We previously found that hypertonicity (high NaCl) causes DNA damage and oxidative stress both in cell culture and in kidney medullas in vivo. DNA damage and oxidative stress are associated with cellular senescence, most striking in aging and in cancer. We also found that high salt causes cellular senescence in tissue culture and that age-associated accumulation of a senescent cells is accelerated in kidney medullas of normal mice, as well as in C. Elegans exposed to high salt. Thus, hyperosmolality not only causes DNA damage and oxidative stress, but also causes cellular senescence. Dehydration with aging is attributed to decreased urine concentrating ability and thirst. We further investigated by comparing urine concentration and water balance in 3, 18 and 27 months old mice, consuming equal amounts of water. During water restriction, 3 months mice concentrate their urine sufficiently to maintain water balance (stable weight). 18 months mice concentrate their urine as well, but still lose weight (negative water balance). 27 months mice do not concentrate their urine as well and lose even more weight than the 18 months mice, indicating a larger negative water balance. Negative water balance in older mice is accompanied by increased vasopressin excretion, providing further evidence of dehydration. All 3 groups maintain water balance while consuming only the water in gel food containing 56% water. However, both older groups excrete a smaller volume of urine of higher osmolality, indicating greater extra urinary water loss. Since their feces also contain less water, the excess water lost by the older mice apparently is through other routes, presumably insensible loss through the respiratory tract and skin. The greater insensible water loss occurs at an earlier age (18 months) than decreased urine concentrating ability (27 months). We propose that insensible water loss through skin and respiration increases with age, making a major contribution to aging related dehydration. Mre11 is a critical participant in upkeep of nuclear DNA, its repair, replication, meiosis and maintenance of telomeres. The upkeep of mitochondrial DNA (mtDNA) is less well characterized and whether Mre11 participates had been unknown. We previously found that high NaCl causes some of the Mre11 to leave the nucleus, but we did not then attempt to localize it within the cytoplasm. In the present studies we find Mre11 in mitochondria isolated from primary renal cells and show that the amount of Mre11 in mitochondria increases with elevation of extracellular NaCl. We additionally confirm the presence of Mre11 in the mitochondria of cells by confocal microscopy and show that some of the Mre11 colocalizes with mtDNA. Bleomycin, which is known to damage mtDNA, increases colocalization of Mre11 with mtDNA. Abundant Mre11 is also present in tissue sections from normal mouse kidneys, colocalized with mitochondria of proximal tubule and thick ascending limb cells. To explore whether distribution of Mre11 changes with cell differentiation we used an experimental model of tubule formation by culturing primary kidney cells in Matrigel matrix. In non-differentiated cells Mre11 is mostly in the nucleus, but it becomes mostly cytoplasmic upon cell differentiation. We conclude that Mre11 is present in mitochondria as well as in nuclei, and that the amount in mitochondria varies, depending on cellular stress and differentiation. Our results suggest a new role for Mre11 in the maintenance of genome integrity in mitochondria, in addition to previously known role in maintenance of nuclear DNA. High concentration of NaCl increases DNA breaks both in cell culture and in vivo. The breaks remain elevated as long as NaCl concentration remains high and are rapidly repaired when the concentration is lowered. The exact nature of the breaks, as well as their location, had not been entirely clear, nor had it been evident how cells survive, replicate and maintain genome integrity in environments like the renal inner medulla in which the additional breaks persist because the cells are constantly exposed to high NaCl concentration. Repair of the breaks after NaCl is reduced is accompanied by formation of foci containing phosphorylated H2AX (gammaH2AX). This histone modification, which occurs around DNA double-strand breaks, contributes to their repair. We find (PNAS, in press)that gammaH2AX foci that occur during repair of high NaCl-induced DNA breaks are non-randomly distributed in the mouse genome. By chromatin immunoprecipitation using anti-gamma H2AX antibody, followed by massive parallel sequencing (ChIP-Seq), we find that during repair of double strand breaks induced by high NaCl, gammaH2AX is predominantly localized to regions of the genome devoid of genes (gene deserts), indicating that the high NaCl-induced double-strand breaks are located there. Localization to gene deserts helps explain why the DNA breaks are less harmful than are the random breaks induced by genotoxic agents such as UV radiation, ionizing radiation and oxidants. We propose that the universal presence of NaCl around animal cells has directly influenced the evolution of the structure of their genomes.
|Dmitrieva, Natalia I; Burg, Maurice B (2014) Secretion of von Willebrand factor by endothelial cells links sodium to hypercoagulability and thrombosis. Proc Natl Acad Sci U S A 111:6485-90|
|Zhou, Xiaoming; Burg, Maurice B; Ferraris, Joan D (2012) Water restriction increases renal inner medullary manganese superoxide dismutase (MnSOD). Am J Physiol Renal Physiol 303:F674-80|
|Dmitrieva, Natalia I; Malide, Daniela; Burg, Maurice B (2011) Mre11 is expressed in mammalian mitochondria where it binds to mitochondrial DNA. Am J Physiol Regul Integr Comp Physiol 301:R632-40|
|Dmitrieva, Natalia I; Burg, Maurice B (2011) Increased insensible water loss contributes to aging related dehydration. PLoS One 6:e20691|
|Dmitrieva, Natalia I; Cui, Kairong; Kitchaev, Daniil A et al. (2011) DNA double-strand breaks induced by high NaCl occur predominantly in gene deserts. Proc Natl Acad Sci U S A 108:20796-801|