ERYTHROCYTE SENESCENCE

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The mechanism responsible for erythrocyte senescence remains an unsolved problem. Even though the study of genetically determined hemolytic anemias has demonstrated a variety of defects which can shorten the erythrocyte life span, no clear cut answer has emerged as to the normal mechanisms which determine erythrocyte survival. The examination of this aging process has been hindered by the technical difficulty of isolating aged cells. Historically a variety of physical techniques have been utilized for obtaining these senescent erythrocytes but most are not satisfactory. This proposal employs a procedure for the isolation of senescent erythrocytes from the rabbit in which the cells are covalently labeled with biotin by reaction with N-hydroxysuccinimido biotin. The bound biotin has been shown to not affect in vivo survival when the derivatized cells are re-infused into the rabbit. At various times after derivatization and in vivo aging, the biotinylated erythrocytes can be selectively recovered by their affinity for an avidin support. With this procedure, erythrocytes are routinely isolated within 5 to 10 days of their expected death. This proposal will examine the involvement of lipid peroxidation in the red cell aging process by quantitating the level of malondialdehyde-protein adducts in senescent cells, by quantitating the level of lipid hydroperoxides as a function of red cell age and by determining the amount of heme and non-heme iron associated with the senescent cell membrane. Additionally, the state of several membrane components in aged red cell will be evaluated including the levels of sialic acid, the integrity of band 3 and the level of complement components on the cell surface. Also, the mechanism responsible for the age-dependent loss of adenosine 5'-monophosphate deaminase activity in the red cell will be examined. A careful examination of these parameters will facilitate evaluation of specific hypotheses related to the red cell aging process and result in a better understanding of those factors which control the normal senescence of erythrocytes.