Lillian D. Traber

Heparin nebulization attenuates acute lung injury in sepsis following smoke inhalation in sheep.

Pseudomonas pneumonia is a common complication of smoke inhalation injury. Airway casts formed from clotted mucous occur frequently in this condition. A recent report shows that intravenous heparin improves oxygenation and reduces lung damage in a sheep model of smoke inhalation. We hypothesized that nebulized heparin could be an effective means of reducing cast formation. Female sheep (n = 19) were surgically prepared for a study of acute lung injury (ALI). After a tracheotomy, 48 breaths of cotton smoke (<40°C) were inflated into the airway. Afterwards, live Pseudomonas aeruginosa (5 × 1011 CFU) was instilled into the lung. All sheep were mechanically ventilated with 100% O2 and were divided into four groups: a heparin-nebulized group (n = 5; animals received aerosolized heparin [10,000 I.U.] 1 h after the bacterial instillation and subsequently every 4 h thereafter), an intravenous heparin group (n = 5,300U/kg/23 h, infusion was started 1 h after the injury), a saline-nebulization group (n = 5; animals received inhaled nebulized saline), and a sham injury group (n = 4, treated in the same fashion, but no injury). The animals were sacrificed after 24 h of mechanical ventilation, and lung samples were harvested. Sheep exposed to lung injury presented with typical hyperdynamic cardiovascular changes and a corresponding drop in PaO2. These changes were significantly attenuated in the heparin groups. Histological changes consisting of cellular infiltrates, lung edema, congestion, and cast formation were reduced by heparin. These data suggest that nebulized inhaled heparin is a beneficial therapy for sepsis-induced ALI.

The pathophysiology of smoke inhalation injury in a sheep model

This study describes an experimental model of smoke inhalation injury in sheep, in which the same pathophysiologic alterations occur as with clinical inhalation in man. Both the patients and the experimental sheep develop diffuse pulmonary mucosal sloughing, pulmonary edema, and a decrease in systemic oxygen tension. The results of this study indicate that the pulmonary edema is the result of an increase in microvascular permeability, characterized by increases in lung lymph flow (Qlym), lymph-to-plasma protein concentration ratio (L/P), and transvascular protein flux (Qlym X lung lymph protein concentration), while pulmonary vascular pressures remain constant. Neutrophil degranulation may contribute to the increased microvascular permeability.

Effects of nitric oxide synthesis inhibition in hyperdynamic endotoxemia

ObjectiveTo investigate the effects of Nω-nitro-L-arginine methyl ester, an inhibitor of nitric oxide synthesis, on hemodynamics, gas exchange and oxygen transport in an ovine model of hyperdynamic sepsis. DesignProspective, nonrandomized, controlled study, with repeated measurements. SettingUniversity research laboratory. SubjectsTwenty healthy adult sheep (weighing 20 to 45 kg) were divided into two groups of 12 treated sheep and eight control sheep and studied. InterventionsTwenty awake, chronically instrumented sheep received a continuous infusion of endotoxin (10 ng/kg/min) over 48 hrs. Twenty-four hours after the start of the endotoxin infusion, 12 sheep (treatment group) received a bolus of the nitric oxide synthesis inhibitor Nω-nitro-L-arginine methyl ester (25 mg/kg), while the other eight animals (control group) received the carrier (0.9% NaCl). Measurements and Main ResultsTwenty-four hours after the start of the endotoxin infusion, both groups exhibited a hyperdynamic state with increased cardiac indices, decreased systemic vascular resistance indices, impaired oxygenation, and increased pulmonary shunt fractions. In both groups, oxygen delivery was significantly increased, while oxygen consumption remained virtually unchanged, resulting in a decreased oxygen extraction ratio. In the control group, the significant alterations in systemic hemodynamics, lung function and oxygen transport persisted for the remainder of the study. Administration of Nω-nitro-L-arginine methyl ester normalized cardiac index and systemic vascular resistance index, increased mean arterial blood pressure, and decreased heart rate. Although oxygen delivery significantly decreased after administration of Nω-nitro-L-arginine methyl ester, oxygen consumption did not change, resulting in a normalization of oxygen extraction ratio. Despite a significant reduction of pulmonary shunt fraction, oxygenation did not improve. Pulmonary arterial pressure and pulmonary vascular resistance index showed a peak 2 hrs after administration of the nitric oxide synthesis inhibitor and then tended to decrease. In contrast, the effects of Nω-nitro-L-arginine methyl ester on the systemic circulation persisted for the remainder of the study. ConclusionsThe data support the assumption that augmented nitric oxide production in a major cause of the hemodynamic alterations seen in hyperdynamic endotoxemia. Administration of the nitric oxide synthesis inhibitor Nω-nitro-L-arginine methyl ester normalized the endotoxin-induced hyperdynamic state, but did not impair oxygen consumption, indicating adequate tissue perfusion of metabolically active organs. Inhibition of nitric oxide synthesis may be a therapeutic option in the treatment of hyperdynamic septic patients when conventional therapy fails to maintain a minimum of cardiovascular performance. (Crit Care Med 1994; 22:306–312)

Inhalation injury in burned patients: effects and treatment.

Pulmonary pathology in major thermal injury is found in 30-80 per cent of burn fatalities. The incidence and mortality from inhalation injury increases both with age and increasing burn size. Toxic smoke inhalation injury, characterized by increased lung microvascular permeability, is attenuated by increasing cardiac output to normal levels, indicating that fluid restriction after inhalation injury may lead to excessive lung fluid formation and hypoxia. Fluid administration of approximately 2 ml/kg/% area burned above the calculated resuscitation volume is required following an inhalation injury to provide adequate support for the systemic circulation and maintain cardiac output at normal levels. This additional volume does not contribute to the development of pulmonary oedema, but may decrease its formation by increasing shear forces thus reducing polymorphonuclear leucocyte deposition in the pulmonary microcirculation. Nasotracheal intubation is preferred when airway integrity is compromised by inhalation injury. The advantages are non-operative placement, ease of discontinuation, minimal bacterial contamination and leaving neck burns undisturbed. The administration of the appropriate antibiotics for documented infection is recommended, while steroids have been shown to be of no benefit.

Correction of blood pH attenuates changes in hemodynamics and organ blood flow during permissive hypercapnia

OBJECTIVES To determine whether changes in cardiac output, regional blood flow, and intracranial pressure during permissive hypercapnia are blood pH-dependent and can be attenuated by correction of intravascular acidemia. DESIGN Prospective, controlled study. SETTING Research laboratory. SUBJECTS Female Marino ewes. INTERVENTIONS Animals were instrumented with a pulmonary artery catheter, femoral arterial and venous catheters, a catheter in the third cerebral ventricle, and ultrasonic flow probes on the left carotid, superior mesenteric, and left renal arteries 1 wk before experimentation. At initiation of the protocol, ewes underwent endotracheal intubation and mechanical ventilation under general anesthesia. Minute ventilation was reduced to induce hypercapnia with a target PaCO2 of 80 torr (10.7 kPa). In the pH-uncorrected group (n = 6), arterial blood pH was allowed to decreased without treatment. In the pH-corrected group (n = 5), 14.4 mEq/kg of sodium bicarbonate was given intravenously as a bolus to correct arterial blood pH toward a target arterial pH of 7.40 (dose calculated by the Henderson-Hasselbalch equation). MEASUREMENTS AND MAIN RESULTS Arterial blood pH, PCO2, cardiac output, intracranial pressure, and carotid, superior mesenteric, and renal artery blood flow rates were measured at normocapnic baseline and at every hour during hypercapnia for 6 hrs. In the pH-uncorrected group, arterial blood pH decreased from 7.41 +/- 0.03 at normocapnia to 7.14 +/- 0.01 (p < .01 vs. normocapnia) as blood PCO2 increased to 81.2 +/- 1.8 torr (10.8 +/- 0.2 kPa). In the pH-corrected group, arterial blood pH was 7.42 +/- 0.02 at normocapnia and was maintained at 7.37 +/- 0.01 while PaCO2 was increased to 80.3 +/- 0.9 torr (10.7 +/- 0.1 kPa). Significant increases in cardiac output occurred with the initiation of hypercapnia for both groups (pH-uncorrected group: 4.3 +/- 0.6 L/min at normocapnia vs. 6.8 +/- 1.0 L/min at 1 hr [p < .05]; pH-corrected group: 4.1 +/- 0.4 at normocapnia vs. 5.7 +/- 0.4 L/min at 1 hr [p < .05]). However, this increase was sustained only in the uncorrected group. Changes in carotid and mesenteric artery blood flow rates, as a percent of baseline values, showed sustained significant increases in the pH-uncorrected groups (p < .05) and only transient (carotid at 1 hr) or no (superior mesenteric) significant change in the pH-corrected groups. Conversely, significant increases in renal artery blood flow were seen only in the pH-uncorrected group during the last 2 hrs of the experiment (p < .05). Organ blood flow, as a percent of cardiac output, did not change significantly in either group. Intracranial pressure increased significantly in the pH-uncorrected group (9.0 +/- 1.5 mm Hg at normocapnia vs. 26.8 +/- 5.1 at 1 hr, p < .05), and remained increased, while showing no significant change in the pH-corrected group (8.5 +/- 1.6 mm Hg at normocapnia to 7.7 +/- 4.2 at 1 hr). CONCLUSIONS Acute hypercapnia, induced within 1 hr, is associated with significant increases in cardiac output, organ blood flow, and intracranial pressure. These changes can be significantly attenuated by correction of blood pH with the administration of sodium bicarbonate, without adverse effects on hemodynamics.

Burn resuscitation: crystalloid versus colloid versus hypertonic saline hyperoncotic colloid in sheep.

OBJECTIVES The present study was undertaken to assess the fluid balance and hemodynamic effects during the first 8 hrs of resuscitation in animals with a large body surface area burn, using lactated Ringers solution, 6% hetastarch, and hypertonic saline dextran. DESIGN A prospective, blinded, controlled, terminal study, using anesthetized animals. The initial baseline period was followed by scald injury, and three different treatment regimens were administered from coded bags to achieve a physiologic end point. SETTING University laboratory. SUBJECTS Eighteen female sheep (35 to 45 kg) were anesthetized with isoflurane. INTERVENTIONS Test solutions (10 mL/kg of either lactated Ringers solution, hetastarch, or hypertonic saline dextran) were infused 30 mins after the scald injury at a rate to restore and maintain the baseline oxygen delivery (DO2) value. MEASUREMENTS AND MAIN RESULTS Cardiovascular hemodynamics, plasma sodium concentration, plasma colloid osmotic pressure, and fluid balance were measured before and after scalding and resuscitation. After the initial 10-mL/kg test solution dose was given, lactated Ringers solution was infused to achieve the same end point of baseline DO2 for the remainder of the 8 hrs. The scald caused an initial 30% reduction in cardiac output, a 20% reduction in mean arterial pressure, and 10% to 15% increase in hematocrit. All three test solutions restored and maintained baseline DO2 within 1 hr. However, hetastarch and hypertonic saline dextran reduced the net fluid volume over 8 hrs by 48% and 74%, respectively, compared with lactated Ringers solution. Edema in the burn wound was not affected by treatment, while hypertonic saline dextran reduced edema in nonburned skin compared with both lactated Ringers solution and hypertonic saline dextran. Plasma colloid osmotic pressure was significantly higher in the hetastarch and hypertonic saline dextran groups. A continuous decrease in plasma sodium concentrations from baseline values (140 to 145 mmol/L) was measured in the lactated Ringers solution and hetastarch groups (130 to 133 mmol/L) over 8 hrs. Plasma sodium concentrations in the group receiving hypertonic saline dextran were increased (150 to 155 mmol/L) at 4 hrs, but returned toward baseline by 8 hrs. CONCLUSIONS Net volume loading can be reduced markedly by initial resuscitation of large body surface area burn injury using a colloid (hetastarch), and can be further reduced by use of hypertonic saline colloid. Hyponatremia was apparent in the isotonic crystalloid- and colloid-treated animals, but not in those animals treated with hypertonic saline colloid.

Recombinant human activated protein C improves pulmonary function in ovine acute lung injury resulting from smoke inhalation and sepsis

Objective:To investigate the effects of recombinant human activated protein C (rhAPC) on pulmonary function in acute lung injury (ALI) resulting from smoke inhalation in association with a bacterial challenge. Design:Prospective, randomized, controlled, experimental animal study with repeated measurements. Setting:Investigational intensive care unit at a university hospital. Subjects:Eighteen sheep (37.2 ± 1.0 kg) were operatively prepared and randomly allocated to either the sham, control, or rhAPC group (n = 6 each). After a tracheotomy had been performed, ALI was produced in the control and rhAPC group by insufflation of 4 sets of 12 breaths of cotton smoke. Then, a 30 mL suspension of live Pseudomonas aeruginosa bacteria (containing 2–5 × 1011 colony forming units) was instilled into the lungs according to an established protocol. The sham group received only the vehicle, i.e., 4 sets of 12 breaths of room air and instillation of 30 mL normal saline. The sheep were studied in the awake state for 24 hrs and were ventilated with 100% oxygen. RhAPC (24 &mgr;g/kg/hr) was intravenously administered. The infusion was initiated 1 hr post-injury and lasted until the end of the experiment. The animals were resuscitated with Ringers lactate solution to maintain constant pulmonary artery occlusion pressure. Measurements and Main Results:In comparison with nontreatment in controls, the infusion of rhAPC significantly attenuated the fall in Pao2/Fio2 ratio (control group values were 521 ± 22 at baseline [BL], 72 ± 5 at 12 hrs, and 74 ± 7 at 24 hrs, vs. rhAPC group values of 541 ± 12 at BL, 151 ± 29 at 12 hours [p < .05 vs. control], and 118 ± 20 at 24 hrs), and significantly reduced the increase in pulmonary microvascular shunt fraction (Qs/Qt; control group at BL, 0.14 ± 0.02, and at 24 hrs, 0.65 ± 0.08; rhAPC group at BL, 0.24 ± 0.04, and at 24 hrs, 0.45 ± 0.02 [p < .05 vs. control]) and the increase in peak airway pressure (mbar; control group at BL, 20 ± 1, and at 24 hrs, 36 ± 4; rhAPC group at BL, 21 ± 1, and at 24 hrs, 28 ± 2 [p < .05 vs. control]). In addition, rhAPC limited the increase in lung 3-nitrotyrosine (after 24 hrs [%]: sham, 7 ± 2; control, 17 ± 1; rhAPC, 12 ± 1 [p < .05 vs. control]), a reliable indicator of tissue injury. However, rhAPC failed to prevent lung edema formation. RhAPC-treated sheep showed no difference in activated clotting time or platelet count but exhibited less fibrin degradation products (1/6 animals) than did controls (4/6 animals). Conclusions:Recombinant human activated protein C attenuated ALI after smoke inhalation and bacterial challenge in sheep, without bleeding complications.

Aerosolized tissue plasminogen inhibitor improves pulmonary function in sheep with burn and smoke inhalation

Acute respiratory distress syndrome is a major complication in patients with thermal injury. The obstruction of the airway by cast material, composed in part of fibrin, contributes to deterioration of pulmonary gas exchange. We tested the effect of aerosol administration of tissue plasminogen activator, which lyses fibrin clots, on acute lung injury in sheep that had undergone combined burn/smoke inhalation injury. Anesthetized sheep were given a 40% total body surface, third degree burn and were insufflated with cotton smoke. Tissue plasminogen activator (TPA) was nebulized every 4 h at 1 or 2 mg for each nebulization, beginning 4 h after injury. Injured but untreated control sheep developed multiple symptoms of acute respiratory distress syndrome: decreased pulmonary gas exchange, increased pulmonary edema, and extensive airway obstruction. These control animals also showed increased pulmonary transvascular fluid flux and increased airway pressures. These variables were all stable in sham animals. Nebulization of saline or 1 mg of TPA only slightly improved measures of pulmonary function. Treatment of injured sheep with 2 mg of TPA attenuated all the pulmonary abnormalities noted above. The results provide evidence that clearance of airway obstructive cast material is crucial in managing acute respiratory distress syndrome resulting from combined burn and smoke inhalation injury.

Inhibition of poly (ADP-ribose) polymerase attenuates acute lung injury in an ovine model of sepsis.

It is known that in various pathophysiological conditions, reactive oxidants cause DNA strand breakage and subsequent activation of the nuclear enzyme poly(ADP ribose) polymerase (PARP). Activation of PARP results in cellular dysfunction. We hypothesized that pharmacological inhibition of PARP reduces the damage in the ovine model of acute lung injury (ALI). After smoke inhalation, Pseudomonas aeruginosa (5 × 109 cfu/kg) was instilled into both lungs. All of the animals were mechanically ventilated with 100% O2. The infusion of the PARP inhibitor (INO-1001, n = 6) began 1 h after the injury and thereafter through 24 h (3 mg bolus + 0.3 mg/kg/h, i.v.). Control animals (n = 6) were treated with saline. Sham injury animals (n = 8) received sham smoke and were mechanically ventilated in the same fashion. One-half of those sham animals (n = 4) were given the same dose of INO-1001. PaO2/FiO2 ratio at 24 h in saline and in the INO-1001-treated groups were 95 ± 22 and 181 ± 22, respectively (P < 0.05). Peak airway pressure at 24 h in the saline- and INO-1001-treated groups was 32.6 ± 3.0 and 24.4 ± 2.2, respectively (P < 0.05). Pulmonary shunt fraction was also significantly attenuated. INO-1001 treatment reduced pulmonary histological injury and attenuated poly (ADP-ribose) accumulation in the lung. In conclusion, inhibition of PARP improved the ALI after smoke inhalation and pneumonia. The results suggest that the activation of PARP plays a role in the pathophysiology of ALI in sheep.

Human mesenchymal stem cells reduce the severity of acute lung injury in a sheep model of bacterial pneumonia

Background Human bone marrow-derived mesenchymal stem (stromal) cells (hMSCs) improve survival in mouse models of acute respiratory distress syndrome (ARDS) and reduce pulmonary oedema in a perfused human lung preparation injured with Escherichia coli bacteria. We hypothesised that clinical grade hMSCs would reduce the severity of acute lung injury (ALI) and would be safe in a sheep model of ARDS. Methods Adult sheep (30–40 kg) were surgically prepared. After 5 days of recovery, ALI was induced with cotton smoke insufflation, followed by instillation of live Pseudomonas aeruginosa (2.5×1011 CFU) into both lungs under isoflurane anaesthesia. Following the injury, sheep were ventilated, resuscitated with lactated Ringers solution and studied for 24 h. The sheep were randomly allocated to receive one of the following treatments intravenously over 1 h in one of the following groups: (1) control, PlasmaLyte A, n=8; (2) lower dose hMSCs, 5×106 hMSCs/kg, n=7; and (3) higher-dose hMSCs, 10×106 hMSCs/kg, n=4. Results By 24 h, the PaO2/FiO2 ratio was significantly improved in both hMSC treatment groups compared with the control group (control group: PaO2/FiO2 of 97±15 mm Hg; lower dose: 288±55 mm Hg (p=0.003); higher dose: 327±2 mm Hg (p=0.003)). The median lung water content was lower in the higher-dose hMSC-treated group compared with the control group (higher dose: 5.0 g wet/g dry [IQR 4.9–5.8] vs control: 6.7 g wet/g dry [IQR 6.4–7.5] (p=0.01)). The hMSCs had no adverse effects. Conclusions Human MSCs were well tolerated and improved oxygenation and decreased pulmonary oedema in a sheep model of severe ARDS. Trail registration number: NCT01775774 for Phase 1. NCT02097641 for Phase 2.