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The nuclear membrane in multidrug resistance: microinjection of epirubicin into bladder cancer cell lines

The nuclear membrane in multidrug resistance: microinjection of epirubicin into bladder cancer cell lines
The nuclear membrane in multidrug resistance: microinjection of epirubicin into bladder cancer cell lines
Objevtive: to assess whether microinjecting epirubicin into cells showing multidrug resistance (MDR, common to many cancers, including bladder cancer, with resistance to, e.g. anthracyclines and mitomycin C) spares the nucleus, as when these drugs accumulate, distribution in MDR cells characteristically spares the nucleus, suggesting that the nuclear membrane is responsible for excluding cytotoxic drugs from MDR nuclei.

Materials and methods: nuclear exclusion of drugs is an important feature of resistance in MDR cells, as many MDR-susceptible drugs have cytotoxic actions within the nucleus. Drug accumulation in 'classical' P-glycoprotein-mediated MDR cells is greatly reduced by efflux. Microinjection of epirubicin into the cytoplasm of MDR cells bypasses the P-glycoprotein efflux pump on the plasma membrane. Nuclear sparing would directly implicate the nuclear membrane in this phenomenon. Because of their fluorescence properties, which allow study by confocal microscopy and flow cytometry, anthracyclines have also been used extensively to investigate MDR. Thus sensitive (MGH-U1 and RT112) and MDR (MGH-U1R and MGH-U1-MMC) bladder cancer cell lines were used. Adherent cells from each cell line were individually microinjected with epirubicin (0.5 mg/mL) and a 77 kDa fluorescein isothiocyanate (FITC)-dextran (0.5 mg/mL). The pattern of nuclear epirubicin uptake in injected cells was then evaluated by confocal microscopy. The 77 kDa FITC-dextran allowed easier identification of injected cells and was also excluded from their nuclei.

Results: sensitive bladder cancer cell lines all showed a nuclear accumulation pattern of epirubicin, consistent with their normal uptake after exposure to epirubicin. The MDR cell lines showed the characteristic nuclear-sparing pattern of epirubicin uptake, similar to the normal uptake pattern after epirubicin exposure. The 77 kDa FITC-dextran showed clearly which cells had been microinjected, and was excluded from the nuclei of all injected cells. Cell viability was confirmed by acridine-orange staining after initial visualization of injected cells.

Conclusion: the nuclear membrane is responsible for the nuclear exclusion of epirubicin in MDR cells. Further work is necessary to determine the mechanisms involved.
bladder cancer, multidrug resistance, epirubicin, nuclear membrance, microinjection
1464-4096
1091-1098
Featherstone, J.F
f24534ce-9bda-4a2a-8aea-540ffd1035d6
Speers, A.G
aab49fc2-adf1-4ca2-a912-2747cbb6bd04
Lwaleed, B.A
e7c59131-82ad-4a14-a227-7370e91e3f21
Hayes, C.M
9a5b655e-586c-427c-9299-a1aaeb030023
Cooper, A.J
ff520c5d-ecfa-4473-8ed9-da59434b1c90
Birch, B.R
f2699d05-ec2b-4be5-8d86-5b0663bb3cfc
Featherstone, J.F
f24534ce-9bda-4a2a-8aea-540ffd1035d6
Speers, A.G
aab49fc2-adf1-4ca2-a912-2747cbb6bd04
Lwaleed, B.A
e7c59131-82ad-4a14-a227-7370e91e3f21
Hayes, C.M
9a5b655e-586c-427c-9299-a1aaeb030023
Cooper, A.J
ff520c5d-ecfa-4473-8ed9-da59434b1c90
Birch, B.R
f2699d05-ec2b-4be5-8d86-5b0663bb3cfc

Featherstone, J.F, Speers, A.G, Lwaleed, B.A, Hayes, C.M, Cooper, A.J and Birch, B.R (2005) The nuclear membrane in multidrug resistance: microinjection of epirubicin into bladder cancer cell lines. BJU International, 95 (7), 1091-1098. (doi:10.1111/j.1464-410X.2005.05473.x).

Record type: Article

Abstract

Objevtive: to assess whether microinjecting epirubicin into cells showing multidrug resistance (MDR, common to many cancers, including bladder cancer, with resistance to, e.g. anthracyclines and mitomycin C) spares the nucleus, as when these drugs accumulate, distribution in MDR cells characteristically spares the nucleus, suggesting that the nuclear membrane is responsible for excluding cytotoxic drugs from MDR nuclei.

Materials and methods: nuclear exclusion of drugs is an important feature of resistance in MDR cells, as many MDR-susceptible drugs have cytotoxic actions within the nucleus. Drug accumulation in 'classical' P-glycoprotein-mediated MDR cells is greatly reduced by efflux. Microinjection of epirubicin into the cytoplasm of MDR cells bypasses the P-glycoprotein efflux pump on the plasma membrane. Nuclear sparing would directly implicate the nuclear membrane in this phenomenon. Because of their fluorescence properties, which allow study by confocal microscopy and flow cytometry, anthracyclines have also been used extensively to investigate MDR. Thus sensitive (MGH-U1 and RT112) and MDR (MGH-U1R and MGH-U1-MMC) bladder cancer cell lines were used. Adherent cells from each cell line were individually microinjected with epirubicin (0.5 mg/mL) and a 77 kDa fluorescein isothiocyanate (FITC)-dextran (0.5 mg/mL). The pattern of nuclear epirubicin uptake in injected cells was then evaluated by confocal microscopy. The 77 kDa FITC-dextran allowed easier identification of injected cells and was also excluded from their nuclei.

Results: sensitive bladder cancer cell lines all showed a nuclear accumulation pattern of epirubicin, consistent with their normal uptake after exposure to epirubicin. The MDR cell lines showed the characteristic nuclear-sparing pattern of epirubicin uptake, similar to the normal uptake pattern after epirubicin exposure. The 77 kDa FITC-dextran showed clearly which cells had been microinjected, and was excluded from the nuclei of all injected cells. Cell viability was confirmed by acridine-orange staining after initial visualization of injected cells.

Conclusion: the nuclear membrane is responsible for the nuclear exclusion of epirubicin in MDR cells. Further work is necessary to determine the mechanisms involved.

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Published date: May 2005
Keywords: bladder cancer, multidrug resistance, epirubicin, nuclear membrance, microinjection

Identifiers

Local EPrints ID: 19199
URI: http://eprints.soton.ac.uk/id/eprint/19199
ISSN: 1464-4096
PURE UUID: 9a9009d2-e9c4-4ad8-aeee-852295320ca3

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Date deposited: 20 Jan 2006
Last modified: 15 Mar 2024 06:13

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Contributors

Author: J.F Featherstone
Author: A.G Speers
Author: B.A Lwaleed
Author: C.M Hayes
Author: A.J Cooper
Author: B.R Birch

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