TY - JOUR
T1 - Cytotoxic manifestations of the interaction between hyperthermia and TNF
T2 - DNA fragmentation
AU - Tomasovic, S. P.
AU - Vasey, T. A.
AU - Story, M. D.
AU - Stephens, L. Clifton
AU - Klostergaard, J.
N1 - Funding Information:
This investigation was supported in part by grant CA-47786 (S.P.T./J.K.) from the US Public Health Service.
PY - 1994
Y1 - 1994
N2 - The relationship of DNA fragmentation to the greatly enhanced cytotoxicity seen in vitro against tumour cells when recombinant human tumour necrosis factor-α (TNF-α is combined with hyperthermia was investigated. The TNF-αsensitive L929 and -resistant EMT6 cells were treated with 8.8 and 16 ng of TNF-α per ml, respectively, and then heated at 40.5°C for 24 h (L929) or at 43°C for 1 h (L929) or 1.5 h (EMT-6) beginning 1 h later. For both cell lines at both temperatures, the addition of heating to the TNF-α treatment significantly decreased viability and increased DNA fragmentation at earlier time points than seen with either TNF-α or heat alone. DNA fragmentation was further studied using agarose gel electrophoresis to examine the size distribution of the DNA fragments and the ability of intracellular calcium buffering agents BAPTA and quin-2 to inhibit fragmentation. At 4.5 h after L929 cells were treated with TNF-α at 43°C, the size distribution of DNA fragments more closely resembled the oligonucleosome sized apoptotic DNA fragmentation, as seen in irradiated rat thymocytes, than the spectrum of DNA fragments seen in necrotic fragmentation. However, while BAPTA and quin-2 inhibited the calcium-dependent apoptotic fragmentation seen in thymocytes they did not inhibit the DNA fragmentation in L929 cells. In addition, the loss of membrane integrity in both L929 and EMT-6 cells preceded or approximated the appearance of DNA fragmentation, whereas loss of membrane integrity usually follows DNA fragmentation in apoptosis. However, morphological studies showed that apoptotic bodies were present in L929 cell cultures treated with TNF-α and heat, and were distinguishable from necrosing cells. We conclude that both types of DNA fragmentation are operant in some cell lines exhibiting a cytotoxic response to TNF-α and heat treatments, and that increased fragmentation reflects the greatly enhanced cytotoxic interactions seen with combination treatments in those cells.
AB - The relationship of DNA fragmentation to the greatly enhanced cytotoxicity seen in vitro against tumour cells when recombinant human tumour necrosis factor-α (TNF-α is combined with hyperthermia was investigated. The TNF-αsensitive L929 and -resistant EMT6 cells were treated with 8.8 and 16 ng of TNF-α per ml, respectively, and then heated at 40.5°C for 24 h (L929) or at 43°C for 1 h (L929) or 1.5 h (EMT-6) beginning 1 h later. For both cell lines at both temperatures, the addition of heating to the TNF-α treatment significantly decreased viability and increased DNA fragmentation at earlier time points than seen with either TNF-α or heat alone. DNA fragmentation was further studied using agarose gel electrophoresis to examine the size distribution of the DNA fragments and the ability of intracellular calcium buffering agents BAPTA and quin-2 to inhibit fragmentation. At 4.5 h after L929 cells were treated with TNF-α at 43°C, the size distribution of DNA fragments more closely resembled the oligonucleosome sized apoptotic DNA fragmentation, as seen in irradiated rat thymocytes, than the spectrum of DNA fragments seen in necrotic fragmentation. However, while BAPTA and quin-2 inhibited the calcium-dependent apoptotic fragmentation seen in thymocytes they did not inhibit the DNA fragmentation in L929 cells. In addition, the loss of membrane integrity in both L929 and EMT-6 cells preceded or approximated the appearance of DNA fragmentation, whereas loss of membrane integrity usually follows DNA fragmentation in apoptosis. However, morphological studies showed that apoptotic bodies were present in L929 cell cultures treated with TNF-α and heat, and were distinguishable from necrosing cells. We conclude that both types of DNA fragmentation are operant in some cell lines exhibiting a cytotoxic response to TNF-α and heat treatments, and that increased fragmentation reflects the greatly enhanced cytotoxic interactions seen with combination treatments in those cells.
KW - Apoptosis
KW - Hyperthermia
KW - Necrosis
KW - TNF-α
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U2 - 10.3109/02656739409009346
DO - 10.3109/02656739409009346
M3 - Article
C2 - 8064183
AN - SCOPUS:0028294624
SN - 0265-6736
VL - 10
SP - 247
EP - 262
JO - International Journal of Hyperthermia
JF - International Journal of Hyperthermia
IS - 2
ER -