Abstract
Upon transforming growth factor-β (TGF-β) treatment, Ramos cells, a B-cell lymphoma cell line, undergo apoptosis, as measured by annexin V labeling, DNA fragmentation, and propidium iodide staining. Apoptosis could be observed by 24 h after TGF-β exposure and occurred before the development of a significant blockage of cell cycle progression. TGF-β-mediated apoptosis was also accompanied by a strong induction of caspase-3 subfamily activity. Incubation of cells with the caspase inhibitor Z-VAD.FMK at 20 μM, but not at 10 μM, prevented TGF-β-induced apoptosis from occurring. By comparison, caspase-3 subfamily activity was 87% inhibited at 10 μM Z-VAD.FMK and completely inhibited at 20 μM. Because of TGF-β's well-established role of regulating gene transcription, the mRNA levels for proteins associated with apoptosis (Fas- and Fas-associated proteins, Bcl-2 family members, IAP proteins, and IκB) were also studied. After 24 h of TGF-β treatment, the most significant mRNA changes occurred with Bcl-X L(twofold decrease) and Bik (twofold increase). TGF-β treatment also resulted after 48 h in a fivefold decrease in Bcl-X Lprotein levels, based on immunoblotting analysis. Therefore, TGF-β-mediated apoptosis involves the activation of caspases. In addition, TGF-β transcriptionally regulates Bcl-2 family members, Bcl-X Land Bik, to further influence the apoptosis process.
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