Distinct functional neutrophil phenotypes in sepsis patients correlate with disease severity.

Purpose: Sepsis is a clinical syndrome defined as life-threatening organ dysfunction caused by a dysregulated host response to infection. Sepsis is a highly heterogeneous syndrome with distinct phenotypes that impact immune function and response to infection. To develop targeted therapeutics, immunophenotyping is needed to identify distinct functional phenotypes of immune cells. In this study, we utilized our Organ-on-Chip assay to categorize sepsis patients into distinct phenotypes using patient data, neutrophil functional analysis, and proteomics. Methods: Following informed consent, neutrophils and plasma were isolated from sepsis patients in the Temple University Hospital ICU (n=45) and healthy control donors (n=7). Human lung microvascular endothelial cells (HLMVEC) were cultured in the Organ-on-Chip and treated with buffer or cytomix ((TNF/IL-1ß/IFNγ). Neutrophil adhesion and migration across HLMVEC in the Organ-on-Chip were used to categorize functional neutrophil phenotypes. Quantitative label-free global proteomics was performed on neutrophils to identify differentially expressed proteins. Plasma levels of sepsis biomarkers and neutrophil extracellular traps (NETs) were determined by ELISA. Results: We identified three functional phenotypes in critically ill ICU sepsis patients based on ex vivo neutrophil adhesion and migration patterns. The phenotypes were classified as: Hyperimmune characterized by enhanced neutrophil adhesion and migration, Hypoimmune that was unresponsive to stimulation, and Hybrid with increased adhesion but blunted migration. These functional phenotypes were associated with distinct proteomic signatures and differentiated sepsis patients by important clinical parameters related to disease severity. The Hyperimmune group demonstrated higher oxygen requirements, increased mechanical ventilation, and longer ICU length of stay compared to the Hypoimmune and Hybrid groups. Patients with the Hyperimmune neutrophil phenotype had significantly increased circulating neutrophils and elevated plasma levels NETs. Conclusion: Neutrophils and NETs play a critical role in vascular barrier dysfunction in sepsis and elevated NETs may be a key biomarker identifying the Hyperimmune group. Our results establish significant associations between specific neutrophil functional phenotypes and disease severity and identify important functional parameters in sepsis pathophysiology that may provide a new approach to classify sepsis patients for specific therapeutic interventions.

Impairments in Blood Pressure Regulation and Cardiac Baroreceptor Sensitivity Among Patients With Heart Failure Supported With Continuous-Flow Left Ventricular Assist Devices.

BACKGROUND: Continuous-flow (CF) left ventricular assist devices (LVADs) improve outcomes for patients with advanced heart failure (HF). However, the lack of a physiological pulse predisposes to side-effects including uncontrolled blood pressure (BP), and there are little data regarding the impact of CF-LVADs on BP regulation. METHODS: Twelve patients (10 males, 60±11 years) with advanced heart failure completed hemodynamic assessment 2.7±4.1 months before, and 4.3±1.3 months following CF-LVAD implantation. Heart rate and systolic BP via arterial catheterization were monitored during Valsalva maneuver, spontaneous breathing, and a 0.05 Hz repetitive squat-stand maneuver to characterize cardiac baroreceptor sensitivity. Plasma norepinephrine levels were assessed during head-up tilt at supine, 30o and 60o. Heart rate and BP were monitored during cardiopulmonary exercise testing. RESULTS: Cardiac baroreceptor sensitivity, determined by Valsalva as well as Fourier transformation and transfer function gain of Heart rate and systolic BP during spontaneous breathing and squat-stand maneuver, was impaired before and following LVAD implantation. Norepinephrine levels were markedly elevated pre-LVAD and improved-but remained elevated post-LVAD (supine norepinephrine pre-LVAD versus post-LVAD: 654±437 versus 323±164 pg/mL). BP increased during cardiopulmonary exercise testing post-LVAD, but the magnitude of change was modest and comparable to the changes observed during the pre-LVAD cardiopulmonary exercise testing. CONCLUSIONS: Among patients with advanced heart failure with reduced ejection fraction, CF-LVAD implantation is associated with modest improvements in autonomic tone, but persistent reductions in cardiac baroreceptor sensitivity. Exercise-induced increases in BP are blunted. These findings shed new light on mechanisms for adverse events such as stroke, and persistent reductions in functional capacity, among patients supported by CF-LVADs. Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT03078972.