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Retroviral-mediated transfer and functional expression of multidrug resistance gene in human placenta mesenchymal
Li-ying Han1, Ming-zhu Ye, Ya-ping Li
1Department of Obstetrics and Gynecology, Second Hospital, Jilin University, Changchun, Jilin, China.
Background:
Most of gynecologic malignancies are sensitive to chemotherapy. Myelosuppression is the main dose-related toxicity of many chemotherapeutic drugs. The human multidrug resistance (mdr1) gene is well known for its ability to confer drug resistance. This study aimed to explore the feasibility of expression and resistance of mdr1 gene transduction into human placenta mesenchymal stem cells (P-MSCs) by retrovirus vector.
Methods:
Human P-MSCs were isolated from trypsin-digested term placentas, and their immunophenotypes and differentiation potential were evaluated. Human P-MSCs were transduced by reconstructed retroviral vector containing the mdr1 gene and green fluorescent protein (GFP) reporter gene. The integration and expression of the mdr1 gene were observed indirectly by the expression of GFP, and fluorescence-activated cell sorter was used to evaluate the functional activity of permeability glycoprotein (P-gp) encoded by the mdr1 gene. The stimulating test was made in vitro to show pleiotropic drug resistance of transfected cells.
Results:
The isolated, cultured and expanded P-MSCs expressed stem cell markers such as CD29, CD44 and CD73, and showed osteogenic and adipogenic differentiation potentials under appropriate conditions. The expression of P-gp in the non-transfected P-MSCs cells was (0.4 +/- 0.1)%, but increased to (28.1 +/- 4.7)% after gene transfection (P < 0.01). And positive staining of P-gp located mainly at cell membrane and cytoplasm. Accumulation and extrusion assays showed that P-gp expressed by the transfected cells had pump-functional activity and could efflux daunomycin out of cells. The analysis of cell survival confirmed that transfected P-MSCs had a characteristic of multidrug resistance with a significant increase in the resistance to anticancer agents.
Conclusions:
Transfer and expression of human mdr1 gene mediated by retrovirus vector conferred P-MSCs drug resistance. It might provide a new alternative to chemoprotection strategies.
Insights
This study successfully transferred the human multidrug resistance (mdr1) gene into placenta mesenchymal stem cells (P-MSCs) using a retrovirus vector. Transfected P-MSCs exhibited significant multidrug resistance, offering a potential new strategy for chemoprotection.
Area of Science:
- Stem cell biology
- Gene therapy
- Cancer research
Background:
- Gynecologic cancers often respond to chemotherapy, but myelosuppression is a common dose-limiting toxicity.
- The human multidrug resistance (mdr1) gene confers resistance to various chemotherapeutic agents.
- Placenta-derived mesenchymal stem cells (P-MSCs) are a potential candidate for therapeutic applications.
Purpose of the Study:
- To investigate the feasibility of transducing the mdr1 gene into human P-MSCs using a retroviral vector.
- To evaluate the expression and drug resistance conferred by the mdr1 gene in P-MSCs.
Main Methods:
- Human P-MSCs were isolated, characterized for immunophenotype and differentiation potential, and transduced with a retroviral vector containing the mdr1 and green fluorescent protein (GFP) genes.
- GFP expression was used to indirectly assess mdr1 gene integration and expression.
- Fluorescence-activated cell sorting (FACS) and in vitro assays were employed to evaluate P-glycoprotein (P-gp) function and multidrug resistance in transfected cells.
Main Results:
- Transduced P-MSCs expressed stem cell markers and retained differentiation potential.
- MDR1 gene transfection significantly increased P-gp expression in P-MSCs from 0.4% to 28.1% (P < 0.01).
- Functional assays confirmed P-gp's ability to efflux daunomycin and demonstrated multidrug resistance in transfected P-MSCs.
Conclusions:
- Retroviral-mediated transfer and expression of the human mdr1 gene successfully conferred multidrug resistance to P-MSCs.
- This approach may offer a novel strategy for chemoprotection against chemotherapy-induced toxicities.
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