Donor heart valves: electron microscopic and morphometric assessment of cellular injury induced by warm ischemia

Cryopreserved allograft valves are increasingly being used as valvular replacements. Leaflet fibroblast viability has been suggested to influence clinical durability. The warm ischemic time is thought to be a critical determinant of this cell viability. The purpose of this study was to apply quantitative morphometric methods to characterize, by transmission electron microscopy, valvular cellular injury resulting from progressive warm ischemic time. Porcine aortic valves were harvested with a spectrum of warm ischemic times (40 minutes and 2, 6, 12, 24, and 36 hours; five valves per warm ischemic time; n = 30) and processed by standard electron microscopic methods. To ensure randomized tissue selection within each warm ischemic time interval, we randomly selected one thin section from each leaflet. The first ten cells in each thin section were photographed and cellular injury was assessed (cell disruption, dilation of endoplasmic reticulum, cytoplasmic edema, nuclear and mitochondrial changes). Nine hundred micrographs have been analyzed by Cochran-Mantel-Haenszel statistics to determine if a significant association between warm ischemic time and cellular injury exists. Our findings indicate a significant association between reversible cell injury through 24 hours of warm ischemic injury (p less than 0.0001). Furthermore, a significant association between irreversible cell injury and progressive warm ischemia through 36 hours was also found. These findings indicate that the ischemic interval after donor death is associated with progressive leaflet cell injury. Cellular damage begins shortly after donor death and continues incrementally throughout 36 hours. After 2 hours of warm ischemic injury 37% of the cells had morphologic evidence of injury. After 6 hours of warm ischemic injury the number of injured cells increased to 73%. By 36 hours 22% of the cells appeared normal. Irreversible cell injury increases with prolonged ischemia and becomes quantitatively impressive at 24 hours, by which time 26% of cells are so affected. Conversely, some cells are resistant to irreversible injury for a prolonged ischemic interval.