Yusuke Yamamoto

University of Texas Medical Branch at Galveston, Galveston, Texas, United States

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Publications (12)31.41 Total impact

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    ABSTRACT: Pulmonary coagulopathy has become an important therapeutic target in adult respiratory distress syndrome (ARDS). We hypothesized that combining intravenous recombinant human antithrombin (rhAT), nebulized heparin, and nebulized tissue plasminogen activator (TPA) more effectively improves pulmonary gas exchange compared with a single rhAT infusion, while maintaining the anti-inflammatory properties of rhAT in ARDS. Therefore, the present prospective, randomized experiment was conducted using an established ovine model. Following burn and smoke inhalation injury (40% of total body surface area, third-degree flame burn, and 4 × 12 breaths of cold cotton smoke), 18 chronically instrumented sheep were randomly assigned to receive intravenous saline plus saline nebulization (control), intravenous rhAT (6 IU/kg/h) started 1 hour after injury plus saline nebulization (AT i.v.) or intravenous rhAT combined with nebulized heparin (10,000 IU every 4 hours, started 2 hours after injury), and nebulized TPA (2 mg every 4 hours, started 4 hours after injury) (triple therapy, n = 6 each). All animals were mechanically ventilated and fluid resuscitated according to standard protocols during the 48-hour study period. Both treatment approaches attenuated ARDS compared with control animals. Notably, triple therapy was associated with an improved PaO2/FiO2 ratio (p = 0.007), attenuated pulmonary obstruction (p = 0.02) and shunting (p = 0.025), as well as reduced ventilatory pressures (p < 0.05 each) versus AT i.v. at 48 hours. However, the anti-inflammatory effects of sole AT i.v., namely, the inhibition of neutrophil activation (neutrophil count in the lymph and pulmonary polymorphonuclear cells, p < 0.05 vs. control each), pulmonary transvascular fluid flux (lymph flow, p = 0.004 vs. control), and systemic vascular leakage (cumulative net fluid balance, p < 0.001 vs. control), were abolished in the triple therapy group. Combining intravenous rhAT with nebulized heparin and nebulized TPA more effectively restores pulmonary gas exchange, but the anti-inflammatory effects of sole rhAT are abolished with the triple therapy. Interferences between the different anticoagulants may represent a potential explanation for these findings.
    The journal of trauma and acute care surgery. 01/2014; 76(1):126-33.
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    ABSTRACT: To test the hypothesis that restoration of antithrombin plasma concentrations attenuates vascular leakage by inhibiting neutrophil activation through syndecan-4 receptor inhibition in an established ovine model of acute lung injury. Randomized controlled laboratory experiment. University animal research facility. Eighteen chronically instrumented sheep. Following combined burn and smoke inhalation injury (40% of total body surface area, third-degree flame burn; 4 × 12 breaths of cold cotton smoke), chronically instrumented sheep were randomly assigned to receive an IV infusion of 6 IU/kg/hr recombinant human antithrombin III or normal saline (n = 6 each) during the 48-hour study period. In addition, six sham animals (not injured, continuous infusion of vehicle) were used to obtain reference values for histological and immunohistochemical analyses. Compared to control animals, recombinant human antithrombin III reduced the number of neutrophils per hour in the pulmonary lymph (p < 0.01 at 24 and 48 hr), alveolar neutrophil infiltration (p = 0.04), and pulmonary myeloperoxidase activity (p = 0.026). Flow cytometric analysis revealed a significant reduction of syndecan-4-positive neutrophils (p = 0.002 vs control at 24 hr). Treatment with recombinant human antithrombin III resulted in a reduction of pulmonary nitrosative stress (p = 0.002), airway obstruction (bronchi: p = 0.001, bronchioli: p = 0.013), parenchymal edema (p = 0.044), and lung bloodless wet-to-dry-weight ratio (p = 0.015). Clinically, recombinant human antithrombin III attenuated the increased pulmonary transvascular fluid flux (12-48 hr: p ≤ 0.001 vs control each) and the deteriorated pulmonary gas exchange (12-48 hr: p < 0.05 vs control each) without increasing the risk of bleeding. The present study provides evidence for the interaction between antithrombin and neutrophils in vivo, its pathophysiological role in vascular leakage, and the therapeutic potential of recombinant human antithrombin III in a large animal model of acute lung injury.
    Critical care medicine 10/2013; · 6.37 Impact Factor
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    ABSTRACT: INTRODUCTION: We hypothesized that maintaining physiological plasma levels of antithrombin attenuates myocardial dysfunction and inflammation as well as vascular leakage associated with burn and smoke inhalation injury. Therefore, the present prospective, randomized experiment was conducted using an established ovine model. METHODS: Following 40% of total body surface area, 3rd degree flame burn and 4x12 breaths of cold cotton smoke, chronically instrumented sheep were randomly assigned to receive an intravenous infusion of 6 IUkg-1h-1 recombinant human antithrombin (rhAT) or normal saline (control group; n=6 each). In addition, six sheep were designated as sham animals (not injured, continuous infusion of vehicle). During the 48h study period the animals were awake, mechanically ventilated and fluid resuscitated according to standard formulas RESULTS: Compared to the sham group, myocardial contractility was severely impaired in control animals, as suggested by lower stroke volume and left ventricular stroke work indexes. As a compensatory mechanism heart rate increased, thereby increasing myocardial oxygen consumption. In parallel, myocardial inflammation was induced via nitric oxide production, neutrophil accumulation (myeloperoxidase activity) and activation of the p38-mitogen-activated protein kinase pathway resulting in cytokine release (tumor necrosis factor-alpha, interleukin-6) in control vs. sham animals. rhAT-treatment significantly attenuated these inflammatory changes leading to a myocardial contractility and myocardial oxygen consumption comparable to sham animals. In control animals, systemic fluid accumulation progressively increased over time resulting in a cumulative positive fluid balance of about 4000ml at the end of the study period. Contrary, in rhAT-treated animals there was only an initial fluid accumulation until 24h that was reversed back to the level of sham animals during the second day. CONCLUSIONS: Based on these findings, the supplementation of rhAT may represent a valuable therapeutic approach for cardiovascular dysfunction and inflammation after burn and smoke inhalation injury.
    Critical care (London, England) 05/2013; 17(3):R86. · 4.72 Impact Factor
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    ABSTRACT: More than 20,000 burn injury victims suffer from smoke inhalation injury in the United States annually. In an ovine model of acute lung injury, γ-tocopherol had a beneficial effect when nebulized into the airway. We hypothesize that γ-tocopherol scavenges reactive oxygen species (ROS) and reactive nitrogen species resulting from burn and smoke inhalation injury and that these ROS/reactive nitrogen species activate the arginase pathway, leading to increased collagen deposition and decreased pulmonary function. To test this hypothesis, ewes were operatively prepared for chronic study, then they were randomly divided into groups (n = 8): uninjured, injured, or injured with nebulization (γ-tocopherol [950 mg/g] and α-tocopherol [40 mg/g] from hours 3 to 48 after the injury). The injury, under deep anesthesia, consisted of a 20% total body surface burn and 36 breaths of cotton smoke; all animals were killed after 3 weeks. Treatment increased lung γ-tocopherol at 3 weeks after γ-tocopherol nebulization compared with injured sheep (1.75 ± 0.62 nmol/g vs. 0.45 ± 0.06, P < 0.05). The expression of dimethylarginine dimethylaminohydrolase-2, which degrades asymmetrical dimethylarginine, a nitric oxide synthase inhibitor, significantly increases with γ-tocopherol treatment compared with injured sheep (P < 0.05). Arginase activity (0.15 ± 0.02 μM urea/μg protein vs. 0.24 ± 0.009, P < 0.05), ornithine aminotransferase (11,720 ± 888 vs. 13,170 ± 1,775), and collagen deposition (0.62 ± 0.12 μM hydroxyproline/μg protein vs. 1.02 ± 0.13, P < 0.05) significantly decrease with γ-tocopherol compared with injured animals without γ-tocopherol. The decreases in arginase and collagen with γ-tocopherol are associated with significantly increased diffusion capacity (P < 0.05) and decreased lung wet-to-dry ratio (P < 0.05). Smoke-induced chronic pulmonary dysfunction is mediated through the ROS/asymmetrical dimethylarginine/arginase pathway, and ROS scavengers such as γ-tocopherol may be a potential therapeutic management of burn patients with inhalation injury.
    Shock (Augusta, Ga.) 12/2012; 38(6):671-6. · 2.87 Impact Factor
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    ABSTRACT: Vasopressin analogues are used as a supplement to norepinephrine in septic shock. The isolated effects of vasopressin agonists on sepsis-induced vascular dysfunction, however, remain controversial. Since V2-receptor stimulation induces vasodilation and procoagulant effects, a higher V1a- vs. V2-receptor selectivity might be advantageous. We therefore hypothesized that a sole, titrated infusion of the selective V1a-agonist Phe2-Orn8-Vasotocin (POV) is more effective than the mixed V1a-/V2-agonist arginine vasopressin (AVP) for the treatment of vascular and cardiopulmonary dysfunction in MRSA pneumonia-induced, ovine sepsis. After the onset of hemodynamic instability, awake, chronically instrumented, mechanically ventilated and fluid resuscitated sheep were randomly assigned to receive continuous infusions of either POV, AVP or saline solution (control, each n=6). AVP and POV were titrated to maintain mean arterial pressure above baseline-10mmHg. Compared to control animals, AVP and POV reduced neutrophil migration (myeloperoxidase activity, alveolar neutrophils) and plasma levels of nitric oxide resulting in higher mean arterial pressures and a reduced vascular leakage (net fluid balance, chest and abdominal fluid, pulmonary bloodless wet-to-dry-weight-ratio, alveolar and septal edema). However, POV stabilized hemodynamics at lower doses than AVP. In addition, POV, but not AVP, reduced myocardial and pulmonary tissue concentrations of 3-nitrotyrosine, vascular endothelial growth factor and angiopoietin-2, thereby leading to an abolishment of cumulative fluid accumulation (POV: 9±15mL•kg-1 vs. AVP: 110±13mL•kg-1 vs. control: 213±16mL•kg-1, p<0,001 each) and an attenuated cardiopulmonary dysfunction (left ventricular stroke work index, PaO2/FiO2 ratio) vs. control animals. Highly selective V1a-agonism appears to be superior to unselective vasopressin analogues for the treatment of sepsis-induced vascular dysfunction.
    AJP Heart and Circulatory Physiology 09/2012; · 4.01 Impact Factor
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    ABSTRACT: Smoke inhalation injury frequently increases the risk of pneumonia and mortality in burn patients. The pathophysiology of acute lung injury secondary to burn and smoke inhalation is well studied, but long-term pulmonary function, especially the process of lung tissue healing following burn and smoke inhalation, has not been fully investigated. By contrast, early burn excision has become the standard of care in the management of major burn injury. While many clinical studies and small-animal experiments support the concept of early burn wound excision, and show improved survival and infectious outcomes, we have developed a new chronic ovine model of burn and smoke inhalation injury with early excision and skin grafting that can be used to investigate lung pathophysiology over a period of 3 weeks. Eighteen female sheep were surgically prepared for this study under isoflurane anesthesia. The animals were divided into three groups: an Early Excision group (20% TBSA, third-degree cutaneous burn and 36 breaths of cotton smoke followed by early excision and skin autografting at 24h after injury, n=6), a Control group (20% TBSA, third-degree cutaneous burn and 36 breaths of cotton smoke without early excision, n=6) and a Sham group (no injury, no early excision, n=6). After induced injury, all sheep were placed on a ventilator and fluid-resuscitated with Lactated Ringers solution (4 mL/% TBS/kg). At 24h post-injury, early excision was carried out to fascia, and skin grafting with meshed autografts (20/1000 in., 1:4 ratio) was performed under isoflurane anesthesia. At 48 h post-injury, weaning from ventilator was begun if PaO(2)/FiO(2) was above 250 and sheep were monitored for 3 weeks. At 96 h post-injury, all animals were weaned from ventilator. There are no significant differences in PaO(2)/FiO(2) between Early Excision and Control groups at any points. All animals were survived for 3 weeks without infectious complication in Early Excision and Sham groups, whereas two out of six animals in the Control group had abscess in lung. The percentage of the wound healed surviving area (mean ± SD) was 74.7 ± 7.8% on 17 days post-surgery in the Early Excision group. Lung wet-to-dry weight ratio (mean ± SD) was significantly increased in the Early Excision group vs. Sham group (p<0.05). The calculated net fluid balance significantly increased in the early excision compared to those seen in the Sham and Control groups. Plasma protein, oncotic pressure, hematocrit of % baseline, hemoglobin of % baseline, white blood cell and neutrophil were significantly decreased in the Early Excision group vs. Control group. The early excision model closely resembles practice in a clinical setting and allows long-term observations of pulmonary function following burn and smoke inhalation injury. Further studies are warranted to assess lung tissue scarring and measuring collagen deposition, lung compliance and diffusion capacity.
    Burns: journal of the International Society for Burn Injuries 03/2012; 38(6):908-16. · 1.95 Impact Factor
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    ABSTRACT: We hypothesize that the nebulization of γ-tocopherol (g-T) in the airway of our ovine model of acute respiratory distress syndrome will effectively improve pulmonary function following burn and smoke inhalation after 96 h. Adult ewes (n = 14) were subjected to 40% total body surface area burn and were insufflated with 48 breaths of cotton smoke under deep anesthesia, in a double-blind comparative study. A customized aerosolization device continuously delivered g-T in ethanol with each breath from 3 to 48 h after the injury (g-T group, n = 6), whereas the control group (n = 5) was nebulized with only ethanol. Animals were weaned from the ventilator when possible. All animals were killed after 96 h, with the exception of one untreated animal that was killed after 64 h. Lung g-T concentration significantly increased after g-T nebulization compared with the control group (38.5 ± 16.8 vs. 0.39 ± 0.46 nmol/g, P < 0.01). The PaO(2)/FIO(2) ratio was significantly higher after treatment with g-T compared with the control group (310 ± 152 vs. 150 ± 27.0, P < 0.05). The following clinical parameters were improved with g-T treatment: pulmonary shunt fraction, peak and pause pressures, lung bloodless wet-to-dry weight ratios (2.9 ± 0.87 vs. 4.6 ± 1.4, P < 0.05), and bronchiolar obstruction (2.0% ± 1.1% vs. 4.6% ± 1.7%, P < 0.05). Nebulization of g-T, carried by ethanol, improved pulmonary oxygenation and markedly reduced the time necessary for assisted ventilation in burn- and smoke-injured sheep. Delivery of g-T into the lungs may be a safe, novel, and efficient approach for management of acute lung injury patients who have sustained oxidative damage to the airway.
    Shock (Augusta, Ga.) 01/2012; 37(4):408-14. · 2.87 Impact Factor
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    ABSTRACT: We investigated the effect of the angiotensin-converting enzyme (ACE) inhibitor captopril in a clinically relevant ovine model of smoke and burn injury, with special reference to oxidative stress and activation of poly(ADP-ribose) polymerase, in the lung and in circulating leukocytes. Female, adult sheep (28-40 kg) were divided into three groups. After tracheostomy and under deep anesthesia, both vehicle-control-treated (n = 5) and captopril-treated (20 mg/kg per day, i.v., starting 0.5 h before the injury) (n = 5) groups were subjected to 2 × 20%, third-degree burn injury and were insufflated with 48 breaths of cotton smoke. A sham group not receiving burn/smoke was also studied (n = 5). Animals were mechanically ventilated and fluid resuscitated for 24 h in the awake state. Burn and smoke injury resulted in an upregulation of ACE in the lung, evidenced by immunohistochemical determination and Western blotting. Burn and smoke injury resulted in pulmonary dysfunction, as well as systemic hemodynamic alterations. Captopril treatment of burn and smoke animals improved PaO2/FiO2 ratio and pulmonary shunt fraction and reduced the degree of lung edema. There was a marked increase in PAR levels in circulating leukocytes after burn/smoke injury, which was significantly decreased by captopril. The pulmonary level of ACE and the elevated pulmonary levels of transforming growth factor β in response to burn and smoke injury were significantly decreased by captopril treatment. Our results suggest that the ACE inhibitor captopril exerts beneficial effects on the pulmonary function in burn/smoke injury. The effects of the ACE inhibitor may be related to the prevention of reactive oxygen species-induced poly(ADP-ribose)polymerase overactivation. Angiotensin-converting enzyme inhibition may also exert additional beneficial effects by inhibiting the expression of the profibrotic mediator transforming growth factor β.
    Shock (Augusta, Ga.) 06/2011; 36(4):402-9. · 2.87 Impact Factor
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    ABSTRACT: The nuclear enzyme poly(ADP-ribose)polymerase (PARP) plays a significant role in the pathogenesis of various forms of critical illness. DNA strand breaks induced by oxidative and nitrative stress trigger the activation of PARP, and PARP, in turn, mediates cell death and promotes proinflammatory responses. Until recently, most studies focused on the role of PARP in solid organs such as heart, liver, and kidney. We investigated the effect of burn and smoke inhalation on the levels of poly(ADP-ribosylated) proteins in circulating sheep leukocytes ex vivo. Adult female merino sheep were subjected to burn injury (2× 20% each flank, 3 degrees) and smoke inhalation injury (insufflated with a total of 48 breaths of cotton smoke) under deep anesthesia. Arterial and venous blood was collected at baseline, immediately after the injury and 1 to 24 h after the injury. Leukocytes were isolated with the Histopaque method. The levels of poly(ADP-ribosyl)ated proteins were determined by Western blotting. The amount of reactive oxygen species was quantified by the OxyBlot method. To examine whether PARP activation continues to increase ex vivo in the leukocytes, blood samples were incubated at room temperature or at 37°C for 3 h with or without the PARP inhibitor PJ34. To investigate whether the plasma of burn/smoke animals may trigger PARP activation, burn/smoke plasma was incubated with control leukocytes in vitro. The results show that burn and smoke injury induced a marked PARP activation in circulating leukocytes. The activity was the highest immediately after injury and at 1 h and decreased gradually over time. Incubation of whole blood at 37°C for 3 h significantly increased poly(ADP-ribose) levels, indicative of the presence of an ongoing cell activation process. In conclusion, PARP activity is elevated in leukocytes after burn and smoke inhalation injury, and the response parallels the time course of reactive oxygen species generation in these cells.
    Shock (Augusta, Ga.) 02/2011; 36(2):144-8. · 2.87 Impact Factor
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    ABSTRACT: Evidence suggests that lung structure and function are partly maintained by a balance between the competing arginine-metabolizing enzymes arginase and nitric oxide (NO) synthase. Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of NO synthase. It is metabolized by dimethylarginine dimethylaminohydrolase 2 (DDAH-2), which is oxidant-sensitive. The mechanism that induces excess lung collagen deposition in burned patients has not yet been explored. Our objective was to investigate the role of ADMA and the arginase pathway in acute lung injury. An ovine model for burn and smoke inhalation injury was used to assess excess lung collagen deposition. Sheep were deeply anesthetized during the injury, mechanically ventilated, resuscitated with fluid, and killed after either 2 or 3 weeks. Lungs were assessed histologically and biochemically for collagen content, arginase activity, lipid peroxidation product and antioxidant concentration, and protein concentrations. Plasma was assessed for amino acid and nitrate/nitrite concentrations. Burn and inhalation injury resulted in significantly reduced pulmonary function and increased lung collagen deposition. These physiological changes were associated with significantly increased lung arginase activity, collagen synthesis precursor ornithine aminotransferase, and ornithine decarboxylase, which is associated with cell proliferation. Significant decreases in plasma nitrate/nitrite after injury were associated with increased lung ADMA concentrations and decreased DDAH-2 expression. The decreased DDAH-2 expression was associated with significantly increased lipid peroxidation product and decreased antioxidant content in the lung. These data support that excess lung collagen deposition and reduced pulmonary function in acute lung injury after burn and inhalation injury are mediated through the arginase pathway.
    Shock (Augusta, Ga.) 10/2010; 35(3):282-8. · 2.87 Impact Factor
  • Free Radical Biology and Medicine - FREE RADICAL BIOL MED. 01/2010; 49.
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    ABSTRACT: Smoke inhalation injury continues to increase morbidity and mortality in burn patients in both the third world and industrialized countries. The lack of uniform criteria for the diagnosis and definition of smoke inhalation injury contributes to the fact that, despite extensive research, mortality rates have changed little in recent decades. The formation of reactive oxygen and nitrogen species, as well as the procoagulant and antifibrinolytic imbalance of alveolar homeostasis, all play a central role in the pathogenesis of smoke inhalation injury. Further hallmarks include massive airway obstruction owing to cast formation, bronchospasm, the increase in bronchial circulation and transvascular fluid flux. Therefore, anticoagulants, antioxidants and bronchodilators, especially when administered as an aerosol, represent the most promising treatment strategies. The purpose of this review article is to provide an overview of the pathophysiological changes, management and treatment options of smoke inhalation injury based on the current literature.
    Expert Review of Respiratory Medicine 06/2009; 3(3):283-297.