87 results found
Oakley C, Koh M, Baldi R, et al., 2019, Ventilation following established ARDS: a preclinical model framework to improve predictive power, Thorax, Vol: 74, Pages: 1120-1129, ISSN: 1468-3296
Background Despite advances in understanding the pathophysiology of acute respiratory distress syndrome, effective pharmacological interventions have proven elusive. We believe this is a consequence of existing preclinical models being designed primarily to explore biological pathways, rather than predict treatment effects. Here, we describe a mouse model in which both therapeutic intervention and ventilation were superimposed onto existing injury and explored the impact of β-agonist treatment, which is effective in simple models but not clinically.Methods Mice had lung injury induced by intranasal lipopolysaccharide (LPS), which peaked at 48 hours post-LPS based on clinically relevant parameters including hypoxaemia and impaired mechanics. At this peak of injury, mice were treated intratracheally with either terbutaline or tumour necrosis factor (TNF) receptor 1-targeting domain antibody, and ventilated with moderate tidal volume (20 mL/kg) to induce secondary ventilator-induced lung injury (VILI).Results Ventilation of LPS-injured mice at 20 mL/kg exacerbated injury compared with low tidal volume (8 mL/kg). While terbutaline attenuated VILI within non-LPS-treated animals, it was ineffective to reduce VILI in pre-injured mice, mimicking its lack of clinical efficacy. In contrast, anti-TNF receptor 1 antibody attenuated secondary VILI within pre-injured lungs, indicating that the model was treatable.Conclusions We propose adoption of a practical framework like that described here to reduce the number of ultimately ineffective drugs reaching clinical trials. Novel targets should be evaluated alongside interventions which have been previously tested clinically, using models that recapitulate the (lack of) clinical efficacy. Within such a framework, outperforming a failed pharmacologic should be a prerequisite for drugs entering trials.
Soni S, Tirlapur N, O'Dea KP, et al., 2019, Microvesicles as new therapeutic targets for the treatment of the acute respiratory distress syndrome (ARDS), EXPERT OPINION ON THERAPEUTIC TARGETS, ISSN: 1472-8222
Hua R, Edey LF, O'Dea KP, et al., 2019, CCR2 mediates the adverse effects of LPS in the pregnant mouse., Biol Reprod
In our earlier work, we found that intrauterine (i.u.) and intraperitoneal (i.p.) injection of LPS (10 μg serotype 0111:B4) induced preterm labor (PTL) with high pup mortality, marked systemic inflammatory response and hypotension. Here, we used both i.u. and i.p. LPS models in pregnant wild type (wt) and CCR2 knockout (CCR2-/-) mice on E16 to investigate the role played by the CCL2/CCR2 system in the response to LPS. Basally, lower numbers of monocytes and macrophages and higher numbers of neutrophils were found in the myometrium, placenta and blood of CCR2-/- vs. wt mice. After i.u. LPS, parturition occurred at 14 hours in both groups of mice. At 7 hours post injection, 70% of wt pups were dead vs. 10% of CCR2-/- pups, but at delivery 100% of wt and 90% of CCR2-/- pups were dead. Myometrial and placental monocytes and macrophages were generally lower in CCR2-/- mice, but this was less consistent in the circulation, lung and liver. At 7 hours post LPS, myometrial ERK activation was greater and JNK and p65 lower and the mRNA levels of chemokines were higher and of inflammatory cytokines lower in CCR2-/- vs. wt mice. Pup brain and placental inflammation was similar. Using the IP LPS model, we found that all measures of arterial pressure increased in CCR2-/- but declined in wt mice. These data suggest that the CCL2/CCR2 system plays a critical role in the cardiovascular response to LPS and contributes to pup death but does not influence the onset of inflammation-induced PTL.
Zöllner J, Howe LG, Edey LF, et al., 2019, LPS-induced hypotension in pregnancy: The effect of progesterone supplementation, Shock, ISSN: 1073-2322
Our previous work has shown that pregnancy exacerbates the hypotensive response to both infection and LPS. The high levels of progesterone (P4) associated with pregnancy have been suggested to be responsible for the pregnancy-induced changes in the cardiovascular response to infection. Here, we test the hypothesis that P4 supplementation exacerbates the hypotensive response of the maternal cardiovascular to LPS.Female CD1 mice had radiotelemetry probes implanted to measure haemodynamic function non-invasively and were time-mated. From day 14 of pregnancy, mice received either 10 mg of P4 or vehicle alone per day and on day 16, intraperitoneal LPS (10 μg of serotype 0111:B4) was injected. In two identically treated cohorts of mice, tissue and serum (for RNA, protein studies) were collected at 6 and 12 hours.Administration of LPS resulted in a fall in blood pressure in vehicle treated, but not P4 supplemented mice. This occurred with similar changes in the circulating levels of cytokines, vasoactive factors and in both circulating and tissue inflammatory cell numbers, but with reduced left ventricular expression of cytokines in P4-supplemented mice. However, left ventricular expression of markers of cardiac dysfunction and apoptosis were similar.This study demonstrates that P4 supplementation prevented LPS-induced hypotension in pregnant mice in association with reduced myocardial inflammatory cytokine gene expression. These observations suggest that rather than being detrimental, P4 supplementation has a protective effect on the maternal cardiovascular response to sepsis.
Soni S, O'Dea K, Tan YY, et al., 2019, ATP redirects cytokine trafficking and promotes novel membrane TNF signalling via microvesicles, FASEB Journal, ISSN: 0892-6638
Cellular stress or injury induces release of endogenous danger signals such as ATP, which plays a central role in activating immune cells. ATP is essential for the release of nonclassically secreted cytokines such as IL-1β but, paradoxically, has been reported to inhibit the release of classically secreted cytokines such as TNF. Here, we reveal that ATP does switch off soluble TNF (17 kDa) release from LPS-treated macrophages, but rather than inhibiting the entire TNF secretion, ATP packages membrane TNF (26 kDa) within microvesicles (MVs). Secretion of membrane TNF within MVs bypasses the conventional endoplasmic reticulum– and Golgi transport–dependent pathway and is mediated by acid sphingomyelinase. These membrane TNF–carrying MVs are biologically more potent than soluble TNF in vivo, producing significant lung inflammation in mice. Thus, ATP critically alters TNF trafficking and secretion from macrophages, inducing novel unconventional membrane TNF signaling via MVs without direct cell-to-cell contact. These data have crucial implications for this key cytokine, particularly when therapeutically targeting TNF in acute inflammatory diseases.—Soni, S., O’Dea, K. P., Tan, Y. Y., Cho, K., Abe, E., Romano, R., Cui, J., Ma, D., Sarathchandra, P., Wilson, M. R., Takata, M. ATP redirects cytokine trafficking and promotes novel membrane TNF signaling via microvesicles.
Tan YY, O'Dea KP, Soo AP, et al., 2019, Investigation into the Roles of Circulating Microvesicles Within the Pulmonary Vasculature Using Ex Vivo Isolated Perfused Lung, International Conference of the American-Thoracic-Society, Publisher: AMER THORACIC SOC, ISSN: 1073-449X
Matsumoto S, Iki Y, Abe M, et al., 2019, Neutrophil-Derived Microvesicles as a Novel Biomarker in Hyperoxic Bronchopulmonary Dysplasia in Mice, International Conference of the American-Thoracic-Society, Publisher: AMER THORACIC SOC, ISSN: 1073-449X
Gordon AC, Santhakumaran S, Al-Beidh F, et al., 2019, Levosimendan to prevent acute organ dysfunction in sepsis: the LeoPARDS RCT, Efficacy and Mechanism Evaluation, Vol: 5, Pages: 1-94, ISSN: 2050-4365
<jats:sec id="abs1-1"> <jats:title>Background</jats:title> <jats:p>In septic shock, cardiovascular resuscitation using catecholamine vasopressors and inotropes is standard therapy, but catecholamines have important side effects. Levosimendan (Simdax<jats:sup>®</jats:sup>; Orion Pharma, Newbury, UK) is a calcium-sensitising drug with inotropic and other properties that may have a role in sepsis.</jats:p> </jats:sec> <jats:sec id="abs1-2"> <jats:title>Objectives</jats:title> <jats:p>To determine, in adult septic shock, whether or not levosimendan reduces the incidence and severity of acute organ dysfunction, the effect of levosimendan on individual organ function and the safety profile of levosimendan.</jats:p> </jats:sec> <jats:sec id="abs1-3"> <jats:title>Design</jats:title> <jats:p>Multicentre, randomised, double-blind, parallel-group, placebo-controlled study.</jats:p> </jats:sec> <jats:sec id="abs1-4"> <jats:title>Setting</jats:title> <jats:p>UK intensive care units.</jats:p> </jats:sec> <jats:sec id="abs1-5"> <jats:title>Participants</jats:title> <jats:p>Adult patients with sepsis and cardiovascular failure requiring vasopressors to maintain blood pressure despite adequate fluid resuscitation.</jats:p> </jats:sec> <jats:sec id="abs1-6"> <jats:title>Intervention</jats:title> <jats:p>Levosimendan, at a dosage of 0.05–0.2 µg/kg/minute
Antcliffe D, Ward J, Marshall T, et al., 2018, Multivariate analysis of cytokines in septic shock predicts outcome, European Society of Intensive Care Medicine Congress, Publisher: SpringerOpen, ISSN: 2197-425X
Antcliffe D, Wolfer A, O'Dea K, et al., 2018, Profiling inflammatory markers in patients with pneumonia on intensive care, Scientific Reports, Vol: 8, ISSN: 2045-2322
Clinical investigations lack predictive value when diagnosing pneumonia, especially when patients are ventilated and develop ventilator associated pneumonia (VAP). New tools to aid diagnosis are important to improve outcomes. This pilot study examines the potential for a panel of inflammatory mediators to aid in the diagnosis. Forty-four ventilated patients, 17 with pneumonia and 27 with brain injuries, eight of whom developed VAP, were recruited. 51 inflammatory mediators, including cytokines and oxylipins, were measured in patients’ serum using flow cytometry and mass spectrometry. The mediators could separate patients admitted to ICU with pneumonia compared to brain injury with an area under the receiver operating characteristic curve (AUROC) 0.75 (0.61–0.90). Changes in inflammatory mediators were similar in both groups over the course of ICU stay with 5,6-dihydroxyeicosatrienoic and 8,9-dihydroxyeicosatrienoic acids increasing over time and interleukin-6 decreasing. However, brain injured patients who developed VAP maintained inflammatory profiles similar to those at admission. A multivariate model containing 5,6-dihydroxyeicosatrienoic acid, 8,9-dihydroxyeicosatrienoic acid, intercellular adhesion molecule-1, interleukin-6, and interleukin-8, could differentiate patients with VAP from brain injured patients without infection (AUROC 0.94 (0.80–1.00)). The use of a selected group of markers showed promise to aid the diagnosis of VAP especially when combined with clinical data.
Gordon AC, Santhakumaran S, Al-Beidh F, et al., 2018, Levosimendan for the Prevention of Acute oRgan Dysfunction in Sepsis: the LeoPARDS Randomised Controlled Trial, Efficacy and Mechanism Evaluation, ISSN: 2050-4365
Background:In septic shock, cardiovascular resuscitation using catecholamine vasopressors and inotropes is standard therapy but catecholamines have important side-effects. Levosimendan is a calcium-sensitizing drug with inotropic and other properties that may have a role in sepsis.Objectives: In adult septic shock1. Does levosimendan reduce the incidence and severity of acute organ dysfunction ?2. What is the effect of levosimendan on individual organ function ?3. What is the safety profile of levosimendan?Design: Multi-centre, randomised, double-blind, parallel-group, placebo-controlled study.Setting: UK Intensive Care UnitsParticipants: Adult patients who have sepsis and cardiovascular failure requiring vasopressors to maintain blood pressure despite adequate fluid resuscitation.Interventions: Levosimendan 0.05 to 0.2 µg/kg/min vs. placebo for 24 hour, in addition to standard care, within 24 hours of meeting inclusion criteria.Primary outcome measure: Mean SOFA score on ICU after randomisation to a maximum of 28 days.Secondary outcome measures: Time to extubationSurvival upto 6 monthsSerious Adverse EventsResults: 2382 patients were screened at 34 centres, of whom 516 were randomised to treatment, 259 allocated to levosimendan and 257 to placebo. Baseline characteristics were well balanced across treatment arms.There was no significant difference in mean (±SD) SOFA score in the levosimendan group (6.7 ± 4.0) compared with placebo (6.1 ± 3.9); (mean difference 0.61, 95%CI -0.07 to 1.29). 28-day mortality was 34.5% versus 30.9% in the levosimendan and placebo groups respectively (absolute difference 3.6%, 95%CI -4.5 to 11.7). Patients in the levosimendan group were less likely to be successfully extubated over 28 days than the placebo group (hazard ratio 0.77, 95%CI 0.60 to 0.97). More patients in the levosimendan group had supraventricular tachyarrhythmias, (3.1% versus 0.4% absolute difference 2.7%, 95%CI 0.1 to 5.3), but there was no
Edey LF, Georgiou H, O'Dea KP, et al., 2017, Progesterone, the maternal immune system and the onset of parturition in the mouse, Biology of Reproduction, Vol: 98, Pages: 376-395, ISSN: 1529-7268
The role of progesterone (P4) in the regulation of the local (uterine) and systemic innate immune system, myometrial expression of connexin 43 (Cx-43) and cyclooxygenase 2 (COX-2) and the onset of parturition was examined in: 1) naïve mice delivering at term; 2) E16 mice treated with RU486 (P4-antagonist) to induce preterm parturition; and 3) in mice treated with P4 to prevent term parturition.In naïve mice, myometrial neutrophil and monocyte numbers peaked at E18 and declined with the onset of parturition. In contrast, circulating monocytes did not change and although neutrophils were increased with pregnancy, they did not change across gestation. The myometrial mRNA and protein levels of most chemokines/cytokines, Cx-43 and COX-2 increased with, but not before, parturition.With RU486-induced parturition, myometrial and systemic neutrophil numbers increased before and myometrial monocyte numbers increased with parturition only. Myometrial chemokine/cytokine mRNA abundance increased with parturition, but protein levels peaked earlier at between 4.5 and 9h post RU486. Cx-43, but not COX-2, mRNA expression and protein levels increased prior to the onset of parturition.In mice treated with P4, the gestation-linked increase in myometrial monocyte, but not neutrophil, numbers was prevented and expression of Cx-43 and COX-2 was reduced. On E20 of P4 supplementation, myometrial chemokine/cytokine and leukocyte numbers, but not Cx-43 and COX-2 expression, increased.These data show that during pregnancy P4 controls myometrial monocyte infiltration, cytokine and prolabour factor synthesis via mRNA dependent and independent mechanisms and, with prolonged P4 supplementation, P4 action is repressed resulting in increased myometrial inflammation.
Zöllner J, Howe LG, Edey LF, et al., 2017, The response of the innate immune and cardiovascular systems to LPS in pregnant and nonpregnant mice., Biology of Reproduction, Vol: 97, Pages: 258-272, ISSN: 1529-7268
Sepsis is the leading cause of direct maternal mortality, but there are no data directly comparing the response to sepsis in pregnant and nonpregnant (NP) individuals. This study uses a mouse model of sepsis to test the hypothesis that the cardiovascular response to sepsis is more marked during pregnancy. Female CD1 mice had radiotelemetry probes implanted and were time mated. NP and day 16 pregnant CD-1 mice received intraperitoneal lipopolysaccharide (LPS; 10 μg, serotype 0111: B4). In a separate study, tissue and serum (for RNA, protein and flow cytometry studies), aorta and uterine vessels (for wire myography) were collected after LPS or vehicle control administration. Administration of LPS resulted in a greater fall in blood pressure in pregnant mice compared to NP mice. This occurred with similar changes in the circulating levels of cytokines, vasoactive factors, and circulating leukocytes, but with a greater monocyte and lesser neutrophil margination in the lungs of pregnant mice. Baseline markers of cardiac dysfunction and apoptosis as well as cytokine expression were higher in pregnant mice, but the response to LPS was similar in both groups as was the ex vivo assessment of vascular function. In pregnant mice, nonfatal sepsis is associated with a more marked hypotensive response but not a greater immune response. We conclude that endotoxemia induces a more marked hypotensive response in pregnant compared to NP mice. These changes were not associated with a more marked systemic inflammatory response in pregnant mice, although monocyte lung margination was greater. The more marked hypotensive response to LPS may explain the greater vulnerability to some infections exhibited by pregnant women.
Wilson MR, Petrie JE, Shaw MW, et al., 2017, High fat feeding protects mice from ventilator-induced lung injury, via neutrophil-independent mechanisms, Critical Care Medicine, Vol: 45, Pages: e831-e839, ISSN: 1530-0293
Objective:Obesity has a complex impact on acute respiratory distress syndrome patients, being associated with increased likelihood of developing the syndrome, but reduced likelihood of dying. We propose that such observations are potentially explained by a model in which obesity influences the iatrogenic injury that occurs subsequent to intensive care admission. This study therefore investigated whether fat-feeding protected mice from ventilator-induced lung injury (VILI). Design: In vivo study.Setting: University research laboratory.Subjects: Wildtype C57Bl/6 mice or TNF receptor 2 knockout mice, either fed a high fat diet for 12-14 weeks, or age-matched lean controls. Interventions: Anesthetized mice were ventilated with injurious high tidal volume ventilation for periods up to 180 minutes.Measurements and main results:Fat-fed mice showed clear attenuation of VILI in terms of respiratory mechanics, blood gases and pulmonary edema. Leukocyte recruitment and activation within the lungs were not significantly attenuated, nor were a host of circulating or intra-alveolar inflammatory cytokines. However, intra-alveolar matrix metalloproteinase (MMP) activity and levels of the MMP cleavage product sRAGE were significantly attenuated in fat-fed mice. This was associated with reduced stretch-induced CD147 expression on lung epithelial cells. Conclusions:Consumption of a high fat diet protects mice from VILI in a manner independent of neutrophil recruitment, which we postulate instead arises through blunted upregulation of CD147 expression and subsequent activation of intra-alveolar MMPs. These findings may open avenues for therapeutic manipulation in ARDS, and could have implications for understanding the pathogenesis of lung disease in obese patients.
Tatham KC, O'Dea KP, Romano R, et al., 2017, Intravascular donor monocytes play a central role in lung transplant ischaemia-reperfusion injury, Thorax, Vol: 73, Pages: 350-360, ISSN: 1468-3296
Rationale Primary graft dysfunction in lung transplant recipients derives from the initial, largely leukocyte-dependent, ischaemia-reperfusion injury. Intravascular lung-marginated monocytes have been shown to play key roles in experimental acute lung injury, but their contribution to lung ischaemia-reperfusion injury post transplantation is unknown.Objective To define the role of donor intravascular monocytes in lung transplant-related acute lung injury and primary graft dysfunction.Methods Isolated perfused C57BL/6 murine lungs were subjected to warm ischaemia (2 hours) and reperfusion (2 hours) under normoxic conditions. Monocyte retention, activation phenotype and the effects of their depletion by intravenous clodronate-liposome treatment on lung inflammation and injury were determined. In human donor lung transplant samples, the presence and activation phenotype of monocytic cells (low side scatter, 27E10+, CD14+, HLA-DR+, CCR2+) were evaluated by flow cytometry and compared with post-implantation lung function.Results In mouse lungs following ischaemia-reperfusion, substantial numbers of lung-marginated monocytes remained within the pulmonary microvasculature, with reduced L-selectin and increased CD86 expression indicating their activation. Monocyte depletion resulted in reductions in lung wet:dry ratios, bronchoalveolar lavage fluid protein, and perfusate levels of RAGE, MIP-2 and KC, while monocyte repletion resulted in a partial restoration of the injury. In human lungs, correlations were observed between pre-implantation donor monocyte numbers/their CD86 and TREM-1 expression and post-implantation lung dysfunction at 48 and 72 hours.Conclusions These results indicate that lung-marginated intravascular monocytes are retained as a ‘passenger’ leukocyte population during lung transplantation, and play a key role in the development of transplant-associated ischaemia-reperfusion injury.
Tan YY, O'Dea KP, Soni S, et al., 2017, Enhanced Recognition and Internalisation of Microvesicles by Lung-Marginated, Ly-6C(high) Monocytes During Endotoxaemia, Annual Meeting of the American-Society-for-Pharmacology-and-Experimental-Therapeutics (ASPET) at Experimental Biology Meeting, Publisher: FEDERATION AMER SOC EXP BIOL, ISSN: 0892-6638
Davies R, O'Dea KP, Soni S, et al., 2017, P362: Vasopressin alone and with noradrenaline attenuates TNF-α production in an in-vitro model of monocyte priming and deactivation, 37th International Symposium on Intensive Care and Emergency Medicine, Publisher: BioMed Central, ISSN: 1364-8535
Wilson MR, Wakabayashi K, Bertok S, et al., 2017, Inhibition of TNF receptor p55 by a domain antibody attenuates the initial phase of acid-induced lung injury in mice, Frontiers in Immunology, Vol: 8, ISSN: 1664-3224
Background: Tumor necrosis factor-α (TNF) is strongly implicated in the development ofacute respiratory distress syndrome (ARDS), but its potential as a therapeutic target has beenhampered by its complex biology. TNF signals through two receptors, p55 and p75, whichplay differential roles in pulmonary edema formation during ARDS. We have recentlyshown that inhibition of p55 by a novel domain antibody (dAb™) attenuated ventilator36induced lung injury. In the current study we explored the efficacy of this antibody in mousemodels of acid-induced lung injury, to investigate the longer consequences of treatment.Methods: We employed two acid-induced injury models, an acute ventilated model and aresolving spontaneously breathing model. C57BL/6 mice were pretreated intratracheally orintranasally with p55-targeting dAb or non-targeting ‘dummy’ dAb, 1 or 4 hours before acidinstillation.Results: Acid instillation in the dummy dAb group caused hypoxemia, increased respiratorysystem elastance, pulmonary inflammation and edema in both the ventilated and resolvingmodels. Pretreatment with p55-targeting dAb significantly attenuated physiological markersof ARDS in both models. p55-targeting dAb also attenuated pulmonary inflammation in theventilated model, with signs that altered cytokine production and leukocyte recruitmentpersisted beyond the very acute phase.Conclusions: These results demonstrate that the p55-targeting dAb attenuates lung injury andedema formation in models of ARDS induced by acid aspiration, with protection from asingle dose lasting up to 24 hours. Together with our previous data, the current study lends support towards the clinical targeting of p55 for patients with, or at risk of ARDS.
O'Dea KP, Porter J, TIrlapur N, et al., 2016, Circulating microvesicles are elevated acutely following major burns injury and associated with clinical severity, PLOS One, Vol: 11, ISSN: 1932-6203
Microvesicles are cell-derived signaling particles emerging as important mediators and biomarkers of systemic inflammation, but their production in severe burn injury patients has not been described. In this pilot investigation, we measured circulating microvesicle levels following severe burns, with severe sepsis patients as a comparator group.We hypothesized that levels of circulating vascular cell-derived microvesicles are elevated acutely following burns injury, mirroring clinical severity due to the early onset and prevalence of systemic inflammatory response syndrome (SIRS) in these patients. Blood samples were obtained from patients with moderate to severe thermal injury burns, with severe sepsis, and from healthy volunteers. Circulating microvesicles derived from total leukocytes, granulocytes, monocytes, and endothelial cells were quantified in plasma by flow cytometry.All circulating microvesicle subpopulations were elevated in burns patients on day of admission (day 0) compared to healthy volunteers (leukocyte-microvesicles: 3.5-fold, p=0.005; granulocyte-microvesicles: 12.8-fold, p<0.0001; monocyte-microvesicles: 20.4-fold, p<0.0001; endothelial- microvesicles: 9.6-fold, p=0.01), but decreased significantly by day 2. Microvesicle levels were increased with severe sepsis, but less consistently between patients. Leukocyte- and granulocyte-derived microvesicles on day 0 correlated with clinical assessment scores and were higher in burns ICU non-survivors compared to survivors (leukocyte MVs 4.6 fold, p=0.002; granulocyte MVs 4.8 fold, p=0.003). Mortality prediction analysis of area under receiver operating characteristic curve was 0.92 (p=0.01) for total leukocyte microvesicles and 0.85 (p=0.04) for granulocyte microvesicles. These findings demonstrate, for the first t
Edey LF, O'Dea KP, Herbert BR, et al., 2016, The local and systemic immune response to intrauterine LPS in the prepartum mouse, Biology of Reproduction, Vol: 95, Pages: 1-10, ISSN: 1529-7268
Inflammation plays a key role in human term and preterm labor (PTL). Intrauterine LPS has been widely used to model inflammation-induced complications of pregnancy, including PTL. It has been shown to induce an intense myometrial inflammatory cell infiltration, but the role of LPS-induced inflammatory cell activation in labor onset and fetal demise is unclear. We investigated this using a mouse model of PTL, where an intrauterine injection of 10 μg of LPS (serotype 0111:B4) was given at E16 of CD1 mouse pregnancy. This dose induced PTL at an average of 12.7 h postinjection in association with 85% fetal demise. Flow cytometry showed that LPS induced a dramatic systemic inflammatory response provoking a rapid and marked leucocyte infiltration into the maternal lung and liver in association with increased cytokine levels. Although there was acute placental inflammatory gene expression, there was no corresponding increase in fetal brain inflammatory gene expression until after fetal demise. There was marked myometrial activation of NFκB and MAPK/AP-1 systems in association with increased chemokine and cytokine levels, both of which peaked with the onset of parturition. Myometrial macrophage and neutrophil numbers were greater in the LPS-injected mice with labor onset only; prior to labor, myometrial neutrophils and monocytes numbers were greater in PBS-injected mice, but this was not associated with an earlier onset of labor. These data suggest that intrauterine LPS induces parturition directly, independent of myometrial inflammatory cell infiltration, and that fetal demise occurs without fetal inflammation. Intrauterine LPS provokes a marked local and systemic inflammatory response but with limited inflammatory cell infiltration into the myometrium or placenta.
Sivakumar S, Taccone FS, Desai KA, et al., 2016, ESICM LIVES 2016: part two : Milan, Italy. 1-5 October 2016, Intensive Care Medicine Experimental, Vol: 4, ISSN: 2197-425X
Takata M, Soni S, Wilson MR, et al., 2016, Alveolar macrophage-derived microvesicles mediate acute lung injury, Thorax, Vol: 71, Pages: 1020-1029, ISSN: 1468-3296
Background Microvesicles (MVs) are important mediators of intercellular communication, packaging a variety of molecular cargo. They have been implicated in the pathophysiology of various inflammatory diseases; yet, their role in acute lung injury (ALI) remains unknown.Objectives We aimed to identify the biological activity and functional role of intra-alveolar MVs in ALI.Methods Lipopolysaccharide (LPS) was instilled intratracheally into C57BL/6 mice, and MV populations in bronchoalveolar lavage fluid (BALF) were evaluated. BALF MVs were isolated 1 hour post LPS, assessed for cytokine content and incubated with murine lung epithelial (MLE-12) cells. In separate experiments, primary alveolar macrophage-derived MVs were incubated with MLE-12 cells or instilled intratracheally into mice.Results Alveolar macrophages and epithelial cells rapidly released MVs into the alveoli following LPS. At 1 hour, the dominant population was alveolar macrophage-derived, and these MVs carried substantive amounts of tumour necrosis factor (TNF) but minimal amounts of IL-1β/IL-6. Incubation of these mixed MVs with MLE-12 cells induced epithelial intercellular adhesion molecule-1 (ICAM-1) expression and keratinocyte-derived cytokine release compared with MVs from untreated mice (p<0.001). MVs released in vitro from LPS-primed alveolar macrophages caused similar increases in MLE-12 ICAM-1 expression, which was mediated by TNF. When instilled intratracheally into mice, these MVs induced increases in BALF neutrophils, protein and epithelial cell ICAM-1 expression (p<0.05).Conclusions We demonstrate, for the first time, the sequential production of MVs from different intra-alveolar precursor cells during the early phase of ALI. Our findings suggest that alveolar macrophage-derived MVs, which carry biologically active TNF, may play an important role in initiating ALI.
Tatham KC, O'Dea KP, Romana R, et al., 2016, Retention And Activation Of Donor Vascular Monocytes In Transplanted Lungs Suggests A Central Role In Primary Graft Dysfunction, American Thoracic Society Conference 2016
Tatham KC, O'Dea KP, Romano R, et al., 2016, Retention And Activation Of Donor Vascular Monocytes In Transplanted Lungs Suggests A Central Role In Primary Graft Dysfunction, International Conference of the American-Thoracic-Society (ATS), Publisher: AMER THORACIC SOC, ISSN: 1073-449X
Wilson M, Petrie J, Shaw M, et al., 2016, High Fat Feeding Protects Mice From Ventilator-Induced Lung Injury Via A Neutrophil-Independent Mechanism, International Conference of the American-Thoracic-Society (ATS), Publisher: AMER THORACIC SOC, ISSN: 1073-449X
Tirlapur N, O'Dea KP, Takata M, 2016, Human Neutrophil-Derived Microvesicles Activate Pulmonary Endothelial Cells In An In Vitro Model Of Pulmonary Microvascular Inflammation, International Conference of the American-Thoracic-Society (ATS), Publisher: AMER THORACIC SOC, ISSN: 1073-449X
Patel BV, Tatham KC, Wilson MR, et al., 2015, In vivo compartmental analysis of leukocytes in mouse lungs, American Journal of Physiology-Lung Cellular and Molecular Physiology, Vol: 309, Pages: L639-L652, ISSN: 1522-1504
The lung has a unique structure consisting of three functionally different compartments (alveolar, interstitial, and vascular) situated in an extreme proximity. Current methods to localize lung leukocytes using bronchoalveolar lavage and/or lung perfusion have significant limitations for determination of location and phenotype of leukocytes. Here we present a novel method using in vivo antibody labelling to enable accurate compartmental localization/quantification and phenotyping of mouse lung leukocytes. Anesthetized C57BL/6 mice received combined in vivo intravenous and intratracheal labelling with fluorophore-conjugated anti-CD45 antibodies, and lung single cell suspensions were analyzed by flow cytometry. The combined in vivo intravenous and intratracheal CD45 labelling enabled robust separation of the alveolar, interstitial, and vascular compartments of the lung. In naive mice, the alveolar compartment consisted predominantly of resident alveolar macrophages. The interstitial compartment, gated by events negative for both intratracheal and intravenous CD45 staining, showed two conventional dendritic cell populations, as well as a Ly6C(lo) monocyte population. Expression levels of MHCII on these interstitial monocytes were much higher than the vascular Ly6C(lo) monocyte populations. In mice exposed to acid-aspiration induced lung injury, this protocol also clearly distinguished the three lung compartments showing the dynamic trafficking of neutrophils and exudative monocytes across the lung compartments during inflammation and resolution. This simple in vivo dual labelling technique substantially increases the accuracy and depth of lung flow cytometric analysis, facilitates a more comprehensive examination of lung leukocyte pools, and enables the investigation of previously poorly defined 'interstitial' leukocyte populations during models of inflammatory lung diseases.
O'Callaghan DJ, O'Dea KP, Scott AJ, et al., 2015, Monocyte Tumor Necrosis Factor-α-Converting Enzyme Catalytic Activity and Substrate Shedding in Sepsis and Noninfectious Systemic Inflammation., Critical Care Medicine, Vol: 43, Pages: 1375-1385, ISSN: 1530-0293
OBJECTIVES: To determine the effect of severe sepsis on monocyte tumor necrosis factor-α-converting enzyme baseline and inducible activity profiles. DESIGN: Observational clinical study. SETTING: Mixed surgical/medical teaching hospital ICU. PATIENTS: Sixteen patients with severe sepsis, 15 healthy volunteers, and eight critically ill patients with noninfectious systemic inflammatory response syndrome. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Monocyte expression of human leukocyte antigen-D-related peptide, sol-tumor necrosis factor production, tumor necrosis factor-α-converting enzyme expression and catalytic activity, tumor necrosis factor receptor 1 and 2 expression, and shedding at 48-hour intervals from day 0 to day 4, as well as p38-mitogen activated protein kinase expression. Compared with healthy volunteers, both sepsis and systemic inflammatory response syndrome patients' monocytes expressed reduced levels of human leukocyte antigen-D-related peptide and released less sol-tumor necrosis factor on in vitro lipopolysaccharide stimulation, consistent with the term monocyte deactivation. However, patients with sepsis had substantially elevated levels of basal tumor necrosis factor-α-converting enzyme activity that were refractory to lipopolysaccharide stimulation and this was accompanied by similar changes in p38-mitogen activated protein kinase signaling. In patients with systemic inflammatory response syndrome, monocyte basal tumor necrosis factor-α-converting enzyme, and its induction by lipopolysaccharide, appeared similar to healthy controls. Changes in basal tumor necrosis factor-α-converting enzyme activity at day 0 for sepsis patients correlated with Acute Physiology and Chronic Health Evaluation II score and the attenuated tumor necrosis factor-α-converting enzyme response to lipopolysaccharide was associated with increased mortality. Similar changes in monocyte tumor necrosis factor-α-converting enz
Woods SJ, Waite AAC, O'Dea KP, et al., 2015, Kinetic profiling of in vivo lung cellular inflammatory responses to mechanical ventilation, AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY, Vol: 308, Pages: L912-L921, ISSN: 1040-0605
Patel BV, Tatham KC, Wilson MR, et al., 2015, In Vivo Compartmental Labeling of the Mouse Lung, American Thoracic Society, Publisher: ATS Journals, Pages: A3926-A3926, ISSN: 1073-449X
ShareModerators: G.N. Maksym, PhD, R.S. Harris, MD, J.C. Sieren, PhDSession Info: Mini Symposium, [C18] STATE OF PLAY IN RESPIRATORY STRUCTURE AND FUNCTION: SEEING IS BELIEVINGDay/Date: Tuesday, May 19, 2015Session Time: 9:30 AM - 11:30 AMRoom: Mile High Ballroom 2C/3C (Lower Level)Location: Colorado Convention CenterIn Vivo Compartmental Labeling of the Mouse Lung, [Publication Page: A3926]B.V. Patel, MBBS MRCP FRCA PhD, K.C. Tatham, MBBS, M.R. Wilson, PhD, K.P. O'Dea, PhD, M. Takata, MDLondon/UKRationaleCurrent methods for compartmental analyses of lungs using bronchoalveolar lavage (BAL) and/or lung perfusion have significant limitations for accurate determination of location and phenotype of lung leukocytes. BAL retrieves only <25% of intra-alveolar cells (1) and lung perfusion fails to remove vascular marginated cell populations (2). We present a novel method of in vivo antibody labelling to facilitate the accurate compartmental investigation of mouse lung leukocytes by flow cytometry.MethodsAnesthetized C57BL6 mice underwent tracheostomy and venous cannulation. An anti-CD45 antibody (PE-conjugated) was injected intravenously and allowed to circulate for 5 minutes. Mice were then exsanguinated and lungs immediately flooded intratracheally with another anti-CD45 antibody (PE-Cy5-conjugated). Lungs were harvested, and single cell suspensions prepared for flow cytometric analysis using a panel of myeloid markers. This in vivo labelling protocol was also applied to mice exposed to acid-aspiration induced lung injury (3). To confirm the separation between the vascular and interstitial cell populations, a group of mice underwent intravenous labelling alone followed by lung perfusion for 15 minutes using an isolated-perfused lung apparatus (2).ResultsThe combined in vivo intravenous and intratracheal CD45 labelling enabled robust separation of the alveolar, interstitial, and vascular compartments of the lung by flow cytometry, with <0.3% of leukocytes staining
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