ACQUIRED-RESISTANCE OF A MAMMALIAN-CELL LINE TO HYPOXIA-REOXYGENATIONTHROUGH COTRANSFECTION OF KIR6.2 AND SUR1 CLONES

Reoxygenation after transient hypoxia is a common clinical condition t hat often causes greater tissue damage than persistent hypoxia itself. This warrants the development of a means to protect cells against hyp oxia-reoxygenation injury. Adenosine triphosphate (ATP)-sensitive K+ ( K-ATP,) channels have been proposed to play an essential role in the m echanisms of endogenous cellular protection. Thus far, however, K-ATP channel proteins have not been exploited to generate an injury-resista nt cellular phenotype by delivering K-ATP, channel genes into injury-p rone cells. A first step in this direction is the evaluation of the ou tcome of transferring genes encoding K-ATP, channels into a K-ATP chan nel-deficient cell type exposed to metabolic stress. Untransfected COS -7 monkey kidney cells, which natively lack K-ATP channels, were found to be vulnerable to hypoxia-reoxygenation injury, which induced cytos olic Ca2+ loading, as measured by digital epifluorescent imaging. COS- 7 cells cotransfected with K-ATP, channel genes, Kir6.2 and SUR1, gain ed resistance to hypoxia-reoxygenation. This acquired resistance was a bolished by glyburide, the K-ATP channel antagonist. We have previousl y shown that Kir6.2 and SUR1 physically associate to form a functional K-ATP channel, not reconstituted by either of the subunits alone. Tra nsfection with individual channel subunits, Kir6.2 or SUR1, failed to produce resistance to hypoxia-reoxygenation induced Ca2+ loading. This is a first demonstration that transfer of K-ATP channel subunits can generate an injury-resistant cellular phenotype. The findings from thi s study may, thus, provide a framework for future therapeutic strategi es based on gene delivery of K-ATP channel subunits in cells and tissu es vulnerable to hypoxia-reoxygenation insults.