Biomedical subjects
Publications and source records attributed to K Konno.
We have isolated two kinds of cDNAs and the gene encoding human peptide YY and determined their nucleotide sequences. The human peptide YY gene is composed of four exons and three introns spanning approx. 1.2 kbp. Two mRNA species are generated from the gene by alternative splicing of the third intron.
Xanthine derivatives with several functional groups at the 1- or 7-position were synthesized, and their pharmacological activities in guinea pigs were studied. In general, the in vitro tracheal relaxant action and positive chronotropic action of 3-propylxanthines were increased by substitutions with nonpolar functional groups at the 1-position, but decreased by any substitution at the 7-position. On the other hand, because positive chronotropic actions of substituents with allyl, aminoalkyl, alkoxyalkyl, and normal alkyl groups were much less than tracheal muscle became very high with substitutions of 3'-butenyl, (dimethylamino)ethyl, 2'-ethoxyethyl, 3'-methoxypropyl, and n-propyl groups at the 1-position and of 2'-ethoxyethyl, 2'-oxopropyl, and n-propyl groups at the 7-position, compared with theophylline and the corresponding unsubstituted xanthines, 3-propylxanthine and 1-methyl-3-propylxanthine. When compounds were intraduodenally administered to the guinea pig, 1-(2'-ethoxyethyl)-, 1-(3'-methoxypropyl)-, 1-(3'-butenyl)-, and 1-[(dimethylamino)-ethyl]-3-propylxanthines, 1-methyl-7-(2'-oxopropyl)-3-propylxanthine, and denbufylline (1,3-di-n-butyl-7-(2'-oxopropyl)xanthine) effectively inhibited the acetylcholine-induced bronchospasm without heart stimulation or central nervous system-stimulation at the effective dosage range. Particularly, the bronchodilatory effect of 1-(2'-ethoxyethyl)-3-propylxanthine was much stronger and more continuous than those of theophylline and pentoxifylline. On the other hand, there were certain relationships among the in vitro tracheal relaxant activities of these compounds, their affinities for adenosine (A1) receptors in the brain membrane, and their inhibition of cyclic AMP-phosphodiesterase (PDE) in the tracheal muscle. The affinity for A2 receptors of these compounds was very low or negligible. This suggests that both the action on A1 receptors or interaction with adenosine and the cyclic AMP-PDE inhibitory activity contribute to the bronchodilator action of 1- and 7-substituted xanthines. This study indicates that the substitutions with none or low polar functional groups at the 1-position could improve the selectivity and duration of the bronchodilator effects of xanthines.
To clarify the precise role of epidermal growth factor (EGF) on fetal lung development, rabbit anti-mouse EGF (anti-mEGF) antiserum was administered to pregnant mice from days 10 to 17 during late gestation. Control mice were administered either normal rabbit serum (NRS) or physiological saline (PS). Serum EGF was not detected in fetuses from anti-mEGF antiserum treated mothers, but the level in NRS treated control animals was 4.73 +/- 0.66 ng/mL. One day prior to birth, the fetuses were removed and their body and lung weights were measured. There was no difference between body weights and lung weights of anti-mEGF antiserum treated animals and NRS-treated control animals. On light microscopic morphometry, there was no obvious difference between pulmonary architecture of anti-mEGF antiserum treated animals and NRS treated control animals. On transmission electron microscopy, osmiophillic lamellar inclusion bodies were less prominent in the type II epithelial cells in anti-mEGF antiserum treated animals. Electron microscopic morphometric study revealed that the osmiophillic lamellar inclusion bodies in type II epithelial cells of anti-mEGF antiserum treated animals were fewer in number and had decreased area fraction. These findings support the previous finding that EGF promotes epithelial cell differentiation of the fetal lung without affecting body weight and lung weight.
Our study was aimed at elucidating the effect of strenuous maternal exercise (running at a speed of 20 m/min) on fetal somatic growth and lung development. Dams were separated into three groups: (1) exercising during the entire course of pregnancy; (2) exercising from the 16th to the 20th gestational day; and (3) controls treated in the same manner as the exercising animals, but not exercised. There were no differences in the mother's body weight (mother's body weight--total body weight of the fetuses) among the three groups. Overall growth and lung development in fetuses from mothers that exercised during the entire course of pregnancy showed markedly decreased size and weight of body and lung, smaller air spaces and thicker future alveolar walls, and fewer osmiophilic lamellated bodies in alveolar type II cells. The morphometric data of fetuses from mothers that had exercised only during late gestation were similar to those of control animals. The ratio of fetal lung volume to body weight was the same in all three groups. These findings indicate that long-term strenuous maternal exercise induces retardation of fetal lung development, possibly accompanied by a disturbance of overall growth. Maternal exercise represents another means of manipulating lung development.
Evidence that L-glutamate is a neurotransmitter of corticofugal fibers was sought by measuring changes in several biochemical markers of neurotransmitter function in discrete regions of spinal cord after ablation of sensorimotor cortex in monkeys. One and five weeks after unilateral cortical ablation, samples from six areas of spinal cord (ventral, lateral and dorsal regions of the left and right sides) were analysed for choline acetyltransferase (ChAT) activity and contents of amino acid transmitter candidates-glutamic acid (Glu), aspartic acid (Asp), glycine (Gly), taurine (Tau) and gamma-aminobutyric acid (GABA). During one to five weeks after unilateral cortical ablation of the monkey, prolonged hemiplegia in the contralateral side was observed. Histological examination of the spinal cord 5 weeks after unilateral (left) cortical ablation showed no apparent change in either control (ipsilateral, left) or affected (contralateral, right) sides of the cord as examined by the Klüver-Barrera method. The ChAT activity as a cholinergic marker was scarcely changed in any region of either left (control) or right (affected) side of the spinal cord at one and five weeks after unilateral (left side) ablation of the motor cortex. Amino acid levels in each region of the spinal cord were not significantly changed one week after unilateral ablation of the motor cortex. However, a significant decrease of Glu content was observed in the lateral column of the affected (right) side compared to the control (left) side of cervical and lumbar cord five weeks after cortical ablation of the left motor area. No concomitant alterations of other amino acids were detected.(ABSTRACT TRUNCATED AT 250 WORDS)
The effects of catecholamines on ciliary motility of canine bronchial epithelium was studied by a photoelectric method in vitro. Addition of beta-adrenoceptor agonists increased ciliary beat frequency, a rank order of potency being salbutamol > or = BRL 37344, a selective beta 3-adrenoceptor agonist > norepinephrine. The response to BRL 37344 was relatively resistant to the blockade of beta 1- and beta 2-adrenoceptors but was competitively antagonized by the beta 3-adrenoceptor antagonist cyanopindolol, with the pA2 value being lower than that when salbutamol was used as an agonist. These results suggest that beta 3-adrenoceptors exist in canine bronchial epithelium and that stimulation of this receptor subtype may enhance ciliary motility and, hence, mucociliary transport in the airway.
To elucidate the effect of Beraprost, a stable prostaglandin (PG) I2 analogue, on airway smooth muscle functions and its mechanism of action, we studied canine bronchial segments under isometric conditions in vitro. Addition of PGI2 and its analogues dose-dependently relaxed bronchial smooth muscle precontracted with acetylcholine, with the rank order of potency being Beraprost (1) > or = Hoprost (0.65) > PGI2 (0.04), accompanied by the corresponding increase in intracellular cyclic AMP levels. The Beraprost- and PGI2-induced muscle relaxations were significantly inhibited by each of the PG antagonist diphloretin phosphate, the adenylate cyclase inhibitor SQ 22,536, and the Na(+)-K(+)-ATPase inhibitor ouabain. Beraprost and PGI2 at concentrations insufficient to cause muscle relaxation reduced the contractile responses to electrical field stimulation, whereas they were without effect on those to exogenous acetylcholine. These results suggest that Beraprost not only potently relaxes airway smooth muscle through cyclic AMP production and the subsequent stimulation of Na(+)-K(+)-ATPase but also reduces neurally mediated contraction by inhibiting the release of acetylcholine from the cholinergic nerve terminals.
Several naphthalenedisulfonic acid derivatives were found to be selective inhibitors of human cytomegalovirus (CMV) replication in MRC-5 cells. Among the test compounds, the bis-naphthalenedisulfonic acid derivative having an hexamethylene spacer emerged as the most potent inhibitor of CMV replication. Its 50% antivirally effective concentration (EC50) for AD-169 strain was 12 microM, whereas the compound did not affect the growth of mock-infected MRC-5 cells at concentrations up to 500 microM. The naphthalenedisulfonic acid derivatives were also inhibitory to CMV clinical isolates. Virus yield reduction assay revealed that the compounds significantly reduced virus growth in CMV-infected MRC-5 cells. The bis-naphthalenedisulfonic acid derivatives with an hexamethylene spacer suppressed the expression of CMV-induced immediate early, and early antigens at a concentration of 20 microM, whereas the anti-CMV nucleoside ganciclovir did not do so even at the concentration that was 10-fold higher than its EC50 for CMV-induced plaque formation. Furthermore, naphthalenedisulfonic acid derivatives had to be present at the time of virus infection to exert their anti-CMV activity. These results suggest that the compounds are targeted at an early event in the virus replicative cycle, presumably, virus adsorption.
Substructure of the myosin rod and its correlation to the filament formation were investigated by using fish myosin rod. It was found that fish rod contains a unique chymotrypsin susceptible site, 40 kDa from the COOH-terminus or 20 kDa downstream from the subfragment-2/light meromyosin junction (S-2/LMM junction). Cleavage at this new site produced subfragment-2 possessing 95 kDa subunit (95k S-2) and light meromyosin possessing 40 kDa subunit (40k LMM). The latter is the shortest unit ever reported to exhibit filament formation. Moreover, the 40k LMM was able to form filaments independently of the presence of Mg2+, while filament formation of rod and ordinary LMM (70k LMM) was promoted by Mg2+ addition. These results indicated that the Mg2+ binding sites are present within the NH2-terminal 20 kDa region of the 70k LMM. We concluded that the COOH-terminal 40 kDa portion of rod is responsible for the self-assembly ability of myosin, while the NH2-terminal 20 kDa region of the 70k LMM is the regulatory domain for thick filament formation through Mg(2+)-binding.
We have studied ciliary beat frequency (CBF) of rabbit cultured tracheal epithelium by a photoelectric method in-vitro. Addition of erythromycin and roxithromycin increased CBF in a dose-dependent fashion, whereas clarithromycin was without effect. The rank order potency of macrolide was roxithromycin > erythromycin >> clarithromycin. The roxithromycin-induced increase in CBF was not altered by propranolol, AA-861, or verapamil, but partially attenuated by indomethacin. Roxithromycin increased intracellular cAMP concentrations. These results suggest that certain macrolides can stimulate airway ciliary motility probably via prostaglandin- and cAMP-dependent regulatory pathways, which may affect mucociliary transport function in the respiratory tract.
OBJECTIVE: The effect of azelastine on airway mucociliary transport function was studied by measuring ciliary motility of human bronchial epithelium in vitro with a photoelectric method. METHOD: Bronchial epithelial cells were obtained by fibreoptic bronchoscopy, mounted in a Rose chamber, and perfused with Krebs-Henseleit solution. The preparations were placed on a microscope stage equipped with an illuminator, and the variations of light intensity caused by ciliary beating were detected by a photometer. RESULTS: The addition of azelastine to the perfusate increased ciliary beat frequency (CBF) in a dose dependent manner without ciliary discoordination. The mean (SE) maximal increase from the baseline value and the concentration required to produce a half maximal effect were 27.0 (4.2)% and 9.2 x 10(-6) mol/l, respectively. Exposure of the cells to the perfusate containing 3 ppm sulphur dioxide rapidly decreased CBF by 59.2 (5.0)%, and was accompanied by a reduction in intracellular cyclic AMP levels from 38.1 (4.3) to 10.1 (2.4) pmol/mg protein. This effect was prevented by pretreatment of cells with azelastine in a dose dependent manner. CONCLUSIONS: Azelastine not only stimulates ciliary motility of airway epithelium and hence mucociliary transport function, but may also protect against sulphur dioxide induced ciliary dysfunction, probably by inhibiting intracellular cyclic AMP loss.
To determine whether atypical beta-adrenoceptors (beta 3-adrenoceptors) exist in the airway smooth muscle, we studied isolated bronchial segments from dogs under isometric conditions in vitro. Addition of beta-adrenoceptor agonists produced a concentration-dependent relaxation of tissues precontracted with 10(-5) M acetylcholine, rank-order potency being isoproterenol (1) > or = salbutamol (0.95) > or = BRL 37344, a beta 3-selective adrenoceptor agonist (0.83) > norepinephrine (0.10). Under the condition that alpha- and beta 1-adrenoceptors had been blocked, the relaxant response to salbutamol was competitively antagonized by the beta 2-adrenoceptor antagonist ICI 118551 and the pA2 was 7.01 +/- 0.25 (SE), whereas the response to BRL 37344 was resistant, with an apparent pA2 of 5.66. However, cyanopindolol, an antagonist for atypical beta-adrenoceptors, antagonized the BRL-induced relaxation in a competitive manner, with a pA2 of 6.74 +/- 0.11. This pA2 was lower than that when salbutamol was used as an agonist (P < 0.05). Intracellular adenosine 3',5'-cyclic monophosphate (cAMP) levels were increased by BRL 37344 in a concentration-dependent fashion. These results suggest that beta 3-adrenoceptors may exist in canine bronchial smooth muscle and that the stimulation of this type of receptor causes a bronchodilation through a cAMP-dependent pathway.
The effects of surfactant apoprotein A (SP-A) on the superoxide production of rat alveolar macrophages (AM) were studied. Superoxide production was measured by the ferricytochrome c reduction method. When AM were incubated with SP-A only during the measurement of superoxide production, superoxide production was not influenced by SP-A. However, when AM were preincubated with SP-A at a concentration of 1, 2, and 10 micrograms/ml, superoxide production by AM was significantly inhibited (P < 0.05, P < 0.01, P < 0.01, respectively). The superoxide production of AM stimulated by PMA was significantly inhibited by SP-A at a concentration of 1 microgram/ml (P < 0.01), and superoxide production stimulated by zymosan was also inhibited by SP-A at a concentration of 10 micrograms/ml (P < 0.05). Suppression of superoxide production of unstimulated and PMA-stimulated AM was significantly inhibited by anti-SP-A antibody. Superoxide generation by the xanthine and xanthine oxidase system was not affected by the presence of SP-A. Our results suggest that superoxide production of AM can be inhibited by SP-A and that this inhibitory effect on AM is due to a specific effect of SP-A. From these results, it is speculated that SP-A may have a protective role for oxidant injury by AM in the lung.
To determine the effects of the Kampo drugs Saiboku-to, Bakumondo-to and Orengedoku-to on beta-adrenergic function in airway smooth muscle, we studied isolated canine bronchial segments under isometric conditions in vitro. Incubation of tissues with these drugs did not alter the contractile responses to acetylcholine and histamine. The relaxation of tissues precontracted with acetylcholine induced by isoproterenol was augmented by Saiboku-to and Bakumondo-to, so that the concentration of isoproterenol required to produce a half-maximal effect (IC50) was decreased from 4.6 +/- 0.5 x 10(-8) M to 1.9 +/- 1.0 x 10(-8) M (P < 0.05) and to 1.0 +/- 0.8 x 10(-8) M (P < 0.01), respectively, whereas Orengedoku-to had no effect. These effects were concentration-dependent with the threshold concentrations being 0.3 mg/ml for Saiboku-to and 0.1 mg/ml for Bakumondo-to. Saiboku-to and Bakumondo-to did not affect cyclic AMP levels in airway smooth muscle per se but potentiated the isoproterenol-induced cyclic AMP accumulation. These results suggest that Saiboku-to and Bakumondo-to but not Orengedoku-to potentiate beta-adrenergic function in airway smooth muscle, which may reflect the efficacy of these drugs on airway hyperresponsiveness and asthma.
To determine whether atypical beta-adrenoceptor (beta 3-adrenoceptor) agonist, BRL 37344, stimulates airway ciliary motility and, if so, what its mechanism of action is, we studied rabbit tracheal epithelium in vitro. Epithelial cells cultured for 7 days were mounted in the Rose chamber and ciliary beat frequency (CBF) was measured by a photoelectric method. Addition of BRL 37344 rapidly increased CBF from 15.5 +/- 0.6 to 20.3 +/- 1.4 Hz (mean +/- SE, p < 0.01), which remained elevated for at least 20 min. This effect was concentration-dependent, the maximal increase from the baseline value and the EC50 being 27.2 +/- 0.4% (mean +/- SE, p < 0.01) and 1.75 x 10(-7) M, respectively. Other beta-adrenoceptor agonists likewise increased CBF, where the rank order of potency was salbutamol (1) > BRL 37344 (0.57) >> norepinephrine (1.3 x 10(-3)). Preincubation of cells with propranolol inhibited the increase in CBF produced by salbutamol, and BRL 37344 significantly raised intracellular cyclic AMP contents. These results suggest that beta 3-adrenoceptor probably exists in rabbit tracheal epithelium and that stimulation of this type of receptor increases ciliary motility through a cyclic AMP-dependent mechanism, an effect that might be of value in enhancing mucociliary transport function in the airway.
To determine the effect of somatostatin (SRIF) on airway smooth muscle contractile and relaxant responses and its mechanism of action, we studied rabbit tracheal smooth muscle under isometric conditions in vitro. SRIF did not change the contractile responses to electrical field stimulation and acetylcholine, but it inhibited the relaxant responses to isoproterenol in a concentration-dependent manner, an effect that was reversed by treatment of tissues with pertussis toxin (PTX). In contrast, the forskolin- and verapamil-induced relaxations were not altered by SRIF. SRIF also attenuated the increase in intracellular cyclic AMP levels in response to isoproterenol. These results suggest that SRIF may decrease beta-adrenoceptor-mediated muscle relaxation by acting at the site proximal to cyclic AMP synthesis, especially at PTX-sensitive GTP-binding regulatory protein, Gi, coupled to adenylate cyclase.
The relationships between emphysema, bronchiolar lesions, and pulmonary function tests were studied in very mild centrilobular emphysematous lungs. The severity of emphysema was significantly related to VC, FEV1, FEV1/FVC, the slope of phase III, CC/TLC and static elastic lung recoil. Emphysema was not correlated with DLco. Bronchiolar lesions were poorly correlated with pulmonary function tests. Emphysema was related to bronchiolar lesions, such as smooth muscle hypertrophy and goblet cell metaplasia. These findings suggest that alterations of the mechanical property of the lung occur in very mild centrilobular emphysema. Measurements of these abnormalities may be useful to detect early emphysema.
To determine whether activations of K-channel and Na-K-ATPase are involved in the action of bronchodilating agents, we studied the effects of various K-channel blockers and ouabain, a Na-K-ATPase inhibitor, on the relaxant responses of canine airway smooth muscle in vitro. Addition of charybdotoxin (ChTx), a Ca-activated potassium channel blocker, and ouabain had no effect on the responses to verapamil and BRL 38227, but inhibited those to salbutamol, forskolin, DBcAMP, and nitroprusside. Apamin, a low-conductance K-channel blocker, and glibenclamide, an ATP-sensitive K-channel blocker, had no effect on the smooth muscle relaxation induced by any agents. The inhibitory effect of ChTx was predominant when the concentration of bronchodilator was low, whereas the effect of ouabain was observed in the presence of a high concentration of bronchodilator. These results suggest that Ca-activated K-channel and Na-K-ATPase may participate in the relaxation induced by bronchodilators that can raise intracellular cyclic nucleotides, and that the contribution of each component differs under different concentrations of the drugs.