Mitochondrial DNA and TLR9 Signaling Is Not Involved in Mechanical Ventilation-Induced Inflammation.

Exogenous administration of mitochondrial DNA (mtDNA) causes inflammatory lung injury in a toll-like receptor (TLR) 9-dependent manner. We investigated whether mechanical ventilation results in endogenous release of mtDNA and whether TLR9 plays a role in the pulmonary inflammatory response induced by mechanical ventilation.Wild-type and TLR9/ C57bl/6 mice were ventilated with low (8 mL/kg) and high (32 mL/kg) tidal volumes for 4 hours. Levels of nuclear DNA and mtDNA in bronchoalveolar lavage fluid, as well as pulmonary concentrations of keratinocyte-derived chemokine, interleukin-1ß, and interleukin-6, were determined.Cytokine and nuclear DNA, but not mtDNA, levels were increased after mechanical ventilation with both tidal volumes. Cytokine concentrations were similar between wild-type and TLR9/ mice. Mechanical ventilation does not result in the release of mtDNA, and TLR9 is not involved in mechanical ventilation-induced inflammation.

Plasma levels of danger-associated molecular patterns are associated with immune suppression in trauma patients.

PURPOSE: Danger-associated molecular patterns (DAMPs) released of trauma could contribute to an immune suppressed state that renders patients vulnerable towards nosocomial infections. We investigated DAMP release in trauma patients, starting in the prehospital phase, and assessed its relationship with immune suppression and nosocomial infections. METHODS: Blood was obtained from 166 adult trauma patients at the trauma scene, emergency room (ER), and serially afterwards. Circulating levels of DAMPs and cytokines were determined. Immune suppression was investigated by determination of HLA-DRA gene expression and ex vivo lipopolysaccharide-stimulated cytokine production. RESULTS: Compared with healthy controls, plasma levels of nuclear DNA (nDNA) and heat shock protein-70 (HSP70) but not mitochondrial DNA were profoundly increased immediately following trauma and remained elevated for 10 days. Plasma cytokines were increased at the ER, and levels of anti-inflammatory IL-10 but not of pro-inflammatory cytokines peaked at this early time-point. HLA-DRA expression was attenuated directly after trauma and did not recover during the follow-up period. Plasma nDNA (r = -0.24, p = 0.006) and HSP70 (r = -0.38, p < 0.0001) levels correlated negatively with HLA-DRA expression. Ex vivo cytokine production revealed an anti-inflammatory phenotype already at the trauma scene which persisted in the following days, characterized by attenuated TNF-α and IL-6, and increased IL-10 production. Finally, higher concentrations of nDNA and a further decrease of HLA-DRA expression were associated with infections. CONCLUSIONS: Plasma levels of DAMPs are associated with immune suppression, which is apparent within minutes/hours following trauma. Furthermore, aggravated immune suppression during the initial phase following trauma is associated with increased susceptibility towards infections.

Behavior of neuropathic pain in mice following chronic constriction injury comparing silk and catgut ligatures.

INTRODUCTION: Neuropathic pain is defined as pain arising as a direct consequence of a lesion or disease affecting the somatosensory system and is common after surgery. Neuropathic pain can persist without an obvious injury. In this study we aim to validate a murine chronic constriction injury model as a model for neuropathic pain research and determine if silk or catgut ligatures induced most stable neuropathic pain behavior. METHODS: In this study mice underwent chronic constriction or sham surgery. Mice were tested on cutaneous hyperalgesia with the cumulative reaction time in the acetone test, on allodynia with the cumulative reaction time and number of lifts in the cold plate test and the maximal force before withdrawal in von Frey test. RESULTS: In the acetone test neuropathic pain was seen in CCI mice, but not in sham mice. Hyperalgesia was present postoperatively in CCI mice compared with preoperatively. In the cold plate test cumulative reaction time and number of lifts were higher in the ipsilateral hind paw than in the contralateral hind paw and sham mice. Postoperative measurements were higher than preoperatively. In the von Frey test the postoperative measurements were lower in the ipsilateral hind paw than preoperatively, while the contralateral hind paw showed an increase in maximal force before withdrawal. The contralateral hind paw showed more difference with sham mice than the ipsilateral hind paw. Silk ligatures showed more stable neuropathic pain behavior. In the acetone test, the cold plate test and the von Frey test the mice scored higher on neuropathic pain having silk ligatures, compared with catgut ligatures. CONCLUSION: In this study we validated a murine CCI model for neuropathic pain behavior. In the murine CCI model it appears that silk ligatures demonstrate more stable neuropathic pain behaviors than catgut ligatures in de CCI model.

Soluble urokinase-type plasminogen activator levels are related to plasma cytokine levels but have low predictive value for mortality in trauma patients.

INTRODUCTION: Soluble urokinase-type plasminogen activator (suPAR) represents a marker for immune activation and has predictive value in critically ill patients. The kinetics of suPAR and its correlation with the immune response and outcome in trauma patients are unknown. METHODS: Plasma concentrations of inflammatory cytokines and suPAR were determined in adult trauma patient (n = 69) samples obtained by the Helicopter Emergency Medical Services at arrival at the emergency department (ED) and at days 1, 3, 5, 7, 10, and 14. RESULTS: Initial suPAR levels were unrelated to injury severity score and higher in nonsurvivors compared with survivors, although no difference was observed between early and late mortality. The area under the receiver operating characteristic curve to predict mortality was 0.6 (95% confidence interval, 0.48-0.72). Soluble urokinase-type plasminogen activator levels increased over time in 94% of patients, although suPAR increase did not precede death. Tumor necrosis factor α at the ED correlated with suPAR at that time point, whereas concentrations of other proinflammatory cytokines at the ED correlated with suPAR levels at days 1 and 5. CONCLUSIONS: After trauma, initial suPAR plasma concentrations are higher in nonsurvivors compared with survivors, but its predictive value is low. Soluble urokinase-type plasminogen activator levels increase over time after trauma, and concentrations at later time points are related to cytokine levels at the ED.

Circulating iFABP Levels as a marker of intestinal damage in trauma patients.

Both the initial trauma and the subsequent hemodynamic instability may contribute to intestinal damage, which is of great importance in (immunological) posttrauma complications. This study assesses intestinal damage using the biomarker intestinal Fatty Acid Binding Protein (iFABP) in trauma patients during the first days of their hospital admission and the risk factors involved. Plasma iFABP levels were measured in blood samples obtained from adult multiple trauma patients (n = 93) at the trauma scene by the Helicopter Emergency Medical Services, at arrival at the emergency department (ED), and at days 1, 3, 5, 7, 10, and 14 after trauma and related to injury severity and hemodynamic parameters. Plasma iFABP concentrations showed highest levels immediately after trauma at time points Helicopter Emergency Medical Services and ED. Nonsurvivors demonstrated higher iFABP levels at the ED compared with survivors. Furthermore, iFABP values at the ED correlated with Injury Severity Scores, and patients suffering from abdominal trauma demonstrated significantly higher iFABP concentrations in comparison with patients with other types of trauma or healthy controls. Also, patients presenting with a mean arterial pressure (MAP) less than 70 mmHg at the ED demonstrated significantly higher plasma iFABP concentrations in comparison with patients with a normal (70-99 mmHg) or high (>100 mmHg) MAP or healthy controls. Finally, patients with a low hemoglobin (Hb) (<80% of reference value) displayed significantly higher iFABP concentrations in comparison with patients with a normal Hb or healthy controls. Plasma iFABP levels, indicative of intestinal injury, are increased immediately after trauma in patients with abdominal trauma, low MAP, or low Hb and are related to the severity of the trauma. As intestinal injury is suggested to be related to late complications, such as multiorgan dysfunction syndrome or sepsis in trauma patients, strategies to prevent intestinal damage after trauma could be of benefit to these patients.

Mechanical ventilation of mice under general anesthesia during experimental procedures.

Mechanical ventilation is frequently used in patients under general anesthesia during invasive procedures. Invasive animal experiments similarly require the maintenance of normal hemodynamic and pulmonary parameters during long-term general anesthesia. The authors describe a method for mechanical ventilation of mice. Mice were ventilated and monitored for up to 8 h of general anesthesia during surgery. Hemodynamic and pulmonary parameters remained within the normal ranges. The authors believe that this ventilation technique can be of great value for experimental procedures in mice that require general anesthesia.

Blueprints of signaling interactions between pattern recognition receptors: implications for the design of vaccine adjuvants.

Innate immunity activation largely depends on recognition of microorganism structures by Pattern Recognition Receptors (PRRs). PRR downstream signaling results in production of pro- and anti-inflammatory cytokines and other mediators. Moreover, PRR engagement in antigen-presenting cells initiates the activation of adaptive immunity. Recent reports suggest that for the activation of innate immune responses and initiation of adaptive immunity, synergistic effects between two or more PRRs are necessary. No systematic analysis of the interaction between the major PRR pathways were performed to date. In this study, a systematical analysis of the interactions between PRR signaling pathways was performed. PBMCs derived from 10 healthy volunteers were stimulated with either a single PRR ligand or a combination of two PRR ligands. Known ligands for the major PRR families were used: Toll-like receptors (TLRs), C-type lectin receptors (CLRs), NOD-like receptors (NLRs), and RigI-helicases. After 24 h of incubation, production of tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1ß), IL-6, and IL-10 was measured in supernatants by enzyme-linked immunosorbent assay (ELISA). The consistency of the PRR interactions (both inhibitory and synergistic) between the various individuals was assessed. A number of PRR-dependent signaling interactions were found to be consistent, both between individuals and with regard to multiple cytokines. The combinations of TLR2 and NOD2, TLR5 and NOD2, TLR5 and TLR3, and TLR5 and TLR9 acted as synergistic combinations. Surprisingly, inhibitory interactions between TLR4 and TLR2, TLR4 and Dectin-1, and TLR2 and TLR9 as well as TLR3 and TLR2 were observed. These consistent signaling interactions between PRR combinations may represent promising targets for immunomodulation and vaccine adjuvant development.

IL-1ß processing in mechanical ventilation-induced inflammation is dependent on neutrophil factors rather than caspase-1.

PURPOSE: Mechanical ventilation can cause ventilator-induced lung injury, characterized by a sterile inflammatory response in the lungs resulting in tissue damage and respiratory failure. The cytokine interleukin-1ß (IL-1ß) is thought to play an important role in the pathogenesis of ventilator-induced lung injury. Cleavage of the inactive precursor pro-IL-1ß to form bioactive IL-1ß is mediated by several types of proteases, of which caspase-1, activated within the inflammasome, is the most important. Herein, we studied the roles of IL-1ß, caspase-1 and neutrophil factors in the mechanical ventilation-induced inflammatory response in mice. METHODS: Untreated wild-type mice, IL-1αß knockout and caspase-1 knockout mice, pralnacasan (a selective caspase-1 inhibitor)-treated mice, anti-keratinocyte-derived chemokine (KC)-treated mice and cyclophosphamide-treated neutrophil-depleted wild-type mice were ventilated using clinically relevant ventilator settings (tidal volume 8 ml/kg). The lungs and plasma were collected to determine blood gas values, cytokine profiles and neutrophil influx. RESULTS: Mechanical ventilation resulted in increased pulmonary concentrations of IL-1ß and KC and increased pulmonary neutrophil influx compared with non-ventilated mice. Ventilated IL-1αß knockout mice did not demonstrate this increase in cytokines. No significant differences were observed between wild-type and caspase-1-deficient or pralnacasan-treated mice. In contrast, in anti-KC antibody-treated mice and neutropenic mice, inflammatory parameters decreased in comparison with ventilated non-treated mice. CONCLUSIONS: Our results illustrate that IL-1 is indeed an important cytokine in the inflammatory cascade induced by mechanical ventilation. However, the inflammasome/caspase-1 appears not to be involved in IL-1ß processing in this type of inflammatory response. The attenuated inflammatory response observed in ventilated anti-KC-treated and neutropenic mice suggests that IL-1ß processing in mechanical ventilation-induced inflammation is mainly mediated by neutrophil factors.

Toll-like receptor 4 signaling in ventilator-induced diaphragm atrophy.

BACKGROUND: Mechanical ventilation induces diaphragm muscle atrophy, which plays a key role in difficult weaning from mechanical ventilation. The signaling pathways involved in ventilator-induced diaphragm atrophy are poorly understood. The current study investigated the role of Toll-like receptor 4 signaling in the development of ventilator-induced diaphragm atrophy. METHODS: Unventilated animals were selected for control: wild-type (n = 6) and Toll-like receptor 4 deficient mice (n = 6). Mechanical ventilation (8 h): wild-type (n = 8) and Toll-like receptor 4 deficient (n = 7) mice.Myosin heavy chain content, proinflammatory cytokines, proteolytic activity of the ubiquitin-proteasome pathway, caspase-3 activity, and autophagy were measured in the diaphragm. RESULTS: Mechanical ventilation reduced myosin content by approximately 50% in diaphragms of wild-type mice (P less than 0.05). In contrast, ventilation of Toll-like receptor 4 deficient mice did not significantly affect diaphragm myosin content. Likewise, mechanical ventilation significantly increased interleukin-6 and keratinocyte-derived chemokine in the diaphragm of wild-type mice, but not in ventilated Toll-like receptor 4 deficient mice. Mechanical ventilation increased diaphragmatic muscle atrophy factor box transcription in both wild-type and Toll-like receptor 4 deficient mice. Other components of the ubiquitin-proteasome pathway and caspase-3 activity were not affected by ventilation of either wild-type mice or Toll-like receptor 4 deficient mice. Mechanical ventilation induced autophagy in diaphragms of ventilated wild-type mice, but not Toll-like receptor 4 deficient mice. CONCLUSION: Toll-like receptor 4 signaling plays an important role in the development of ventilator-induced diaphragm atrophy, most likely through increased expression of cytokines and activation of lysosomal autophagy.

Effects of vagus nerve stimulation and vagotomy on systemic and pulmonary inflammation in a two-hit model in rats.

Pulmonary inflammation contributes to ventilator-induced lung injury. Sepsis-induced pulmonary inflammation (first hit) may be potentiated by mechanical ventilation (MV, second hit). Electrical stimulation of the vagus nerve has been shown to attenuate inflammation in various animal models through the cholinergic anti-inflammatory pathway. We determined the effects of vagotomy (VGX) and vagus nerve stimulation (VNS) on systemic and pulmonary inflammation in a two-hit model. Male Sprague-Dawley rats were i.v. administered lipopolysaccharide (LPS) and subsequently underwent VGX, VNS or a sham operation. 1 hour following LPS, MV with low (8 mL/kg) or moderate (15 mL/kg) tidal volumes was initiated, or animals were left breathing spontaneously (SP). After 4 hours of MV or SP, rats were sacrificed. Cytokine and blood gas analysis was performed. MV with 15, but not 8 mL/kg, potentiated the LPS-induced pulmonary pro-inflammatory cytokine response (TNF-α, IL-6, KC: p<0.05 compared to LPS-SP), but did not affect systemic inflammation or impair oxygenation. VGX enhanced the LPS-induced pulmonary, but not systemic pro-inflammatory cytokine response in spontaneously breathing, but not in MV animals (TNF-α, IL-6, KC: p<0.05 compared to SHAM), and resulted in decreased pO(2) (p<0.05 compared to sham-operated animals). VNS did not affect any of the studied parameters in both SP and MV animals. In conclusion, MV with moderate tidal volumes potentiates the pulmonary inflammatory response elicited by systemic LPS administration. No beneficial effects of vagus nerve stimulation performed following LPS administration were found. These results questions the clinical applicability of stimulation of the cholinergic anti-inflammatory pathway in systemically inflamed patients admitted to the ICU where MV is initiated.

Hemorrhagic shock-induced endothelial cell activation in a spontaneous breathing and a mechanical ventilation hemorrhagic shock model is induced by a proinflammatory response and not by hypoxia.

INTRODUCTION: The interaction between neutrophils and activated endothelium is essential for the development of multiple organ dysfunction in patients with hemorrhagic shock (HS). Mechanical ventilation frequently is used in patients with HS. The authors sought to investigate the consequences of mechanical ventilation of mice subjected to HS on microvascular endothelial activation in the lung and kidney. METHODS: Anesthetized wild type C57BL/6 male mice were subjected to controlled hemorrhage; subgroups of mice were mechanically ventilated during the HS insult. To study the effect of acute hypoxia on the mice, the animals were housed in hypoxic cages. Gene expression levels was assessed by quantitative real-time polymerase chain reaction. Protein expression was assessed by immunohistochemistry and enzyme-linked immunosorbent assay. RESULTS: Ninety minutes after the shock induction, a vascular bed-specific, heterogeneous proinflammatory endothelial activation represented by E-selectin, vascular cell adhesion molecule 1, and intercellular adhesion molecule 1 expression was seen in kidney and lung. No differences in adhesion molecules between the spontaneously breathing and mechanically ventilated mice were found. Concentrations of the proinflammatory cytokines chemokine (C-X-C motif) ligand 1 (11.0-fold) and interleukin-6 (21.7-fold) were increased after 90 min of HS. Two hours of 6% oxygen did not induce the expression of E-selectin, vascular cell adhesion molecule 1, and intercellular adhesion molecule 1 in the kidneys and the lung. CONCLUSIONS: Hemorrhagic shock leads to an early and reversible proinflammatory endothelial activation in kidney and lung. HS-induced endothelial activation is not changed by mechanical ventilation during the shock phase. Hypoxia alone does not lead to endothelial activation. The observed proinflammatory endothelial activation is mostly ischemia- or reperfusion-dependent and not related to hypoxia.

A single recruitment maneuver in ventilated critically ill children can translocate pulmonary cytokines into the circulation.

INTRODUCTION: Recruitment maneuvers (RMs) are advocated to prevent pulmonary collapse during low tidal volume ventilation and improve oxygenation. However, convincing clinical evidence for improved outcome is lacking. Recent experimental studies demonstrate that RMs translocate pulmonary inflammatory mediators into the circulation. To determine whether a single RM in ventilated children affects pulmonary and systemic cytokine levels, we performed a prospective intervention study. METHODS: Cardiorespiratory stable ventilated patients (0.5-45 months, n = 7) with acute lung injury were subjected to an RM determining opening and closing pressures (peak inspiratory pressure < or =45 cmH(2)O, positive end expiratory pressure (PEEP) < or =30 cmH(2)O). Before and after RM, cardiorespiratory parameters and ventilator settings were recorded, blood gas analysis performed, and bronchoalveolar lavage fluid and plasma TNF-alpha, IL-1beta, IL-6, IL-8, and IL-10 concentrations were determined. RESULTS: Fifteen minutes after the RM, an increase was observed in plasma tumor necrosis factor-alpha (400% +/- 390% of baseline, P = .04), IL-6 (120% +/- 35%, P = .08), and IL-1beta (520% +/- 535%, P = .04), which decreased at T = 60 minutes, hence indicative of translocation. Recruitment maneuver did not change the plasma levels of the anti-inflammatory IL-10 (105% +/- 12%, P = .5). Apart from a nonsignificant increase of IL-8 after 360 minutes (415% +/- 590%,P = .1), bronchoalveolar cytokine levels were not influenced by the RM. No increase in oxygenation or improvement of lung kinetics was observed. CONCLUSIONS: A single RM can translocate pro-inflammatory cytokines from the alveolar space into the systemic circulation in ventilated critically ill children.

Mechanical ventilation induces a Toll/interleukin-1 receptor domain-containing adapter-inducing interferon beta-dependent inflammatory response in healthy mice.

BACKGROUND: Mechanical ventilation (MV) can induce lung injury. Proinflammatory cytokines have been shown to play an important role in the development of ventilator-induced lung injury. Previously, the authors have shown a role for Toll-like receptor 4 signaling. The current study aims to investigate the role of Toll/interleukin-1 receptor domain-containing adapter-inducing interferon-beta (TRIF), a protein downstream of Toll-like receptors, in the development of the inflammatory response after MV in healthy mice. METHODS: Wild-type C57BL6 and TRIF mutant mice were mechanically ventilated for 4 h. Lung tissue and plasma was used to investigate changes in cytokine profile, leukocyte influx, and nuclear factor-kappaB activity. In addition, experiments were performed to assess the role of TRIF in changes in cardiopulmonary physiology after MV. RESULTS: MV significantly increased messenger RNA expression of interleukin (IL)-1beta in wild-type mice, but not in TRIF mutant mice. In lung homogenates, MV increased levels of IL-1alpha, IL-1beta, and keratinocyte-derived chemokine in wild-type mice. In contrast, in TRIF mutant mice, only a minor increase in IL-1beta and keratinocyte-derived chemokine was found after MV. Nuclear factor-kappaB activity after MV was significantly lower in TRIF mutant mice compared with wild-type mice. In plasma, MV increased levels of IL-6 and keratinocyte-derived chemokine. In TRIF mutant mice, no increase of IL-6 was found after MV, and the increase in keratinocyte-derived chemokine appeared less pronounced. TRIF deletion did not affect cardiopulmonary physiology after MV. CONCLUSIONS: The current study supports a prominent role for TRIF in the development of the pulmonary and systemic inflammatory response after MV.

The oxygenation ratio during mechanical ventilation in children: the role of tidal volume and positive end-expiratory pressure.

OBJECTIVE: The objective of this study is to analyze the role of tidal volume (Vt) and positive end-expiratory pressure on the oxygenation ratio (OR) (Pao(2)/Fio(2)) during mechanical ventilation (MV) in children with a normal pulmonary gas exchange on admission. METHODS: A retrospective cohort study of children with an admission OR greater than 300 mm Hg and duration of MV greater than 48 hours (n = 96) was done. We analyzed Vt, Fio(2), Pao(2), and positive end-expiratory pressure and calculated Vt (mL/kg) and Pao(2)/Fio(2) based on the measured Vt and weight. Patients were divided into group 1, Vt less than 9 mL/kg (n = 24); 2, Vt 9 to 12 mL/kg (n = 58); and 3, Vt 12 mL/kg or higher (n = 14). RESULTS: Baseline characteristics and OR were comparable. Forty-one percent of patients developed OR less than 300 mm Hg. The proportion of patients developing an OR less than 300 mm Hg was lowest in group 1 and highest in group 3, and differences became more pronounced with longer MV duration: 56%, 58%, and 89% on day 5; 29%, 65%, and 100% on day 7 (P = .05); 0%, 40%, and 100% on day 10 (P = .03). In patients maintaining an OR greater than 300 mm Hg during 10 days of MV, Vt was 9.3 +/- 1.0 vs 12.7 +/- 4.8 mL/kg in patients developing an OR less than 300 mm Hg (P = .05). Mechanical ventilation duration was longer in children developing OR less than 300 mm Hg (P < .01). Positive end-expiratory pressure levels were not significantly different between groups. CONCLUSION: In ventilated children, Vt was greater than 9 mL/kg were associated with increased development of an OR less than 300 mm Hg and longer duration of MV.

Low-tidal-volume mechanical ventilation induces a toll-like receptor 4-dependent inflammatory response in healthy mice.

BACKGROUND: Mechanical ventilation (MV) can induce ventilator-induced lung injury. A role for proinflammatory pathways has been proposed. The current studies analyzed the roles of Toll-like receptor (TLR) 4 and TLR2 involvement in the inflammatory response after MV in the healthy lung. METHODS: Wild-type (WT) C57BL6, TLR4 knockout (KO), and TLR2 KO mice were mechanically ventilated for 4 h. Bronchoalveolar lavage fluid was analyzed for presence of endogenous ligands. Lung homogenates were used to investigate changes in TLR4 and TLR2 expression. Cytokines were measured in lung homogenate and plasma, and leukocytes were counted in lung tissue. RESULTS: MV significantly increased endogenous ligands for TLR4 in bronchoalveolar lavage fluid and relative messenger RNA expression of TLR4 and TLR2 in lung tissue. In lung homogenates, MV in WT mice increased levels of keratinocyte-derived chemokine, interleukin (IL)-1alpha, and IL-1beta. In TLR4 KO mice, MV increased IL-1alpha but not IL-1beta, and the increase in keratinocyte-derived chemokine was less pronounced. In plasma, MV in WT mice increased levels of IL-6, keratinocyte-derived chemokine, and tumor necrosis factor alpha. In TLR4 KO mice, MV did not increase levels of IL-6 or tumor necrosis factor alpha, and the response of keratinocyte-derived chemokine was less pronounced. MV in TLR2 KO mice did not result in different cytokine levels compared with WT mice. In WT and TLR2 KO mice, but not in TLR4 KO mice, MV increased the number of pulmonary leukocytes. CONCLUSIONS: The current study supports a role for TLR4 in the inflammatory reaction after short-term MV in healthy lungs. Increasing the understanding of the innate immune response to MV may lead to future treatment advances in ventilator-induced lung injury, in which TLR4 may serve as a therapeutic target.

Hypercapnic acidosis attenuates the pulmonary innate immune response in ventilated healthy mice.

BACKGROUND: Mechanical ventilation with small tidal volumes reduces the development of ventilator-induced lung injury and mortality, but may increase PaCO2. It is not clear whether the beneficial effect of a lung-protective strategy results from reduced ventilation pressures/tidal volumes or is mediated by the effects of hypercapnic acidosis on the inflammatory response involved in the pathogenesis of ventilator-induced lung injury. OBJECTIVE: To analyze whether hypercapnic acidosis affects lung tissue cytokine levels and leukocyte influx in healthy ventilated mice. STUDY DESIGN: Analysis of lung tissue and plasma concentrations of interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, IL-6, IL-10, and keratocyte-derived chemokine after 2 hrs of mechanical ventilation (V(t) 8 mL/kg, positive end-expiratory pressure 4 cm H2O) with 0.06% CO2 (room air), 2% CO2, or 4% CO2. SUBJECTS: Healthy C57BL6 mice (n = 40). MEASUREMENTS/RESULTS: PaCO2 and pH were within normal range when ventilated with 0.06% CO2 and significantly changed with 2% and 4% CO2: (mean +/- SD) pH 7.23 +/- 0.06 and 7.15 +/- 0.04, PaCO2 7.9 +/- 1.4 and 10.8 +/- 0.7 kPa, respectively (p < 0.005). Blood pressure remained within normal limits in all animals. Quantitative microscopic analysis showed a 4.7 +/- 3.7-fold increase (p < 0.01) in pulmonary leukocyte influx in normocapnic ventilated animals and a significant reduction in leukocyte influx of 57 +/- 32% (p < 0.01) and 67 +/- 22% (p < 0.01) when ventilated with 2% and 4% CO2, respectively. Normocapnic ventilation induced a significant elevation of lung tissue IL-1beta (1516 +/- 119 ng/mL), TNF-alpha (344 +/- 88 ng/mL), IL-6 (6310 +/- 807 ng/mL), IL-10 (995 +/- 152 ng/mL), and keratocyte-derived chemokine (36,966 +/- 15,294 ng/mL) (all p-values <0.01). Hypercapnic acidosis with 2% respectively 4% CO2 significantly attenuated this increase with 25 +/- 32% and 54 +/- 32% (IL-1beta, p < 0.01); 17 +/- 36% and 58 +/- 33% (TNF-alpha, p < 0.02); 22 +/- 34% and 89 +/- 6% (IL-6, p < 0.01); 20 +/- 31% and 67 +/- 17% (IL-10, p < 0.01) and 16 +/- 44% and 45 +/- 30% (keratocyte-derived chemokine, p = 0.07). CONCLUSION: Hypercapnic acidosis attenuates the mechanical ventilation-induced immune response independent from reduced tidal volumes/pressures and may protect the lung from ventilator induced lung injury.

Mechanical ventilation in healthy mice induces reversible pulmonary and systemic cytokine elevation with preserved alveolar integrity: an in vivo model using clinical relevant ventilation settings.

BACKGROUND: Mechanical ventilation (MV) may activate the innate immune system, causing the release of cytokines. The resulting proinflammatory state is a risk factor for ventilator-induced lung injury. Cytokine increase results from direct cellular injury but may also result from cyclic stretch alone as demonstrated in vitro: mechanotransduction. To study mechanotransduction in vivo, the authors used an animal MV model with clinically relevant ventilator settings, avoiding alveolar damage. METHODS: Healthy C57BL6 mice (n = 82) were ventilated (tidal volume, 8 ml/kg; positive end-expiratory pressure, 4 cm H2O; fraction of inspired oxygen, 0.4) for 30, 60, 120, and 240 min. Assigned animals were allowed to recover for 2 days after MV. Both pulmonary tissue and plasma interleukin (IL)-1alpha, IL-1beta, tumor necrosis factor alpha, IL-6, IL-10, and keratinocyte-derived chemokine levels were measured. Histopathologic appearance of lung tissue was analyzed by light microscopy and electron microscopy. RESULTS: In lung tissue, all measured cytokines and keratinocyte-derived chemokine levels increased progressively with MV duration. Light microscopy showed increased leukocyte influx but no signs of alveolar leakage or albumin deposition. Electron microscopy revealed intact epithelial cell and basement membranes with sporadically minimal signs of partial endothelial detachment. In plasma, increased levels of IL-1alpha, tumor necrosis factor alpha, IL-6, and keratinocyte-derived chemokine were measured after MV. In the recovery animals, cytokine levels had normalized and no histologic alterations could be found. CONCLUSIONS: Mechanical ventilation induces reversible cytokine increase and leukocyte influx with preserved tissue integrity. This model offers opportunities to study the pathophysiologic mechanisms behind ventilator-induced lung injury and the contribution of MV to the "multiple-hit" concept.

Impairments as measured by ISS do not greatly change between one and eight years after CRPS 1 diagnosis.

BACKGROUND: Complex Regional Pain Syndrome type 1 (CRPS 1) is a potentially incapacitating complication in which pain seems to be the most disabling factor. We performed a late follow up study of a well-defined CRPS 1 population more than eight years after diagnosis. The relationships between early and late impairments were studied with a view to outcome prediction and to investigate possible differences in long-term impairments according to initial CRPS 1 subdiagnosis (i.e. "warm" or "cold", diagnosed according to skin temperature measured via infrared thermometer). METHODS: We again measured patients using the Impairment Level SumScore (ISS) (T8). These data were compared with earlier ISS measurements at CRPS diagnosis (T0) and after one year's treatment (T1). Correlations were determined between these measures. RESULTS: Forty-five patients participated in the present study. Total median ISS improved by 55% (statistically/clinically significant) after one year's treatment (T1), and worsened (non-significantly) by 14% from T1 to T8 - without differences according to original subdiagnosis. ISS correlations were stronger for T1 vs. T8 than for T0 vs. T1 or T0 vs. T8, being strongest for the ISS factors related to pain. CONCLUSIONS: Considerable impairments, as measured by ISS, are still present over eight years after first CRPS 1 diagnosis. These do not greatly change between one and eight years post-diagnosis. ISS outcomes are similar for "cold" and "warm" CRPS 1 diagnostic subgroups. Component ISS scores associated with pain appear to possess greatest predictive power.