Anthony J. Lewis

Biology and Metabolism of Sepsis: Innate Immunity, Bioenergetics, and Autophagy.

Sepsis is a complex, heterogeneous physiologic condition that represents a significant public health concern. While many insights into the pathophysiology of sepsis have been elucidated over the past decades of research, important questions remain. This article serves as a review of several important areas in sepsis research. Understanding the innate immune response has been at the forefront as of late, especially in the context of cytokine-directed therapeutic trials. Cellular bioenergetic changes provide insight into the development of organ dysfunction in sepsis. Autophagy and mitophagy perform crucial cell housekeeping and stress response functions. Finally, age-related changes and their potential impact on the septic response are reviewed.

Use of Biotelemetry to Define Physiology-Based Deterioration Thresholds in a Murine Cecal Ligation and Puncture Model of Sepsis.

Objectives:Murine models of critical illness are commonly used to test new therapeutic interventions. However, these interventions are often administered at fixed time intervals after the insult, perhaps ignoring the inherent variability in magnitude and temporality of the host response. We propose to use wireless biotelemetry monitoring to define and validate criteria for acute deterioration and generate a physiology-based murine cecal ligation and puncture model that is more similar to the conduct of human trials of sepsis. Design:Laboratory and animal research. Setting:University basic science laboratory. Subjects:Male C57BL/6 mice. Interventions:Mice underwent cecal ligation and puncture, and an HD-X11 wireless telemetry monitor (Data Sciences International) was implanted that enabled continuous, real-time measurement of heart rate, core temperature, and mobility. We performed a population-based analysis to determine threshold criteria that met face validity for acute physiologic deterioration. We assessed construct validity by temporally matching mice that met these acute physiologic deterioration thresholds with mice that had not yet met deterioration threshold. We analyzed matched blood samples for blood gas, inflammatory cytokine concentration, cystatin C, and alanine aminotransferase. Measurements and Main Results:We observed that a 10% reduction in both heart rate and temperature sustained for greater than or equal to 10 minutes defined acute physiologic deterioration. There was significant variability in the time to reach acute deterioration threshold across mice, ranging from 339 to 529 minutes after cecal ligation and puncture. We found adequate construct validity, as mice that met criteria for acute deterioration had significantly worse shock, systemic inflammation (elevated tumor necrosis factor-&agr;, p = 0.003; interleukin-6, p = 0.01; interleukin-10, p = 0.005), and acute kidney injury when compared with mice that had not yet met acute deterioration criteria. Conclusions:We defined a murine threshold for acute physiologic deterioration after cecal ligation and puncture that has adequate face and construct validity. This model may enable a more physiology-based model for evaluation of novel therapeutics in critical illness.

Blue light reduces organ injury from ischemia and reperfusion

Significance It is well established that light regulates mammalian biology. And yet, we have been unable to define and thus harness the underlying mechanisms so as to apply them to alter the course of human disease. In this study we determine that the spectrum of light is a critical determinant of its effect on critical illness. We show that an acute and short (24 h) exposure to high-illuminance (1,400 lx) blue spectrum (peak 442 nm) light prior to ischemia/reperfusion (I/R) significantly attenuates the degree of organ injury. Our characterization of the biological mechanisms through which blue light beneficially alters the cellular response to I/R provides an opportunity to develop novel therapeutics for the prevention and treatment of many diseases. Evidence suggests that light and circadian rhythms profoundly influence the physiologic capacity with which an organism responds to stress. However, the ramifications of light spectrum on the course of critical illness remain to be determined. Here, we show that acute exposure to bright blue spectrum light reduces organ injury by comparison with bright red spectrum or ambient white fluorescent light in two murine models of sterile insult: warm liver ischemia/reperfusion (I/R) and unilateral renal I/R. Exposure to bright blue light before I/R reduced hepatocellular injury and necrosis and reduced acute kidney injury and necrosis. In both models, blue light reduced neutrophil influx, as evidenced by reduced myeloperoxidase (MPO) within each organ, and reduced the release of high-mobility group box 1 (HMGB1), a neutrophil chemotactant and key mediator in the pathogenesis of I/R injury. The protective mechanism appeared to involve an optic pathway and was mediated, in part, by a sympathetic (β3 adrenergic) pathway that functioned independent of significant alterations in melatonin or corticosterone concentrations to regulate neutrophil recruitment. These data suggest that modifying the spectrum of light may offer therapeutic utility in sterile forms of cellular injury.

Calcium/calmodulin-dependent protein kinase regulates the PINK1/Parkin and DJ-1 pathways of mitophagy during sepsis.

During sepsis and shock states, mitochondrial dysfunction occurs. Consequently, adaptive mechanisms, such as fission, fusion, and mitophagy, are induced to eliminate damaged portions or entire dysfunctional mitochondria. The regulatory PINK1/Parkin and DJ‐1 pathways are strongly induced by mitochondrial depolarization, although a direct link between loss of mitochondrial membrane potential (ΔΨ) and mitophagy has not been identified. Mitochondria also buffer Ca2+, and their buffering capacity is dependent on ΔΨ. Here, we characterize a role for calcium/calmodulin‐dependent protein kinase (CaMK) I in the regulation of these mechanisms. Loss of ΔΨ with either pharmacologic depolarization or LPS leads to Ca2+‐dependent mitochondrial recruitment and activation of CaMKI that precedes the colocalization of PINK1/Parkin and DJ‐1. CaMKI is required and serves as both a PINK1 and Parkin kinase. The mechanisms operate in both immune and nonimmune cells and are induced in in vivo models of endotoxemia, sepsis, and hemorrhagic shock. These data support the idea that CaMKI links mitochondrial stress with the PINK1/Parkin and DJ‐1 mechanisms of mitophagy.—Zhang, X., Yuan, D., Sun, Q., Xu, L., Lee, E., Lewis, A. J., Zuckerbraun, B. S., Rosengart, M. R. Calcium/calmodulin‐dependent protein kinase regulates the PINK1/Parkin and DJ‐1 pathways of mitophagy during sepsis. FASEB J. 31, 4382‐4395 (2017). www.fasebj.org

1379: BLUE LIGHT THERAPY MODULATES THE SYSTEMIC RESPONSE TO SEPSIS IN MICE AND HUMANS.

Crit Care Med 2016 • Volume 44 • Number 12 (Suppl.) clinical outcomes and add significantly to healthcare burden and cost. Little data is available on CDI complicating patients hospitalized with septic shock (SS). We sought to determine the prevalence of CDI in hospitalized patients with SS and the impact of CDI on clinical outcomes in these patients. Methods: The National Inpatient Sample (NIS) was used to identify hospitalizations from 2007 to 2013 of adults (age > 18 years) with SS. Septic shock was identified by either ICD9-CM code for SS or vasopressor use associated with ICD-9 codes of sepsis, severe sepsis, septicemia, bacteremia, or fungemia. CDI was identified by secondary diagnosis ICD-9-CM code. Demographic data was obtained from NIS database. Primary outcome was prevalence of CDI among patients with SS. Secondary outcomes were: mortality and hospital length of stay (LOS) associated with CDI. Results: There were 2,031,739 hospitalizations with SS between 2007 and 2013. CDI was present in 8.2% of patients with SS. The prevalence of CDI in SS remained stable between 2007 and 2013. Patients with CDI were older (mean age 69.8 (+ 14.7 vs 66.8 (+ 15.9) years, p <0.001). The in-hospital mortality of SS complicated by CDI was comparable to those without CDI (37.1% versus 37.0%, p=0.48). The median LOS was significantly longer for patients with CDI (13 vs 9 days, p<0.001). Prolonged LOS (>75th percentile of LOS for SS cases, 17 days) was observed in 36.9% of SS patients with CDI vs 22.7% without CDI (p<0.001). After adjusting for age, gender, race, and Charlson Comorbidity Index; the odds of prolonged LOS with presence of CDI was significantly greater (OR 2.11, 95%CI 2.06 – 2.15, p<0.001). Conclusions: CDI complicating septic shock is associated with a significant increase in length of stay but no increase in in-hospital mortality.

Reducing Animal Use with a Biotelemetry-Enhanced Murine Model of Sepsis

Animal models of sepsis exhibit considerable variability in the temporal development of the physiologic response, which reduces the power of studies, particularly if interventions are tested at arbitrary time points. We developed a biotelemetry-based model of cecal ligation and puncture (CLP) that standardizes the testing of time-sensitive therapies to specific criteria of physiologic deterioration. In this study we seek to further define the variability in physiologic response to CLP sepsis and conduct a cost analysis detailing the potential for reducing animal usage. We have further characterized the variability in physiologic response after CLP in mice and determined peaks in the temporal distribution of points of physiologic decline. Testing therapies at physiologic thresholds reduces the variability found in historical fixed time-based models. Though initial cost is higher with biotelemetry, this is eventually offset by the significantly reduced number of mice needed to conduct physiologically relevant sepsis experiments.