Poly (ADP-ribose) (PAR) is rapidly synthesized by PAR polymerases (PARPs) upon activation by DNA single- and double-strand breaks. In this study, we examined the quantitative amount of PAR in HeLa cells cultured within the physiological temperatures below 41 degrees C for verification of the effect of shifting up or -down the temperature from 37.0 degrees C on the DNA breaks, whether the temperature-shift caused breaks that could be monitored by the level of PAR. While PAR level did not change significantly when HeLa cells were cultured at 33.5 degrees C or 37.0 degrees C, it was significantly increased 2- and 3-fold when cells were cultured for 12 h and 24 h, respectively, at 40.5 degrees C as compared to 37.0 degrees C. Similar to the results with HeLa cells, PAR level was increased 2-fold in CHO-K1 cells cultured at 40.5 degrees C for 24 h as compared to 37.0 degrees C. As the cellular levels of PAR polymerasel (PARP1) and PAR glycohydrolase (PARG), a major degradation enzyme for PAR, did not seem to change significantly, this increase could be caused by activation of PARP1 by DNA strand breaks. In fact, gamma H2AX, claimed to be a marker of DNA double-strand breaks, was found in cell extracts of HeLa cells and CHO-Kl cells at elevated temperature vs. 37.0 degrees C, and these gamma H2AX signals were intensified in the presence of 3-aminobenzamide, a PARP inhibitor. The gamma H2AX immunohistochemistry results in HeLa cells were consistent with Western blot analyses. In HeLa cells, proliferation was significantly suppressed at 40.5 degrees C in 72 h-continuous cultures and decreased viabilities were also observed after 24-72 h at 40.5 degrees C. Flow cytometric analyses showed that the HeLa cells were arrested at G2/M after temperature shift-up to 40.5 degrees C. These physiological changes were potentiated in the presence of 3-aminobenzamide. Decrease in growth rates, increased cytotoxicity and G2/M arrest, were associated with the temperature-shift to 40.5 degrees C and are indirect evidence of DNA breaks. In addition to gamma H2AX, PAR could be a sensitive marker for DNA single- and double-strand breaks. These two molecular markers provide evidence of physiological changes occurring within cells. (C) 2016 Published by Elsevier Inc.