Rami A. Namas

Sepsis: Something old, something new, and a systems view

Sepsis is a clinical syndrome characterized by a multisystem response to a microbial pathogenic insult consisting of a mosaic of interconnected biochemical, cellular, and organ-organ interaction networks. A central thread that connects these responses is inflammation that, while attempting to defend the body and prevent further harm, causes further damage through the feed-forward, proinflammatory effects of damage-associated molecular pattern molecules. In this review, we address the epidemiology and current definitions of sepsis and focus specifically on the biologic cascades that comprise the inflammatory response to sepsis. We suggest that attempts to improve clinical outcomes by targeting specific components of this network have been unsuccessful due to the lack of an integrative, predictive, and individualized systems-based approach to define the time-varying, multidimensional state of the patient. We highlight the translational impact of computational modeling and other complex systems approaches as applied to sepsis, including in silico clinical trials, patient-specific models, and complexity-based assessments of physiology.

Temporal Patterns of Circulating Inflammation Biomarker Networks Differentiate Susceptibility to Nosocomial Infection Following Blunt Trauma in Humans.

BACKGROUND Severe traumatic injury can lead to immune dysfunction that renders trauma patients susceptible to nosocomial infections (NI) and prolonged intensive care unit (ICU) stays. We hypothesized that early circulating biomarker patterns following trauma would correlate with sustained immune dysregulation associated with NI and remote organ failure. METHODS In a cohort of 472 blunt trauma survivors studied over an 8-year period, 127 patients (27%) were diagnosed with NI versus 345 trauma patients without NI. To perform a pairwise, case-control study with 1:1 matching, 44 of the NI patients were compared with 44 no-NI trauma patients selected by matching patient demographics and injury characteristics. Plasma obtained upon admission and over time were assayed for 26 inflammatory mediators and analyzed for the presence of dynamic networks. RESULTS Significant differences in ICU length of stay (LOS), hospital LOS, and days on mechanical ventilation were observed in the NI patients versus no-NI patients. Although NI was not detected until day 7, multiple mediators were significantly elevated within the first 24 hours in patients who developed NI. Circulating inflammation biomarkers exhibited 4 distinct dynamic patterns, of which 2 clearly distinguish patients destined to develop NI from those who did not. Mediator network connectivity analysis revealed a higher, coordinated degree of activation of both innate and lymphoid pathways in the NI patients over the initial 24 hours. CONCLUSIONS These studies implicate unique dynamic immune responses, reflected in circulating biomarkers that differentiate patients prone to persistent critical illness and infections following injury, independent of mechanism of injury, injury severity, age, or sex.

Central Role for MCP-1/CCL2 in Injury-Induced Inflammation Revealed by In Vitro , In Silico , and Clinical Studies

The translation of in vitro findings to clinical outcomes is often elusive. Trauma/hemorrhagic shock (T/HS) results in hepatic hypoxia that drives inflammation. We hypothesize that in silico methods would help bridge in vitro hepatocyte data and clinical T/HS, in which the liver is a primary site of inflammation. Primary mouse hepatocytes were cultured under hypoxia (1% O2) or normoxia (21% O2) for 1–72 h, and both the cell supernatants and protein lysates were assayed for 18 inflammatory mediators by Luminex™ technology. Statistical analysis and data-driven modeling were employed to characterize the main components of the cellular response. Statistical analyses, hierarchical and k-means clustering, Principal Component Analysis, and Dynamic Network Analysis suggested MCP-1/CCL2 and IL-1α as central coordinators of hepatocyte-mediated inflammation in C57BL/6 mouse hepatocytes. Hepatocytes from MCP-1-null mice had altered dynamic inflammatory networks. Circulating MCP-1 levels segregated human T/HS survivors from non-survivors. Furthermore, T/HS survivors with elevated early levels of plasma MCP-1 post-injury had longer total lengths of stay, longer intensive care unit lengths of stay, and prolonged requirement for mechanical ventilation vs. those with low plasma MCP-1. This study identifies MCP-1 as a main driver of the response of hepatocytes in vitro and as a biomarker for clinical outcomes in T/HS, and suggests an experimental and computational framework for discovery of novel clinical biomarkers in inflammatory diseases.

Hemoadsorption reprograms inflammation in experimental gram-negative septic peritonitis: insights from in vivo and in silico studies.

Improper compartmentalization of the inflammatory response leads to systemic inflammation in sepsis. Hemoadsorption (HA) is an emerging approach to modulate sepsis-induced inflammation. We sought to define the effects of HA on inflammatory compartmentalization in Escherichia coli-induced fibrin peritonitis in rats. Hypothesis: HA both reprograms and recompartmentalizes inflammation in sepsis. Sprague Dawley male rats were subjected to E. coli peritonitis and, after 24 h, were randomized to HA or sham treatment (sepsis alone). Venous blood samples collected at 0, 1, 3 and 6 h (that is, 24–30 h of total experimental sepsis), and peritoneal samples collected at 0 and 6 h, were assayed for 14 cytokines along with NO2−/NO3−. Bacterial counts were assessed in the peritoneal fluid at 0 and 6 h. Plasma tumor necrosis factor (TNF)-α, interleukin (IL)-6, CXCL-1, and CCL2 were significantly reduced in HA versus sham. Principal component analysis (PCA) suggested that inflammation in sham was driven by IL-6 and TNF-α, whereas HA-associated inflammation was driven primarily by TNF-α, CXCL-1, IL-10 and CCL2. Whereas peritoneal bacterial counts, plasma aspartate transaminase levels and peritoneal IL-5, IL-6, IL-18, interferon (IFN)-γ and NO2−/NO3− were significantly lower, both CXCL-1 and CCL2 as well as the peritoneal-to-plasma ratios of TNF-α, CXCL-1 and CCL2 were significantly higher in HA versus sham, suggesting that HA-induced inflammatory recompartmentalization leads to the different inflammatory drivers discerned in part by PCA. In conclusion, this study demonstrates the utility of combined in vivo/in silico methods and suggests that HA exerts differential effects on mediator gradients between local and systemic compartments that ultimately benefit the host.

Inducible protein-10, a potential driver of neurally controlled interleukin-10 and morbidity in human blunt trauma.

Objective:Blunt trauma and traumatic spinal cord injury induce systemic inflammation that contributes to morbidity. Dysregulated neural control of systemic inflammation postinjury is likely exaggerated in patients with traumatic spinal cord injury. We used in silico methods to discern dynamic inflammatory networks that could distinguish systemic inflammation in traumatic spinal cord injury from blunt trauma. Design:Retrospective study. Settings:Tertiary care institution. Patients:Twenty-one severely injured thoracocervical traumatic spinal cord injury patients and matched 21 severely injured blunt trauma patients without spinal cord injury. Intervention:None. Measurements and Main Results:Serial blood samples were obtained from days 1 to 14 postinjury. Twenty-four plasma inflammatory mediators were quantified. Statistical significance between the two groups was determined by two-way analysis of variance. Dynamic Bayesian network inference was used to suggest dynamic connectivity and central inflammatory mediators. Circulating interleukin-10 was significantly elevated in thoracocervical traumatic spinal cord injury group versus non–spinal cord injury group, whereas interleukin-1&bgr;, soluble interleukin-2 receptor-&agr;, interleukin-4, interleukin-5, interleukin-7, interleukin-13, interleukin-17, macrophage inflammatory protein 1&agr; and 1&bgr;, granulocyte-macrophage colony-stimulating factor, and interferon-&ggr; were significantly reduced in traumatic spinal cord injury group versus non–spinal cord injury group. Dynamic Bayesian network suggested that post-spinal cord injury interleukin-10 is driven by inducible protein-10, whereas monocyte chemotactic protein-1 was central in non–spinal cord injury dynamic networks. In a separate validation cohorts of 356 patients without spinal cord injury and 85 traumatic spinal cord injury patients, individuals with plasma inducible protein-10 levels more than or equal to 730 pg/mL had significantly prolonged hospital and ICU stay and days on mechanical ventilator versus patients with plasma inducible protein-10 level less than 730 pg/mL. Conclusion:This is the first study to compare the dynamic systemic inflammatory responses of traumatic spinal cord injury patients versus patients without spinal cord injury, suggesting a key role for inducible protein-10 in driving systemic interleukin-10 and morbidity and highlighting the potential utility of in silico tools to identify key inflammatory drivers.

Trauma in silico: Individual-specific mathematical models and virtual clinical populations

A mathematical model of the human response to trauma replicates individual patient outcomes but predicts unexpected results in populations. A virtual large sample size Severe trauma or bleeding evokes an all-hands-on-deck immune response. When properly orchestrated, the myriad cytokines and peptides help to heal the patient. However, this process can easily go awry, and administering the right drug to compensate is a challenge. Brown and colleagues mathematically modeled the complicated immune responses from the data of 33 blunt trauma patients and then generated a larger cohort of 10,000 virtual trauma patients. This large virtual cohort predicted the reactions of smaller validation cohorts, but the surprise was that understanding the response details in a single patient did not predict how the population would act. The author’s virtual clinical trial indicated that inhibition of interleukin-6 (IL-6) produced a small survival benefit, whereas IL-1β inhibition did not help much and tumor necrosis factor–α made things worse. Trauma-induced critical illness is driven by acute inflammation, and elevated systemic interleukin-6 (IL-6) after trauma is a biomarker of adverse outcomes. We constructed a multicompartment, ordinary differential equation model that represents a virtual trauma patient. Individual-specific variants of this model reproduced both systemic inflammation and outcomes of 33 blunt trauma survivors, from which a cohort of 10,000 virtual trauma patients was generated. Model-predicted length of stay in the intensive care unit, degree of multiple organ dysfunction, and IL-6 area under the curve as a function of injury severity were in concordance with the results from a validation cohort of 147 blunt trauma patients. In a subcohort of 98 trauma patients, those with high–IL-6 single-nucleotide polymorphisms (SNPs) exhibited higher plasma IL-6 levels than those with low IL-6 SNPs, matching model predictions. Although IL-6 could drive mortality in individual virtual patients, simulated outcomes in the overall cohort were independent of the propensity to produce IL-6, a prediction verified in the 98-patient subcohort. In silico randomized clinical trials suggested a small survival benefit of IL-6 inhibition, little benefit of IL-1β inhibition, and worse survival after tumor necrosis factor–α inhibition. This study demonstrates the limitations of extrapolating from reductionist mechanisms to outcomes in individuals and populations and demonstrates the use of mechanistic simulation in complex diseases.

Insights into the Role of Chemokines, Damage-Associated Molecular Patterns, and Lymphocyte-Derived Mediators from Computational Models of Trauma-Induced Inflammation

SIGNIFICANCE Traumatic injury elicits a complex, dynamic, multidimensional inflammatory response that is intertwined with complications such as multiple organ dysfunction and nosocomial infection. The complex interplay between inflammation and physiology in critical illness remains a challenge for translational research, including the extrapolation to human disease from animal models. RECENT ADVANCES Over the past decade, we and others have attempted to decipher the biocomplexity of inflammation in these settings of acute illness, using computational models to improve clinical translation. In silico modeling has been suggested as a computationally based framework for integrating data derived from basic biology experiments as well as preclinical and clinical studies. CRITICAL ISSUES Extensive studies in cells, mice, and human blunt trauma patients have led us to suggest (i) that while an adequate level of inflammation is required for healing post-trauma, inflammation can be harmful when it becomes self-sustaining via a damage-associated molecular pattern/Toll-like receptor-driven feed-forward circuit; (ii) that chemokines play a central regulatory role in driving either self-resolving or self-maintaining inflammation that drives the early activation of both classical innate and more recently recognized lymphoid pathways; and (iii) the presence of multiple thresholds and feedback loops, which could significantly affect the propagation of inflammation across multiple body compartments. FUTURE DIRECTIONS These insights from data-driven models into the primary drivers and interconnected networks of inflammation have been used to generate mechanistic computational models. Together, these models may be used to gain basic insights as well as serving to help define novel biomarkers and therapeutic targets.

Impact of Injury Severity on Dynamic Inflammation Networks Following Blunt Trauma

ABSTRACT Introduction: Clinical outcomes following trauma depend on the extent of injury and the host’s response to injury, along with medical care. We hypothesized that dynamic networks of systemic inflammation manifest differently as a function of injury severity in human blunt trauma. Study Design: From a cohort of 472 blunt trauma survivors studied following institutional review board approval, three Injury Severity Score (ISS) subcohorts were derived after matching for age and sex: mild ISS (49 patients [33 males and 16 females, aged 42 ± 1.9 years; ISS 9.5 ± 0.4]); moderate ISS (49 patients [33 males and 16 females, aged 42 ± 1.9; ISS 19.9 ± 0.4]), and severe ISS (49 patients [33 males and 16 females, aged 42 ± 2.5 years; ISS 33 ± 1.1]). Multiple inflammatory mediators were assessed in serial blood samples. Dynamic Bayesian Network inference was utilized to infer causal relationships based on probabilistic measures. Results: Intensive care unit length of stay, total length of stay, days on mechanical ventilation, Marshall Multiple Organ Dysfunction score, prevalence of prehospital hypotension and nosocomial infection, and admission lactate and base deficit were elevated as a function of ISS. Multiple circulating inflammatory mediators were significantly elevated in severe ISS versus moderate or mild ISS over both the first 24 h and out to 7 days after injury. Dynamic Bayesian Network suggested that interleukin 6 production in severe ISS was affected by monocyte chemotactic protein 1/CCL2, monokine inducible by interferon &ggr; (MIG)/CXCL9, and IP-10/CXCL10; by monocyte chemotactic protein 1/CCL2 and MIG/CXCL9 in moderate ISS; and by MIG/CXCL9 alone in mild ISS over 7 days after injury. Conclusions: Injury Severity Score correlates linearly with morbidity, prevalence of infection, and early systemic inflammatory connectivity of chemokines to interleukin 6.

Prehospital Hypotension Is Associated With Altered Inflammation Dynamics and Worse Outcomes Following Blunt Trauma in Humans

Objective:To define the impact of prehospital hypotension on the dynamic, systemic acute inflammatory response to blunt trauma. Design:Retrospective study. Settings:Tertiary care institution. Patients:Twenty-two hypotensive blunt trauma patients matched with 28 normotensive blunt trauma patients. Intervention:None. Measurements and Main Results:From a cohort of 472 blunt trauma survivors studied following institutional review board approval, two stringently matched subcohorts were derived. Twenty-two patients who sustained prehospital hypotension following blunt trauma (15 males and 7 females; age, 45 ± 3.8; Injury Severity Score, 20.7 ± 1.8) were matched with 28 normotensive trauma patients (20 males and 8 females; age, 46.1 ± 2.5; Injury Severity Score, 20.8 ± 1.3). Serial blood samples (three samples within the first 24 hr and then from days 1 to 7 postinjury) were assessed for 24 inflammatory mediators using Luminex, and No2–/No3– was measured using the nitrate reductase/Griess assay. Two-way analysis of variance was used to compare groups. Dynamic Bayesian Network inference was used to infer causal relationships based on probabilistic measures. Statistically significant differences were observed in ICU length of stay, total length of stay, days on mechanical ventilator, and Marshall Multiple Organ Dysfunction score between hypotensive and normotensive patients. Shock markers (shock index, pH, lactate, and base deficit) were significantly altered in hypotensive patients. Plasma levels of chemokines (monocyte chemotactic protein-1/CCL2, inducible protein-10/CXCL10, macrophage inflammatory protein-1&agr;/CCL3, and interleukin-8/CCL8) and cytokines (interleukin-6, interleukin-10, interleukin-17, granulocyte-macrophage colony-stimulating factor, interleukin-1&bgr;, and interleukin-7) as well as soluble interleukin-2 receptor-&agr; were significantly elevated over the first 7 days postinjury in the hypotensive versus normotensive patients. Dynamic Bayesian Network suggested that the chemokines monocyte chemotactic protein-1/CCL2 and monokine induced by gamma interferon/CXCL9 in the hypotensive and normotensive patients, respectively, affect plasma interleukin-6 levels differentially in the initial 24 hours postinjury. Conclusions:Studies in stringently matched patient cohorts suggest that an episode of prehospital hypotension post trauma leads to early, dynamic reprogramming of systemic inflammation (including differential upstream regulation of interleukin-6), which is associated with worse outcomes.

Racial disparities and sex-based outcomes differences after severe injury.

BACKGROUND Controversy exists about the mechanisms responsible for sex-based outcomes differences post-injury. X-chromosome-linked immune response pathway polymorphisms represent a potential mechanism resulting in sex-based outcomes differences post-injury. The prevalence of these variants is known to differ across race. We sought to characterize racial differences and the strength of any sex-based dimorphism post-injury. STUDY DESIGN A retrospective analysis was performed using data derived from the National Trauma Data Bank 7.1 (2002-2006). Blunt-injured adult (older than 15 years) patients, surviving >24 hours and with an Injury Severity Score >16 were analyzed (n = 244,371). Patients were stratified by race (Caucasian, black, Hispanic, Asian) and multivariable regression analysis was used to characterize the risk of mortality and the strength of protection associated with sex (female vs male). RESULTS When stratified by race, multivariable models demonstrated Caucasian females had an 8.5% lower adjusted risk of mortality (odds ratio [OR] = 0.91; 95% CI, 0.88-0.95; p < 0.001) relative to Caucasian males, with no significant association found for Hispanics or blacks. An exaggerated survival benefit was afforded to Asian females relative to Asian males, having a >40% lower adjusted risk of mortality (OR = 0.59; 95% CI, 0.44-78; p < 0.001). Asian males had a >75% higher adjusted risk of mortality relative to non-Asian males (OR = 1.77; 95% CI, 1.5-2.0; p < 0.001), and no significant difference in the mortality risk was found for Asian females relative to non-Asian females. CONCLUSIONS These results suggest that Asian race is associated with sex-based outcomes differences that are exaggerated, resulting from worse outcomes for Asian males. These racial disparities suggest a negative male X-chromosome-linked effect as the mechanism responsible for these sex-based outcomes differences.