Rationale: The plasma lipidome has the potential to reflect many facets of the host status during severe infection. Previous work is limited to specific lipid groups or was focused on lipids as prognosticators. Objectives: To map the plasma lipidome during sepsis due to community-acquired pneumonia (CAP) and determine the disease specificity and associations with clinical features. Methods: We analyzed 1,833 lipid species across 33 classes in 169 patients admitted to the ICU with sepsis due to CAP, 51 noninfected ICU patients, and 48 outpatient controls. In a paired analysis, we reanalyzed patients still in the ICU 4 days after admission (n = 82). Measurements and Main Results: A total of 58% of plasma lipids were significantly lower in patients with CAP-attributable sepsis compared with outpatient controls (6% higher, 36% not different). We found strong lipid class–specific associations with disease severity, validated across two external cohorts, and inflammatory biomarkers, in which triacylglycerols, cholesterol esters, and lysophospholipids exhibited the strongest associations. A total of 36% of lipids increased over time, and stratification by survival revealed diverging lipid recovery, which was confirmed in an external cohort; specifically, a 10% increase in cholesterol ester levels was related to a lower odds ratio (0.84; P = 0.006) for 30-day mortality (absolute mortality, 18 of 82). Comparison with noninfected ICU patients delineated a substantial common illness response (57.5%) and a distinct lipidomic signal for patients with CAP-attributable sepsis (37%). Conclusions: Patients with sepsis due to CAP exhibit a time-dependent and partially disease-specific shift in their plasma lipidome that correlates with disease severity and systemic inflammation and is associated with higher mortality.

The lipidome of immune cells during infection has remained unexplored, although evidence of the importance of lipids in the context of immunity is mounting. In this study, we performed untargeted lipidomic analysis of blood monocytes and neutrophils from patients hospitalized for pneumonia and age- and sex-matched noninfectious control volunteers. We annotated 521 and 706 lipids in monocytes and neutrophils, respectively, which were normalized to an extensive set of internal standards per lipid class. The cellular lipidomes were profoundly altered in patients, with both common and distinct changes between the cell types. Changes involved every level of the cellular lipidome: differential lipid species, class-wide shifts, and altered saturation patterns. Overall, differential lipids were mainly less abundant in monocytes and more abundant in neutrophils from patients. One month after hospital admission, lipidomic changes were fully resolved in monocytes and partially in neutrophils. Integration of lipidomic and concurrently collected transcriptomic data highlighted altered sphingolipid metabolism in both cell types. Inhibition of ceramide and sphingosine-1-phosphate synthesis in healthy monocytes and neutrophils resulted in blunted cytokine responses upon stimulation with lipopolysaccharide. These data reveal major lipidomic remodeling in immune cells during infection, and link the cellular lipidome to immune functionality.

Rationale: Lymphopenia in coronavirus disease (COVID-19) is associated with increased mortality. Objectives: To explore the association between lymphopenia, host response aberrations, and mortality in patients with lymphopenic COVID-19. Methods: We determined 43 plasma biomarkers reflective of four pathophysiological domains: endothelial cell and coagulation activation, inflammation and organ damage, cytokine release, and chemokine release. We explored if decreased concentrations of lymphocyte-derived proteins in patients with lymphopenia were associated with an increase in mortality. We sought to identify host response phenotypes in patients with lymphopenia by cluster analysis of plasma biomarkers. Measurements and Main Results: A total of 439 general ward patients with COVID-19 were stratified by baseline lymphocyte counts: normal (>1.0 × 109/L; n = 167), mild lymphopenia (>0.5 to ≤1.0 × 109/L; n = 194), and severe lymphopenia (≤0.5 × 109/L; n = 78). Lymphopenia was associated with alterations in each host response domain. Lymphopenia was associated with increased mortality. Moreover, in patients with lymphopenia (n = 272), decreased concentrations of several lymphocyte-derived proteins (e.g., CCL5, IL-4, IL-13, IL-17A) were associated with an increase in mortality (at P < 0.01 or stronger significance levels). A cluster analysis revealed three host response phenotypes in patients with lymphopenia: “hyporesponsive” (23.2%), “hypercytokinemic” (36.4%), and “inflammatory-injurious” (40.4%), with substantially differing mortality rates of 9.5%, 5.1%, and 26.4%, respectively. A 10-biomarker model accurately predicted these host response phenotypes in an external cohort with similar mortality distribution. The inflammatory-injurious phenotype showed a remarkable combination of relatively high inflammation and organ damage markers with high antiinflammatory cytokine levels yet low proinflammatory cytokine levels. Conclusions: Lymphopenia in COVID-19 signifies a heterogenous group of patients with distinct host response features. Specific host responses contribute to lymphopenia-associated mortality in COVID-19, including reduced CCL5 levels.

Alterations in platelet function have been implicated in the pathophysiology of COVID-19 since the beginning of the pandemic. While early reports linked hyperactivated platelets to thromboembolic events in COVID-19, subsequent investigations demonstrated hyporeactive platelets with a procoagulant phenotype. Mitochondria are important for energy metabolism and the function of platelets. Here, we sought to map the energy metabolism of platelets in a cohort of noncritically ill COVID-19 patients and assess platelet mitochondrial function, activation status, and responsiveness to external stimuli. We enrolled hospitalized COVID-19 patients and controls between October 2020 and December 2021. Platelets function and metabolism was analyzed by flow cytometry, metabolomics, glucose fluxomics, electron and fluorescence microscopy and western blot. Platelets from COVID-19 patients showed increased phosphatidylserine externalization indicating a procoagulant phenotype and hyporeactivity to ex vivo stimuli, associated with profound mitochondrial dysfunction characterized by mitochondrial depolarization, lower mitochondrial DNA–encoded transcript levels, an altered mitochondrial morphology consistent with increased mitochondrial fission, and increased pyruvate/lactate ratios in platelet supernatants. Metabolic profiling by untargeted metabolomics revealed NADH, NAD⁺, and ATP among the top decreased metabolites in patients’ platelets, suggestive of energy metabolism failure. Consistently, platelet fluxomics analyses showed a strongly reduced utilization of ¹³C-glucose in all major energy pathways together with a rerouting of glucose to de novo generation of purine metabolites. Patients’ platelets further showed evidence of oxidative stress, together with increased glutathione oxidation and synthesis. Addition of plasma from COVID-19 patients to normal platelets partially reproduced the phenotype of patients’ platelets and disclosed a temporal relationship between mitochondrial decay and (subsequent) phosphatidylserine exposure and hyporeactivity. These data link energy metabolism failure in platelets from COVID-19 patients with a prothrombotic platelet phenotype with features matching cell death.

Introduction: Monocytes play a pivotal role in the regulation of inflammation and pathogen clearance during infection. Knowledge of the human monocyte proteome during infection is limited. We present a comprehensive proteome profile of blood monocytes from patients with community-acquired pneumonia (CAP), one of the most common infectious diseases, and controls without infection. Methods: Monocytes were purified from blood of patients with CAP within 16 h of admission to a general hospital ward and from controls matched for sex, age and comorbidities. Monocyte proteins were measured using liquid chromatography/mass spectrometry. The transcriptome was analysed in the same samples by RNA sequencing. Monocytes were stimulated with lipopolysaccharide for 24 h, after which cytokines were measured in the supernatant. Results: We analysed the monocyte proteome of 34 CAP patients and 23 controls. Out of 7315 annotated proteins, 1340 (18.3%) were differentially abundant between groups. Functional enrichment analyses revealed a marked downregulation of mitochondrial respiration processes in patient monocytes; pathways pertaining to cell cycle, cytokine signalling and cell death were upregulated in patient monocytes. Monocyte mRNA levels correlated poorly with the abundance of corresponding proteins and associated functional pathways, raising caution regarding interpretation of functionality estimates based on transcriptome analyses. Differential expression of monocyte proteins had functional and clinical implications as indicated by associations with cytokine production capacity, disease severity and time to clinical stability. Conclusion: This study provides a publicly available monocyte protein atlas that can serve as a resource for future research on monocyte functions during infection.

Introduction: Platelets play a crucial role, amongst others, in the host response to infection through the release of proinflammatory and procoagulant proteins. Pneumonia represents a major disease burden, which has further increased since the onset of the coronavirus disease 2019 (COVID-19) pandemic. We aimed to determine platelets’ protein release profiles in hospitalised patients with non-COVID community-acquired pneumonia (CAP) or COVID-19.Methods: Platelets from patients with CAP or COVID-19 and from control subjects were incubated with platelet receptor agonists stimulating either glycoprotein (GP)VI or protease-activated receptor 1 (PAR1), or medium (spontaneous release); nine platelet products were measured in supernatants. Platelet activation was also assessed by measuring P-selectin and active GPIIbIIIa surface expression by flow cytometry.Results: We enrolled 32 patients with CAP, 104 patients with COVID-19 and 47 control subjects. Platelets from CAP patients spontaneously released less chemokines as compared to controls, which sharply contrasted with the platelets’ release profile in COVID-19, which was characterised by enhanced release of chemokines, plasminogen activator inhibitor 1 (PAI-1) and CD40 ligand relative to both controls and CAP patients. These differences were partially maintained uponex vivostimulation of GPVI or PAR1. Enhanced GPVI-induced platelet release of P-selectin, CD40 ligand, PAI-1 and CCL5 was associated with an increased risk for the need of (non)invasive ventilation and/or mortality in COVID-19 patients. Platelet expression of P-selectin and GPIIbIIIa did not differ between groups.Conclusion: Platelet release profiles differ strongly between CAP and COVID-19 patients, wherein platelets from COVID-19 patients released more bioactive products, linking with an adverse clinical outcome.