Raquel M. Forsythe

The role of lymph factors in lung injury, bone marrow suppression, and endothelial cell dysfunction in a primate model of trauma-hemorrhagic shock.

Studies in rodent models of trauma-hemorrhagic shock (T/HS) have shown that factors contained in the intestinal lymph are responsible for acute lung injury and bone marrow suppression, and that they contribute to a systemic inflammatory state. Because results observed in rodent T/HS models may not fully reflect the response of injured patients, it is necessary to determine if these results can be replicated in primates before the institution of invasive studies in humans. Thus, the three goals of this study were to determine if diversion of thoracic duct lymph reduced T/HS-induced lung injury; to compare the biologic activity of thoracic duct lymph from baboons subjected to T/HS or trauma sham-shock (T/SS); and to compare the biologic activity and composition of plasma from baboons subjected to T/SS, T/HS, and T/HS with thoracic duct lymph drainage. Three groups of baboons were studied: T/SS plus lymph diversion via a thoracic duct catheter, T/HS, and T/HS plus lymph diversion (T/HS-LD). The trauma component consisted of a neck dissection with resection of the proximal clavicle plus a laparotomy. HS was to a mean arterial pressure of 40 mmHg and was maintained at 40 mmHg until the base excess reached −5 mEq or the total shock period reached 3 h. Volume resuscitation was carried out by reinfusing the shed blood plus crystalloids. Before, during, and after the T/HS or T/SS period, blood and lymph samples were obtained for analysis, and lung samples were harvested for measurement of lung wet-to-dry ratio at 5 h after the end of the shock period. Diversion of thoracic duct lymph prevented T/HS-induced lung injury as reflected in lung wet-to-dry weight ratios (T/SS = 4.6 ± 0.5; T/HS+LD = 4.8 ± 0.7; T/HS = 5.4 ± 0.6; P < 0.05). Lymph from the T/HS group collected during the early postshock period was cytotoxic for human endothelial cells (HUVECs; 16% vs. 100% survival in T/SS lymph) and increased HUVEC monolayer permeability almost 2-fold (P < 0.01). T/HS lymph and plasma also suppressed red blood cell (erythroid burst-forming unit) and white blood cell (granulocyte-monocyte colony-forming unit) progenitor cell growth of human bone marrow to ∼50% of control, whereas T/SS lymph and plasma were not suppressive (P < 0.05). Plasma cytokine levels were increased to a similar degree in the two T/HS groups. Thus, in a primate model of T/HS, gut-derived factors carried in the lymph potentiates lung injury and endothelial dysfunction, and suppresses bone marrow progenitor cell growth.

Lipopolysaccharide-induced enterocyte-derived nitric oxide induces intestinal monolayer permeability in an autocrine fashion.

Studies indicate that endotoxin (LPS) causes intestinal injury, increases inducible nitric oxide synthase (iNOS) activity, leads to increased NO production, and promotes bacterial translocation (BT). To investigate the mechanism by which LPS causes gut injury and to test the hypothesis that NO produced by enterocytes promotes gut injury in an autocrine fashion, rat intestinal epithelial cell (IEC-6) monolayers were tested. IEC-6 monolayers grown in a bicameral system were incubated with media or with LPS (25 &mgr;g/mL) and tested for permeability to phenol red, BT, and nitrate/nitrite (NO2/NO3) production. To determine the direct effect of NO on permeability, monolayers were incubated with the NO donor S-nitroso-acetylpenicillinamide (SNAP; 1 mM) and tested for permeability. Next, the protective effects of two NOS inhibitors (L-NMMA and L-NIL) were tested. Finally, to determine if LPS-induced permeability occurs via a poly (ADP-ribose) synthetase- (PARS) dependent pathway, monolayers incubated with LPS alone or with the PARS inhibitor, INH2BP (100 &mgr;M) were tested. LPS significantly increased IEC-6 permeability to phenol red, as well as increased NO2/NO3 by 20-fold (P < 0.001) and increased BT 10-fold (P < 0.001). SNAP mimicked the effect of LPS and significantly increased both permeability to phenol red and BT. Inhibition of iNOS significantly decreased the LPS-induced increase in monolayer permeability and BT (P < 0.05). Monolayers incubated with INH2BP had significantly decreased permeability to phenol red and BT, suggesting that LPS-induced NO production increases monolayer permeability at least in part via a PARS-dependent mechanism. In summary, LPS-induced disruption of monolayer barrier function appears to be related, at least in part, to enterocyte produced NO. This supports the hypothesis that NO produced by LPS-stimulated enterocytes promotes injury in an autocrine fashion and highlights the fact that enterocytes can be a target as well as a producer of NO.

Systolic blood pressure criteria in the National Trauma Triage Protocol for geriatric trauma: 110 is the new 90.

BACKGROUND Undertriage is a concern in geriatric patients. The National Trauma Triage Protocol (NTTP) recognized that systolic blood pressure (SBP) less than 110 mm Hg may represent shock in those older than 65 years. The objective was to evaluate the impact of substituting an SBP of less than 110 mm Hg for the current SBP of less than 90 mm Hg criterion within the NTTP on triage performance and mortality. METHODS Subjects undergoing scene transport in the National Trauma Data Bank (2010–2012) were included. The outcome of trauma center need was defined as Injury Severity Score (ISS) greater than 15, intensive care unit admission, urgent operation, or emergency department death. Geriatric (age > 65 years) and adult (age, 16–65 years) cohorts were compared. Triage characteristics and area under the curve (AUC) were compared between SBP of less than 110 mm Hg and SBP of less than 90 mm Hg. Hierarchical logistic regression was used to determine whether geriatric patients newly triaged positive under this change (SBP, 90–109 mm Hg) have a risk of mortality similar to those triaged positive with SBP of less than 90 mm Hg. RESULTS There were 1,555,944 subjects included. SBP of less than 110 mm Hg had higher sensitivity but lower specificity in geriatric (13% vs. 5%, 93% vs. 99%) and adult (23% vs. 10%, 90% vs. 98%) cohorts. AUC was higher for SBP of less than 110 mm Hg individually in both geriatric and adult (p < 0.01) cohorts. Within the NTTP, the AUC was similar for SBP of less than 110 mm Hg and SBP of less than 90 mm Hg in geriatric subjects but was higher for SBP of less than 90 mm Hg in adult subjects (p < 0.01). Substituting SBP of less than 110 mm Hg resulted in an undertriage reduction of 4.4% with overtriage increase of 4.3% in the geriatric cohort. Geriatric subjects with SBP of 90 mm Hg to 109 mm Hg had an odds of mortality similar to those of geriatric patients with SBP of less than 90 mm Hg (adjusted odds ratio, 1.03; 95% confidence interval, 0.88–1.20; p = 0.71). CONCLUSION SBP of less than 110 mm Hg increases sensitivity. SBP of less than 110 mm Hg has discrimination as good as that of SBP of less than 90 mm Hg, with superior improvements in undertriage relative to overtriage in geriatric patients. Geriatric patients newly triaged to be positive under this change have a risk of mortality similar to those under the current SBP criterion. This change in SBP criteria may be merited in geriatric patients, warranting further study to consider elevation to a Step 1 criterion in the NTTP. LEVEL OF EVIDENCE Diagnostic study, level IV.

The immunomodulatory effects of damage control abdominal packing on local and systemic neutrophil activity.

Background: Damage control laparotomy (DCL) with abdominal packing has become commonplace after major trauma, but the immune consequences of DCL are unknown. Methods: We collected 37 fluid samples from laparotomy pads (LPF) removed from 28 patients 1 hour to 7 days after DCL. Samples from eight patients who underwent serial packing were assayed for their mediator content and effects on neutrophil (PMN) function. Respiratory burst (RB) to N-formyl-methionyl-leucyl-phenylalanine and phorbol myristate acetate (PMA), as well as PMN calcium ([Ca 2+ ] i ) mobilization by GRO-a and platelet-activating factor were studied using dihydrorhodamine and fura-2-acetoxymethyl ester fluorescence. Brief exposure to 20% LPF (LPF20) modeled LPF acting on peritoneal PMNs and 2% LPF (LPF2) modeled the systemic effects on PMNs. Endotoxin (ETX), GRO-a, and leukotriene B4 were assayed by enzyme-linked immunosorbent assay. Data analysis was by analysis of variance with Dunns comparisons or the Mann-Whitney test when indicated. Results: LPF increased N-formyl-methionyl-leucyl-phenylalanine-induced RB from 0.4 ± 0.1 x 10 3 counts per second (control) to 0.7 ± 0.1 (LPF2) to 1.3 ± 0.3 (LPF20) (p < 0.05), with LPF2 increasingly active at later times after injury. PMA-elicited RB was primed only by LPF2 from < 24 hours. Both LPF2 and LPF20 markedly suppressed GRO-a [Ca 2+ ] i flux. Suppression by LPF2 was maximal at < 24 hours, abating after 48 hours. Suppression of GRO-a response was dose dependent: 150 ± 8 nmol/L in control PMNs, 97 ± 19 after LPF2, and 59 ± 4 after LPF20 (all p < 0.05). [Ca 2+ ] i flux after 1 nmol/L platelet-activating factor was only suppressed (from 181 ± 14 nmol/L to 149 ± 15 nmol/L,p < 0.05) by LPF20. LPF contained ETX, GRO-a, and leukotriene B4 at 10- to 20-fold plasma concentration in trauma patients. Conclusion: DCL results in peritoneal ETX and mediator accumulation even when cultures are sterile. LPF exposure primes PMN RB elicited by nonreceptor- (PMA) or receptor-coupled agonists that resist receptor desensitization. Conversely, LPF suppresses PMN responses to agonists that undergo receptor desensitization at high mediator concentrations. PMN dysfunction in such circumstances probably reflects a concomitant priming of some cell functions (e.g., RB) and desensitization of other (receptor-dependent) functions after an exposure to concentrated mediators. Peritoneal mediator production after DCL may be ETX driven, and may contribute to systemic inflammatory response syndrome. DCL trades early hemostasis for later inflammation. This should be considered in planning management strategies.

Hematopoietic failure after hemorrhagic shock is mediated partially through mesenteric lymph

ObjectiveTo determine whether hemorrhagic shock-induced bone marrow failure is mediated by the gut through the production of toxic mesenteric lymph and whether shock-induced bone marrow failure could be prevented by division of the mesenteric lymphatics. DesignProspective, controlled study. SettingUniversity surgical research laboratory. SubjectsMale Sprague-Dawley rats. InterventionsRats were divided into five groups: unmanipulated controls (n = 12), hemorrhagic shock with laparotomy (n = 8), hemorrhagic shock with mesenteric lymph duct ligation (n = 10), sham shock with laparotomy (n = 6), and sham shock with mesenteric lymph duct ligation (n = 7). At either 3 or 6 hrs after resuscitation, bone marrow was obtained for determination of early (cobblestone forming cells) and late (granulocyte-macrophage colony forming unit and erythroid burst forming unit) hematopoietic progenitor cell growth. Parallel cultures were plated with plasma (1% and 2% v/v) from all groups to determine the effect of lymphatic ligation on hematopoiesis. Measurements and Main Results Bone marrow cellularity, cobblestone forming cells, granulocyte-macrophage colony forming unit, and erythroid burst forming unit growth in rats subjected to hemorrhagic with lymph duct ligation were similar to those observed in sham-treated animals and significantly greater than in rats subjected to shock and laparotomy without lymphatic duct ligation. Plasma from rats subjected to shock without lymph ligation was inhibitory to hematopoietic progenitor cell growth. In contrast, this shock-induced inhibition was not observed with plasma obtained from shocked rats that underwent mesenteric lymph ligation. ConclusionsHemorrhagic shock suppresses bone marrow hematopoiesis as measured by a decrease in early and late progenitor cell growth. This suppression appears mediated through mesenteric lymph as the effect is abrogated by mesenteric lymph duct ligation. These data clearly demonstrate a link between the gut and bone marrow failure after hemorrhagic shock

Massive Transfusion: An Evidence-Based Review of Recent Developments

The design and implementation of massive transfusion protocols with ratio-based transfusion of blood and blood products are important and active areas of investigation. A significant yet controversial body of literature exists to support the use of hemostatic resuscitation in massive transfusion and new data to support the use of adjuncts, such as recombinant factor VIIa and tranexamic acid. We review the developments in massive transfusion research during the past 5 years, including protocol implementation, hemostatic resuscitation, the use of tranexamic acid, and goal-directed therapy for coagulopathy. Furthermore, we provide a level of evidence analysis of the data surrounding the use of component therapy and recombinant factor VIIa in massive transfusion, summary recommendations for the various agents of resuscitation, and new methods of goal-directed therapy.

LYMPH FROM A PRIMATE BABOON TRAUMA HEMORRHAGIC SHOCK MODEL ACTIVATES HUMAN NEUTROPHILS

ABSTRACT We have reported that toxic factors in intestinal lymph are responsible for acute lung injury and bone marrow suppression and that they contribute to a systemic inflammatory state based on studies in rodent models of trauma-hemorrhagic shock. Rodent models may not completely reflect the responses of injured patients. Thus, it is important to confirm these findings in primates before applying them to injured human patients with trauma. Thus, we have recently established baboon trauma-hemorrhagic shock (T/HS) and trauma-sham shock (T/SS) models that showed that gut-derived factors carried in the lymph potentiates lung injury and causes human endothelial dysfunction and suppresses human bone marrow progenitor cell growth. Here, we further investigated the effects of these primate lymph samples on human neutrophils. We hypothesized that toxic factors in baboon lymph may prime and/or activate human polymorphonuclear leukocyte (PMN) leading to overproduction of superoxide, thereby contributing to the development of adult respiratory distress syndrome and multiple organ failure. To this effect, we have examined the priming effect of baboon T/HS and T/SS lymph on PMN respiratory burst and expression of adhesion molecule in human neutrophils. The results of these studies indicate that PMN treated with baboon T/HS lymph showed significantly induced respiratory burst responses compared with PMN treated with T/SS lymph or medium when phorbol myristate acetate PMA was applied after lymph pretreatment. Secondly, we found that the expression of CD11b adhesion molecule was increased by incubation with T/HS lymph. These results suggest that baboon lymph from T/HS models can increase respiratory burst and adhesion molecule expression in human PMN, thereby potentially contributing to PMN-mediated organ injury.

Evidence-based improvement of the National Trauma Triage Protocol: The Glasgow Coma Scale versus Glasgow Coma Scale motor subscale

BACKGROUND Ideal triage uses simple criteria to identify severely injured patients. Glasgow Coma Scale motor (GCSm) may be easier for field use and was considered for the National Trauma Triage Protocol (NTTP). This study evaluated performance of the NTTP if GCSm is substituted for the current GCS score ⩽ 13 criterion. METHODS Subjects in the National Trauma Data Bank undergoing scene transport were included. Presence of NTTP physiologic (Step 1) and anatomic (Step 2) criteria was determined. GCSm score ⩽ 5 was defined as a positive criterion. Trauma center need (TCN) was defined as Injury Severity Score (ISS) > 15, intensive care unit admission, urgent operation, or emergency department death. Test characteristics were calculated to predict TCN. Area under the curve was compared between GCSm and GCS scores, individually and within the NTTP. Logistic regression was used to determine the association of GCSm score ⩽ 5 and GCS score ⩽ 13 with TCN after adjusting for other triage criteria. Predicted versus actual TCN was compared. RESULTS There were 811,143 subjects. Sensitivity was lower (26.7% vs. 30.3%), specificity was higher (95.1% vs. 93.1%), and accuracy was similar (66.1% vs. 66.3%) for GCSm score ⩽ 5 compared with GCS score ⩽ 13. Incorporated into the NTTP Steps 1 + 2, GCSm score ⩽ 5 traded sensitivity (60.4% vs. 62.1%) for specificity (67.1% vs. 65.7%) with similar accuracy (64.2% vs. 64.2%) to GCS score ⩽ 13. There was no difference in the area under the curve between GCSm score ⩽ 5 and GCS score ⩽ 13 when incorporated into the NTTP Steps 1 + 2 (p = 0.10). GCSm score ⩽ 5 had a stronger association with TCN (odds ratio, 3.37; 95% confidence interval, 3.27–3.48; p < 0.01) than GCS score ⩽ 13 (odds ratio, 3.03; 95% confidence interval, 2.94–3.13; p < 0.01). GCSm had a better fit of predicted versus actual TCN than GCS at the lower end of the scales. CONCLUSION GCSm score ⩽ 5 increases specificity at the expense of sensitivity compared with GCS score ⩽ 13. When applied within the NTTP, there is no difference in discrimination between GCSm and GCS. GCSm score ⩽ 5 is more strongly associated with TCN and better calibrated to predict TCN. Further study is warranted to explore replacing GCS score ⩽ 13 with GCSm score ⩽ 5 in the NTTP. LEVEL OF EVIDENCE Prognostic study, level III.

Hypothermia in massive transfusion: Have we been paying enough attention to it?

OBJECTIVE The development of acidosis, coagulopathy, and hypothermia has been shown to adversely affect survival after injury. Significant attention has focused on the correction of the early coagulopathy in those requiring massive transfusion (MT). We sought to characterize the importance of temperature as a risk factor for poor outcome relative to the changes in MT resuscitation that have occurred. METHODS Data were obtained from a multicenter prospective cohort study of adults with blunt injury with hemorrhagic shock. MT was defined as 10 U or more of packed red blood cell (PRBC) during 24 hours. The lowest 24-hour temperature was categorized into groups (<34.0°C, 34.1–35.0°C, 35.1–36.0°C, and >36°C). A Kaplan-Meier analysis and a multivariate logistic regression were used to analyze temperature survival differences over time and independent risks of mortality after controlling for all important confounders. RESULTS In the MT cohort (n = 604), as temperature decreased, shock parameters, early coagulopathy, injury severity, and blood component transfusion requirements significantly increased. A Kaplan-Meier comparison revealed a dose-response relationship with a temperature lower than 34°C resulting in the greatest mortality. Logistic regression analysis demonstrated that a temperature lower than 34°C was associated with a greater independent risk of mortality of more than 80% after controlling for differences in shock, coagulopathy, injury severity, and transfusion requirements (odds ratio, 1.87; 95% confidence interval, 1.18–3.0; p = 0.007). When the cohort was stratified into high or low plasma to red blood cell transfusion ratio groups (high fresh frozen plasma [FFP]/PRBC, ≥1:2 vs. low FFP/PRBC, <1:2), regression modeling demonstrated that a temperature lower than 34°C was associated with a twofold higher independent risk of mortality, only in the low FFP/PRBC transfusion group. CONCLUSION A temperature of 34°C seems to define a clinically significant hypothermia in MT. The independent risks of mortality were greatest in those who received a low FFP/PRBC transfusion ratio. These data suggest that the prevention of hypothermia may be as important as addressing early coagulopathy. Further research is required to verify if the prevention or correction of hypothermia improves the outcome of patients requiring MT. LEVEL OF EVIDENCE Prognostic study, level III.

Injury-enhanced calcium mobilization in circulating rat neutrophils models human PMN responses.

G-protein coupled (GPC) chemoattractants are important neutrophil (PMN) activators in human shock and sepsis, acting in part by increasing cytosolic calcium ([Ca2+]i). Rats are widely used as laboratory models of shock and sepsis, but reports of [Ca2+]i flux in circulating rat PMN are rare. Moreover, the [Ca2+]i values reported often differ markedly from human systems. We developed study methods where basal [Ca2+]i values in circulating rat PMN were comparable to human PMN, but rat PMN still mobilized calcium poorly after stimulation. Trauma (laparotomy) did not change rat PMN basal [Ca2+]i, but induced brisk [Ca2+]i responses to chemokine and lipid mediators that approximated human PMN responses. This was associated with marked loading of microsomal calcium stores. Formyl peptides still mobilized calcium less well in rat than human PMN. Normal rat PMN appear to circulate in a less mature or primed form than human PMN. A very limited injury rapidly converts rat PMN to a more activated phenotype. PMN thus activated act quite similar to human PMN in terms of GPC receptor-mediated calcium mobilization. Trauma enhances rat PMN responses to GPC agonists at least in part by loading cell calcium stores.