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[Effect of Recombinant Adenovirus AdE-SH2-Caspase 8 on the Apoptosis of Imatinib-resistant K562/G01 Cell Line]
Lin Wang1,2, Chang Fei1, Zheng-Lan Huang2
1College of Laboratorial Medicine, Hunan University of Medicine, Huaihua 418000, Hunan Province, China.
Objective:
To investigate the effect of SH2-Caspase 8 fusion protein expressed by recombinant adenovirus AdE-SH2-Caspase8-HA-GFP (SC) on the apoptosis of K562/G01 cell line, which is a BCR/ABL positive chronic myeloid leukemia cell line and resistant to imatinib.
Methods:
The K562/G01 cell line was infected with AdE-SH2-Caspase 8-HA-GFP adenovirus (SC), then the cells were divided into 3 groups: AdE-SH2m-Caspase 8-HA-GFP (SmC) group, AdE-GFP (CMV) group and PBS group as control. The infection efficiency was observed under fluorescent microscopy and by flow cytometry. The expression of fusion protein SH2-Caspase 8-HA was measured by Western blot. The morphology of the cells detected by Wright's staining. The apoptosis of the cells were detected by flow cytometry and DNA ladder. The expression of Caspase 3 and PARP were detected by Western blot.
Result:
The infection efficiency of SC on K562/G01 cells was high which was confirmed by fluorescent microscopy and FCM. SH2-Caspase 8-HA fusion protein were expressed correctly in K562/G01 cells. After treatment with SC the apoptosis of K562/G01 cells could be observed by microscopy. The result of FCM showed that early apoptosis of K562/G01 cells increased significantly as compared with control groups (P < 0.05). DNA ladder showed that the classic DNA ladders appeared in K562/G01 cells after treatment with SC. The wester blot detection showed that the expression level of apoptosis-related protein Caspase 3 and PARP increased.
Conclusion:
The recombinant adenovirus SC expressing SH2-Caspase 8 fusion protein can induces the apoptosis of K562/G01 cells.
Insights
Recombinant adenovirus expressing SH2-Caspase 8 fusion protein effectively induces apoptosis in imatinib-resistant K562/G01 chronic myeloid leukemia cells, increasing Caspase 3 and PARP expression.
Area of Science:
- Molecular biology
- Cancer research
- Virology
Context:
- K562/G01 cells are a BCR/ABL positive chronic myeloid leukemia cell line resistant to imatinib.
- Recombinant adenovirus AdE-SH2-Caspase8-HA-GFP (SC) was used to deliver the SH2-Caspase 8 fusion protein.
Purpose:
- To investigate the effect of the SH2-Caspase 8 fusion protein on the apoptosis of K562/G01 cells.
- To assess the potential of SC as a therapeutic agent against imatinib-resistant CML.
Summary:
- High infection efficiency of SC on K562/G01 cells was confirmed.
- Expression of SH2-Caspase 8-HA fusion protein was verified, leading to observable apoptosis.
- Flow cytometry and DNA ladder assays demonstrated significant early apoptosis induction.
- Western blot analysis revealed increased levels of apoptosis-related proteins Caspase 3 and PARP.
Impact:
- SC effectively induces apoptosis in imatinib-resistant K562/G01 cells.
- This study provides a potential new strategy for treating CML resistant to imatinib.
- The findings highlight the therapeutic potential of targeted gene therapy using recombinant adenoviruses.
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