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Retrovirus-mediated siRNA targeting TRPM7 gene induces apoptosis in RBL-2H3 cells
1Clinical Department of School of Medicine, Shenzhen University, Shenzhen, China.
Background And Objective:
Calcium signaling is important for both normal physiologic processes and pathology of various diseases. Transient receptor potential melastatin 7 (TRPM7) gene has been reported to be a potential candidate for calcium influx. The present study aimed to investigate the possible role of TRPM7 channels in apoptosis in rat basophilic leukemia mast cell line (RBL-2H3), which is widely used in mast cell-associated studies.
Materials And Methods:
A recombinant retrovirus vector siRNA targeting rat TRPM7 gene was constructed and identified. Cellular survival was assessed by MTT. Cell apoptosis was evaluated by flow cytometry and TUNEL-FITC/Hoechst 33258 staining.
Results:
The transfection efficiency by retrovirus vector was about 60%-70%. Transfection with TRPM7 siRNA significantly reduced TRPM7 expression both at mRNA and protein levels. Suppression of TRPM7 expression by siRNA led to significantly decreased cellular survival rates and increased apoptosis rates in RBL-2H3 cells.
Conclusions:
This study indicates that TRPM7 is involved in the apoptosis process in RBL-2H3 cells.
Insights
Transient receptor potential melastatin 7 (TRPM7) channels play a role in apoptosis. Suppressing TRPM7 expression in rat basophilic leukemia mast cells (RBL-2H3) decreased cell survival and increased apoptosis.
Area of Science:
- Molecular Biology
- Cell Biology
- Immunology
Background:
- Calcium signaling is crucial for physiological processes and disease pathology.
- Transient receptor potential melastatin 7 (TRPM7) is implicated in calcium influx.
- Rat basophilic leukemia mast cell line (RBL-2H3) is a model for mast cell studies.
Purpose of the Study:
- Investigate the role of TRPM7 channels in apoptosis.
- Examine TRPM7's function in RBL-2H3 cells.
Main Methods:
- Constructed and identified a recombinant retrovirus vector siRNA targeting rat TRPM7.
- Assessed cellular survival using MTT assay.
- Evaluated apoptosis via flow cytometry and TUNEL-FITC/Hoechst 33258 staining.
Main Results:
- Achieved 60%-70% transfection efficiency with the retrovirus vector.
- Significantly reduced TRPM7 expression at both mRNA and protein levels by siRNA.
- Observed decreased cellular survival rates and increased apoptosis rates in RBL-2H3 cells upon TRPM7 suppression.
Conclusions:
- TRPM7 channels are involved in the apoptosis process in RBL-2H3 cells.
- TRPM7 suppression induces apoptosis in mast cells.
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