Complicated effects of Gfi-1 on proliferation and apoptosis of murine 32D cells

Abstract

To explore the relationship between growth factor independence 1 (Gfi-1) and apoptosis as well as proliferation in 32D cell and observe the effects of Gfi-1 and N382S on 32D cell so as to further explore its possible mechanism of apoptosis and proliferation in hemopoietic stem cells. 32D cells expressed a high level of Gfi-1 or its mutant N382S by plasmid transduction, cell apoptosis was examined by flow cytometry and the levels of Bax, signal transducer and activator of transcription 3 (STAT3) and Gfi-1 protein by Western blot. The apoptotic cell ratios were lower in the 32D/Gfi-1 groups than those in the 32D/puro groups (4.0% ± 1.8%, 7.0% ± 1.6%, 10.0% ± 2.4% vs 6.0% ± 1.0%, 11.0% ± 1.5% and 23.0% ± 1.5% respectively, all P < 0.05)after culture for 48, 96 and 144 h. The apoptotic rates were compared between the 32D/N382S and 32D/puro groups.However, there were no marked statistical difference (5.0% ± 1.3%, 12.0% ± 2.1%, 21.0% ± 2.2% vs 6.0% ± 1.0%, 11.0% ± 1.5%, 23.0% ± 1.5% respectively, P > 0.05).Interestingly, the number of 32D cells expressed Gfi-1 decreased compared to the controls (0.96 × 10⁶, 2.03 × 10⁶, 3.47 × 10⁶ vs 1.42 × 10⁶, 3.26 × 10⁶, 6.12 × 10⁶ respectively, all P < 0.05). But no marked difference existed between 32D/N382S groups and the controls (1.51 × 10⁶, 3.36 × 10⁶, 6.54 × 10⁶ vs 1.42 × 10⁶, 3.26 × 10⁶ and 6.12 × 10⁶ respectively, P > 0.05) after culturing for 48, 96 and 144 h. The expression level of Bax decreased significantly in 32D/Gfi-1 groups (1.090 ± 0.062 vs 1.707 ± 0.025, P < 0.05), but showed no marked in 32D/N382S groups (1.710 ± 0.062 vs 1.707 ± 0.025, P > 0.05), compared to 32D/puro groups. The expression level of STAT3 increased significantly in 32D/Gfi-1 groups (1.723 ± 0.168 vs 1.203 ± 0.168, P < 0.05) and 32D/N382S groups (1.797 ± 0.292 vs 1.203 ± 0.168, P < 0.05) compared to 32D/puro groups. Gfi-1 inhibits cell apoptosis and proliferation.Such an effect is associated with Bax down-regulation.Zinc finger structure of Gfi-1 has no marked interaction with the expression of STAT3. And Gfi-1 is both unique and complicated in hemopoietic stem cells.