Hirofumi Fujita

Effect of Antioxidants on Hyperoxia-Induced ICAM-1 Expression in Human Endothelial Cells

The regulating mechanism of hyperoxia-induced ICAM-1 expression has not been elucidated. We studied the effect of antioxidants, including superoxide dismutase (SOD), catalase and N-acetylcysteine (NAC), on hyperoxia-induced ICAM-1 expression in human pulmonary artery endothelial cells (HPAEC) and human umbilical vein endothelial cells (HUVEC). Cells were cultured to confluence and exposed to either hyperoxic or normoxic gas with or without various kinds of antioxidants. The levels of ICAM-1 expression in the endothelial cells and the concentrations of reduced (GSH) and oxidized glutathione (GSSG) in the media were examined by flow cytometry and by spectrophotometry, respectively. After 48-hour exposure to hyperoxia, ICAM-1 expression was increased (HPAEC; 161 +/- 21% and HUVEC; 163 +/- 16%) and total glutathione concentration in the media was decreased as compared with normoxia. SOD did not change the GSH and GSSG concentrations in the media. Catalase dose-dependently decreased the supernatant GSSG concentration in both HPAEC and HUVEC, while the GSH concentration was nearly constant. NAC dose-dependently increased the supernatant GSH concentrations in both HPAEC and HUVEC. There was no difference in the supernatant GSSG concentrations between the NAC-treated HPAEC and HUVEC. There was no difference in ICAM-1 expression in either HPAEC or HUVEC with SOD treatment. ICAM-1 expressions in 100 U/ml (236 +/- 20%) and 1,000 U/ml (315 +/- 36%) of catalase were increased in HPAEC, and that in 1,000 U/ml (440 +/- 209%) of catalase was increased in HUVEC. Five and 10 U/ml of NAC decreased ICAM-1 expression in HPAEC (141 +/- 26% and 113 +/- 11%) and HUVEC (119 +/- 23% and 106 +/- 7%), respectively. These results suggest that extracellular glutathione may play a role in regulating hyperoxia-induced ICAM-1 expression in HPAEC and HUVEC.

Constriction and dilatation of pulmonary arterial ring by hydrogen peroxide - Importance of prostanoids

Reactive oxygen species (ROS) have been considered as one of the important factors causing acute lung injury associated with alveolar flooding, especially in the case of sepsis, fat embolism, hyperoxic lung damage as well as reperfusion injury (Hefner and Repine, 1989). Among them, hydrogen peroxide (H2O2) and its derivatives are relatively stable as compared to other ROS and are taken notice as the substance altering pulmonary hemodynamics accompanied by alveolar flooding (Archer et al., 1989; Gurtner and Burke-Wolin, 1991; Barnard et al., 1992). Effects of H2O2 on pulmonary circulation is much complicated, namely vasoactive action of H2O2 is dependent on the vascular tone. In other words, H2O2 relaxes the pulmonary vessel when the tone is high but constricts it when the tone is low (Burke-Wolin et al., 1991). Gurtner and Burke-Wolin (1991) reported that vasoconstrictive effect of H2O2 would be mainly mediated through thromboxane A2 (TXA2) generated in the endothelial cells of pulmonary vasculature. However, several authors (Madden et al., 1986; Smith, 1986; Takayasu et al., 1990) showed that arachidonate metabolism might exist not only in endothelial cells but also in smooth muscle cells and fibroblasts in vascular walls, indicating that the conclusion derived by Gurtner and Burke-Wolin (1991) should be reassessed.

Ventilation-perfusion inequality and diffusion impairment in acutely injured lungs

To assess the significant role of diffusion impairment and its unequal distribution in acutely injured lungs with alveolar flooding, oleic acid was intravenously injected into twenty-five mongrel dogs. The animals were divided into two groups, A and B. 0.1% CO in air was delivered, as an inspired gas, to the animals of group A. Simultaneously, saline containing a trace amount of six foreign inert gases was infused through a peripheral vein. While allowing the animals in group B to breathe air, saline containing ethylene, acetylene and freon 22 was infused. After injection of oleic acid, group A revealed increase in intrapulmonary shunt accompanied by a marked broadening of ventilation-perfusion (VA/Q) and diffusing capacity-perfusion (G/Q) distributions. A considerable amount of total cardiac output was received by the lung areas with low G/Q ratios where significant diffusion limitation was predicted to occur. Group B showed that excretion of freon 22 (gas with lower diffusivity) in injured lungs was considerably distorted as compared to those of ethylene and acetylene (gases with higher diffusivities), again ascertaining the importance of diffusion limitation in lungs with exudate in alveolar regions.

Impact of pharmacokinetics and pharmacogenetics on the efficacy of pranlukast in Japanese asthmatics

Background and objective:  Wide inter‐individual variability in therapeutic effects limits the efficacy of leukotriene (LT) receptor antagonists in the treatment of asthma. We have reported that genetic variability in the expression of LTC4 synthase is associated with responsiveness to pranlukast in Japanese asthmatic patients. However, the effects of pharmacokinetic variability are less well known. This was an analysis of the pharmacokinetics of pranlukast in a population of adult asthmatics, and its effect on clinical responses. Other factors that may be related to the therapeutic effects of pranlukast, including LTC4 synthase gene polymorphisms, were also investigated.

Attenuation of live E. coli-induced acute lung injury by X-ray irradiation in guinea pigs.

Abstract. Irradiation is suspected to injure inflammatory cells, such as neutrophils and mononuclear phagocytes, cells known to contribute to the development of acute lung injury (ALI). This study examined whether preexposure to x-ray irradiation modifies ALI induced by E. coli injected intravenously in guinea pig. Thirty animals were divided into two control and two irradiated subgroups: the first control group received saline only (n= 8), and the second control group received E. coli, 2 × 109/kg body weight, suspended in saline (n= 6), IV. The first irradiated group received a single 12-Gy dose + saline (n= 6), and the second irradiated group received a single 12-Gy dose +E. coli (n= 10). The lung wet-to-dry-weight ratio (W/D) and 125I-albumin lung tissue/plasma ratio (T/P) were measured as markers of lung injury. W/D was significantly higher in the control E. coli group than in the other groups. T/P in the control E. coli group was also increased compared with T/P measured in the other groups. In the control E. coli group, a marked increase in bronchoalveolar lavage (BAL) neutrophils was observed compared with the control saline group. However, no significant difference in BAL neutrophil counts was observed between the control and irradiated E. coli groups. In contrast, BAL macrophages were significantly reduced in the irradiated E. coli groups compared with the control E. coli group. These findings suggest that x-ray irradiation attenuates E. coli–induced ALI in guinea pigs, an effect explained, at least in part, by a reduction in the number of alveolar macrophages.

Behavior of Stimulated Leukocytes in the Pulmonary Microcirculation of Perfused Rat Lungs

To investigate the dynamics of activated leukocytes and the roles of CD18-ICAM-1 pathway, we examined the effects of rat IL-8 and monoclonal antibodies (mAbs) against CD18 and ICAM-1 on the behavior of leukocytes in microvessels of perfused rat lungs. Specific pathogen free male Sprague-Dawley rats were used. Perfused rat lungs were prepared so as to obtain stable physiological shear rates. We used a confocal laser scanning microscope equipped with a high speed video analysis system to visualize pulmonary microcirculation. Rat leukocytes were activated with rat IL-8. No rolling leukocytes were observed in either pulmonary arterioles or venules, and leukocytes were sequestered in capillaries. The majority of unstimulated capillary leukocytes moved smoothly. About 50% of stimulated leukocytes, however, showed a transient cessation of movement in pulmonary capillaries. Rat IL-8 decreased the relative leukocyte velocities against mean blood velocities in capillaries (45%) and venules (65%), and increased intracapillary neutrophils. Anti-CD18 and anti-ICAM-1 mAbs attenuated these changes. These results suggest that unique features exist in the interaction between activated leukocytes and pulmonary microvessels, and that CD18-ICAM-1-dependent capillary sequestration is one of the major mechanisms by which activated leukocytes accumulate in lungs.

Endotoxin-induced acute lung injury in guinea pigs.

SDZ MRL 953 (SDZ), a novel immunostimulatory lipid A analog, has been reported to have immunopharmacological activities similar to those of lipopolysaccharide (LPS) but to have little of the toxicity of LPS. We investigated the effects of pretreatment with SDZ on Escherichia coli endotoxin-induced acute lung injury in guinea pigs. Four experimental groups consisted of saline control (n = 16), SDZ (-12 h) plus LPS (2 mg/kg of SDZ per kg of body weight injected intravenously 12 h before intravenous injection of 2 mg of LPS per kg; n = 15), SDZ (-10 min) plus LPS (SDZ injected 10 min before LPS injection; n = 10), and LPS alone (n = 16). The animals were sacrificed, and lung tissue was sampled 4 h after LPS or saline infusion. Lung injury was assessed by measuring the wet weight-to-dry weight ratio and the level of 125I-labeled albumin accumulation in bronchoalveolar lavage fluid relative to that in plasma. In the SDZ (-12 h) plus LPS group, these two parameters of acute lung injury were decreased compared with those in the LPS alone group. However, they were not decreased in the SDZ (-10 min) plus LPS group. We conclude that SDZ attenuates endotoxin-induced acute lung injury when it is administered 12 h before LPS injection. The attenuating effects of SDZ are speculated to be due to down regulation of the response to endotoxin rather than to receptor blocking.