Vitronectin protects alveolar macrophages from silica toxicity.

Silicosis is an interstitial lung disease caused by the inhalation of crystalline silicon dioxide. Current concepts suggest that a crucial step in the development of silicosis is silica-induced injury of alveolar macrophages (AM). The adhesive protein vitronectin is a natural constituent of the lung, in which its function is largely unexplored. This study investigated a possible role for vitronectin in protecting AM from silica exposure. In this study, the concentration of vitronectin was shown to be increased in the bronchoalveolar lavage fluid of silica-treated rats. Vitronectin affinity for silica was shown both in vitro and in vivo by immunostaining. Vitronectin reduced silica-induced injury to cultured AM as determined with the (51)Cr release assay. Vitronectin reduced silica-induced free radical production as determined with a cell-free thiobarbituric acid assay. Additionally, vitronectin reduced the silica-induced respiratory burst in AM as determined with chemiluminescence. This study suggests that vitronectin may protect AM during the initial exposure to silica.

Pneumocystis carinii attachment increases expression of fibronectin-binding integrins on cultured lung cells.

Pneumocystis carinii is an extracellular pathogen which requires attachment to alveolar epithelial cells for growth and replication. Previous studies have demonstrated that the extracellular matrix protein fibronectin (Fn) facilitates attachment of P. carinii to lung cells. This study addresses the role of cell surface Fn receptors (integrins) as mediators of P. carinii attachment and demonstrates the effect of P. carinii attachment on integrin expression on cultured lung cells. To determine the role of Fn-binding integrins in P. carinii attachment, attachment of 51Cr-labelled P. carinii organisms to the lung epithelial cell line A549 was quantified in the presence or absence of anti-integrin antibodies. Antibodies to the alpha v and alpha 5 integrin subunits significantly inhibited P. carinii attachment, while addition of antibody to the alpha subunit of a non-Fn-binding integrin, alpha 2, did not affect P. carinii attachment. To further investigate the role of Fn-binding integrins in P. carinii attachment, the effect of P. carinii attachment on expression of the alpha v and alpha 5 integrin subunits was determined. A549 cells incubated with either P. carinii organisms or with the P. carinii major surface glycoprotein gp120 demonstrated a 3- to 10-fold increase in expression of the alpha 5 integrin subunit; however, neither P. carinii nor gp120 affected the expression of alpha v integrin. Furthermore, the effects of P. carinii on A549 cell alpha 5 integrin expression were attenuated by the addition of an anti-gp120 antibody which blocks P. carinii attachment to A549 cells. Therefore, P. carinii attachment to lung epithelial cells appears to be mediated by alpha v- and alpha 5-containing integrins expressed on the epithelial cell surface, and P. carinii attachment results in increased expression of the alpha 5 integrin subunit.

Fast and sensitive laser-based enzymatic detection of the lactose operon in microorganisms.

This work investigates the ability of laser-based, time-resolved fluorometry to detect the lactose operon genetic marker in microorganisms and to study protein-DNA interactions. In the first study, rapid detection of the Escherichia coli lacZY operon inserted in two strains of Pseudomonas proposed as fungal biological control organisms was achieved. Optimization of incubation time, immobilization apparatus size, and reagent volumes, along with the laser-based instrumentation, yielded an assay capable of detecting 10(4) immobilized lac+ Pseudomonas fluorescens cells within a 30-min incubation time. In the second study, the synthesis of E. coli beta-galactosidase was monitored in "real-time" with observable enzymatic activity beginning 4 to 5 min after induction with isopropylthiogalactoside.