PNNL-13501 - Pacific Northwest National Laboratory

Study Control Number: PN99036/1364
Isoprostane-Mediated Tumor Promotion
Thomas J. Weber
Exposure to radiation has been associated with life-span shortening and cancer. The response of a living organism to
radiation can be modified by many factors. The toxic response to ionizing radiation, as well as to numerous chemical
contaminants at DOE sites, is associated with the generation of an oxidative stress. However, the cellular and molecular
mechanisms underlying the carcinogenic response to reactive oxygen species is not known.
Project Description
The toxic response to ionizing radiation, as well as
numerous contaminants identified on DOE sites, is
consistently associated with the generation of an oxidative
stress. Thus, defining the effects of oxidative stress in
mammalian systems is of central interest to DOE.
Oxidative stress is known to play a role in physiologic
and pathophysiologic processes such as cancer, aging, and
neuronal degeneration. It is not clear whether low-level
environmental exposure to radiation sources, or toxicants
whose mechanism of action includes an oxidative stress
component, will increase the risk of cancer.
In the present study, we have investigated whether
prototypical isoprostanes modulate anchorage-dependent
and -independent growth of several in vitro model
systems as an index of tumor promoting activity. We
have found that the isoprostanes modulate anchoragedependent
and -independent growth at extremely low
concentrations (pM-nM), suggesting the biological
activity of these compounds is physiologically relevant.
Introduction
Oxygen free radicals have been implicated in the
pathophysiology of a number of human diseases,
including cancer, atherosclerosis, neurodegenerative
disorders, and aging. Oxidative stress is also central to
the adverse effects of ionizing radiation. Recent
experimental evidence indicates that arachidonic acid may
be converted by reactive oxygen species into a new class
of oxidized lipid metabolites, termed the isoprostanes. In
several model systems, the isoprostanes have been shown
to be biologically active and coupled to signal
transduction pathways associated with carcinogenic
processes. Therefore, it will be important to determine
whether the formation of isoprostanes in response to
ionizing radiation and other environmental associated
toxic insults may increase cancer risk.
The isoprostanes have been shown to signal through the
inositol phospholipid/protein kinase C pathway, which in
turn, is widely implicated in carcinogenic processes.
Thus, the isoprostanes may represent a novel class of
physiologically relevant tumor promoters.
To test this hypothesis, we conducted preliminary studies
to determine whether isoprostanes exhibit tumor
promoting activity using an in vitro model of tumor
promotion (JB6 cell). In addition, we have investigated
whether prototypical isoprostanes modulate anchoragedependent
and -independent growth of several in vitro
model systems as an index of tumor promoting activity.
We have found that the isoprostanes modulate anchoragedependent
and -independent growth at extremely low
concentrations (pM-nM), suggesting the biological
activity of these compounds is physiologically relevant.
Results and Accomplishments
We previously showed that 8-iso-PGF2α increased the
anchorage-independent growth of JB6 cells. We have
extended our investigation to several other in vitro model
systems and a second prototypical isoprostane (8-iso-
PGE2) to determine whether the biological activity of the
isoprostanes was generalizable. Increasing experimental
evidence indicates that vascular smooth muscle cells are
responsive to isoprostanes, and the biological response
isoprostanes may be mediated by a thromboxane A2
(TXA2) receptor (TP), or a putative novel isoprostane
receptor. Studies were conducted to determine whether
8-iso-PGF2α and DDM-PGE2 modulate the proliferation
of smooth muscle cells in vitro. Rat aortic smooth muscle
cells were treated with a range of prostanoid
concentrations and cell proliferation determined using a
picogreen DNA quantitation assay and multiwell
fluorescent plate reader (Cytofluor 4000) according to
manufacturer’s directions. DNA synthesis was increased
in a concentration-dependent fashion by 8-iso-PGF2α, but
Biosciences and Biotechnology 81