Patients admitted to the intensive care unit (ICU) are in need of continuous organ replacement strategies and specialized care, for example because of neurological dysfunction, cardio‐pulmonary instability, liver or kidney failure, trauma, hemorrhagic or septic shock or even preterm birth. The 24‐h nursing and care interventions provided to critically ill patients significantly limit resting and/or recovery phases. Consecutively, the patient's endogenous circadian rhythms are misaligned and disrupted, which in turn may interfere with their critical condition. A more thorough understanding of the complex interactions of circadian effectors and tissue‐specific molecular clocks could therefore serve as potential means for enhancing personalized treatment in critically ill patients, conceivably restoring their circadian network and thus accelerating their physical and neurocognitive recovery. This review addresses the overarching issue of how circadian rhythms are affected and disturbed in critically ill newborns and adults in the ICU, and whether the conflicting external or environmental cues in the ICU environment further promote disruption and thus severity of illness. We direct special attention to the influence of cell‐type specific molecular clocks on with severity of organ dysfunctions such as severity of brain dysfunction, pneumonia‐ or ventilator‐associated lung inflammation, cardiovascular instability, liver and kidney failure, trauma, and septic shock. Finally, we address the potential of circadian rhythm stabilization to enhance and accelerate clinical recovery.

Introduction: Lung-protective ventilation reduced acute respiratory distress syndrome (ARDS) mortality. To minimize ventilator-induced lung injury (VILI), tidal volume is limited, high plateau pressures are avoided, and positive end-expiratory pressure (PEEP) is applied. However, the impact of specific ventilatory patterns on VILI is not well defined. Increasing inspiratory time and thereby the inspiratory/expiratory ratio (I:E ratio) may improve oxygenation, but may also be harmful as the absolute stress and strain over time increase. We thus hypothesized that increasing inspiratory time and I:E ratio aggravates VILI. Methods: VILI was induced in mice by high tidal-volume ventilation (HV_T 34 ml/kg). Low tidal-volume ventilation (LV_T 9 ml/kg) was used in control groups. PEEP was set to 2 cm H₂O, FiO₂ was 0.5 in all groups. HV_T and LV_T mice were ventilated with either I:E of 1:2 (LV_T 1:2, HV_T 1:2) or 1:1 (LV_T 1:1, HV_T 1:1) for 4 hours or until an alternative end point, defined as mean arterial blood pressure below 40 mm Hg. Dynamic hyperinflation due to the increased I:E ratio was excluded in a separate group of animals. Survival, lung compliance, oxygenation, pulmonary permeability, markers of pulmonary and systemic inflammation (leukocyte differentiation in lung and blood, analyses of pulmonary interleukin-6, interleukin-1β, keratinocyte-derived chemokine, monocyte chemoattractant protein-1), and histopathologic pulmonary changes were analyzed. Results: LV_T 1:2 or LV_T 1:1 did not result in VILI, and all individuals survived the ventilation period. HV_T 1:2 decreased lung compliance, increased pulmonary neutrophils and cytokine expression, and evoked marked histologic signs of lung injury. All animals survived. HV_T 1:1 caused further significant worsening of oxygenation, compliance and increased pulmonary proinflammatory cytokine expression, and pulmonary and blood neutrophils. In the HV_T 1:1 group, significant mortality during mechanical ventilation was observed. Conclusion: According to the “baby lung” concept, mechanical ventilation-associated stress and strain in overinflated regions of ARDS lungs was simulated by using high tidal-volume ventilation. Increase of inspiratory time and I:E ratio significantly aggravated VILI in mice, suggesting an impact of a “stress/strain × time product” for the pathogenesis of VILI. Thus increasing the inspiratory time and I:E ratio should be critically considered.

The skin is the largest human organ with a circadian clock that regulates its function. Although circadian rhythms in specific functions are known, rhythms in the proximal clock output, gene expression, in human skin have not been thoroughly explored. This work reports 24 h gene expression rhythms in two skin layers, epidermis and dermis, in a cohort of young, healthy adults, who maintained natural, regular sleep-wake schedules. 10% of the expressed genes showed such diurnal rhythms at the population level, of which only a third differed between the two layers. Amplitude and phases of diurnal gene expression varied more across subjects than layers, with amplitude being more variable than phases. Expression amplitudes in the epidermis were larger and more subject-variable, while they were smaller and more consistent in the dermis. Core clock gene expression was similar across layers at the population-level, but were heterogeneous in their variability across subjects. We also identified small sets of biomarkers for internal clock phase in each layer, which consisted of layer-specific non-core clock genes. This work provides a valuable resource to advance our understanding of human skin and presents a novel methodology to quantify sources of variability in human circadian rhythms.

Rationale: Mechanical ventilation is life-saving but may evoke ventilator-induced lung injury. Objectives: To explore how the circadian clock modulates severity of murine ventilator-induced lung injury via the core clock component BMAL1 in myeloid cells. Methods: Myeloid cell BMAL1-deficient (LysM^Bmal1-/-) or wild type control (LysM^Bmal1+/+) mice were subjected to 4 hours mechanical ventilation (34 mL/kg bodyweight) to induce lung injury. Ventilation was initiated at dawn or dusk, or in complete darkness (circadian time 0 or 12) to determine diurnal and circadian effects. Lung injury was quantified by lung function, pulmonary permeability, blood gas analysis, neutrophil recruitment, inflammatory markers and histology. Neutrophil activation and oxidative burst were analyzed ex vivo. Measurements and Main Results: In diurnal experiments, mice ventilated at dawn exhibited higher permeability and neutrophil recruitment compared to dusk. Experiments at circadian time showed deterioration of pulmonary function, worsening of oxygenation and increased mortality at circadian time 0 compared to circadian time 12. Wild type neutrophils isolated at dawn showed higher activation and ROS production compared to dusk, while these day-night differences were dampened in LysM^Bmal1-/-neutrophils. In LysM^Bmal1-/-mice, circadian variations in VILI severity were dampened and VILI-induced mortality at circadian time 0 was reduced compared with LysM^Bmal1+/+mice. Conclusion: Inflammatory response and lung barrier dysfunction upon mechanical ventilation exhibit diurnal variations, regulated by the circadian clock. LysM^Bmal1-/-mice are less susceptible to ventilation-induced pathology and lack circadian variation of severity compared with LysM^Bmal1+/+mice. Our data suggest that the internal clock in myeloid cells is an important modulator of ventilator-induced lung injury.

Pneumolysin (PLY) is a pore-forming toxin of Streptococcus pneumoniae that contributes substantially to the inflammatory processes underlying pneumococcal pneumonia and lung injury. Host responses against S. pneumoniae are regulated in part by neutrophils and platelets, both individually and in cooperative interaction. Previous studies have shown that PLY can target both neutrophils and platelets, however, the mechanisms by which PLY directly affects these cells and alters their interactions are not completely understood. In this study, we characterize the effects of PLY on neutrophils and platelets and explore the mechanisms by which PLY may induce neutrophil–platelet interactions. In vitro studies demonstrated that PLY causes the formation of neutrophil extracellular traps (NETs) and the release of extracellular vesicles (EVs) from both human and murine neutrophils. In vivo, neutrophil EV (nEV) levels were increased in mice infected with S. pneumoniae. In platelets, treatment with PLY induced the cell surface expression of P-selectin (CD62P) and binding to annexin V and caused a significant release of platelet EVs (pl-EVs). Moreover, PLY-induced nEVs but not NETs promoted platelet activation. The pretreatment of nEVs with proteinase K inhibited platelet activation, indicating that the surface proteins of nEVs play a role in this process. Our findings demonstrate that PLY activates neutrophils and platelets to release EVs and support an important role for neutrophil EVs in modulating platelet functions in pneumococcal infections.

The skin is the largest human organ with a circadian clock that regulates its function. Although circadian rhythms in specific functions are known, rhythms in the proximal clock output, gene expression, in human skin have not been thoroughly explored. This work reports circadian gene expression in two skin layers, epidermis and dermis, in a cohort of young, healthy adults, who maintained natural, regular sleep schedules. 10% of the expressed genes showed rhythms at the population level, of which only a third differed between the two layers. Broadly, expression magnitudes of circadian genes were consistent across subjects in each layer. Amplitude and phases of circadian gene expression, however, varied more across subjects than layers, with amplitude being more variable than phases. Expression amplitudes in the epidermis were larger and more subject-variable, while they were smaller and more consistent in the dermis. Core clock gene expression was similar across layers at the population-level, but were heterogeneous in the their variability across subjects. We used this data to identify small sets of biomarkers for internal clock phase in each layer, which consisted of layer-specific non-core clock genes. This work provides a valuable resource to advance our understanding of human skin to realize the potential of circadian medicine as well as a novel methodology to quantify sources of variability in human circadian rhythms.