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Cells in the kidney medulla are subject to variable and often extreme osmotic stress during concentration of the urine. Previous studies showed that renal inner medullary epithelial (IME) cells respond to hypertonicity by G(2) arrest. The purpose of the present study was to investigate the mechanisms involved in initiation and maintenance of G(2) arrest. Rapid initiation of G(2) arrest after UV radiation is mediated by p38 kinase. Here we find that p38 kinase is responsible for rapid initiation of the G(2) delay in IME cells after the hypertonic stress created by adding NaCl. High NaCl, but not high urea, rapidly initiates G(2) arrest. Inhibition of p38 kinase by SB202190 (10 microM) blocks the rapid initiation of this checkpoint both in an immortalized cell line (mIMCD3) and in second-passage IME cells from mouse renal inner medulla. p38 inhibition does not affect exit from G(2) arrest. The rapid initiation of G(2) arrest is followed by inhibition of cdc2 kinase, which is also prevented by SB202190. To assess the possible protective role of G(2) arrest, we measured DNA strand breaks as reflected by immunostaining against phospho-histone H2AX, which becomes phosphorylated on Ser-139 associated with DNA breaks. Abrogation of rapid G(2)/M checkpoint activation by SB202190 increases the histone H2AX phosphorylation in G(2)/M cells. We propose that the rapid initiation of G(2) delay by p38 kinase after hypertonicity protects the cells by decreasing the level of DNA breaks caused by aberrant mitosis entry.

proceedings of the national academy of sciences of the united states of america

Rapid activation of G <sub>2</sub> /M checkpoint after hypertonic stress in renal inner medullary epithelial (IME) cells is protective and requires p38 kinase

Rapid activation of G 2 /M checkpoint after hypertonic stress in renal inner medullary epithelial (IME) cells is protective and requires p38 kinase

Rapid activation of G ₂ /M checkpoint after hypertonic stress in renal inner medullary epithelial (IME) cells is protective and requires p38 kinase