sensitive approach to assess polyclonal T cell reactivity in diverse COVID-19 patient populations to better understand its contribution to disease pathogenesis and outcome prediction.

This study enrolled 55 COVID-19 patients admitted to the University Hospital Regensburg and 42 healthy controls. Patients were stratified into non-ventilated and ventilated groups (further divided into survivors and non-survivors). A sensitive method was employed to analyze polyclonal T cell reactivity by measuring downstream effects on responder cells (basophils, pDCs, monocytes, and neutrophils) in whole blood samples. This approach offered greater sensitivity and consistency compared to traditional methods using PBMCs. Flow cytometry was used to quantify the expression of surface markers on these responder cells after stimulation with anti-CD3 antibody. Cytokine release from activated T cells was also measured using ELISA. To determine whether T cell hyporeactivity was intrinsic or extrinsic, washed whole blood from a healthy donor was incubated with plasma from COVID-19 patients and healthy controls. The effects of various interventions, such as IL-2 addition, IL-10 blockade, and L-tryptophan supplementation, on T cell reactivity were investigated. Immunophenotyping of fresh blood samples was also performed to analyze innate cell populations (basophils, pDCs, monocytes, and neutrophils). Finally, a predictive score was developed based on immunological parameters to predict mortality in ventilated patients. Statistical analyses included one-way ANOVA, two-tailed unpaired t-tests, and calculation of predictive values (sensitivity, specificity, positive predictive value, negative predictive value).

The study revealed significant T cell hyporeactivity in COVID-19 patients, particularly those requiring mechanical ventilation. This hyporeactivity was strongly associated with prolonged viral replication and poor clinical outcomes. The hyporeactivity was determined to be T cell-extrinsic and mediated by plasma components, independent of immunosuppressive medication. Gender-specific differences were observed, with males exhibiting stronger monocyte responses. IL-2 partially restored T cell activation. A novel predictive score, incorporating basophil counts and CD123/CD11b upregulation on monocytes and neutrophils respectively, accurately predicted mortality in ventilated patients. Immunophenotyping of fresh blood revealed reduced basophil and pDC counts, as well as altered monocyte marker expression in severe COVID-19. The study demonstrated the reversibility of T cell hyporeactivity in patients with favorable outcomes, contrasting with persistent hyporeactivity in those with fatal outcomes. In summary, the study revealed significant T cell hyporeactivity in ventilated COVID-19 patients, associated with prolonged viral persistence and poor outcomes. This hyporeactivity is caused by plasma components and is partially reversible with clinical recovery. A novel predictive score based on immunological markers was developed to predict mortality in these patients.

The findings highlight the importance of T cell hyporeactivity as a key factor in the pathogenesis of severe COVID-19. The novel method used in this study, focusing on downstream effects of T cell activation on responder cells, proved to be more sensitive than traditional methods and revealed subtle but significant differences between patient groups. The identification of plasma components as the mediators of T cell hyporeactivity suggests potential therapeutic targets. The predictive score developed in this study could be a valuable tool for clinicians to identify patients at high risk of mortality and guide therapeutic interventions. Future studies should focus on larger clinical trials to validate the score and explore the therapeutic potential of strategies aimed at overcoming T cell hyporeactivity.

This study provides strong evidence for the role of T cell hyporeactivity in the severity and outcome of COVID-19, particularly in ventilated patients. The development of a predictive score and the identification of plasma-mediated T cell suppression offer new avenues for improving patient care. Future research should focus on validating the predictive score in larger cohorts and exploring the efficacy of targeted therapies to restore T cell function in critically ill patients.

The study's relatively small sample size limits the generalizability of the findings. Further research with larger and more diverse populations is needed to confirm the findings and refine the predictive score. The study was conducted during the early stages of the pandemic, and the emergence of new SARS-CoV-2 variants might affect the results. The mechanisms underlying the plasma-mediated suppression of T cell activity require further investigation.