Selective susceptibility of transformed T lymphocytes to induction of apoptosis by PSC 833, an inhibitor of P-glycoprotein.

P-glycoprotein is a cellular efflux pump. The P-glycoprotein inhibitor PSC 833 causes apoptosis of cancer cells and induces a rise in the intracellular levels of ceramide. Our aims were to determine whether a cause and effect relationship exists between these two actions of PSC 833, and to assess whether the PSC 833-induced apoptosis is restricted to transformed cells. Apoptosis was determined by flow cytometry and radioactive quantitation of DNA fragmentation. PSC 833 induced apoptosis in the human T leukemia cell lines: Molt-4 and Jurkat. Analysis of the apoptosis in Molt-4 and Jurkat cells revealed that PSC 833 induced a rise in the cellular ceramide levels (as measured by the DG kinase assay). PSC 833-induced apoptosis was significantly reduced by specific inhibitors of ceramide de novo synthesis (i.e., fumonisin B1 and L-cycloserine). On the other hand, PSC 833 did not induce apoptosis in normal peripheral blood T cells regardless of whether these cells were quiescent, activated, or proliferating. Our results suggest that PSC 833 induces apoptotic death in human transformed T lymphocytes through an increase in ceramide de novo synthesis. In addition, normal lymphocytes are not susceptible to induction of apoptosis by PSC 833. This difference between normal lymphocytes and leukemia cells presents a potential target for chemotherapy.

Aspirin enhances multidrug resistance gene 1 expression in human Molt-4 T lymphoma cells.

BACKGROUND: We recently found that aspirin induces the expression of P-glycoprotein (P-gp), a protein mediating drug resistance, in human prostate cancer cells. The purpose of this study was to evaluate the effect of aspirin on the expression of P-gp in a different human cancer type, i.e., T lymphoma. Furthermore, we analyzed this effect at the level of the gene encoding P-gp, MDR1, and of the transcription factor (NF-IL6), regulating this gene. MATERIALS AND METHODS: NF-IL6 was assayed by the electrophoretic mobility shift assay, MDR1 mRNA was assayed by the reverse transcriptase polymerase chain reaction (RT-PCR), and P-gp was assayed by Western blotting. RESULTS: aspirin, at plasma attainable levels, induced NF-IL6 DNA-binding activity, and increased MDR1 mRNA expression (by up to 140%), as well as the expression of P-gp, in Molt-4 cells. CONCLUSIONS: This study suggests that treatment with aspirin induces a cellular signal culminating in the enhancement of P-gp expression in T lymphoma Molt-4 cells.

Overexpression of peptide-methionine sulfoxide reductase in Saccharomyces cerevisiae and human T cells provides them with high resistance to oxidative stress.

The yeast peptide-methionine sulfoxide reductase (MsrA) was overexpressed in a Saccharomyces cerevisiae null mutant of msrA by using a high-copy plasmid harboring the msrA gene and its promoter. The resulting strain had about 25-fold higher MsrA activity than its parent strain. When exposed to either hydrogen peroxide, paraquat, or 2,2'-azobis-(2-amidinopropane) dihydrochloride treatment, the MsrA overexpressed strain grew better, had lower free and protein-bound methionine sulfoxide and had a better survival rate under these conditions than did the msrA mutant and its parent strain. Substitution of methionine with methionine sulfoxide in a medium lacking hydrogen peroxide had little effect on the growth pattern, which suggests that the oxidation of free methionine in the growth medium was not the main cause of growth inhibition of the msrA mutant. Ultraviolet A radiation did not result in obvious differences in survival rates among the three strains. An enhanced resistance to hydrogen peroxide treatment was shown in human T lymphocyte cells (Molt-4) that were stably transfected with the bovine msrA and exposed to hydrogen peroxide. The survival rate of the transfected strain was much better than its parent strain when grown in the presence of hydrogen peroxide. These results support the proposition that the msrA gene is involved in the resistance of yeast and mammalian cells to oxidative stress.