Figure 3

FHL1C overexpression induced apoptosis in Jurkat cells. (A) Jurkat cells were transiently transfected with pEGFP or pEGFP-FHL1C by using the Nucleofection method. Apoptosis in the GFP⁻ and GFP⁺ fractions of cells was determined by AnnexinV staining followed by FACS 48 h post-transfection. (B) Percentages of apoptotic cells (Annexin V⁺) in GFP⁻ and GFP⁺ cell fractions in (A) were compared. (C) Jurkat cells were transfected with pEGFP or pEGFP-FHL1C by using the Nucleofection method. Early and late apoptotic cells were depicted 48 h post-transfection by using Annexin V and PI staining followed by FACS. (D) GFP⁺ cells in early and late apoptotic phases in (C) were compared. (E) Jurkat cells were transiently transfected with pEGFP or pEGFP-FHL1C by using the Nucleofection method. Cells were stained with Hoechst 24 h post-transfection and nuclei were observed under a fluorescence microscope. Arrow heads indicate Hoechst-positive apoptotic nuclei. (F) Typical cell apoptosis in (E) was depicted under TEM. Intact cell membrane, organelles and normal nuclear morphology were observed in vector-transfected cells, whereas incomplete membrane and condensed nuclei were observed in cells overexpressing FHL1C (magnification, × 9900). (G) Total RNA was prepared from cells in (E) 24 h post-transfection. The mRNA levels of the apoptosis-related molecules were determined by real time RT-PCR, with β-actin as a reference. (H) Cell lysates were prepared from cells in (E) 24 h post-transfection. The level of Caspase3 was determined by Western blot analysis. Bars = means ± S.D (n = 3), *P < 0.05; NS, not significant.