Sublobular Distribution Of Cytochromes In Cold-Stored Rat Liver

Jeffrey J. Kelly, Katherine A. Kelly and Clyde H. Barlow

Department of Chemistry, The Evergreen State College, Olympia, WA 98505
Barlow Scientific, Inc., Olympia, WA 98502

J.J. Kelly, K.A. Kelly and C.H. Barlow  "Sublobular distribution of cytochromes in cold-stored rat liver".  25th Annual Conference of the International Society on Oxygen Transport to Tissue.  Milwaukee, Wisconsin.  August, 1997.  This study examines the distribution of mitochondrial cytochromes between pericentral and periportal regions of rat liver by high resolution imaging. A comparison is made between images of oxidized and reduced mitochondrial cytochrome a and cytochrome c in coupled and uncoupled (dinitrophenol) cold-stored liver during transitions in oxygen delivery. The effect of acetaminophen-induced hepatotoxicity on cytochrome redox states was also examined.

Spectroscopic images at 18x18-micron resolution of cytochrome reflectance (-log R) from 4.5x4.5-mm regions of the surface of cold (4C) livers were obtained through bandpass filters using a cooled CCD camera. The gas over the cold-stored liver was varied from between air, 100% nitrogen and 100% oxygen.

Cytochrome a difference images for air to nitrogen transitions produced reflectance changes that were 63% as large as those for oxygen to nitrogen transitions. Cytochrome c measurements taken for the same transitions were 94% as large. From these differences we concluded that the longer wavelengths of light used to measure cytochrome a penetrate and are backscattered from a tissue depth that is deeper than that reached by oxygen diffusion furnished by air, while the shorter wavelengths used to measure cytochrome c penetrated to about the same depths as oxygen diffusion from air.

Between lobules in healthy livers exposed to first oxygen and then nitrogen, the mean increase in cytochrome a reflectance was 0.043 with pericentral increase of 0.039 and periportal of 0.047. For cyctochrome c the means increase in reflectance was 0.037 and pericentral and periportal changes were 0.036 and 0.039 respectively. These results are consistent with mitochondria being more highly concentrated in periportal regions compared to pericentral regions of liver lobules. Periportal to pericentral differences are more pronounced from cytochrome a as compared to cytochrome c in uncoupled livers as well as coupled livers, but a marked difference exists between the behavior of cytochrome c in coupled and uncoupled livers. In the coupled liver the cytochrome c reflectance change is 1/2 that of cytochrome a while in the uncoupled liver they are of the same magnitude.

An explanation can be found in the functioning of the mitochondrial electron transport chain. Cytochrome c is separated from the site of oxygen interaction with the chain at cytochrome a by phosphorylation site III. In coupled livers cytochrome a reacts directly with oxygen while cytochrome c oxidation is dependent upon and limited by low ADP levels found in livers stored at 4C. In uncoupled liver the phosphorylation limitation is removed and cytochrome c is readily oxidized by electron transport to oxygen via cyctochrome a resulting in larger reflectance changes during reduction.

Research was supported by grant DK46756 from the National Institutes of Health.

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