Effect of Pseudomonas aeruginosa rhamnolipids on mucociliary transport and ciliary beating.

Pseudomonas aeruginosa rhamnolipid causes ciliostasis and cell membrane damage to rabbit tissue, is a secretagogue in cats, and inhibits epithelial ion transport in sheep tissue. It could therefore perturb mucociliary clearance. We have investigated the effect of rhamnolipid on mucociliary transport in the anesthetized guinea pig and guinea pig and human respiratory epithelium in vitro. Application of rhamnolipid to the guinea pig tracheal mucosa reduced tracheal mucus velocity (TMV) in vivo in a dose-dependent manner: a 10-microgram bolus caused cessation of TMV without recovery; a 5-micrograms bolus reduced TMV over a period of 2 h by 22.6% (P = 0.037); a 2.5-microgram bolus caused no overall changes in TMV. The ultrastructure of guinea pig tracheal epithelium exposed to 10 micrograms of rhamnolipid in vivo was normal. Application of 1,000 micrograms/ml rhamnolipid had no effect on the ciliary beat frequency (CBF) of guinea pig tracheal rings in vitro after 30 min, but 250 micrograms/ml stopped ciliary beating after 3 h. Treatment with 100 micrograms/ml rhamnolipid caused immediate slowing of the CBF (P less than 0.01) of human nasal brushings (n = 7), which was maintained for 4 h. Mono- and dirhamnolipid had equivalent effects. The CBF of human nasal turbinate organ culture was also slowed by 100 micrograms/ml rhamnolipid, but only after 4 h (CBF test, 9.87 +/- 0.41 Hz; control, 11.48 +/- 0.27 Hz; P less than 0.05, n = 6), and there was subsequent recovery by 14 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Sulfated O-glycoproteins secreted by guinea pig trachea in organ culture.

Organ culture of guinea pig trachea was performed in the presence of [35S]sulfate in order to characterize the sulfated glycoproteins released from the respiratory epithelium and mucosa. The sulfated macromolecules that were synthesized during a 6-h incorporation were separated by CsBr density-gradient centrifugation and gel-filtration chromatography successively. Most of the sulfated secreted macromolecules corresponded to a population of glycoproteins sensitive to reductive beta-elimination but resistant to both chondroitinase ABC and heparinase. These glycoproteins had different buoyant densities (ranging from 1.48 g/ml to 1.16 g/ml) and could be subfractionated according to molecular mass. A major part of the radioactivity was incorporated into high-molecular-mass mucins that were excluded from a Sepharose CL-2B column and did not penetrate into polyacrylamide gel in PAGE. However, a mixture of sulfated O-glycoproteins of much lower molecular mass was also characterized in addition to low amounts of chondroitin sulfate. Epithelial goblet cells are the predominant mucin-containing cells of the respiratory guinea pig trachea. Our results suggest that a wide range of sulfated O-glycoproteins are secreted by the guinea pig tracheal mucosa.

Effect of pyocyanin and 1-hydroxyphenazine on in vivo tracheal mucus velocity.

Products of the bacterium Pseudomonas aeruginosa have been shown to slow the beating of human respiratory tract cilia in vitro. We have tested the effects of two of these compounds, pyocyanin and 1-hydroxyphenazine (given as a bolus dose dissolved in 2 microliters Ringer solution), on tracheal mucus velocity of radiolabeled erythrocytes in anesthetized guinea pigs. 1-Hydroxyphenazine (200 ng) caused a rapid slowing of tracheal mucus velocity (maximum fall 47% at 20 min) with recovery by 1 h. The effect of pyocyanin was slower in onset, 600 ng causing 60% reduction in tracheal mucus velocity at 3 h, and no recovery occurred. A combination of pyocyanin and 1-hydroxyphenazine produced an initial rapid slowing equivalent to the same dose of 1-hydroxyphenazine given alone, but the later slowing attributed to pyocyanin was greater than the same dose administered alone. This study demonstrates one mechanism by which products of P. aeruginosa may facilitate its colonization of the respiratory tract.

The effect of acetylsalicylate on aggregation and arachidonate metabolism by human platelets suspended in plasma or buffer.

Acetylsalicylate inhibits prostaglandin and thromboxane production by human platelets suspended in plasma or buffer. Acetylsalicylate inhibits arachidonate-induced aggregation of human platelets suspended in plasma, but the effect of acetylsalicylate on arachidonate-induced aggregation of human washed platelets in buffer has not been reported. We have therefore studied this in relation to arachidonate metabolism in human platelets suspended in plasma or buffer. Platelets suspended in plasma and in buffer were both prepared from each donor, who had not taken acetylsalicylate or like-acting drugs for at least 7 days. Acetylsalicylate was 1500 times less potent in inhibiting arachidonate-induced aggregation in buffer (IC50 = 27.3 +/- 7.5 (s.e.m.)mM) than it was in plasma (IC50 = 18.3 +/- 6.0 microM); whereas it was only 4 times less potent in inhibiting thromboxane production in buffer (IC50 = 110 +/- 51.0 microM) than in plasma (IC50 = 25.3 +/- 8.9 microM). The acetylsalicylate concentration required to inhibit aggregation in buffer was sufficient to inhibit 12-hydroxyeicosatetraenoic acid production whereas the concentration that inhibited thromboxane production in buffer was not. These results indicate that arachidonate-induced aggregation of platelets in buffer may depend on product(s) of lipoxygenase rather than of cyclooxygenase, and is hence insensitive to inhibition by acetylsalicylate compared with arachidonate-induced aggregation of platelets in plasma.

Comparisons between the abilities of various human and ovine plasmas to inhibit prostaglandin synthesis.

The ability of various human and ovine blood plasmas to inhibit prostaglandin synthesis in vitro has been tested. Human plasmas were significantly more potent in their ability to inhibit prostaglandin synthesis than their counterpart ovine plasmas. In general, female plasma had greater inhibitory activities than male plasmas and adult plasmas were more active than fetal plasmas. There was no simple correlation between the activity of plasmas as inhibitors of prostaglandin synthesis and their respective albumin or haptoglobin contents.

Prostaglandin synthesis inhibition by maternal and fetal sheep plasma and its relation to haptoglobin and albumin levels.

Mammalian plasmas and sera have been reported to contain endogenous inhibitors of prostaglandin synthesis (EIPS), but the identity and role of these suggested inhibitors is as yet undetermined. Albumin and haptoglobin have been proposed as possible inhibitors, and it has been suggested that EPIS may have a part to play in the control of PG production during pregnancy and in the neonatal period. As part of a series of studies aimed at elucidating the identity and role of EIPS, maternal and fetal blood samples were collected from chronically catheterized pregnant ewes, and plasma levels of albumin, haptoglobin and EIPS activity determined. Pregnant ewe plasma possessed high EIPS activity and fetal lamb plasma little or no EIPS activity. Levels of albumin and/or haptoglobin did not consistently parallel that of EIPS activity. A post-operative rise (4 sheep studied) and a pre-parturition nadir (1 sheep studied) in maternal plasma EIPS activity were also noted. The possible physiological significance of these results is discussed.

Paracetamol potentiates acetylsalicylate in inhibiting prostaglandin synthesis.

Lower concentrations of paracetamol stimulated, but a higher concentration inhibited prostaglandin synthesis by bull seminal vesicle homogenate in the absence of added co-factors. Admixed with acetylsalicylate or indomethacin, paracetamol strongly potentiated the inhibition of prostaglandin synthesis in vesicle homogenate, and weakly potentiated inhibition in rat gastric fundus strip. We propose that, by acting as a phenolic co-factor, paracetamol stimulates prostaglandin synthesis and thus renders the cyclo-oxygenase more vulnerable to acetylsalicylate or indomethacin.

Endogenous inhibitor of prostaglandin synthetase.

Mammalian serum and plasma contain an endogenous inhibitor of prostaglandin synthetase (EIPS). Human plasma fractions rich in EIPS show anti-inflammatory activity in vivo. In rats, glucocorticoids raise EIPS activity of plasma and serum. These findings suggest the existence of a natural mechanism of controlling prostaglandin synthesis, possibly related to corticosteroid action.

The local antinociceptive and topical anti-inflammatory effects of propyl gallate in rodents.

1 In common with several anti-inflammatory, analgesic, local anaesthetic and antioxidant drugs, propyl gallate in vitro inhibited the biosynthesis of prostaglandin E2 and F2alpha from arachidonic acid by a prostaglandin synthetase from bull seminal vesicles. 2 In common with analgesic drugs, propyl gallate reduced the ability of arachidonic acid, acetylcholine or acetic acid to cause abdominal constriction in mice. 3 Using a new method for evaluating anti-inflammatory activity, we demonstrated the effectiveness of aspirin or indomethacin given subcuteneously before u.v. irradiation of guinea-pig ears, the prophylactic action of topically applied sunscreen agents and the therapeutic value of bufexamac and propyl gallate applied after irradiation.

Stimulation of prostaglandin biosynthesis by drugs: effects in vitro of some drugs affecting gut function.

Low concentrations of several emetic, purgative or irritant drugs in the absence of added co-factors stimulated conversion of arachidonic acid to prostaglandin E2 and F2alpha by prostaglandin synthetase extracted from bull seminal vesicles (BSV prostaglandin synthetase). Their effect was dependent on concentration and time. Stimulation of BSV prostaglandin synthetase by apomorphine, aloes, tyramine or zingerone was increased several-fold by addition of reduced glutathione to the incubation medium, whereas hydroquinone, a phenolic co-factor of prostaglandin synthetase caused slight depression. From this finding and from the observation that many of the stimulant drugs possess a phenolic group, whereas their inactive relatives lack such a group, it is suggested that these stimulant drugs act as co-factors for prostaglandin synthetase in place of hydroquinone. Aloes, tyramine, ethanol and quipazine also produced a dose-related increase in resting tone of the isolated fundus of the rat stomach. This increase occurred at concentrations comparable to those effective in stimulating BSV prostaglandin synthetase, and was abolished by acetylsalicylate. These findings support the view that certain drugs exert some of their pharmacological effects by stimulating prostaglandin synthetase.

Inhibition of prostaglandin biosynthesis by sulphasalazine and its metabolites.

Sulphasalazine (SZ) inhibits prostaglandin (PG) biosynthesis in vitro with a potency comparable to that of aceylsalicylate. The metabolites of SZ, sulphapyridine and 5-aminosalicylic acid, were of considerably lower potency as inhibitors of PG biosynthesis in the synthetase preparations used. Th inhibition of prostaglandin production by SZ could at least partly account for the clinical utility of sulphasalazine in ulcerative colitis. Sulphapyridine may help to maintain inhibitory concentrations of SZ by restraining bacterial breakdown of the active drug.