Potent and specific inhibition of the biological activity of the type-II transmembrane serine protease matriptase by the cyclic microprotein MCoTI-II.

Matriptase is a type-II transmembrane serine protease involved in epithelial homeostasis in both health and disease, and is implicated in the development and progression of a variety of cancers. Matriptase mediates its biological effects both via as yet undefined substrates and pathways, and also by proteolytic cleavage of a variety of well-defined protein substrates, several of which it shares with the closely-related protease hepsin. Development of targeted therapeutic strategies will require discrimination between these proteases. Here we have investigated cyclic microproteins of the squash Momordica cochinchinensis trypsin-inhibitor family (generated by total chemical synthesis) and found MCoTI-II to be a high-affinity (Ki 9 nM) and highly selective (> 1,000-fold) inhibitor of matriptase. MCoTI-II efficiently inhibited the proteolytic activation of pro-hepatocyte growth factor (HGF) by matriptase but not by hepsin, in both purified and cell-based systems, and inhibited HGF-dependent cell scattering. MCoTI-II also selectively inhibited the invasion of matriptase-expressing prostate cancer cells. Using a model of epithelial cell tight junction assembly, we also found that MCoTI-II could effectively inhibit the re-establishment of tight junctions and epithelial barrier function in MDCK-I cells after disruption, consistent with the role of matriptase in regulating epithelial integrity. Surprisingly, MCoTI-II was unable to inhibit matriptase-dependent proteolytic activation of prostasin, a GPI-anchored serine protease also implicated in epithelial homeostasis. These observations suggest that the unusually high selectivity afforded by MCoTI-II and its biological effectiveness might represent a useful starting point for the development of therapeutic inhibitors, and further highlight the role of matriptase in epithelial maintenance.

A multidrug efflux phenotype mutant of Streptococcus pyogenes.

We describe a mutant of Streptococcus pyogenes NCTC 8198 with a multidrug efflux phenotype. A mutant selected with ethidium bromide showed a four-fold rise in MIC of norfloxacin, a 16-fold rise in MIC of ethidium bromide and an eight-fold rise in MIC of acriflavine when compared with the parent strain. The MICs were unaffected by the efflux pump inhibitors reserpine, rescinnamine and verapamil. The mutant's ethidium bromide MIC was reduced two-fold by norfloxacin. Ethidium bromide accumulation after 10 min was 58% lower in the mutant compared with the parent. This difference was not affected by carbonyl cyanide m-chlorophenylhydrazone.

Magnesium sulfate in exacerbations of chronic obstructive pulmonary disease.

BACKGROUND: Acute exacerbations of chronic obstructive pulmonary disease are commonly seen and difficult to treat. We sought to determine the bronchodilator efficacy of magnesium sulfate in this situation, as this compound is helpful in acute asthma. METHODS: Subjects who came to either of two Veterans Affairs emergency departments were randomized in a double-blind fashion to receive either 1.2 g of magnesium sulfate or placebo over 20 minutes after they first received albuterol, 2.5 mg by nebulization. Peak expiratory flow, dyspnea scores, arterial hemoglobin oxygen saturation by pulse oximetry, maximal inspiratory and expiratory pressures, and vital signs were monitored for 45 minutes after the start of magnesium sulfate or placebo treatment. RESULTS: Seventy-two individuals were studied. The peak expiratory flow increased 16.6% +/- 27.7% (mean +/- SD) in both groups after the initial albuterol treatment, from 121.2 +/- 55.7 L/min to 136.9 +/- 63.9 L/min. The peak expiratory flow increased from 136.7 +/- 69.7 L/min at the start of the infusion to 162.3 +/- 76.6 L/min at 30 minutes and 161.3 +/- 78.7 L/min at 45 minutes with magnesium sulfate treatment. The peak expiratory flow was 137.0 +/- 58.6 L/min on initiation of the intravenous infusion, 143.0 +/- 72.7 L/min at 30 minutes, and 143.3 +/- 70.5 L/min at 45 minutes in the placebo group. The difference in peak expiratory flow from initiation of the infusion to 30 and 45 minutes later (calculated as means of the 30- and 45-minute values) was significantly different for the two groups (25.1 +/- 35.7 L/min vs 7.4 +/- 33.3 L/min; P = 0.3); the difference was also significant when expressed as percentage increase (22.4% +/- 28.5% vs 6.1% +/- 24.4%; P = .01). There was a statistically nonsignificantly trend toward a reduced need for hospitalization in the magnesium sulfate group as compared with the placebo group (28.1% vs 41.9%; P = .25). There were no significant changes in the other parameters with either treatment. CONCLUSION: Magnesium sulfate, 1.2 g over 20 minutes after beta-agonist administration, is safe and modestly efficacious in the treatment of acute exacerbations of chronic obstructive pulmonary disease, and its bronchodilator effect is greater than that of a beta-agonist given alone and lasts beyond the period of magnesium sulfate administration.