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Capillary electrophoresis monitors enhancement in subcellular reactive oxygen species production upon treatment with doxorubicin.

ABSTRACT: This study investigated the role of doxorubicin (DOX) accumulation in reactive oxygen species (ROS) production detected in individually electrophoresed organelles, including mitochondria, acidic organelles, and peroxisomes. While bulk measurements of ROS production in cells and organelles are not capable of discriminating between the effects of preparative procedures on measured ROS production, capillary electrophoresis with dual laser-induced detection of individual organelles demonstrated a difference in the measured ROS production as a result of various preparative procedures. Using this technique, the three different types of detected organelles (i) produce ROS and do not have detectable levels of DOX, (ii) contain detectable DOX but do not produce ROS, or (iii) produce ROS and accumulate DOX. The third type displays two subpopulations of organelles, one of which demonstrated a direct relationship between DOX uptake and subsequent ROS production, corresponding most likely to mitochondria, and a second one with low DOX uptake but large variation in ROS production, corresponding most likely to acidic organelles.

SUBMITTER: Eder AR

PROVIDER: S-EPMC2626132 | biostudies-literature | 2006 Sep

REPOSITORIES: biostudies-literature

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Capillary electrophoresis monitors enhancement in subcellular reactive oxygen species production upon treatment with doxorubicin.

Eder Angela R AR Arriaga Edgar A EA

Chemical research in toxicology 20060901 9

This study investigated the role of doxorubicin (DOX) accumulation in reactive oxygen species (ROS) production detected in individually electrophoresed organelles, including mitochondria, acidic organelles, and peroxisomes. While bulk measurements of ROS production in cells and organelles are not capable of discriminating between the effects of preparative procedures on measured ROS production, capillary electrophoresis with dual laser-induced detection of individual organelles demonstrated a di ...[more]

PMID: 16978019

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Project description:Multipotent stromal cells (MSCs) can be differentiated into osteoblasts and chondrocytes, making these cells candidates to regenerate cranio-facial injuries and lesions in long bones. A major problem with cell replacement therapy, however, is the loss of transplanted MSCs at the site of graft. Reactive oxygen species (ROS) and nonspecific inflammation generated at the ischemic site have been hypothesized to lead to MSCs loss; studies in vitro show MSCs dying both in the presence of ROS or cytokines like FasL. We questioned whether MSCs themselves may be the source of these death inducers, specifically whether MSCs produce ROS under cytokine challenge. On treating MSCs with FasL, we observed increased ROS production within 2 h, leading to apoptotic death after 6 h of exposure to the cytokine. N-acetyl cysteine, an antioxidant, is able to protect MSCs from FasL-induced ROS production and subsequent ROS-dependent apoptosis, though the MSCs eventually succumb to ROS-independent death signaling. Epidermal growth factor (EGF), a cell survival factor, is able to protect cells from FasL-induced ROS production initially; however, the protective effect wanes with continued FasL exposure. In parallel, FasL induces upregulation of the uncoupling protein UCP2, the main uncoupling protein in MSCs, which is not abrogated by EGF; however, the production of ROS is followed by a delayed apoptotic cell death despite moderation by UCP2. FasL-induced ROS activates the stress-induced MAPK pathways JNK and p38MAPK as well as ERK, along with the activation of Bad, a proapoptotic protein, and suppression of survivin, an antiapoptotic protein; the latter two key modulators of the mitochondrial death pathway. FasL by itself also activates its canonical extrinsic death pathway noted by a time-dependent degradation of c-FLIP and activation of caspase 8. These data suggest that MSCs participate in their own demise due to nonspecific inflammation, holding implications for replacement therapies.

Dataset Information

Capillary electrophoresis monitors enhancement in subcellular reactive oxygen species production upon treatment with doxorubicin.

Publications

Capillary electrophoresis monitors enhancement in subcellular reactive oxygen species production upon treatment with doxorubicin.

Similar Datasets

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