Cardiopulmonary effects of low-dose arginine vasopressin in ovine acute lung injury.

OBJECTIVE: To elucidate the effects of low-dose arginine vasopressin on cardiopulmonary functions and nitrosative stress using an established model of acute lung injury. DESIGN: Prospective, randomized, controlled laboratory experiment. SETTING: Investigational intensive care unit. SUBJECTS: Eighteen chronically instrumented sheep. INTERVENTIONS: Sheep were randomly assigned to a sham group without injury or treatment, an injury group without treatment (40% total body surface area third-degree burn and 48 breaths of cold cotton smoke), or an injured group treated with arginine vasopressin (0.02 IU·min⁻¹) from 1 hr after injury until the end of the 24-hr study period (each n = 6). All sheep were mechanically ventilated and fluid resuscitated using an established protocol. MEASUREMENTS AND MAIN RESULTS: There were no differences among groups at baseline. The injury was characterized by a severe deterioration of cardiopulmonary function (left ventricular stroke work indexes and Pao2/Fio2 ratio; p < .01 each vs. sham). Compared with controls, arginine vasopressin infusion improved myocardial function, as suggested by higher stroke volume indexes and left ventricular stroke work indexes (18-24 hrs and 6-24 hrs, respectively; p < .05 each). In addition to an improved gas exchange (higher Pao2/Fio2 ratios from 6 to 24 hrs, p < .01 each), pulmonary edema (bloodless wet-to-dry-weight ratio; p = .018), bronchial obstruction (p = .01), and pulmonary shunt fraction (12-24 hrs; p ≤ .001 each) were attenuated in arginine vasopressin-treated animals compared with controls. These changes occurred along with reduced nitrosative stress, as indicated by lower plasma levels of nitrate/nitrite (12-24 hrs, p < .01 each), as well as lower myocardial and pulmonary tissue concentrations of 3-nitrotyrosine (p = .041 and p = .042 vs. controls, respectively). At 24 hrs, pulmonary 3-nitrotyrosine concentrations were negatively correlated with Pao2/Fio2 ratio (r = -.882; p < .001) and myocardial 3-nitrotyrosine content with stroke volume indexes (r = -.701; p = .004). CONCLUSIONS: Low-dose arginine vasopressin reduced nitrosative stress and improved cardiopulmonary functions in sheep with acute lung injury secondary to combined burn and smoke inhalation injury.

Molecular biological effects of selective neuronal nitric oxide synthase inhibition in ovine lung injury.

Neuronal nitric oxide synthase is critically involved in the pathogenesis of acute lung injury resulting from combined burn and smoke inhalation injury. We hypothesized that 7-nitroindazole, a selective neuronal nitric oxide synthase inhibitor, blocks central molecular mechanisms involved in the pathophysiology of this double-hit insult. Twenty-five adult ewes were surgically prepared and randomly allocated to 1) an uninjured, untreated sham group (n = 7), 2) an injured control group with no treatment (n = 7), 3) an injury group treated with 7-nitroindazole from 1-h postinjury to the remainder of the 24-h study period (n = 7), or 4) a sham-operated group subjected only to 7-nitroindazole to judge the effects in health. The combination injury was associated with twofold increased activity of neuronal nitric oxide synthase and oxidative/nitrosative stress, as indicated by significant increases in plasma nitrate/nitrite concentrations, 3-nitrotyrosine (an indicator of peroxynitrite formation), and malondialdehyde lung tissue content. The presence of systemic inflammation was evidenced by twofold, sixfold, and threefold increases in poly(ADP-ribose) polymerase, IL-8, and myeloperoxidase lung tissue concentrations, respectively (each P < 0.05 vs. sham). These molecular changes were linked to tissue damage, airway obstruction, and pulmonary shunting with deteriorated gas exchange. 7-Nitroindazole blocked, or at least attenuated, all these pathological changes. Our findings suggest 1) that nitric oxide formation derived from increased neuronal nitric oxide synthase activity represents a pivotal reactive agent in the patho-physiology of combined burn and smoke inhalation injury and 2) that selective neuronal nitric oxide synthase inhibition represents a goal-directed approach to attenuate the degree of injury.

The therapeutic test: an ancient malady in the 21st century.

The response to treatment was the diagnostic mainstay in ancient times when diseases were poorly understood. Now that the bases of most diseases are known, appropriate diagnostic means are available. However, many physicians still rely on therapeutic tests to establish diagnoses. Since most illnesses are self-limited and because of the placebo effect, many physicians and patients attribute the improvement to the medication and believe that the correct diagnosis was made. However, inappropriate therapeutic tests often lead to diagnostic delays, rapid emergence of antibiotic-resistant bacterial pathogens, increased risks of adverse drug reactions, and unnecessary expenses. To reduce the frequency of unwarranted therapeutic tests, health-care professionals and educators must take steps to rectify the problem.

Time course of nitric oxide synthases, nitrosative stress, and poly(ADP ribosylation) in an ovine sepsis model.

INTRODUCTION: Different isoforms of nitric oxide synthases (NOS) and determinants of oxidative/nitrosative stress play important roles in the pathophysiology of pulmonary dysfunction induced by acute lung injury (ALI) and sepsis. However, the time changes of these pathogenic factors are largely undetermined. METHODS: Twenty-four chronically instrumented sheep were subjected to inhalation of 48 breaths of cotton smoke and instillation of live Pseudomonas aeruginosa into both lungs and were euthanized at 4, 8, 12, 18, and 24 hours post-injury. Additional sheep received sham injury and were euthanized after 24 hrs (control). All animals were mechanically ventilated and fluid resuscitated. Lung tissue was obtained at the respective time points for the measurement of neuronal, endothelial, and inducible NOS (nNOS, eNOS, iNOS) mRNA and their protein expression, calcium-dependent and -independent NOS activity, 3-nitrotyrosine (3-NT), and poly(ADP-ribose) (PAR) protein expression. RESULTS: The injury induced severe pulmonary dysfunction as indicated by a progressive decline in oxygenation index and concomitant increase in pulmonary shunt fraction. These changes were associated with an early and transient increase in eNOS and an early and profound increase in iNOS expression, while expression of nNOS remained unchanged. Both 3-NT, a marker of protein nitration, and PAR, an indicator of DNA damage, increased early but only transiently. CONCLUSIONS: Identification of the time course of the described pathogenetic factors provides important additional information on the pulmonary response to ALI and sepsis in the ovine model. This information may be crucial for future studies, especially when considering the timing of novel treatment strategies including selective inhibition of NOS isoforms, modulation of peroxynitrite, and PARP.

Inhibition of neuronal nitric oxide synthase in ovine model of acute lung injury.

OBJECTIVE: Acute respiratory distress syndrome/acute lung injury is a serious complication of burn patients with concomitant smoke inhalation injury. Nitric oxide has been shown to play a major role in pulmonary dysfunction from thermal damage. In this study, we have tested the hypothesis that inhibition of neuronal nitric oxide synthase could ameliorate the severity of acute lung injury using our well-established ovine model of cutaneous burn and smoke inhalation. DESIGN: Prospective, randomized, controlled, experimental animals study. SETTING: Investigational intensive care unit at university hospital. SUBJECTS: Adult female sheep. INTERVENTIONS: Female sheep (n = 16) were surgically prepared for the study. Seven days after surgery, all sheep were randomly allocated into three study groups: sham (noninjured, nontreated, n = 6); control (injured, treated with saline, n = 6); and neuronal nitric oxide synthase (injured, treated with specific neuronal nitric oxide synthase inhibitor, ZK 234238 (n = 4). Control and neuronal nitric oxide synthase groups were given a cutaneous burn (40% of total body surface, third degree) and insufflated with cotton smoke (48 breaths, <40 degrees C) under halothane anesthesia. Animals in sham group received fake injury also under halothane anesthesia. After injury or fake injury procedure, all sheep were placed on ventilators and resuscitated with lactated Ringer's solution. Neuronal nitric oxide synthase group was administered with continuous infusion of ZK 234238 started 1 hr postinjury with a dose of 100 microg/kg/hr. Sham and control groups received same amount of saline. MEASUREMENTS AND MAIN RESULTS: Cardiopulmonary hemodynamics monitored during the 24-hr experimental time period was stable in the sham group. Control sheep developed multiple signs of acute lung injury. This pathophysiology included decreased pulmonary gas exchange and lung compliance, increased pulmonary edema, and inflammatory indices, such as interleukin-8. Treatment of injured sheep with neuronal nitric oxide synthase inhibitor attenuated all the observed pulmonary pathophysiology. CONCLUSIONS: The results provide definitive evidence that inhibition of neuronal nitric oxide synthase-derived excessive nitric oxide may be a novel and beneficial treatment strategy for pulmonary pathology in burn victims with smoke inhalation injury.

Pulmonary expression of nitric oxide synthase isoforms in sheep with smoke inhalation and burn injury.

Previous studies have indicated increased plasma levels of inducible nitric oxide synthase in lung. This study further examines the pulmonary expression of nitric oxide synthase (NOS) isoforms in an ovine model of acute lung injury induced by smoke inhalation and burn injury (S+B injury). Female range bred sheep (4 per group) were sacrificed at 4, 8, 12, 24, and 48 hours after injury and immunohistochemistry was performed in tissues for various NOS isoforms. The study indicates that in uninjured sheep lung, endothelial (eNOS) is constitutively expressed in the endothelial cells associated with the airways and parenchyma, and in macrophages. Similarly, neuronal (nNOS) is constitutively present in the mucous cells of the epithelium and in neurons of airway ganglia. In uninjured lung, inducible (iNOS) was present in bronchial secretory cells and macrophages. In tissue after S+B injury, new expression of iNOS was evident in bronchial ciliated cells, basal cells, and mucus gland cells. In the parenchyma, a slight increase in iNOS immunostaining was seen in type I cells at 12 and 24 hours after injury only. Virtually no change in eNOS or nNOS was seen after injury.

Inhibition of lung permeability changes after burn and smoke inhalation by an anti-interleukin-8 antibody in sheep.

PURPOSE: To evaluate the effects of a monoclonal antibody against interleukin-8 (K2.2) on the microvascular fluid flux after combined injury by burn and smoke inhalation. METHODS: Fourteen sheep were prepared surgically by placing a lung lymph catheter and a flank lymph catheter to examine the microvascular fluid flux. After a recovery period, they were subjected to a combined injury of 40% third-degree burns on the flank and smoke inhalation. RESULTS: This combined injury induced a rapid increase in burned tissue lymph flow (b-Q(L)) and a delayed-onset increase in lung lymph flow (l-Q(L)). The initial increase in b-Q(L) was associated with an elevation of the lymph-to-plasma oncotic pressure ratio, which led to a predominant increase in the burned tissue permeability index (b-PI). Pretreatment with K2.2 had no effect on the permeability change seen in the burned tissue; however, the lung permeability changes were attenuated by pretreatment with K2.2. CONCLUSION: These findings indicate that the pathogenesis of the increase in microvascular fluid flux seen after the combined injury differs in burned tissue and the lung.

Karl Otto Landsteiner (1868-1943). Physician-biochemist-immunologist.

Karl Landsteiner applied the sciences of biochemistry, pathology, microbiology, and immunology in medical research to great success during the first half of the 20th century. Although he is principally known for elucidating the major blood group antigens A and B and their isoantibodies for which he was awarded the Nobel Prize in Physiology or Medicine, Landsteiner made many other important medical discoveries. In that respect, he ascertained that paralytic poliomyelitis was due to a virus, the pancreas was damaged in cystic fibrosis, simple chemicals called haptens were able to combine with antibodies, and the Rh antigen that was later found to be the principal cause of hemolytic anemia of the newborn was found in most humans. Moreover, Landsteiner's book "The Specificity of Serological Reactions" was a precursor to the molecular revolution in immunology that occurred after Second World War. Finally, he was one of the leaders of the American Association of Immunology and of the Journal of Immunology.

gamma-Tocopherol nebulization by a lipid aerosolization device improves pulmonary function in sheep with burn and smoke inhalation injury.

Fire accident victims who sustain both thermal injury to skin and smoke inhalation have gross evidence of systemic and pulmonary oxidant damage and acute lung injury. We hypothesized that gamma-tocopherol (gT), a reactive O(2) and N(2) scavenger, when delivered into the airway, would attenuate lung injury induced by burn and smoke inhalation. Acute lung injury was induced in chronically prepared, anesthetized sheep by 40% total burn surface area, third-degree skin burn and smoke insufflation (48 breaths of cotton smoke, <40 degrees C). The study groups were: (1) Sham (not injured, flaxseed oil (FO)-nebulized, n=6); (2) SA-neb (injured, saline-nebulized, n=6); (3) FO-neb (injured, FO-nebulized, n=6); and (4) gT+FO-neb (injured, gT and FO-nebulized, n=6). Nebulization was started 1 h postinjury, and 24 ml of FO with or without gT (51 mg/ml) was delivered into airways over 47 h using our newly developed lipid aerosolization device (droplet size: 2.5-5 microm). The burn- and smoke inhalation-induced pathological changes seen in the saline group were attenuated by FO nebulization; gT addition further improved pulmonary function. Pulmonary gT delivery along with a FO source may be a novel effective treatment strategy in management of patients with acute lung injury.

Neuronal nitric oxide synthase inhibition attenuates cardiopulmonary dysfunctions after combined burn and smoke inhalation injury in sheep.

OBJECTIVE: We hypothesized that nitric oxide derived from the neuronal nitric oxide synthase (NOS) is responsible for much of the injury resulting from skin burn and smoke inhalation. Therefore, we aimed to examine the effects of selective neuronal NOS inhibition on cardiopulmonary functions and cellular injury in sheep with acute respiratory distress syndrome secondary to combined burn and smoke inhalation injury. DESIGN: Prospective, randomized, controlled laboratory experiment. SETTING: Investigational intensive care unit. SUBJECTS: A total of 22 chronically instrumented adult ewes. INTERVENTIONS: Sheep were randomly assigned to either healthy controls (sham), injured controls (40% third-degree flame burn; 48 breaths of cotton smoke), or an injury group treated with the specific neuronal NOS inhibitor 7-nitroindazole (1 mg x kg(-1) x hr(-1)) from 1 hr postinjury to the end of the 48-hr study period. Hypoxic pulmonary vasoconstriction was assessed as decrease in left pulmonary blood flow in response to single-lung hypoxic challenges (100% nitrogen) at baseline, 24 hrs, and 48 hrs. MEASUREMENTS AND MAIN RESULTS: The combination injury contributed to a approximately 90% loss of hypoxic pulmonary vasoconstriction and was associated with significant pulmonary shunting and death of one animal. The increase in nitrate/nitrite plasma levels in injured controls (12 hrs: 17 +/- 2 vs. 6 +/- 1 microM in sham animals; p < .001) was linked to increases in inducible NOS messenger RNA and 3-nitrotyrosine formation in lung tissue (48 hrs: 22 +/- 1 vs. 0.8 +/- 0.3 nM in sham animals; p < .001). 7-Nitroindazole treatment prevented the injury-associated changes in inducible NOS messenger RNA, nitrate/nitrite, and 3-nitrotyrosine, thereby attenuating the loss of hypoxic pulmonary vasoconstriction and improving gas exchange. In addition, 7-nitroindazole decreased lung tissue concentrations of hemoxygenase-1 and ameliorated myocardial depression, airway obstruction, pulmonary edema, ventilatory pressures, and histopathologic changes seen in injured controls. CONCLUSIONS: The present study provides evidence that neuronal NOS-derived nitric oxide plays a pivotal role in the pathogenesis of acute respiratory distress syndrome resulting from combined burn and smoke inhalation injury.

Ilya Ilich Metchnikoff (1845-1915) and Paul Ehrlich (1854-1915): the centennial of the 1908 Nobel Prize in Physiology or Medicine.

Ilya Metchnikoff and Paul Ehrlich shared the 1908 Nobel Prize in Physiology or Medicine - Metchnikoff for discovering the major types and functions of phagocytes and Ehrlich for discovering the types of blood leukocytes, helping to uncover how to generate and use antibodies to protect against bacterial toxins, and formulating the receptor concept of antibodies binding to antigens. In 1908 phagocytic and humoral defences were thought to be unrelated but it was realized much later that they influence one other. Thus, it is fitting that the 1908 Nobel Laureates in Physiology or Medicine remain closely connected in the minds of modern immunologists. Metchnikoff and Ehrlich shared qualities of natural curiosity and tenacity coupled with remarkable inductive-mechanistic thinking and a zest for experimentation. However, their approaches to and methods of research were decidedly different - Metchnikoff's by evolutionary biology and an approach to experimentation via microscopy and Ehrlich's by an imaginative side-chain theory and organic chemistry.

Abraham Lincoln's Gettysburg illness.

When Abraham Lincoln delivered the Gettysburg Address, he was weak and dizzy; his face had a ghastly colour. That evening on the train to Washington, DC, he was febrile and weak, and suffered severe headaches. The symptoms continued; back pains developed. On the fourth day of the illness, a widespread scarlet rash appeared that soon became vesicular. By the tenth day, the lesions itched and peeled. The illness lasted three weeks. The final diagnosis, a touch of varioloid, was an old name for smallpox that was later used in the 20th century to denote mild smallpox in a partially immune individual. It was unclear whether Lincoln had been immunized against smallpox. Indeed, this review suggests that Lincoln had unmodified smallpox and that Lincoln's physicians tried to reassure the public that Lincoln was not seriously ill. Indeed, the successful conclusion of the Civil War and reunification of the country were dependent upon Lincoln's presidency.

The inhibition of inducible nitric oxide synthase in ovine sepsis model.

Excessive NO has been shown to play a major role in the pathogenesis of multiple organ dysfunctions in septic condition. Burn injury, especially if it is associated with smoke inhalation, is often complicated by subsequent development of pneumonia or sepsis that determine the outcome. In the present study, we developed an ovine sepsis model, created by exposing sheep to smoke inhalation followed by instillation of bacteria into the airway, that closely mimics human sepsis and pneumonia. We hypothesized that the inhibition of iNOS-derived excessive NO might be beneficial in treating the cardiopulmonary derangement in this model. Female sheep (n = 18) were surgically prepared for the study and given a tracheostomy. This was followed by insufflation of 48 breaths of cotton smoke (< 40 degrees C) into the airway of each animal and subsequent instillation of live Pseudomonas aeruginosa (5 x 10(11) colony forming units) into each sheep's lung. All sheep were mechanically ventilated using 100% O2. Continuous infusion of BBS-2 (100 microg/kg/h), an iNOS inhibitor, was started 1 h after insult. The administration of BBS-2 improved pulmonary gas exchange (PaO2/FiO2 and pulmonary shunt fraction) and partially reduced airway obstruction and an increase in ventilatory pressures. The lung water content was not affected by iNOS inhibition. The hypotension seen in nontreated animals was not ameliorated either. The increase in plasma concentration of nitrate and nitrite was inhibited by BBS-2. The results of present study show that iNOS may be partially involved in the pathogenesis of acute lung injury induced by smoke inhalation followed by bacterial instillation in the airway.

Franklin Delano Roosevelt's (FDR's) (1882-1945) 1921 neurological disease revisited; the most likely diagnosis remains Guillain-Barré syndrome.

In 2003, we published evidence that the most likely cause of FDR's 1921 neurological disease was Guillain-Barré syndrome. Afterwards, several historians and neurologists stated in their publications that FDR had paralytic poliomyelitis. However, significant criticism of our article or new support for that diagnosis was not revealed. One critic claimed that FDR's cerebrospinal fluid indicated poliomyelitis, but we did not find evidence that a lumbar puncture was performed. The diagnosis of FDR's neurological disease still depends upon documented clinical abnormalities. His age, prolonged symmetric ascending paralysis, transient numbness, protracted dysaesthesia (pain on slight touch), facial paralysis, bladder and bowel dysfunction, and absence of meningismus are typical of Guillain-Barré syndrome and are inconsistent with paralytic poliomyelitis. FDR's prolonged fever was atypical for both diseases. Finally, permanent paralysis, though commoner in paralytic poliomyelitis, is frequent in Guillain-Barré syndrome. Thus, the clinical findings indicate the most likely diagnosis in FDR's case remains Guillain-Barré syndrome.

Cepharanthin, an alkaloid from Stephania cepharantha, inhibits increased pulmonary vascular permeability in an ovine model of sepsis.

Sepsis is a life-threatening event when it occurs in patients suffering from smoke inhalation injury. Pneumonia is one of the most frequent sources of infection in sepsis. Activated leukocytes likely play a role in the pathogenesis of sepsis. Cepharanthin is a biscoclaurine alkaloid that reportedly inhibits the activation of neutrophils. In this study, we investigated the effects of cephranthin on a post-smoke inhalation model of sepsis in sheep. Female sheep (n = 15) were surgically prepared for the study. After 5 days recovery from the operative procedures, tracheostomy was performed in all animals and 48 breaths of cotton smoke (<40 degrees C) were given via a modified bee smoker under halothane anesthesia. After smoke insufflation, Pseudomonas aeruginosa (5 x 109 cfu/kg) was instilled into the airway using a bronchoscope. All of the animals were mechanically ventilated with 100% O(2). Cepharanthin (1.3 mg/kg/h) was infused in five sheep continuously beginning 1 h after the insult and thereafter for the remainder of the 24-h study period. Control animals (n = 6) were treated with 5% dextrose as a vehicle control. Cepharanthin significantly attenuated changes in lung histology as well as in lung wet/dry weight ratio. An in vitro study revealed that cepharanthin inhibited the release of neutrophil elastase from isolated neutrophils stimulated with either formyl-methyl-leucyl-phenylalanine (fMLP) or phorbol myristate acetate with an IC(50) of 60 microM. Cepharanthin also inhibited the fMLP-induced increase in intracellular calcium levels of neutrophils. This result indicates cepharanthin inhibits protein kinase C or a more downstream signaling pathway in neutrophil activation. In conclusion, cepharanthin attenuates acute lung injury and septic shock after smoke inhalation in sheep.

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 degrees C) were inflated into the airway. Afterwards, live Pseudomonas aeruginosa (5 x 10(11) 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 = 5,300 U/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.

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 x 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.

Smoke/burn injury-induced respiratory failure elicits apoptosis in ovine lungs and cultured lung cells, ameliorated with arteriovenous CO2 removal.

STUDY OBJECTIVE: s: The purpose of this study was to examine the effects of two supportive therapies, conventional mechanical ventilation (CMV) and arteriovenous CO(2) removal (AVCO(2)R), during treatment of severe smoke/burn injury-induced ARDS. DESIGN: Sheep were exposed to a smoke/burn injury (lethal dose causing death in 40% of animals); lung tissue and blood was collected prior to injury (control), when an ARDS criterion was met (PaO(2)/fraction of inspired oxygen ratio < 200), then after 72 h of either CMV (group 1) or AVCO(2)R (group 2). Lung tissue was studied by standard histopathologic techniques; cultured lung cells were studied in media supplemented with serum from all four groups. MEASUREMENTS AND RESULTS: In vivo assays demonstrate less apoptotic cell death, and in vitro assays show significantly greater (p < 0.05) cell survival in group 2 (AVCO(2)R) than in group 1 (CMV) or baseline. Differential gene expression demonstrates significantly higher messenger RNA levels of proapoptotic and tumor necrosis factor (TNF)-alpha in cells incubated in baseline media. After exposure of cultured lung cells to conditioned media, protein expression assay of the culture medium revealed no TNF-alpha, TNF receptor (TNFR)-1, or TNFR-2, however, cultured cell lysate reveals elevated levels of TNF-alpha, TNFR-1 and caspase-3 in all groups; most occurred in cells incubated in baseline media (p < 0.05). HOECHST stain, DNA fragmentation, and caspase-3 cleavage show that AVCO(2)R ameliorates apoptosis in this model. CONCLUSIONS: This in vitro work specifically examines cell death in lung cells as a result of smoke/burn injury and effects of therapeutic interventions. Our in vivo studies temporally correlate the clinical pathology to that studied in these lung cells and show that both in vivo and in vitro cell death is predominantly apoptotic and is significantly reduced by AVCO(2)R.

Lipopolysaccharide protection from oxygen toxicity: effect on rat pulmonary selectins.

Sublethal doses of LPS result in increased tolerance to high concentrations of oxygen and this is associated with decreased pulmonary inflammation in a rat model. To investigate the mechanism of decreased neutrophil influx into the lung in this model, we measured levels of mRNA in the lung for the endothelial adhesion molecules, E-selectin and P-selectin. Immunostaining for E-selectin protein was also done in rat lungs, as well as measurement of soluble L-selectin in the blood. These levels were measured in the lungs of adult rats injected with 0.5 mg/kg LPS or placebo at 0 and 24 h and exposed to > 95% O2 for 60 h. Oxygen exposure resulted in significant increases in both E- and P-selectin mRNA and in E-selectin protein after 60 h. LPS resulted in an early rise in E-selectin protein followed by a decline to less than control (saline/O2) levels at 60 h. Messenger RNA for E-selectin followed a similar trend, although there were no differences at 60 h between LPS and control groups exposed to O2. P-selectin mRNA expression did not significantly differ between LPS and control O2 groups. Soluble L-selectin levels decreased by 6 h after LPS infusion and were significantly lower than saline/O2 controls through 24 h, suggesting binding to endothelium. In conclusion, the decrease in E-selectin expression on the surface of pulmonary endothelium after LPS could contribute to decreased inflammation in this model of oxygen toxicity. Soluble L-selectin may serve a further anti-inflammatory role after LPS infusion by binding to pulmonary endothelium.