Acute emergence of the intestinal pathobiome after postinjury pneumonia.

BACKGROUND: Previous preclinical studies have demonstrated sex-specific alterations in the gut microbiome following traumatic injury or sepsis alone; however, the impact of host sex on dysbiosis in the setting of postinjury sepsis acutely is unknown. We hypothesized that multicompartmental injury with subsequent pneumonia would result in host sex-specific dysbiosis. METHODS: Male and proestrus female Sprague-Dawley rats (n = 8/group) were subjected to either multicompartmental trauma (PT) (lung contusion, hemorrhagic shock, cecectomy, bifemoral pseudofracture), PT plus 2-hour daily restraint stress (PT/RS), PT with postinjury day 1 Pseudomonas aeruginosa pneumonia (PT-PNA), PT/RS with pneumonia (PT/RS-PNA), or naive controls. Fecal microbiome was measured on days 0 and 2 using high-throughput 16S rRNA sequencing and Quantitative Insights Into Microbial Ecology 2 bioinformatics analyses. Microbial α-diversity was assessed using Chao1 (number of different unique species) and Shannon (species richness and evenness) indices. ß-diversity was assessed using principal coordinate analysis. Significance was defined as p < 0.05. RESULTS: All groups had drastic declines in the Chao1 (α-diversity) index compared with naive controls ( p < 0.05). Groups PT-PNA and PT/RS-PNA resulted in different ß-diversity arrays compared with uninfected counterparts (PT, PT/RS) ( p = 0.001). Postinjury sepsis cohorts showed a loss of commensal bacteria along with emergence of pathogenic bacteria, with blooms of Proteus in PT-PNA and Escherichia-Shigella group in PT/RS-PNA compared with other cohorts. At day 2, PT-PNA resulted in ß-diversity, which was unique between males and females ( p = 0.004). Microbiome composition in PT-PNA males was dominated by Anaerostipes and Parasuterella , whereas females had increased Barnesiella and Oscillibacter . The PT/RS males had an abundance of Gastranaerophilales and Muribaculaceae . CONCLUSION: Multicompartmental trauma complicated by sepsis significantly diminishes diversity and alters microbial composition toward a severely dysbiotic state early after injury, which varies between males and females. These findings highlight the role of sex in postinjury sepsis and the pathobiome, which may influence outcomes after severe trauma and sepsis.

Multicompartmental trauma alters bone marrow erythroblastic islands.

BACKGROUND: Trauma is associated with widespread inflammation, neuroendocrine activation, and an inadequate bone marrow response to anemia. During late-stage erythropoiesis, erythroid progenitors/erythroblasts form clusters on the surface of specialized bone marrow macrophages where they are supported through terminal differentiation and enucleation. We hypothesized that these erythroblastic islands (EBIs) are adversely impacted by severe trauma. METHODS: Male Sprague-Dawley rats (n = 8/group) were subjected to either multiple injuries (PT) (lung contusion, hemorrhagic shock, cecectomy, and bifemoral pseudofractures), PT plus 2 hours of daily chronic restraint stress (PT/CS), or naive controls. Bone marrow was harvested on days 2 and 7. Nuclear-stained, enriched bone marrow EBIs were fixed and stained for CD71, VCAM-1, and CD163, and confocal images were obtained at 20 times magnification. Numbers of erythroid cells/EBI and ratio of reticulocytes/EBI were counted by a blinded observer. Differences were compared using analysis of variance, with significance defined as p < 0.05. RESULTS: PT and PT/CS had significantly reduced numbers of erythroid cells per EBI on day 2 when compared with naive (PT: 5.9 ± 1.0 cells [ p < 0.05], PT/CS: 6.8 ± 0.8 cells [ p < 0.05] vs. naive: 8.5 ± 0.8 cells). On day 7, the number of erythroid cells/EBI increased following PT (8.3 ± 0.4 cells) but remained reduced following PT/CS (5.9 ± 0.5 cells [ p < 0.05]). This correlated with an increased proportion of reticulocytes/EBI on day 7 following PT, which was not present following PT/CS (PT: 54% [ p < 0.05] vs. PT/CS: 28%). CONCLUSION: Late-stage erythropoiesis was altered following multicompartmental PT early after injury, and these alterations persisted with the addition of daily chronic stress. Alterations in EBI structure and function after severe trauma and critical illness may serve as a promising new area of study to improve mechanistic understanding of persistent anemia after trauma.

NARROWING THE GAP: PRECLINICAL TRAUMA WITH POSTINJURY SEPSIS MODEL WITH INCREASED CLINICAL RELEVANCE.

ABSTRACT: Background : Overall outcomes for trauma patients have improved over time. However, mortality for postinjury sepsis is unchanged. The use of relevant preclinical studies remains necessary to understand mechanistic changes after injury and sepsis at the cellular and molecular level. We hypothesized that a preclinical rodent model of multicompartmental injury with postinjury pneumonia and chronic stress would replicate inflammation and organ injury similar to trauma patients in the intensive care unit. Methods : Male and proestrus female Sprague-Dawley rats ( n = 16/group) were subjected to either polytrauma (PT) (lung contusion, hemorrhagic shock, cecectomy, and bifemoral pseudofracture), PT with daily chronic restraint stress (PT/CS), PT with postinjury day one Pseudomonas pneumonia (PT + PNA), PT/CS with pneumonia (PT/CS + PNA) or naive controls. Weight, white blood cell count, plasma toll-like receptor 4 (TLR4), urine norepinephrine (NE), hemoglobin, serum creatinine, and bilateral lung histology were evaluated. Results : PT + PNA and PT/CS + PNA groups lost more weight compared with those without sepsis (PT, PT/CS) and naive rats ( P < 0.03). Similarly, both PT + PNA and PT/CS + PNA had increased leukocytosis and plasma TLR4 compared with uninfected counterparts. Urine NE was elevated in PT + PNA and PT/CS + PNA compared with naive ( P < 0.03), with PT/CS + PNA exhibiting the highest levels. PT/CS + PNA exhibited worse acute kidney injury with elevated serum creatinine compared with PT/CS ( P = 0.008). PT/CS + PNA right and left lung injury scores were worse than PT + PNA ( P < 0.01). Conclusions : Sepsis, with postinjury pneumonia, induced significant systemic inflammation, organ dysfunction following polytrauma and chronic stress. Advanced animal models that replicate the critically ill human condition will help overcome the classic limitations of previous experimental models and enhance their translational value.

Multicompartmental traumatic injury and the microbiome: Shift to a pathobiome.

BACKGROUND: Previous animal models have demonstrated altered gut microbiome after mild traumatic injury; however, the impact of injury severity and critical illness is unknown. We hypothesized that a rodent model of severe multicompartmental injuries and chronic stress would demonstrate microbiome alterations toward a "pathobiome" characterized by an overabundance of pathogenic organisms, which would persist 1 week after injury. METHODS: Male Sprague-Dawley rats (n = 8 per group) were subjected to either multiple injuries (PT) (lung contusion, hemorrhagic shock, cecectomy, and bifemoral pseudofractures), PT plus daily chronic restraint stress for 2 hours (PT/CS), or naive controls. Fecal microbiome was measured on days 0, 3, and 7 using high-throughput 16S rRNA sequencing and Quantitative Insights Into Microbial Ecology 2 bioinformatics analysis. Microbial α diversity was assessed using Chao1 and Shannon indices, and ß diversity with principle coordinate analysis. Intestinal permeability was evaluated by plasma occludin; ileum and descending colon tissues were reviewed for injury. Analyses were performed in GraphPad (GraphPad Software, La Jolla, CA) and R (R Foundation for Statistical Computing, Vienna, Austria), with significance defined as p < 0.05. RESULTS: There were significant alterations in ß diversity at day 3 and between all groups. By day 3, both PT and PT/CS demonstrated significantly depleted bacterial diversity (Chao1) ( p = 0.01 and p = 0.001, respectively) versus naive, which persisted up to day 7 in PT/CS only ( p = 0.001). Anaerostipes and Rothia dominated PT and Lactobacillus bloomed in PT/CS cohorts by day 7. Plasma occludin was significantly elevated in PT/CS compared with naive ( p = 0.04), and descending colon of both PT and PT/CS showed significantly higher injury compared with naive ( p = 0.005, p = 0.006). CONCLUSIONS: Multiple injuries with and without chronic stress induces significant alterations in microbiome diversity and composition within 3 days; these changes are more prominent and persist for 1 week postinjury with stress. This rapid and persistent transition to a "pathobiome" phenotype represents a critical phenomenon that may influence outcomes after severe trauma and critical illness.

Anemia Recovery after Trauma: A Longitudinal Study.

Background: Post-injury inflammation and its correlation with anemia recovery after severe trauma is poorly described. Severe injury induces a systemic inflammatory response associated with critical illness and organ dysfunction, including disordered hematopoiesis, and anemia. This study sought to characterize the resolution of post-injury inflammation and anemia to identify risk factors associated with persistence of anemia. Patients and Methods: This single-institution study prospectively enrolled 73 trauma patients with an injury severity score >15, hemorrhagic shock, and a lower extremity long bone orthopedic injury. Blood was obtained at enrollment and after 14 days, one, three, and six months. Analytes were compared using Mann-Whitney U tests with correction for multiple comparisons. Results: Median age was 45 years and Injury Severity Score (ISS) was 27, with anemia rates of 97% at two weeks, 80% at one month, 52% at three months, and 30% at six months. Post-injury elevations in erythropoietin, interleukin-6, and C-reactive protein resolved by one month, three months, and six months, respectively. Median granulocyte colony-stimulating factor (G-CSF) and tumor necrosis factor (TNF)-α concentrations remained elevated throughout the six-month follow-up period. Patients with persistent anemia had longer intensive care unit and hospital lengths of stay, more infectious complications, and received more packed red blood cell transfusions compared to those with early anemia recovery. Conclusions: Severe trauma is associated with a prolonged inflammatory response, which is associated with increased transfusion requirements, lengths of stay, and persistent anemia. Further analysis is needed to identify correlations between prolonged inflammation and clinical outcomes after discharge.