Postinjury multiple organ failure in polytrauma: more frequent and potentially less deadly with less crystalloid.

BACKGROUND: Recently, retrospective registry-based studies have reported the decreasing incidence and increasing mortality of postinjury multiple organ failure (MOF). We aimed to describe the current epidemiology of MOF following the introduction of haemostatic resuscitation. METHODS: A 10-year prospective cohort study was undertaken at a Level-1 Trauma Centre-based ending in December 2015. Inclusion criteria age ≥ 16 years, Injury Severity Score (ISS) > 15, Abbreviated Injury Scale (AIS) Head < 3 and survived > 48 h. Demographics, physiological and shock resuscitation parameters were collected. The primary outcome was MOF defined by a Denver Score > 3. SECONDARY OUTCOMES: intensive care unit length of stay (ICU LOS), ventilation days and mortality. RESULTS: Three hundred and forty-seven patients met inclusion criteria (age 48 ± 20; ISS 30 ± 11, 248 (71%) were males and 23 (6.6%) patients died. The 74 (21%) MOF patients (maximum Denver Score: 5.5 ± 1.8; Duration; 5.6 ± 5.8 days) had higher ISS (32 ± 11 versus 29 ± 11) and were older (54 ± 19 versus 46 ± 20 years) than non-MOF patients. Mean daily Denver scores adjusted for age, sex, MOF and ISS did not change over time. Crystalloid usage decreased over the 10-year period (p value < 0.01) and PRBC increased (p value < 0.01). Baseline cumulative incidence of MOF at 28 days was 9% and competing risk analyses showed that incidence of MOF increased over time (subdistribution hazard ratio 1.14, 95% CI 1.04 to 1.23, p value < 0.01). Mortality risk showed no temporal change. ICU LOS increased over time (subdistribution hazard ratio 0.95, 95% CI 0.92 to 0.98, p value < 0.01). Ventilator days increased over time (subdistribution hazard ratio 0.94, 95% CI 0.9 to 0.97, p value < 0.01). CONCLUSION: The epidemiology of MOF continues to evolve. Our prospective cohort suggests an ageing population with increasing incidence of MOF, particularly in males, with little changes in injury or shock parameters, who are being resuscitated with less crystalloids, stay longer on ICU without improvement in survival.

The Diagnostic, Therapeutic and Prognostic Relevance of Neutrophil Extracellular Traps in Polytrauma.

Neutrophil extracellular traps (NETs) represent a recently discovered polymorphonuclear leukocyte-associated ancient defence mechanism, and they have also been identified as part of polytrauma patients' sterile inflammatory response. This systematic review aimed to determine the clinical significance of NETs in polytrauma, focusing on potential prognostic, diagnostic and therapeutic relevance. The methodology covered all major databases and all study types, but was restricted to polytraumatised humans. Fourteen studies met the inclusion criteria, reporting on 1967 patients. Ten samples were taken from plasma and four from whole blood. There was no standardisation of methodology of NET detection among plasma studies; however, of all the papers that included a healthy control NET, proxies were increased. Polytrauma patients were consistently reported to have higher concentrations of NET markers in peripheral blood than those in healthy controls, but their diagnostic, therapeutic and prognostic utility is equivocal due to the diverse study population and methodology. After 20 years since the discovery of NETs, their natural history and potential clinical utility in polytrauma is undetermined, requiring further standardisation and research.

Association Between Blood Donor Demographics and Post-injury Multiple Organ Failure after Polytrauma.

OBJECTIVE: To test the hypothesis that blood donor demographics are associated with transfused polytrauma patients' post-injury multiple organ failure (MOF) status. SUMMARY OF BACKGROUND DATA: Traumatic shock and MOF are preventable causes of death and post-traumatic hemorrhage is a frequent indication for transfusion. The role of blood donor demographics on transfusion recipients is not well known. METHODS: A log-linear analysis accounting for the correlated structure of the data based on our prospective MOF database was utilized. Tests for trend and interaction were computed using a likelihood ratio procedure. RESULTS: A total of 229 critically injured transfused trauma patients were included, with 68% of them being males and a mean age of 45 years. On average 10 units of blood components were transfused per patient. A total of 4379 units of blood components were donated by donors aged 46 years on average, 74% of whom were males. Blood components used were red blood cells (47%), cryoprecipitate (29%), fresh frozen plasma (24%), and platelets (less than 1%). Donor-recipient sex mismatched red blood cells transfusions were more likely to be associated with MOF ( P = 0.0012); fresh frozen plasma and cryoprecipitate recipients were more likely to experience MOF when transfused with a male (vs female) component ( P = 0.0014 and <0.0001, respectively). Donor age was not significantly associated with MOF for all blood components. CONCLUSIONS: Blood components donor sex, but not age, may be an important factor associated with post-injury MOF. Further validation of our findings will help guide future risk mitigation strategies specific to blood donor demographics.

NOT ALL CELL-FREE MITOCHONDRIAL DNA IS EQUAL IN TRAUMA PATIENTS.

ABSTRACT: Mitochondrial DNA (mtDNA) acts as a proinflammatory damage-associated molecular pattern that stimulates innate immune activation via Toll-like receptor 9, similarly to bacterial DNA. A number of clinical studies have measured elevated cell-free mtDNA in the plasma of trauma patients, thought to originate from tissue injury and inflammatory processes; however, the magnitude of this increase, the absolute concentration, and the association with poor outcomes varies considerably across studies. Measurements of cell-free mtDNA in healthy individuals have shown that the majority of "cell-free" mtDNA (>95%) can be centrifuged/filtered from plasma in the size range of 0.45 to 5 µm, suggesting that there are larger forms of mtDNA-containing complexes in the plasma that could be considered cell-free. Whether this is true for trauma patients (and other relevant disease states) and the clinical relevance of the larger forms of mtDNA is unknown. These findings from healthy individuals also suggest that the centrifugation speeds used to generate cell-free plasma (which are rarely consistent among studies) could result in mixed populations of cell-free mtDNA that could confound associations with outcomes. We demonstrate in this study of 25 major trauma patients that the majority of the cell-free mtDNA in trauma patient plasma (>95%) is removed after centrifugation at 16,000g. Despite the larger forms of mtDNA being predominant, they do not correlate with outcomes or expected parameters such as injury/shock severity, multiple organ failure, and markers of inflammation, whereas low-molecular-weight cell-free mtDNA correlates strongly with these variables.

Acute kidney injury development in polytrauma and the safety of early repeated contrast studies: A retrospective cohort study.

BACKGROUND: The role of repeat intravenous contrast doses beyond initial contrast imaging in the development of acute kidney injury (AKI) for multiple injury patients admitted to the intensive care unit (ICU) is not fully understood. We hypothesized that additional contrast doses are potentially modifiable risk factors for worse outcomes. METHODS: An 8-year retrospective study of our institutional prospective postinjury multiple organ failure database was performed. Adult ICU admissions that survived >72 hours with Injury Severity Score (ISS) of >15 were included. Patients were grouped based on number of repeat contrast studies received after initial imaging. Initial vital signs, resuscitation data, and laboratory parameters were collected. Primary outcome was AKI (Kidney Disease: Improving Global Outcomes criteria), and secondary outcomes included contrast-induced acute kidney injury (CI-AKI; >25% or >44 µmol/L increase in creatinine within 72 hours of contrast administration), multiple organ failure, length of stay, and mortality. RESULTS: Six-hundred sixty-three multiple injury patients (age, 45.3 years [SD, 9.1 years]; males, 75%; ISS, 25 (interquartile range, 20-34); mortality, 5.4%) met the inclusion criteria. The incidence of AKI was 13.4%, and CI-AKI was 14.5%. Multivariate analysis revealed that receiving additional contrast doses within the first 72 hours was not associated with AKI (odds ratio, 1.33; confidence interval, 0.80-2.21; p = 0.273). Risk factors for AKI included higher ISS ( p < 0.0007), older age ( p = 0.0109), higher heart rate ( p = 0.0327), lower systolic blood pressure ( p = 0.0007), and deranged baseline blood results including base deficit ( p = 0.0042), creatinine ( p < 0.0001), lactate ( p < 0.0001), and hemoglobin ( p = 0.0085). Acute kidney injury was associated with worse outcomes (ICU length of stay: 8 vs. 3 days, p < 0.0001; mortality: 16% vs. 3.8%, p < 0.0001; MOF: 42% vs. 6.6%, p < 0.0001). CONCLUSION: There is a limited role of repeat contrast administration in AKI development in ICU-admitted multiple injury patients. The clinical significance of CI-AKI is likely overestimated, and it should not compromise essential secondary imaging from the ICU. Further prospective studies are needed to verify our results. LEVEL OF EVIDENCE: Therapeutic/Care Management; Level III.

Stem cell therapy for fracture non-union: The current evidence from human studies.

Non-union is a taxing complication of fracture management for both the patient and their surgeon. Modern fracture fixation techniques have been developed to optimise the biomechanical environment for fracture healing but do not guarantee union. Patient biology has a critical role in achieving union and stem cell therapy has potential for improving fracture healing at a cellular level to treat or avoid non-union. This article reviews the current understanding of non-union, concepts in bone healing and the current literature on the application of stem cells in non-union.

Peripheral inflammation induces long-term changes in tyrosine hydroxylase activation in the substantia nigra.

Inflammation plays a role in neuropathology. We hypothesised that inflammation, induced by a single intraperitoneal injection of lipopolysaccharide (LPS), would induce long-term changes in the regulation of tyrosine hydroxylase (TH) in the rat midbrain. The level of 12 cytokines was initially analysed from one day to six months after LPS injection to confirm that peripheral inflammation led to neuroinflammatory changes in the midbrain. In the substantia nigra (SN), the levels of 8 of the 12 measured cytokines was significantly increased at one day. Granulocyte-macrophage colony-stimulating factor showed a threefold increased level at 6 months. The ventral tegmental area (VTA) showed a completely different pattern, with no increases in the levels of the 12 cytokines at one day and no changes beyond one week. TH activity was determined using a tritiated water release assay, TH protein and phosphorylation levels (Ser19, Ser31 and Ser40) were determined using western blotting. TH-specific activity in the SN was unchanged at one day but was substantially increased at one week and one month with no concomitant increase in TH phosphorylation. Substantial changes in TH activation without changes in TH phosphorylation have not previously been observed in the brain in response to a range of stressors. TH-specific activity was increased in the SN, and in the caudate putamen, at 6 months and was associated with increased TH phosphorylation at Ser19 and Ser40 at both locations. TH-specific activity in the VTA showed only a transient increase at day one associated with increased phosphorylation at Ser19 and Ser31 but thereafter showed no changes. This study shows that inflammation induced by LPS generated two distinct long-term changes in TH activity in the SN that are caused by different mechanisms, but there were no long-term changes in the adjacent VTA.

Peripheral Lipopolysaccharide Challenge Induces Long-Term Changes in Tyrosine Hydroxylase Regulation in the Adrenal Medulla.

Immune activation can alter the activity of adrenal chromaffin cells. The effect of immune activation by lipopolysaccharide (LPS) on the regulation of tyrosine hydroxylase (TH) in the adrenal medulla in vivo was determined between 1 day and 6 months after LPS injection. The plasma levels of eleven cytokines were reduced 1 day after LPS injection, whereas the level for interleukin-10 was increased. The levels of all cytokines remained at control levels until 6 months when the levels of interleukin-6 and -4 were increased. One day after LPS injection, there was a decrease in TH-specific activity that may be due to decreased phosphorylation of serine 31 and 40. This decreased phosphorylation of serine 31 and 40 may be due to an increased activation of the protein phosphatase PP2A. One week after LPS injection, there was increased TH protein and increased phosphorylation of serine 40 that this was not accompanied by an increase in TH-specific activity. All TH parameters measured returned to basal levels between 1 month and 3 months. Six months after injection there was an increase in TH protein. This was associated with increased levels of the extracellular regulated kinase isoforms 1 and 2. This work shows that a single inflammatory event has the capacity to generate both short-term and long-term changes in TH regulation in the adrenal medulla of the adult animal. J. Cell. Biochem. 118: 2096-2107, 2017. © 2016 Wiley Periodicals, Inc.

Catalytic domain surface residues mediating catecholamine inhibition in tyrosine hydroxylase.

Tyrosine hydroxylase (TH) performs the rate-limiting step in catecholamine (CA) synthesis and is a tetramer composed of regulatory, catalytic and tetramerization domains. CAs inhibit TH by binding two sites in the active site; one with high affinity and one with low affinity. Only high affinity CA binding requires the regulatory domain, believed to interact with the catalytic domain in the presence of CA. Without a crystal structure of the regulatory domain, the specific areas involved in this process are largely undefined. It is not clear whether the regulatory domain-catalytic domain interaction is asymmetrical across the tetramer to produce the high and low affinity sites. To investigate this, pure dimeric TH was generated through double substitution of residues at the tetramerization interface and dimerization salt bridge (K170E/L480A). This was shown to be the core regulatory unit of TH for CA inhibition, possessing both high and low affinity CA binding sites, indicating that there is symmetry between dimers of the tetramer. We also examined possible regulatory domain-interacting regions on the catalytic domain that mediate high affinity CA binding. Using site-directed mutagenesis, A297, E362/E365 and S368 were shown to mediate high affinity dopamine inhibition through V(max) reduction and increasing the K(M) for the cofactor.

Mechanism of action of salsolinol on tyrosine hydroxylase.

Tyrosine hydroxylase (TH) is the first and rate-limiting enzyme in dopamine synthesis. Dopamine regulates TH as an end-product inhibitor through its binding to a high and low affinity site, the former being abolished by Ser40 phosphorylation only, and the latter able to bind and dissociate according to intracellular dopamine levels. Here, we have investigated TH inhibition by a dopamine metabolite found in dopaminergic brain regions, salsolinol (SAL). SAL is known to decrease dopamine in the nigrostriatal pathway and mediobasal hypothalamus, and to also decrease plasma catecholamines in rat stress models, however a target and mechanism for the effects of SAL have not been found. We found that SAL inhibits TH activity in the nanomolar range in vitro, by binding to both the high and low affinity dopamine binding sites. SAL produces the same level of inhibition as dopamine when TH is non-phosphorylated. However, it produces 3.7-fold greater inhibition of Ser40-phosphorylated TH compared to dopamine by competing more strongly with tetrahydrobiopterin, the cofactor of this enzymatic reaction. SAL's potent inhibition of phosphorylated TH would prevent TH from being fully activated to synthesise dopamine.

Tyrosine hydroxylase: regulation by feedback inhibition and phosphorylation.

Tyrosine hydroxylase (TH) is the rate-limiting enzyme in the biosynthesis of the catecholamines dopamine, noradrenaline, and adrenaline. In response to short-term stimuli, TH activity is regulated by feedback inhibition by the catecholamines and relief of that inhibition by phosphorylation. This chapter examines the current understanding of these regulatory mechanisms and the roles that they play in different catecholaminergic cells. This chapter also examines hierarchical phosphorylation in TH and how it provides a mechanism for the differential regulation of the major human TH isoforms.

Mutational analysis of catecholamine binding in tyrosine hydroxylase.

Tyrosine hydroxylase (TH) performs the first and rate-limiting step in the synthesis of catecholamines, which feed back to regulate the enzyme by irreversibly binding to a high-affinity site and inhibiting TH activity. Phosphorylation of Ser40 relieves this inhibition by allowing dissociation of catecholamine. We have recently documented the existence of a low-affinity catecholamine binding which is dissociable, is not abolished by phosphorylation, and inhibits TH by competing with the essential cofactor, tetrahydrobiopterin. Here, we have substituted a number of active site residues to determine the structural nature of the low- and high-affinity sites. E332D and Y371F increased the IC(50) of dopamine for the low-affinity site 10-fold and 7 0-fold, respectively, in phosphorylated TH, indicating dramatic reductions in affinity. Only 2-4-fold increases in IC(50) were measured in the nonphosphorylated forms of E332D and Y371F and also in L294A and F300Y. This suggests that while the magnitude of low-affinity site inhibition in wild-type TH remains the same upon TH phosphorylation as previously shown, the active site structure changes to place greater importance on E332 and Y371. Changes to high affinity binding were also measured, including a loss of competition with tetrahydrobiopterin for E332D, A297L, and Y371F and a decreased ability to inhibit catalysis (V(max)) for A297L and Y371F. The common roles of E332 and Y371 indicate that the low- and high-affinity catecholamine binding sites are colocalized in the active site, but due to simultaneous binding, may exist in separate monomers of the TH tetramer.