Increased activity and sensitivity of mitochondrial respiratory enzymes to tumor necrosis factor alpha-mediated inhibition is associated with increased cytotoxicity in drug-resistant leukemic cell lines

Abstract

The drug-resistant leukemic cell lines, CEM/VLB100 and K/DAU600, are more sensitive to tumor necrosis factor alpha (TNFalpha)-mediated cytotoxicity compared with their parental cell lines, CCRF-CEM and K562 cl.6. Drug-resistant leukemic cell lines have more active mitochondrial function, which is associated with a greater susceptibility to TNFalpha- induced respiratory inhibition. TNFalpha blocked electron transfer at three sites, NADH dehydrogenase (complex I), succinate dehydrogenase (complex II), and cytochrome c oxidase (complex IV). Respiratory rate and electron transport chain enzyme activities were significantly inhibited in the drug-resistant, TNF-sensitive cell lines. Respiratory inhibition preceded cell death by at least 5 to 8 hours. The respiratory failure was not compensated for by appropriate up- regulation of the glycolytic pathway. Increasing mitochondrial respiratory rate and enzyme activities by long-term culture with 2 mmol/L adenosine 5′-diphosphate (ADP) and Pi sensitized both drug- sensitive and drug-resistant cells to TNFalpha-induced cytolysis. Intramitochondrial free radicals generated by paraquat only had a limited and delayed effect on respiratory inhibition and cytolysis in comparison with the effect of TNFalpha. We conclude that TNFalpha- induced cytotoxicity in leukemic cells is, at least in part, mediated by inhibition of mitochondrial respiration. Free radical generation by TNFalpha may not directly lead to the observed inhibition of the mitochondrial electron transport and other mechanisms must be involved.

Mitochondrial electron transport chain activity, but not ATP synthesis, is required for drug‐induced apoptosis in human leukaemic cells: a possible novel mechanism of regulating drug resistance

“…1% Triton-X 100. The LDH activity was determined using a LDH assay kit (Amador et al, 1963;Jia et al, 1996).…”

“…Oxygen consumption was calibrated with air-saturated respiratory medium, assuming 390 ng atoms O 2 /ml. Respiratory rate or enzyme activity was expressed in ng atoms O 2 /min/10 7 cells (Granger & Lehninger, 1982;Schoonen et al, 1990;Jia et al, 1996).…”

“…We have previously shown that the daunorubicin-resistant subline K/DAU 600 and the vinblastine-resistant cell line CEM/VLB 100 , which highly express ATP-dependent Pgp, possess more active mitochondrial electron transport chain (ETC) enzyme activities than their parental cell lines K562 cl.6 and CCRF-CEM, whereas glycolytic metabolism between the drug-sensitive and drug-resistant cell lines is consistent. Increased activity of the mitochondrial ETC was associated with increased susceptibility to tumour necrosis factor-a (TNFa) mediated apoptosis ( Jia et al, 1996). It has also been observed that the drug-resistant cell lines K/DAU 600 and CEM/VLB 100 were more susceptible to both etoposide and cytarabine-induced apoptosis than their parental K562 cl.6 and CCRF-CEM cell lines (Allen et al, 1995).…”

“…Mitochondria are one of the targets of anticancer chemotherapy drugs (Kroon & van den Bogert, 1987;Decaudin et al, 1997), TNFa (Schulze-Osthoff et al, 1992;Jia et al, 1996), interferons (Lewis et al, 1996) and reactive oxygen species (ROS) induced apoptosis (Crawford et al, 1997). However, increasing attention is now devoted to mitochondrial function during the apoptotic process (Liu et al, 1996;Marchetti et al, 1996;Zamzami et al, 1996;Yang et al, 1997;Kluck et al, 1997;Higuchi et al, 1997).…”

Constitutive levels of cAMP‐dependent protein kinase activity determine sensitivity of human multidrug‐resistant leukaemic cell lines to growth inhibition and apoptosis by forskolin and tumour necrosis factor alpha

“…Previous studies in our department have found that MDR1-positive leukaemia cells CEM/VLB100 and K562/ Dau600 were more sensitive to TNF-induced cell killing than their parental cell lines (Jia et al, 1995(Jia et al, , 1996. Although these were related in part to decreased manganese superoxide dismutase (MnSOD) activity and increased activity in the enzymes of the mitochondrial electron transport chain in drug-resistant cell lines (Jia et al, 1997), the mechanisms and the signalling pathways related to these is still unclear.…”