In this study， we followed a DNA vector-based RNAi approach to silence the signal transducer and activator of transcription 3 (STAT3) expression in Bel-7402 cells， to explore how the Janus kinase (JAK)/STAT3 signaling pathway influences the apoptosis of hepatocarcinoma cells. According to GenBank's STAT3 cDNA， the plasmid pGCsi.U6/neoRFP STAT3， which was designed for expression of STAT3 small interfering RNA (siRNA)， was constructed and synthesized， and then transfected into Bel-7402 cells using Lipofectamine 2000. Cells with or without siRNA transfection were treated in wells. The apoptotic rate was detected by flow cytometry (FCM) and by staining with the Annexin V/propidium iodide (PI) apoptosis ... More
In this study， we followed a DNA vector-based RNAi approach to silence the signal transducer and activator of transcription 3 (STAT3) expression in Bel-7402 cells， to explore how the Janus kinase (JAK)/STAT3 signaling pathway influences the apoptosis of hepatocarcinoma cells. According to GenBank's STAT3 cDNA， the plasmid pGCsi.U6/neoRFP STAT3， which was designed for expression of STAT3 small interfering RNA (siRNA)， was constructed and synthesized， and then transfected into Bel-7402 cells using Lipofectamine 2000. Cells with or without siRNA transfection were treated in wells. The apoptotic rate was detected by flow cytometry (FCM) and by staining with the Annexin V/propidium iodide (PI) apoptosis detection kit. Simultaneously， the mitochondrial membrane potential (ΔΨm) was visualized by JC-1 fluorescence staining and observed using the inverted fluorescence microscope. Furthermore， the expression of caspase-3 protein was analyzed by Western blotting. The results showed that treatment with STAT3 siRNA displayed effects in the Bel-7402 cells， causing a significantly increased apoptotic ratio (P < 0.05). The mitochondrial membrane potential of the STAT3 siRNA group， observed by the JC-1 fluorescence staining， decreased significantly. The protein expression of active caspase-3 increased with STAT3 siRNA treatment， and was significantly higher than that of the control group (P < 0.05). STAT3 gene-silencing significantly improves the apoptotic effect against Bel-7402 cells.
