Takeyasu Fukushima

The role of nitric oxide in cholinergic neurotransmission in rat trachea

1 We have investigated the role of nitric oxide (NO) in cholinergic contraction in rat trachea. 2 Methylene blue (10 nm to 30 μm) potentiated cholinergic contraction induced by electrical field stimulation (EFS) at 5 Hz in a concentration‐dependent fashion. At a concentration of 30 μm, methylene blue decreased responses to log EFS frequency, producing 50% of maximum contraction from a control value of 0.74 ± 0.09 Hz to 0.30 ± 0.05 Hz without a significant effect on concentration‐response curves to acetylcholine (ACh). 3 NG‐monomethyl‐l‐arginine (l‐NMMA; 100 μm) also potentiated cholinergic contraction induced by EFS at 5 Hz (131.5 ± 4.6% of control) without having any effect against ACh (3 μm)‐induced contractions. Likewise, l‐NMMA (100 μm) significantly increased EFS (5 Hz)‐evoked release of ACh from tracheal segments into the bath solution (51.4 ± 4.0 pmol ml−1 in the presence of l‐NMMA and 35.0 ± 1.8 pmol ml−1 in the absence of l‐NMMA, respectively). 4 Administration of NO (present in acidified soluton of NaNO2) (1 nm to 10 μm) and sodium nitroprusside (100 nm to 10 μm) concentration‐dependently reduced EFS (5 Hz)‐induced cholinergic contractions without having a significant effect on ACh (3 μm)‐induced contractions. These results were unaffected by prior exposure of the tissues to l‐NMMA (100 μm). 5 Dibutyryl cyclic GMP (3 mm) also reduced cholinergic contractions induced by EFS at 5 Hz (70.1 ± 3.6% of control) without any significant effect on ACh (3 μm)‐induced contractions. 6 Pretreatment of tissues with capsaicin (30 μm) or α‐chymotrypsin (1 u ml−1) failed to inhibit methylene blue (30 μm)‐induced potentiation of responses to EFS at 5 Hz. 7 These results suggest that an endogenous NO‐like factor may mediate prejunctional inhibition of cholinergic contraction through a cyclic GMP‐dependent mechanism in rat trachea.

The role of carbon monoxide in lucigenin-dependent chemiluminescence of rat alveolar macrophages

We have investigated the role of carbon monoxide (CO) in lucigenin-dependent chemiluminescence of alveolar macrophages from rat lungs. CO (10 nM to 1 microM) decreased chemiluminescence of alveolar macrophages in a concentration-dependent fashion. At a concentration of 1 microM, CO significantly increased intracellular cyclic GMP levels from a control value of 175 +/- 25 fmol/2 x 10(6) cells to 431 +/- 49 fmol/2 x 10(6) cells. Pretreatment of alveolar macrophages with NG-monomethyl-L-arginine (100 microM) failed to inhibit CO (1 microM)-induced decreases in chemiluminescence of alveolar macrophages (3.7 +/- 0.7 cpm x 10(3) in the presence of NG-monomethyl-L-arginine and 3.4 +/- 0.6 cpm x 10(3) in the absence of NG-monomethyl-L-arginine) and CO (1 microM)-induced increases in intracellular cyclic GMP levels (452 +/- 65 fmol/2 x 10(6) cells in the presence of NG-monomethyl-L-arginine and 419 +/- 58 fmol/2 x 10(6) cells in the absence of NG-monomethyl-L-arginine). Decreases in chemiluminescence of alveolar macrophages induced by CO (1 microM) were concentration-dependently inhibited by methylene blue (from 0.1 microM to 10 microM). Dibutyryl cyclic GMP (db cyclic GMP) (1 mM) also reduced chemiluminescence of alveolar macrophages (1.5 +/- 0.3 cpm x 10(3) in the presence of db cyclic GMP and 3.6 +/- 0.6 cpm x 10(3) in the absence of db cyclic GMP). In contrast to CO and db cyclic GMP, zinc protoporphyrin-9 (10nM to microM), an inhibitor of heme oxygenase potentiated chemiluminescence of alveolar macrophages in a concentration-dependent fashion.(ABSTRACT TRUNCATED AT 250 WORDS)

Tuberculin responses and risk of pneumonia in immobile elderly patients

BACKGROUND Delayed type hypersensitivity (DTH) response to tuberculin is an important marker of T helper 1 (Th1) mediated acquired immunity against tuberculosis. Depressed DTH responses to tuberculin are observed in immobile elderly individuals with reduced activities of daily living (ADL) scores. Immobility in older people increases the risk of pneumonia related mortality. The decline in the competence of the immune system might be a reason for the increased susceptibility to infection in the elderly and the depressed DTH response to tuberculin might be a predictor for increased risk of pneumonia. METHOD The DTH responses to tuberculin were examined in 49 older patients with limited ADL of similar severity, all of whom had a past history of positive DTH responses to tuberculin. Responses with an induration diameter of ⩾10 mm were considered positive. Reactivities of Th1 and Th2 lymphocytes taken from the peripheral blood of each subject were also examined and compared between the positive tuberculin responders and the negative tuberculin responders. The rates of pneumonia in these groups were then compared prospectively for two years. RESULTS The number of CD4 lymphocytes differed significantly between patients with a positive tuberculin response (n=22, mean (SE) 1018 (118) × 106/l) and those with a negative response (n=27, 666 (80) × 106/l, p=0.02). Likewise, the number of Th1 cells was significantly higher in positive tuberculin responders than in negative responders (276 (5.8) × 106/l versus 149 (14) × 106/l, p=0.01). However, the numbers of Th2 cells were similar between patients with positive and negative responses (22 (3) × 106/l versus 19 (3) × 106/l, p=0.41). During the follow up period new pneumonia was diagnosed in 18 (67%) of the 27 negative tuberculin responders and in seven (31%) of the 22 positive tuberculin responders. According to the Cox regression model, a hazard ratio of developing pneumonia in patients with a negative tuberculin response compared with those with a positive tuberculin response was 2.57 (95% CI 1.12 to 6.17, p=0.03). CONCLUSION The diminished DTH response to tuberculin may be a predictor for increased risk of pneumonia in elderly disabled patients.

DEPRESSION OF T HELPER-1 AND TUBERCULIN RESPONSES IN OLDER BED-BOUND PATIENTS

ticularly in older patients, although constipation is controlled. Because sympathetic activity is increased and parasympathetic activity is suppressed in CHF patients,’72 muscular tone of the bowel may become inhibited. Although ileus has been proposed as a complication of CHF, few studies have specifically focused on the relationship between ileus and CHF.3 We examined 109 consecutive patients with CHF who were admitted to our hospital between January 1996 and September 1997. There were 54 men and 55 women (mean age 74 years (SD 12)) in the group. Ileus was diagnosed according to clinical signs and plain abdomen X-ray findings. As an age-matched control group, we examined 114 consecutive patients with fractures of the femoral neck who were hospitalized during the same period, because they were also confined to bed rest for a long period. The controls consisted of 29 men and 85 women (mean age 77 years (SD 11)). The incidence of ileus in CHF patients was 14% (n = 15, 7 men, 8 women), whereas incidence was 3% in the control group. The difference between the two groups was significant ( P < .002). Additional causative factors other than the bedridden state may contribute to the development of ileus in the CHF patients. Table 1 shows the clinical profile of CHF patients with or without ileus. CHF patients with ileus were significantly older than those without ileus ( P < .05). In particular, the incidence of ileus in CHF patients (57 patients) aged 75 years or older was significantly higher than that in younger CHF patients ( P < ,001). The number of severely ill patients who died or required respirator control was higher in the ileus group of CHF patients compared with the non-ileus group ( P < .001). The incidence of chronic renal failure (serum creatinine > 2 mg/dL), diabetes mellitus, or hypertension was not different between the ileus and the non-ileus groups. Serum sodium and potassium levels in the ileus group were almost within normal range. Etiologically, there was no difference in the original diseases between the ileus and the non-ileus groups. In four studies of CHF patients with ileus, no occupied lesion in the intestine was confirmed at autopsy or with colon fiberscopy.

Cytoplasmic motility reflects phagocytic activity in alveolar macrophages from dog lungs

To determine whether cytoplasmic motility relates to phagocytic activity of alveolar macrophages (AM), we measured the remanent field strength (RFS) from the AM containing Fe3O4 particles (5 x 10(6) cells) and the number of phagocytized fluorescent latex spheres in the AM without having Fe3O4 particles (10(6) cells) harvested by broncho-alveolar lavage from dog lungs in vitro. Cytoplasmic motility was estimated from the relaxation rate (lambda o; min-1) calculated from the decay curve of RFS and the number of phagocytized latex spheres was counted using fluorescent microscopy after the addition of latex spheres (5 x 10(7)). The tumor necrosis factor increased both lambda o and the number of phagocytized latex spheres, and cytochalasin D and colchicine decreased them in a concentration-dependent fashion. Increases and decreases in lambda o induced by drugs paralleled the number of phagocytized latex spheres. These results suggest that cytoplasmic motility reflects phagocytic activity of AM and both cytoplasmic movement and phagocytosis may be regulated by a similar mechanism in the cytoskeletal system.

Allergic Sensitization in Elderly Patients with Chronic Obstructive Pulmonary Disease

To study whether allergic sensitization occurs in elderly patients with chronic pulmonary obstructive disease (COPD), we examined serum IgE and skin test reactivity to allergens in three age-matched groups of normal subjects, and in patients with COPD and bronchial asthma (BA). Serum IgE was significantly higher in patients with COPD and BA than in normal subjects (p less than 0.05), and patients with COPD showed serum IgE levels as high as those of patients with BA. However, the skin test scores were significantly higher in patients with BA than in normal subjects and patients with COPD (p less than 0.05). Neither serum IgE nor skin test score significantly correlated with FEV1%, PaO2, PaCO2 or Brinkmans Index in any group (p greater than 0.20). These results suggest that allergic sensitization occurs in elderly patients with COPD and that symptoms associated with COPD may be partly due to allergic inflammation.

Effect of mabuterol on tracheal mucociliary clearance of magnetized iron particles in anesthetized dogs.

We studied the effect of mabuterol on the tracheal mucociliary clearance of magnetized iron particles in anesthetized dogs. A catheter was inserted directly into the trachea through the mouth and 0.05 ml of saline solution with 30 mg of Fe3O4 was instilled into the trachea. After magnetization from outside the chest wall, the remanent magnetic field (RMF) strength generated in the trachea was sequentially measured with a flux-gate magnetometer. The decay of RMF immediately after sequential magnetization showed a clearance of Fe3O4 particles. Mabuterol (10 micrograms/kg) increased clearance as much as isoproterenol (10 micrograms/kg) 30 min after intravenous injection relative to control measurement (p less than 0.01). These results suggest that mabuterol is useful to promote clearance.

Series distribution of mucociliary clearance of magnetized iron particles in anesthetized dogs.

Clearance of magnetic iron particles placed in the trachea, peripheral airway and alveoli was studied in anesthetized dogs. A retrograde catheter was inserted into a peripheral airway 2 mm in diameter. A thin vinyl tube was inserted into the peripheral airway from the chest wall through the retrograde catheter and 0.05 ml of saline containing 30 mg of iron oxide (Fe3O4) was instilled into either the peripheral airway or the trachea. Further, 30 mg of Fe3O4 solution was instilled with an injection needle into the alveoli. After magnetization from outside the chest wall, remanent magnetic fields (RMF) generated in the trachea, peripheral airway and alveoli were sequentially measured with a flux-gate magnetometer. The decay of RMF immediately after sequential magnetization shows clearance of Fe3O4 particles. Relaxation of RMF just before sequential magnetization is related to random misalignment of the particles. The fastest clearance was observed in the trachea, followed by the peripheral airway and the slowest was that in the alveoli. Exposure to isoproterenol resulted in increased clearance in the trachea and peripheral airway. However, relaxation was of similar magnitude and was not altered by isoproterenol throughout the entire airway. These results suggest that mechanisms responsible for clearance and relaxation in vivo are different and that clearance accelerates toward the central airway from alveoli and relaxation may be due to properties of the airway surface lining layer.

Opsonized zymosan decreases cytoplasmic motility of alveolar macrophages in dogs

To examine the mechanisms of changes in alveolar macrophage (AM) activities caused by phagocytic stimulus, we studied the effect of opsonized zymosan (OZ) on cytoplasmic motility (CM) of AM from dog lungs in vitro. Four days after the instillation of ferrimagnetic particles (Fe3O4, 3 mg/kg) into the lower lobe bronchus, AM were harvested by broncho-alveolar lavage. AM were adhered to the bottom of plastic vials (10(6) cells of AM per each vial). Remanent field strength (RFS) from the AM containing Fe3O4 particles was measured immediately after magnetization. RFS decreased with time due to particle rotation (relaxation), which is related to cytoplasmic motility of AM. OZ (1-500 micrograms) decreased lambda 0 (the relaxation rate for the first min) in a concentration-dependent fashion. Neither BW755C (10(-5) M), indomethacin (10(-6) M), leupeptin (10(-5) M), bestatin (10(-5) M), nor superoxide dismutase (1000 U/ml) inhibited OZ (500 micrograms)-induced inhibitory effects on lambda 0, suggesting that cyclooxygenase and lipoxygenase products, serine, thiol enzymes, aminopeptidase and superoxide anion wer not responsible for OZ-induced effects. OZ (500 micrograms) significantly increased the intracellular concentration of Ca2+ (P less than 0.01). Likewise, OZ (500 micrograms)-induced effects on lambda 0 of AM were significantly inhibited by replacement of the medium with a Ca2+ free solution (P less than 0.01). These results imply that opsonized zymosan inhibits cytoplasmic motility of AM via external calcium influx.

INHALATION OF ROAD DUST BY RESIDENTS IN POLLUTED AREAS

From March 1985 to March 1989, pneumomagnetic field strength (PMFS) was measured in 579 healthy subjects who lived in areas where there was substantial road dust pollution. In response to the governments campaign to eliminate the use of studded tires, suspended road dust produced by studded tires during the snowy season in the downtown areas of Sendai, Japan, decreased from 191 micrograms/m3 in March 1985 to 116 micrograms/m3 in March 1989. Suspended road dust in nonpolluted areas varied from 11 to 15 micrograms/m3. Road dust retained in the lungs, which contained 3% iron, was magnetized from the surface of the chest wall, and the PMFS was measured. The proportion of subjects with an abnormally high initial PMFS at the first measurement was 7 to 15% from 1985 to 1989; however, the PMFS of subjects who had an initially high PMFS decreased during each succeeding year. These findings suggest that, despite a government campaign to eliminate studded tires, road dust pollution is still being inhaled by the residents.