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Upregulation of P-glycoprotein in rat hepatoma rho(o) cells: implications for drug-DNA interactions
V Pillay1, R D Martinus, J S Hill
1School of Biochemistry, La Trobe University, Bundoora, Australia.
Abstract:
Rat hepatoma cells lacking mitochondrial DNA (rho(o) cells) were used as a model system to examine the possible roles of mitochondrial DNA as a target for the DNA-acting anticancer drug Adriamycin (doxorubicin). The rho(o) cells were 45-fold less sensitive to Adriamycin than the parental rho+ cells containing mitochondrial DNA. Other non-DNA-acting drugs also exhibited similar behaviour, and this was shown to be due to a multidrug resistance (MDR) phenotype in the rho(o) cells. This was indicated by confocal microscopy where rho+ cells exhibited thirteenfold higher cellular levels of Adriamycin than rho(o) cells. Upregulation (tenfold) of P-glycoprotein in rho(o) cells was also confirmed by Northern dot blot analysis. Since the MDR phenotype is present in rho(o) cells and upregulation of P-glycoprotein is maintained in these cells, rho(o) cells are not a good model system for drug-DNA studies (where the drug is susceptible to extrusion by P-glycoprotein), and any such results obtained with this system must be treated with considerable caution.
Insights
Mitochondrial DNA-lacking cells (rho(o) cells) show reduced sensitivity to Adriamycin due to a multidrug resistance (MDR) phenotype. These cells are not ideal for studying drug-DNA interactions because of P-glycoprotein upregulation.
Area of Science:
- Biochemistry
- Cell Biology
- Pharmacology
Background:
- Mitochondrial DNA (mtDNA) plays a role in cellular processes.
- Adriamycin (doxorubicin) is a DNA-acting anticancer drug.
- Mitochondrial dysfunction can impact drug sensitivity.
Purpose of the Study:
- To investigate the role of mtDNA as a target for Adriamycin.
- To evaluate the utility of rho(o) cells (lacking mtDNA) as a model for drug-DNA interaction studies.
- To characterize drug resistance mechanisms in rho(o) cells.
Main Methods:
- Comparison of Adriamycin sensitivity between rho+ and rho(o) rat hepatoma cells.
- Confocal microscopy to quantify intracellular Adriamycin levels.
- Northern dot blot analysis to assess P-glycoprotein expression.
Main Results:
- Rho(o) cells exhibited a 45-fold decrease in sensitivity to Adriamycin compared to rho+ cells.
- Rho(o) cells displayed a multidrug resistance (MDR) phenotype, affecting both DNA-acting and non-DNA-acting drugs.
- Confocal microscopy revealed 13-fold lower Adriamycin levels in rho(o) cells.
- A tenfold upregulation of P-glycoprotein was confirmed in rho(o) cells.
Conclusions:
- The observed drug resistance in rho(o) cells is primarily mediated by the MDR phenotype and P-glycoprotein.
- Rho(o) cells are unsuitable models for studying drug-DNA interactions due to P-glycoprotein-mediated drug extrusion.
- Results from studies using rho(o) cells for drug-DNA interaction should be interpreted with caution.