Nitric oxide and zinc homeostasis in acute lung injury

Abstract

Among putative small molecules that affect sensitivity to acute lung injury, zinc and nitric oxide are potentially unique by virtue of their interdependence and dual capacities to be cytoprotective or injurious. Nitric oxide and zinc appear to be linked via an intracellular signaling pathway involving S-nitrosation of metallothoinein--itself a small protein known to be an important inducible gene product that may modify lung injury. In the present article, we summarize recent efforts using genetic and fluorescence optical imaging techniques to: (1) demonstrate that S-nitrosation of metallothionein affects intracellular zinc homeostasis in intact pulmonary endothelial cells; and (2) reveal a protective role for this pathway in hyperoxic and LPS-induced injury.

Figure 1.

A schema of a fluorescence resonance energy transfer (FRET) capable reporter molecule (MT) and full spectral report of a single sheep pulmonary artery endothelial cell exposed to the NO donor, L-S-nitrosocysteine ethyl ester (L-SNCEE). In the upper panel, a schema is shown describing the use of a chimera containing cDNA for human MT-IIA sandwiched between enhanced cyan fluorescence protein (ECFP) and enhanced yellow fluorescence protein (EYFP) at N- and C-termini, respectively. Exposure to NO or a metal chelator (EDTA) removes zinc from FRET-MT and causes a conformational change in which the molecule unfolds and the reporter molecules move more distant from each other. In the lower panel, full spectral report is shown by visualizing a single cultured pulmonary artery endothelial cell that was infected with an adenovirus containing cDNA to FRET-MT construct at control and 10 min after exposure to the membrane-permeant NO donor, L-SNCEE. Exposure to L-SNCEE resulted in a decrease in the acceptor and an unquenching of the donor peak. Full spectral reporting was accomplished via the use of a Zeiss 510Meta (Jena, Germany) confocal microscope.