Safety evaluation of adenosine, lidocaine and magnesium (ALM) intranasal therapy toward human nasal epithelial cells in vitro.

Adenosine, lidocaine and Mg2+ (ALM) solution is an emerging therapy that reduces secondary injury after intravenous administration in experimental models of traumatic brain injury (TBI). Intranasal delivery of ALM may offer an alternative route for rapid, point-of-care management of TBI. As a preliminary safety screen, we evaluated whether ALM exerts cytotoxic or inflammatory effects on primary human nasal epithelial cells (pHNEC) in vitro. Submerged monolayers and air-liquid interface cultures of pHNEC were exposed to media only, normal saline only, therapeutic ALM or supratherapeutic ALM for 15 or 60 min. Safety was measured through viability, cytotoxicity, apoptosis, cellular and mitochondrial stress, and inflammatory mediator secretion assays. No differences were found in viability or cytotoxicity in cultures exposed to saline or ALM for up to 60 min, with no evidence of apoptosis after exposure to supratherapeutic ALM concentrations. Despite comparable inflammatory cytokine secretion profiles and mitochondrial activity, cellular stress responses were significantly lower in cultures exposed to ALM than saline. In summary, data show ALM therapy has neither adverse toxic nor inflammatory effects on human nasal epithelial cells, setting the stage for in vivo toxicity studies and possible clinical translation of intranasal ALM therapy for TBI treatment.

Pathophysiology of severe burn injuries: new therapeutic opportunities from a systems perspective.

Severe burn injury elicits a profound stress response with the potential for high morbidity and mortality. If polytrauma is present, patient outcomes appear to be worse. Sex-based comparisons indicate females have worse outcomes than males. There are few effective drug therapies to treat burn shock and secondary injury progression. The lack of effective drugs appears to arise from the current treat-as-you-go approach rather than a more integrated systems approach. In this review, we present a brief history of burns research and discuss its pathophysiology from a systems' perspective. The severe burn injury phenotype appears to develop from a rapid and relentless barrage of damage-associated molecular patterns (DAMPs), pathogen-associated molecular patterns (PAMPs) and neural afferent signals, which leads to a state of hyperinflammation, immune dysfunction, coagulopathy, hypermetabolism and intense pain. We propose that if the central nervous system (CNS) control of cardiovascular function and endothelial-glycocalyx-mitochondrial coupling can be restored early, these secondary injury processes may be minimized. The therapeutic goal is to switch the injury phenotype to a healing phenotype by reducing fluid leak and maintaining tissue O2 perfusion. Currently, no systems-based therapies exist to treat severe burns. We have been developing a small-volume fluid therapy comprising adenosine, lidocaine and magnesium (ALM) to treat hemorrhagic shock, traumatic brain injury and sepsis. Our early studies indicate that the ALM therapy holds some promise in supporting cardiovascular and pulmonary functions following severe burns. Future research will investigate the ability of ALM therapy to treat severe burns with polytrauma and sex disparities, and potential translation to humans.

Adenosine, lidocaine and Mg2+ update: teaching old drugs new tricks.

If a trauma (or infection) exceeds the body's evolutionary design limits, a stress response is activated to quickly restore homeostasis. However, when the injury severity score is high, death is often imminent. The goal of this review is to provide an update on the effect of small-volume adenosine, lidocaine and Mg2+ (ALM) therapy on increasing survival and blunting secondary injury after non-compressible hemorrhagic shock and other trauma and infective/endotoxemic states. Two standout features of ALM therapy are: (1) resuscitation occurs at permissive hypotensive blood pressures (MAPs 50-60 mmHg), and (2) the drug confers neuroprotection at these low pressures. The therapy appears to reset the body's baroreflex to produce a high-flow, hypotensive, vasodilatory state with maintained tissue O2 delivery. Whole body ALM protection appears to be afforded by NO synthesis-dependent pathways and shifting central nervous system (CNS) control from sympathetic to parasympathetic dominance, resulting in improved cardiovascular function, reduced immune activation and inflammation, correction of coagulopathy, restoration of endothelial glycocalyx, and reduced energy demand and mitochondrial oxidative stress. Recently, independent studies have shown ALM may also be useful for stroke, muscle trauma, and as an adjunct to Resuscitative Endovascular Balloon Occlusion of the Aorta (REBOA). Ongoing studies have further shown ALM may have utility for burn polytrauma, damage control surgery and orthopedic surgery. Lastly, we discuss the clinical applications of ALM fluid therapy for prehospital and military far-forward use for non-compressible hemorrhage and traumatic brain injury (TBI).

Changes in plasma alpha-1 acid glycoprotein following hemorrhagic trauma: Possible role in dose differences of ALM drug therapy in rat and pig resuscitation.

INTRODUCTION: The binding of drugs to plasma proteins is an important consideration in drug development. We have reported that the dose of adenosine, lidocaine, and magnesium (ALM) fluid therapy for resuscitation from hemorrhagic shock is nearly 3-times higher for pigs than rats. Since lidocaine strongly binds to serum alpha-1-acid glycoprotein (AGP), the aim of the study was to investigate the effect of hemorrhagic shock on levels of AGP in rats and pigs. MATERIALS AND METHODS: Healthy adult male Sprague-Dawley rats and female crossbred pigs (n = 33 each) underwent tail vein and peripheral ear vein blood sampling, respectively, to collect plasma for AGP measurements. Rats (n = 17) and pigs (n = 16) underwent surgical instrumentation and uncontrolled hemorrhage via liver resection, and were treated with 3% NaCl ± ALM IV bolus followed 60 min later by 4 h 0.9% NaCl ± ALM IV drip. Rats were monitored for 72 h with blood samples taken post-surgery, and at 5.25, 24, and 72 h. Pigs were monitored for 6 h with blood samples taken post-surgery, and at 60 min and 6 h. Plasma AGP was measured with rat- and pig-specific enzyme-linked immunosorbent assay kits. RESULTS: Baseline AGP levels in rats were 3.91 µg/mL and significantly 83-fold lower than in pigs (325 µg/mL). Surgical instrumentation was associated with ~10-fold increases in AGP in rats and a 21% fall in pigs. AGP levels remained elevated in rats after hemorrhage and resuscitation (28-29 µg/mL). In contrast, no significant differences in plasma AGP were found in ALM- or Saline-treated pigs over the monitoring period. CONCLUSIONS: We conclude that the trauma of surgery alone was associated with significant increases in AGP in rats, compared to a contrasting decrease in pigs. Higher levels of plasma AGP in pigs prior to hemorrhagic shock is consistent with the higher ALM doses required to resuscitate pigs compared with rats.

Female rats have a different healing phenotype than males after anterior cruciate ligament rupture with no intervention.

Little is known on the sex-specific healing responses after an anterior cruciate ligament (ACL) rupture. To address this, we compared male and female Sprague-Dawley rats following non-surgical ACL rupture. Hematology, inflammation, joint swelling, range of motion, and pain-sensitivity were analyzed at various times over 31-days. Healing was assessed by histopathology and gene expression changes in the ACL remnant and adjacent joint tissues. In the first few days, males and females showed similar functional responses after rupture, despite contrasting hematology and systemic inflammatory profiles. Sex-specific differences were found in inflammatory, immune and angiogenic potential in the synovial fluid. Histopathology and increased collagen and fibronectin gene expression revealed significant tissue remodeling in both sexes. In the ACL remnant, however, Acta2 gene expression (α-SMA production) was 4-fold higher in males, with no change in females, indicating increased fibroblast-to-myofibroblast transition with higher contractile elements (stiffness) in males. Females had 80% lower Pparg expression, which further suggests reduced cellular differentiation potential in females than males. Sex differences were also apparent in the infrapatellar fat pad and articular cartilage. We conclude females and males showed different patterns of healing post-ACL rupture over 31-days, which may impact timing of reconstruction surgery, and possibly clinical outcome.

Immune dysfunction following severe trauma: A systems failure from the central nervous system to mitochondria.

When a traumatic injury exceeds the body's internal tolerances, the innate immune and inflammatory systems are rapidly activated, and if not contained early, increase morbidity and mortality. Early deaths after hospital admission are mostly from central nervous system (CNS) trauma, hemorrhage and circulatory collapse (30%), and later deaths from hyperinflammation, immunosuppression, infection, sepsis, acute respiratory distress, and multiple organ failure (20%). The molecular drivers of secondary injury include damage associated molecular patterns (DAMPs), pathogen associated molecular patterns (PAMPs) and other immune-modifying agents that activate the hypothalamic-pituitary-adrenal (HPA) axis and sympathetic stress response. Despite a number of drugs targeting specific anti-inflammatory and immune pathways showing promise in animal models, the majority have failed to translate. Reasons for failure include difficulty to replicate the heterogeneity of humans, poorly designed trials, inappropriate use of specific pathogen-free (SPF) animals, ignoring sex-specific differences, and the flawed practice of single-nodal targeting. Systems interconnectedness is a major overlooked factor. We argue that if the CNS is protected early after major trauma and control of cardiovascular function is maintained, the endothelial-glycocalyx will be protected, sufficient oxygen will be delivered, mitochondrial energetics will be maintained, inflammation will be resolved and immune dysfunction will be minimized. The current challenge is to develop new systems-based drugs that target the CNS coupling of whole-body function.

Why are bleeding trauma patients still dying? Towards a systems hypothesis of trauma.

Over the years, many explanations have been put forward to explain early and late deaths following hemorrhagic trauma. Most include single-event, sequential contributions from sympathetic hyperactivity, endotheliopathy, trauma-induced coagulopathy (TIC), hyperinflammation, immune dysfunction, ATP deficit and multiple organ failure (MOF). We view early and late deaths as a systems failure, not as a series of manifestations that occur over time. The traditional approach appears to be a by-product of last century's highly reductionist, single-nodal thinking, which also extends to patient management, drug treatment and drug design. Current practices appear to focus more on alleviating symptoms rather than addressing the underlying problem. In this review, we discuss the importance of the system, and focus on the brain's "privilege" status to control secondary injury processes. Loss of status from blood brain barrier damage may be responsible for poor outcomes. We present a unified Systems Hypothesis Of Trauma (SHOT) which involves: 1) CNS-cardiovascular coupling, 2) Endothelial-glycocalyx health, and 3) Mitochondrial integrity. If central control of cardiovascular coupling is maintained, we hypothesize that the endothelium will be protected, mitochondrial energetics will be maintained, and immune dysregulation, inflammation, TIC and MOF will be minimized. Another overlooked contributor to early and late deaths following hemorrhagic trauma is from the trauma of emergent surgery itself. This adds further stress to central control of secondary injury processes. New point-of-care drug therapies are required to switch the body's genomic and proteomic programs from an injury phenotype to a survival phenotype. Currently, no drug therapy exists that targets the whole system following major trauma.

Anterior Cruciate Ligament Reconstruction Surgery: Creating a Permissive Healing Phenotype in Military Personnel and Civilians for Faster Recovery.

INTRODUCTION: Anterior cruciate ligament (ACL) rupture in military personnel and civilians can be a devastating injury. A service member is 10 times more likely to suffer an ACL injury than their civilian counterparts, and despite successful surgical stabilization, 4%-35% will develop arthrofibrosis, over 50% will not return to full active duty, and up to 50% will develop post-traumatic osteoarthritis (PTOA) within 15 years. Equally concerning, woman are 2 to 8 times more likely to experience ACL injuries than men, which represents a major knowledge gap. MATERIALS AND METHODS: A comprehensive literature search was performed in December 2021 using structured search terms related to prevalence, risk factors, disease progression, and treatment of ACL injury and reconstruction. The literature search was conducted independently by two researchers using PubMed, Cochrane, and Embase databases, with inclusion of articles with military, civilian, and sex relevance, and exclusion of most papers with a publication date greater than 10 years. The resources used for the review reflect the most current data, knowledge, and recommendations associated with research and clinical findings from reliable international sources. RESULTS: Currently, there is no effective system-based drug therapy that creates a "permissive environment" to reduce synovial and cartilage stress after ACL injury and reconstruction and prevent secondary complications. We argue that progress in this area has been hampered by researchers and clinicians failing to recognize that (1) an ACL injury is a system's failure that affects the whole joint, (2) the early molecular events define and perpetuate different injury phenotypes, (3) male and female responses may be different and have a molecular basis, (4) the female phenotype continues to be under-represented in basic and clinical research, and (5) the variable outcomes may be perpetuated by the trauma of surgery itself. The early molecular events after ACL injury are characterized by an overexpression of joint inflammation, immune dysfunction, and trauma-induced synovial stress. We are developing an upstream adenosine, lidocaine, and magnesium therapy to blunt these early molecular events and expedite healing with less arthrofibrosis and early PTOA complications. CONCLUSIONS: ACL injuries continue to be a major concern among military personnel and civilians and represent a significant loss in command readiness and quality of life. The lack of predictability in outcomes after ACL repair or reconstruction underscores the need for new joint protection therapies. The male-female disparity requires urgent investigation.

ALM Induces Cellular Quiescence in the Surgical Margin 3 Days Following Liver Resection, Hemorrhage, and Shock.

INTRODUCTION: The liver has a remarkable capacity to regenerate but not the resected lobe. Our aim was to examine the expression of a number of key genes of metabolism, proliferation, survival, and reprogramming 5 mm inside the resected margin following resuscitation with adenosine, lidocaine, and Mg2+ (ALM) therapy. MATERIALS AND METHODS: Anesthetized adult male Sprague-Dawley rats randomly assigned to ALM treatment (n = 10) or Saline controls (n = 10) underwent liver resection (60% left lateral lobe) and uncontrolled bleeding. After 15 min, 3% NaCl ± ALM bolus was administered, and after 60 min, a 4 h 0.9% NaCl ± ALM stabilization 'drip' was commenced. After 72 h monitoring (or high moribund score), histopathology, inflammatory mediators, and relative expression of key genes of tissue repair were measured in the remaining left lateral liver. RESULTS: ALM animals survived 72 h compared to 23 h for Saline controls (P = 0.002). In the surgical margin, ALM therapy showed preservation of cellular architecture, whereas controls had increased inflammation and diffuse necrosis. Liver proinflammatory cytokines were also 2- to 4-fold higher in Saline controls. ALM therapy dramatically suppressed (∼70%) gene expression of four adenosine receptors, metabolic signaling, autophagy, apoptosis, and cell proliferation compared to controls, including suppression of the Yamanaka factors by up to 85%. CONCLUSIONS: We conclude ALM therapy preserved hepatocyte architecture with less inflammation and necrosis 3 days after resection, hemorrhage, and shock. In addition, ALM induced cellular quiescence in the surgical margin, which may be a strategy for improved barrier protection and healing. Further studies are required to address this question.

Trauma care in the tropics: addressing gaps in treating injury in rural and remote Australia.

In Australia, over half a million people are admitted to hospital every year as a result of injury, and where you live matters. Rural populations have disproportionately higher injury hospitalisation rates (1.5-2.5-fold), higher rates of preventable secondary complications, higher mortality rates (up to fivefold), and higher costs (threefold) than patients injured in major cities. These disparities scale up rapidly with increased remoteness, and shift the service needle from 'scoop and run' to 'continuum of care'. Poorer outcomes, however, are not solely due to longer retrieval distances or delays; they arise from inefficiencies in one or more potentially modifiable factors in the chain of survival. After discussing the burden of injury in Australia, we present a brief history of retrieval services in Queensland and discuss how remoteness requires a different kind of service delivery with many moving parts from point of injury to definitive care. We next address the ongoing challenges for the Australian Trauma Registry, and how centralisation of data from the metropolitan cities masks the inequities in rural and remote trauma. There is an urgent need for accurate data from all service providers around Australia to inform state and federal governments, and we highlight the paucity of trauma data analysis in North Queensland. Last, we identify some major gaps in treating rural and remote polytrauma and en-route patient stabilisation, and discuss the relevance of combat casualty care research and practices. We conclude that a greater emphasis should be placed on collecting more robust trauma patient records, as only accurate data will drive change.

ALM Fluid Therapy Shifts Sympathetic Hyperactivity to Parasympathetic Dominance in the Rat Model of Non-Compressible Hemorrhagic Shock.

ABSTRACT: Excessive sympathetic outflow following trauma can lead to cardiac dysfunction, inflammation, coagulopathy, and poor outcomes. We previously reported that buprenorphine analgesia decreased survival after hemorrhagic trauma. Our aim is to examine the underlying mechanisms of mortality in a non-compressible hemorrhage rat model resuscitated with saline or adenosine, lidocaine, magnesium (ALM). Anesthetized adult male Sprague-Dawley rats were randomly assigned to Saline control group or ALM therapy group (both n = 10). Hemorrhage was induced by 50% liver resection. After 15 min, 0.7 mL/kg 3% NaCl ±â€ŠALM intravenous bolus was administered, and after 60 min, 0.9% NaCl ±â€ŠALM was infused for 4 h (0.5 mL/kg/h) with 72 h monitoring. Animals received 6-12-hourly buprenorphine for analgesia. Hemodynamics, heart rate variability, echocardiography, and adiponectin were measured. Cardiac tissue was analyzed for adrenergic/cholinergic receptor expression, inflammation, and histopathology. Four ALM animals and one Saline control survived to 72 h. Mortality was associated with up to 97% decreases in adrenergic (ß-1, α-1A) and cholinergic (M2) receptor expression, cardiac inflammation, myocyte Ca2+ loading, and histopathology, indicating heart ischemia/failure. ALM survivors had higher cardiac output and stroke volume, a 30-fold increase in parasympathetic/sympathetic receptor expression ratio, and higher circulating adiponectin compared to Saline controls. Paradoxically, Saline cardiac adiponectin hormone levels were higher than ALM, with no change in receptor expression, indicating intra-cardiac synthesis. Mortality appears to be a "systems failure" associated with CNS dysregulation of cardiac function. Survival involves an increased parasympathetic dominance to support cardiac pump function with reduced myocardial inflammation. Increased cardiac α-1A adrenergic receptor in ALM survivors may be significant, as this receptor is highly protective during heart dysfunction/failure.

Comparison of intra-articular administration of adenosine, lidocaine and magnesium solution and tranexamic acid for alleviating postoperative inflammation and joint fibrosis in an experimental model of knee arthroplasty.

BACKGROUND: Dysregulated inflammatory responses are implicated in the pathogenesis of joint stiffness and arthrofibrosis following total knee arthroplasty (TKA). The purpose of this study was to compare the effects of intra-articular (IA) administration of tranexamic acid (TXA), an anti-fibrinolytic commonly used in TKA, and ALM chondroprotective solution on postoperative inflammation and joint tissue healing in a rat model of knee implant surgery. METHODS: Male Sprague-Dawley rats (n = 24) were randomly divided into TXA or ALM treatment groups. The right knee of each rat was implanted with titanium (femur) and polyethylene (tibia) implants. An IA bolus (0.1 ml) of TXA or ALM was administered after implantation and capsule closure, and before skin closure. Postoperative coagulopathy, haematology and systemic inflammatory changes were assessed. Inflammatory and fibrotic markers were assessed in joint tissue, 28 days after surgery. RESULTS: Haemostasis was comparable in animals treated with TXA or ALM after knee implant surgery. In contrast to ALM-treated animals, systemic inflammatory markers remained elevated at day 5 (IL-6, IL-12, IL-10, platelet count) and day 28 (IL-1ß, IL-10) following surgery in TXA-treated animals. At day 28 following surgery, the extension range of motion of operated knees was 1.7-fold higher for ALM-treated animals compared to the TXA group. Key inflammatory mediators (NF-κB, IL-12, IL-2), immune cell infiltration (CD68+ cells) and markers of fibrosis (α-SMA, TGF-ß) were also lower in capsular tissue of ALM-treated knees at day 28. CONCLUSION: Data suggest that IA administration of ALM is superior to TXA for reducing postoperative systemic and joint inflammation and promoting restoration of healthy joint tissue architecture in a rat model of TKA. Further studies are warranted to assess the clinical translational potential of ALM IA solution to improve patient outcomes following arthroplasty.

Major surgery leads to a proinflammatory phenotype: Differential gene expression following a laparotomy.

BACKGROUND: The trauma of surgery is a neglected area of research. Our aim was to examine the differential expression of genes of stress, metabolism and inflammation in the major organs of a rat following a laparotomy. MATERIALS AND METHODS: Anaesthetised Sprague-Dawley rats were randomised into baseline, 6-hr and 3-day groups (n = 6 each), catheterised and laparotomy performed. Animals were sacrificed at each timepoint and tissues collected for gene and protein analysis. Blood stress hormones, cytokines, endothelial injury markers and coagulation were measured. RESULTS: Stress hormone corticosterone significantly increased and was accompanied by significant increases in inflammatory cytokines, endothelial markers, increased neutrophils (6-hr), higher lactate (3-days), and coagulopathy. In brain, there were significant increases in M1 muscarinic (31-fold) and α-1A-adrenergic (39-fold) receptor expression. Cortical expression of metabolic genes increased ∼3-fold, and IL-1ß by 6-fold at 3-days. Cardiac ß-1-adrenergic receptor expression increased up to 8.4-fold, and M2 and M1 muscarinic receptors by 2 to 4-fold (6-hr). At 3-days, cardiac mitochondrial gene expression (Tfam, Mtco3) and inflammation (IL-1α, IL-4, IL-6, MIP-1α, MCP-1) were significantly elevated. Haemodynamics remained stable. In liver, there was a dramatic suppression of adrenergic and muscarinic receptor expression (up to 90%) and increased inflammation. Gut also underwent autonomic suppression with 140-fold increase in IL-1ß expression (3-days). CONCLUSIONS: A single laparotomy led to a surgical-induced proinflammatory phenotype involving neuroendocrine stress, cortical excitability, immune activation, metabolic changes and coagulopathy. The pervasive nature of systemic and tissue inflammation was noteworthy. There is an urgent need for new therapies to prevent hyper-inflammation and restore homeostasis following major surgery.

The role of matrix metalloproteinase-9 and its inhibitor TIMP-1 in burn injury: a systematic review.

Matrix metalloproteinase-9 (MMP-9) and its endogenous inhibitor, tissue inhibitor of metalloproteinase-1 (TIMP-1), are key mediators of acute inflammation and regulators of the wound healing process. The aim of this systematic review was to determine the local and systemic involvement of the MMP-9/TIMP-1 system following burn injury. Two databases (Scopus and MEDLINE) were searched for all studies reporting MMP-9 and/or TIMP-1 after burn injury. Based on our eligibility criteria, we reviewed 24 studies involving 508 burns patients in 11 clinical studies and 367 animals in 13 preclinical studies. Local, systemic, and peripheral gene expression, protein levels and activity of MMP-9 and TIMP-1 were assessed. Increased MMP-9 was reported at the site of injury early after burn trauma in all studies, and remained elevated in non-healing wounds. Increased TIMP-1 expression in burn wounds occurred later than MMP-9, and was persistent in hypertrophic burn scars. Similar to local expression, systemic MMP-9 and TIMP-1 concentrations were significantly elevated after burn injury in response to upregulation of proinflammatory cytokines. While no association was found between systemic MMP-9 concentration and extent of injury or outcome, serum or plasma TIMP-1 showed good correlation with survival and burn severity. This review also found evidence of the MMP-9/TIMP-1 system contributing to secondary tissue damage distant from the burn site, including burn-associated musculoskeletal damage and acute lung injury. In addition, increased MMP-9 synthesis and activity in the brain after peripheral burn may lead to blood-brain barrier dysfunction and cerebral edema, a significant contributor to mortality. This systematic review provides an overview of the available evidence of the role of MMP-9 and TIMP-1 in burn injury pathophysiology and finds that TIMP-1 may be a promising biomarker in outcome prognostication of burns patients. Large-scale studies of both pediatric and adult burns patients with increased female representation and repeated sampling are recommended to validate the reliability of TIMP-1 as a prognostic marker following burn injury.

Identification of defective early immune responses to Burkholderia pseudomallei infection in a diet-induced murine model of type 2 diabetes.

Co-occurrence of bacterial infections with type 2 diabetes (T2D) is a global problem. Melioidosis caused by Burkholderia pseudomallei is 10 times more likely to occur in patients with T2D, than in normoglycemic individuals. Using an experimental model of T2D, we observed that greater susceptibility in T2D was due to differences in proportions of infiltrating leucocytes and reduced levels of MCP-1, IFN-γ and IL-12 at sites of infection within 24 h post-infection. However, by 72 h the levels of inflammatory cytokines and bacteria were markedly higher in visceral tissue and blood in T2D mice. In T2D, dysregulated early immune responses are responsible for the greater predisposition to B. pseudomallei infection.

Specific Pathogen-Free Animals for Civilian and Military Trauma: a Cautionary Note in the Translation of New Drug Therapies.

Specific-pathogen free (SPF) animals were introduced into biomedical research in the early 1960s to reduce the incidence of disease into experimental design. The goal was to provide animals with selected microbiota compatible with sustained health. Sixty years later, SPF status has become a variable itself in biomedical research. Alterations in the gut microbiome-host relationship can profoundly influence basic physiology, immune/inflammatory function, susceptibility to infection and disease, and behavior. In addition, it can influence the translational success of a drug or technology from animal models to humans. We discuss this aspect of SPF status in animal models used for military or civilian trauma and shock research. Currently, there is a broad spectrum of SPF exclusion and inclusion criteria which vary from one supplier or animal husbandry facility. If translation to humans is the end-game of trauma research, we recommend replicating a gut microbiome similar to the wild-type for optimal success. We further suggest that at the end of each publication a URL access be provided on Animal Microbial/Pathogen Exclusion Status that a study was based upon. This may help address the differences in results within a single laboratory or between laboratories around the world and improve translation success.

Traumatic-Induced Coagulopathy as a Systems Failure: A New Window into Hemostasis.

Traumatic-induced coagulopathy (TIC) is often associated with significant bleeding, transfusion requirements, inflammation, morbidity, and mortality. This review considers TIC as a systems failure, not as a single-event manifestation of trauma. After briefly reviewing the meaning of TIC and the bewildering array of fibrinolysis phenotypes, we will discuss the role of platelets and fibrinogen in coagulopathy. Next, we will review the different TIC hypotheses and drill down to a single mechanistic domain comprising (1) thrombin's differential binding to thrombomodulin, (2) the expression of annexin II-S100A10 complex, and (3) the functional integrity of the endothelial glycocalyx. This triad forms the basis of the "switch" hypothesis of TIC. We will next address the potential limitations of current practice in treating a coagulation or fibrinolytic defect, and the next defect, and so on down the line, which often leads to what U.S. surgeon William C. Shoemaker considered "an uncoordinated and sometimes contradictory therapeutic outcome." The treat-as-you-go approach using sequential, single-target treatments appears to be a by-product of decades of highly reductionist thinking and research. Lastly, we will present a unified systems hypothesis of TIC involving three pillars of physiology: the central nervous system (CNS)-cardiovascular system, the endothelial glycocalyx, and mitochondrial integrity. If CNS control of ventriculoarterial coupling is maintained close to unity following trauma, we hypothesize that the endothelium will be protected, mitochondrial energetics will be maintained, and TIC (and inflammation) will be minimized. The Systems Hypothesis of Trauma (SHOT) also helps to answer why certain groups of severely bleeding trauma patients are still dying despite receiving the best care. Currently, no drug therapy exists that targets the whole system.

Living in a Hostile World: Inflammation, New Drug Development, and Coronavirus.

We present a brief history of the immune response and show that Metchnikoff's theory of inflammation and phagocytotic defense was largely ignored in the 20th century. For decades, the immune response was believed to be triggered centrally, until Lafferty and Cunningham proposed the initiating signal came from the tissues. This shift opened the way for Janeway's pattern recognition receptor theory, and Matzinger's danger model. All models failed to appreciate that without inflammation, there can be no immune response. The situation changed in the 1990s when cytokine biology was rapidly advancing, and the immune system's role expanded from host defense, to the maintenance of host health. An inflammatory environment, produced by immune cells themselves, was now recognized as mandatory for their attack, removal and repair functions after an infection or injury. We explore the cellular programs of the immune response, and the role played by cytokines and other mediators to tailor the right response, at the right time. Normally, the immune response is robust, self-limiting and restorative. However, when the antigen load or trauma exceeds the body's internal tolerances, as witnessed in some COVID-19 patients, excessive inflammation can lead to increased sympathetic outflows, cardiac dysfunction, coagulopathy, endothelial and metabolic dysfunction, multiple organ failure and death. Currently, there are few drug therapies to reduce excessive inflammation and immune dysfunction. We have been developing an intravenous (IV) fluid therapy comprising adenosine, lidocaine and Mg2+ (ALM) that confers a survival advantage by preventing excessive inflammation initiated by sepsis, endotoxemia and sterile trauma. The multi-pronged protection appears to be unique and may provide a tool to examine the intersection points in the immune response to infection or injury, and possible ways to prevent secondary tissue damage, such as that reported in patients with COVID-19.

Evaluation of bacteriophage as an adjunct therapy for treatment of peri-prosthetic joint infection caused by Staphylococcus aureus.

Phage therapy offers a potential alternate strategy for the treatment of peri-prosthetic joint infection (PJI), particularly where limited effective antibiotics are available. We undertook preclinical trials to investigate the therapeutic efficacy of a phage cocktail, alone and in combination with vancomycin, to reduce bacterial numbers within the infected joint using a clinically-relevant model of Staphylococcus aureus-induced PJI. Infected animals were randomised to 4 treatment groups, with treatment commencing 21-days post-surgery: bacteriophage alone, vancomycin alone, bacteriophage and vancomycin, and sham. At day 28 post-surgery, animals were euthanised for microbiological and immunological assessment of implanted joints. Treatment with phage alone or vancomycin alone, led to 5-fold and 6.2-fold reductions, respectively in bacterial load within peri-implant tissue compared to sham-treated animals. Compared to sham-treated animals, a 22.5-fold reduction in S. aureus burden was observed within joint tissue of animals that were administered phage in combination with vancomycin, corresponding with decreased swelling in the implanted knee. Microbiological data were supported by evidence of decreased inflammation within the joints of animals administered phage in combination with vancomycin, compared to sham-treated animals. Our findings provide further support for phage therapy as a tolerable and effective adjunct treatment for PJI.