Study Of Aging Mechanism Of Red Blood Cell And Carbonyl Stress

Reactive carbonyl species (RCS) is the common reactive intermediate products of lipid peroxidation and non-enzymatic glycosylation (glycation), which are two most important energy metabolism-related biological side reactions. With high biological reactivity, the RCS can react readily with the thiol and amino group of various biological macromolecules, resulting in cross-linking of biological macromolecules. In blood, these RCS can attack many biological macromolecules, such as attacking erythrocyte membrane protein and causing the structural and functional changes. Therefore, it has been proposed that the accumulation of unpairable damage of macromolecules caused by RCS should be an important biochemical mechanism of biological aging.The aim of this thesis was to explore the relationship between carbonyl stress and aging of erythrocyte with an in vitro erythrocyte rapid aging model of storage blood and an in vivo erythrocyte aging model of different age subjects.Similar to in vivo hemorheological changes of aging process, the blood viscosity of storage blood increased gradually with the extended storage time. However, the detailed biochemical mechanism remains unclear. In order to reveal the mechanism, we determined the alteration of RCS, thiols, carbonylation levels and fluorescence intensity of erythrocyte membrane proteins. Results showed that there were significant increases in blood viscosity and plasma RCS level during storage, significant time-dependent increases in membrane protein carbonylation and protein (tryptophan) fluorescence, and a decrease in the content of thiols. These findings indicated that RCS, by attacking the amino and/or sulfhydryl groups of erythrocyte membrane proteins, induce a series of structural alterations in erythrocytes. These lead to an increase in blood viscosity and reduce the efficacy and safety of storage.To further confirm above results and explore the aging mechanisms of erythrocyte, fresh blood from different age donors were analyzed. Firstly, RCS, reduced thiols and erythrocyte membrane protein carbonylation levels were measured. In accord with above results:there were age-dependent increases in plasma RCS level and membrane protein carbonylation and a decrease in the content of reduced thiols. Secondly, we investigated the relationship of protein carbonylation level of human erythrocyte membrane and mean corpuscular hemoglobin concentration, mean corpuscular hemoglobin content and the major plasma lipid metabolism-related materials. The results showed that protein carbonylation level of erythrocyte membrane has no relevance to the concentration and content of mean corpuscular hemoglobin; however, it was negatively correlated with the content of high-density lipoprotein whereas positively correlated with levels of triglyceride, total cholesterol and total cholesterol/HDL. These results indicated that protein carbonylation level of human erythrocyte membrane was positively correlated with blood lipid content while irrelevant with erythrocyte protein content which indirectly proves the positive correlation of erythrocyte membrane protein carbonylation with RCS. The above results showed that the level of RCS in blood increased with age, they would attack the thiol and amino group of various biomacromolecules, including erythrocyte membrane proteins, and result in changes in their structure and biological function. That may be an important mechanism of aging in erythrocyte.