Shozo Koyama

Cross-talk between lung and systemic circulation during carbon nanotube respiratory exposure. Potential biomarkers.

Nanotechnology is an emerging field that demands urgent development of adequate toxicology and risk assessment. The previous experimental data on carbon nanotube respiratory exposure strongly suggest the need for complex evaluation of potential toxicity. Our work demonstrates that after carbon nanotube deposition in the lung, acute local and systemic responses are activated and characterized by a blood gene and protein expression signature. The approach described here will foster the development of biomarkers for application in human screening of nanoparticle exposure.

A new substance (Yoshixol) with an interesting antibiotic mechanism from wood oil of Japanese traditional tree (Kiso-Hinoki), Chamaecyparis obtusa

1. A neutral wood oil was extracted from Chamaecyparis obtusa (Kiso-Hinoki), which has been trusted nationally and preserved historically in the central part of Japan (Kiso, Nagano). 2. Hinokitiol, or thujaplicin (C10H12O2), which has been believed to exist in Cupressaceae, was not found in this neutral wood oil. Some differences between the extracting processes of the natural products are discussed. 3. A new chemical substance (Yoshixol, 4,4-dimethyl-6-methylene-2-cyclohexen-1-one) was simulated by several criteria (details in the text) as a major candidate of the neutral wood oil from Chamaecyparis obtusa. Thus, Yoshixol was newly synthesized. 4. The antibiotic effects of hinokitiol, the neutral wood oil and Yoshixol on methicillin-resistant Staphylococcus aureus (MRSA) were examined bacteriologically and morphologically. 5. All of the aforementioned three test materials showed complete antibiotic effects on MRSA by the bacteriological examination. However, the morphological findings showed entirely different aspects of cell death. 6. Hinokitiol caused an aggregative, degenerative and/or necrotic aspect, but the neutral wood oil and Yoshixol produced characteristic aspects: separation of contacted cells, blebbing, bugging-like eruption, formation of granules and an extensive reduction of individual cell size of MRSA. 7. Yoshixol was able to enhance those antibiotic effects on MRSA distinctly more than the neutral wood oil. 8. Yoshixol also showed a strong antibiotic effect on Escherichia coli, Mycobacterium chelonei, Pseudomonas aureginosa and Candida albicans. Morphological observations of those bacilli after Yoshixol revealed characteristic aspects of separation of contacted cells, bugging-like swelling, granulation, ballooning and reduction of cell size. 9. A possible mechanism of Yoshixol is discussed in regard to a molecular orbital theory on the basis of its electron orbits and to a thermodynamic interaction with the prokaryotic cell membrane. On the basis of the molecular properties of Yoshixol, future biological interests and possible biological effects of Yoshixol are suggested.

The effects of sevoflurane, enflurane, and isoflurane on baroreceptor-sympathetic reflex in rabbits

This study was designed to determine the effects of sevoflurane, enflurane, and isoflurane ranging from 0.5 to 1.25 minimum alveolar anesthetic concentration (MAC) on spontaneous efferent renal sympathetic nerve activity (RNA) and the baroreceptor-sympathetic reflex in rabbits. Enflurane produced significant decreases in spontaneous RNA by 22.5% +/- 6.6% at 1.0 MAC, while sevoflurane and isoflurane, at the equivalent MAC, did not. All of the anesthetics attenuated the baroreflex gain similarly when mean blood pressure (MBP) was changed by sodium nitroprusside or phenylephrine intravenously. However, the sensitivity of baroreceptors at the aortic wall was not changed by any anesthetic, because no changes in the relationship between aortic nerve activity and MBP were obtained in anesthetic concentration even at 1.25 MAC. Furthermore, these anesthetics suppressed the sympathoinhibitory response to aortic nerve stimulation above 1.0 MAC. In conclusion, enflurane inhibits RNA to a greater degree than sevoflurane or isoflurane. However, all three anesthetics depress the reflex regulation of RNA to the same degree. The suppression on the baroreceptor-sympathetic reflex does not appear to be related to a change in the receptor sensitivity on the aorta, but is mediated by suppression of the central or peripheral sympathetic integrating system. (Anesth Analg 1996;82:342-8)

Participation of baroreceptor reflexes in blood pressure and sympathetic nerve responses to a synthetic human atrial natriuretic peptide in anesthetized dogs

The effects of a synthetic alpha human atrial natriuretic polypeptide (hANP) on systemic blood pressure and renal nerve activity were studied before and after sinoaortic denervation with vagotomy in anesthetized dogs. Intravenous injection of hANP (1 microgram/kg) in animals with the neuraxis intact produced a mild decrease in systemic blood pressure and a significant increase in renal sympathetic nerve activity. However, in animals with sinoaortic denervation and vagotomy, a profound fall in blood pressure occurred after administration of hANP. Renal nerve activity in these animals was reduced significantly, in parallel with decreases in systemic blood pressure. These results indicate that activation of peripheral baroreceptors minimizes the hypotension induced by intravenous injection of hANP, and may mask at least centrally mediated cardiovascular effects of hANP.

Thromboxane A2 analogue contracts predominantly the hepatic veins in isolated canine liver.

Thromboxane A2 (TxA2) is a potent vasoconstrictor and has been implicated as a mediator of liver diseases such as ischemic-reperfusion injury. We determined the effects of TxA2 and the well-known hepatic venoconstrictor histamine, on the vascular resistance distribution and liver weight in isolated canine livers perfused with blood via the portal vein. The stable TxA2 (STA2; 20 micrograms, n = 5) and histamine (5 micrograms, n = 6) similarly increased the hepatic total vascular resistance, 2.5- and 2.4-fold, respectively. The increase in the hepatic venous resistance was significantly greater than that of the portal resistance (threefold vs. 1.9-fold for STA2; threefold vs. 1.8-fold for histamine). Predominant hepatic venoconstriction induced by both agents was confirmed in livers perfused in a reverse direction from the hepatic vein to the portal vein, as shown by marked precapillary vasoconstriction. STA2 transiently increased liver weight loss (-3.6 g/100 g liver weight), followed by a gradual weight gain (9.0 g/100 g). Histamine caused a progressive weight gain (9.1 g/100 g). In conclusion, similar to histamine, TxA2 constricts predominantly the hepatic vein in isolated canine livers.

Contribution of baroreceptor reflexes to blood pressure and sympathetic responses to cholecystokinin and vasoactive intestinal peptide in anesthetized dogs.

The effects of synthetic vasoactive intestinal peptide (VIP) and cholecystokinin (CCK) on systemic blood pressure and renal nerve activity were studied before and after cervical vagotomy, and sino-aortic denervation with vagotomy in anesthetized dogs. Intravenous injection of VIP (5 micrograms/kg) in animals with an intact neuraxis produced a significant decrease in systemic blood pressure and a significant increase in renal nerve activity. These responses to VIP did not change after vagotomy and after complete denervation, VIP did not cause any change in renal nerve activity, even during hypotension. The level of hypotension after complete denervation was greater than that under other conditions. These results indicate that the cardiovascular effects of VIP are reduced by activation of the systemic baroreceptors. Intravenous injection of CCK (10 micrograms/kg) in animals with an intact neuraxis produced significant decreases in blood pressure and renal nerve activity. These responses to CCK were abolished in animals with cervical vagotomy only. However, following complete denervation of the carotid sinus and total section of the vagal nerves, CCK caused a significant increase in blood pressure and renal sympathetic nerve activity. These results indicate that the sympathetic depressor effect of CCK may be mediated by activation of the vagal afferents, and that the sympathetic pressor effect may be due to a direct action of CCK on the central nervous system. Thus, each gastrointestinal peptide may regulate the cardiovascular system through a different mechanism.

Effects of thromboxane A2 analogue on vascular resistance distribution and permeability in isolated blood-perfused dog lungs

This study was designed to determine the effects of thromboxane A2 (TxA2) on the distribution of vascular resistance, lung weight, and microvascular permeability in isolated dog lungs perfused at a constant pressure with autologous blood. The stable TxA2 analogue (STA2; 30 μg, n = 5) caused an increase in pulmonary capillary pressure (Pc) assessed as double-occlusion pressure to 14.0 ± 0.4 mmHg from the baseline of 7.9 ± 0.3 mmHg with progressive lung weight gain. Pulmonary vascular resistance increased threefold exclusively due to pulmonary venoconstriction. Pulmonary venoconstriction was confirmed in lungs perfused in a reverse direction from the pulmonary vein to the artery (n = 5), as evidenced by marked precapillary vasoconstriction and a sustained lung weight loss. Furthermore, in lungs perfused at a constant blood flow (n = 5), STA2 also caused selective pulmonary venoconstriction. Vascular permeability measured by the capillary filtration coefficient and the isogravimetric Pc at 30 and 60 min after STA2 infusion did not change significantly from baseline in any lungs studied. Moreover, elevation of Pc by raising the venous reservoir of the intact lobes (n = 5) to the same level as the STA2 lungs caused a greater or similar weight gain compared with the STA2 lungs. Thus, we conclude that TxA2 constricts selectively the pulmonary vein resulting in an increase in Pc and lung weight gain without significant changes in vascular permeability in isolated blood-perfused dog lungs.

EFFECT OF ONO-5046, A SPECIFIC NEUTROPHIL ELASTASE INHIBITOR, ON THE PHORBOL MYRISTATE ACETATE-INDUCED INJURY IN ISOLATED DOG LUNG

Phorbol myristate acetate (PMA) activates neutrophils and causes acute lung injury. We determined the effect of ONO-5046, a specific neutrophil elastase inhibitor, on the increase in microvascular permeability induced by PMA in isolated dog lung perfused with autologous blood at a constant perfusion flow. The vascular permeability was assessed by the capillary filtration coefficient (Kf, c) and the solvent-drag reflection coefficient (sigma f). PMA (13.3 micrograms) increased vascular permeability, as evidenced by an increase in Kf, c from 0.18 +/- 0.02 to 0.92 +/- 0.14 mL/min/cmH2O/100 g and a decrease in sigma f to 0.35 +/- 0.01 as compared to control values of 0.69 +/- 0.06. The PMA-induced changes in Kf, c and sigma f were dose-dependently attenuated by pretreatment with ONO-5046 (2-20 mg). We conclude that ONO-5046 can effectively attenuate the PMA-induced injury in the isolated blood-perfused dog lungs.

Lecithinized superoxide dismutase attenuates phorbol myristate acetate-induced injury in isolated dog lung

Lecithinized superoxide dismutase, a lecithin derivative bound to recombinant human CuZn superoxide dismutase, has a higher affinity for cells such as polymorphonuclear leukocytes and endothelial cells than recombinant human CuZn superoxide dismutase has. We determined the protective effects of lecithinized superoxide dismutase on the increased microvascular permeability induced by phorbol myristate acetate (PMA) in isolated dog lungs. Microvascular permeability was assessed by the capillary filtration coefficient (Kf,c) and solvent drag reflection coefficient (sigma(f)). PMA (13.3 microg) increased microvascular permeability, as evidenced by an increase in Kf,c and the small sigma(f) value. Lecithinized superoxide dismutase at both low (4800 U) and high doses (48,000 U) inhibited the PMA-induced increase in Kf,c, but only the high dose of lecithinized superoxide dismutase attenuated the decrease in sigma(f). Recombinant human CuZn superoxide dismutase did not affect the PMA-induced increase in vascular permeability at either a low (4800 U) or a high dose (48,000 U). These findings suggest that lecithinized superoxide dismutase has a protective effect against oxygen radical-induced lung injury in isolated dog lungs.

rBPI23 attenuates endotoxin-induced cardiovascular depression in awake rabbits.

We determined the effect of a recombinant N-terminal fragment of bactericidal/permeability-increasing protein (rBPI23) on hemodynamic and renal sympathetic responses to lethal endotoxemia in unanesthetized rabbits. Endotoxin was continuously infused intravenously (200 micrograms/kg/h) over 120 min with simultaneous infusion of either rBPI23 (3 mg/kg bolus followed by 6 mg/kg/h over 120 min; n = 6) or thaumatin (the same dose as rBPI23), a control cationic protein with a molecular weight and isoelectric point similar to that of rBPI23 (n = 9). Tissue blood flow was also determined using colored microspheres to the left ventricle, renal cortex, liver, and skeletal muscle. Seven of nine animals treated with endotoxin and thaumatin died between 45 and 120 min after start of the infusion, whereas all animals with rBPI23 treatment were alive throughout the entire 2 h experimental period. A transient increase in renal sympathetic nerve activity was observed in the thaumatin-treated animals followed by sympathoinhibition with concomitant decreases in heart rate, blood pressure, and cardiac output. Tissue blood flow to all measured organs gradually decreased in animals receiving endotoxin and thaumatin. However, rBPI23 abolished all these deleterious responses to endotoxin. In conclusion, rBPI23 attenuates the acute lethal sympathoinhibitory and hemodynamic effects of endotoxemia in awake rabbits.