American Society of Hematology

Fig. 5.

$Fig. 5. Lonidamine induces apoptosis in TPA-differentiated U937 cells. / Undifferentiated (▴, U937), TPA-differentiated (●, U-TPA), and Bcl-2–overexpressing U937 cells (▪, U-BCL2) were treated for indicated times with 200 μM lonidamine. (A) Apoptotic DNA fragmentation was quantified using a filter elution assay. Values from 1 of 2 independent experiments are shown (mean ± SD of triplicate samples). (B) Western blot analysis of cytochrome c and Smac/Diablo expression in cytosolic fractions from U937, U-TPA, and U-BCL2 cells treated (+) or not treated (−) with 200 μM lonidamine for 48 hours. (C) Mitochondrial membrane depolarization was visualized by flow cytometry in U937, U-TPA, and U-BCL2 cells treated with 200 μM lonidamine for indicated times. Mitochondrial depolarization was identified by an increased level of cytosolic green monomer (FL1-H; dashed lines indicate the controls). (D) Western blot analysis of procaspase-3 and caspase-3 active fragments and PARP expression in whole-cell extracts from U937, U-TPA, and U-BCL2 cells treated (+) or not treated (−) with 200 μM lonidamine for 48 hours. Numbers are molecular weights in kilodaltons. *Cleavage products. Loading was checked with the use of an antihuman HSC70 mAb.$

Lonidamine induces apoptosis in TPA-differentiated U937 cells.

Undifferentiated (▴, U937), TPA-differentiated (●, U-TPA), and Bcl-2–overexpressing U937 cells (▪, U-BCL2) were treated for indicated times with 200 μM lonidamine. (A) Apoptotic DNA fragmentation was quantified using a filter elution assay. Values from 1 of 2 independent experiments are shown (mean ± SD of triplicate samples). (B) Western blot analysis of cytochrome c and Smac/Diablo expression in cytosolic fractions from U937, U-TPA, and U-BCL2 cells treated (+) or not treated (−) with 200 μM lonidamine for 48 hours. (C) Mitochondrial membrane depolarization was visualized by flow cytometry in U937, U-TPA, and U-BCL2 cells treated with 200 μM lonidamine for indicated times. Mitochondrial depolarization was identified by an increased level of cytosolic green monomer (FL1-H; dashed lines indicate the controls). (D) Western blot analysis of procaspase-3 and caspase-3 active fragments and PARP expression in whole-cell extracts from U937, U-TPA, and U-BCL2 cells treated (+) or not treated (−) with 200 μM lonidamine for 48 hours. Numbers are molecular weights in kilodaltons. *Cleavage products. Loading was checked with the use of an antihuman HSC70 mAb.

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