Page 171 - 2014 Printable Abstract Book
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of the NRAS G12D oncogene and deletion of p53. Deletion of HIF-1α sensitized primary sarcomas to RT in
vivo. Moreover, cell lines derived from the primary sarcomas lacking HIF-1α had decreased clonogenic
survival in vitro, demonstrating that HIF-1α promotes radiation resistance in a cell autonomous manner.
In order to determine changes in HIF-1α-regulated gene expression that contribute to cell-autonomous
radiation resistance, we performed sequencing on ribosomal-depleted RNA from primary sarcoma cell
lines with or without HIF-1α at baseline, under hypoxia, and at 8h, 24h, 48h, and 96 hours after 6Gy
irradiation. Although hypoxia induced classical HIF-1α targets, radiation induces other HIF-1α dependent
pathways involving cellular inflammation and immunity. These results implicate HIF-1α as a master
regulator of the cellular immune response to radiation that leads to radiation resistance.
1
(PS2-62) Transgenic mouse model for reducing oxidative damage in bone. Ann-Sofie Schreurs ;
1
1
1
1
1
2
Samantha Torres ; Tiffany Truong ; Akhilesh Kumar ; Josh Alwood ; Charlie Limoli ; and Ruth Globus
2
1
NASA ARC, Mountain View, CA and UC Irvine, Irvine, CA
Exposure to musculoskeletal disuse and radiation result in bone loss; we hypothesized that these
catabolic treatments cause excess reactive oxygen species (ROS), and thereby alter the tight balance
between bone resorption by osteoclasts and bone formation by osteoblasts, culminating in bone loss. To
test this, we used transgenic mice which over-express the human gene for catalase, targeted to
mitochondria (MCAT). Catalase is an anti-oxidant that converts the ROS hydrogen peroxide into water and
oxygen. MCAT mice were shown previously to display reduced mitochondrial oxidative stress and
radiosensitivity of the CNS compared to wild type controls (WT). As expected, MCAT mice expressed the
transgene in skeletal tissue, and in marrow-derived osteoblasts and osteoclast precursors cultured ex vivo,
and also showed greater catalase activity compared to wildtype (WT) mice (3-6 fold). Colony expansion in
marrow cells cultured under osteoblastogenic conditions was 2-fold greater in the MCAT mice compared
to WT mice, while the extent of mineralization was unaffected. MCAT mice had slightly longer tibiae than
WT mice (2%, P<0.01), although cortical bone area was slightly lower in MCAT mice than WT mice (10%,
p=0.09). To challenge the skeletal system, mice were treated by exposure to combined disuse (2 wk
Hindlimb Unloading) and total body irradiation 137 Cs (2 Gy, 0.8 Gy/min), then bone parameters were
analyzed by 2-factor ANOVA to detect possible interaction effects. Treatment caused a 2-fold increase
(p=0.015) in malondialdehyde levels of bone tissue (ELISA) in WT mice, but had no effect in MCAT mice.
These findings indicate that the transgene conferred protection from oxidative damage caused by
treatment. Unexpected differences between WT and MCAT mice emerged in skeletal responses to
treatment. In WT mice, treatment did not alter osteoblastogenesis, cortical bone area, moment of inertia,
or bone perimeter, whereas in MCAT mice, treatment increased these parameters. Taken together, this
typically catabolic treatment (disuse and irradiation) appeared to stimulate cortical expansion in MCAT
mice but not WT mice. In conclusion, these results reveal the importance of mitochondrial ROS generation
in skeletal remodeling and show that MCAT mice provide a useful animal model for bone studies.
169 | P a g e
vivo. Moreover, cell lines derived from the primary sarcomas lacking HIF-1α had decreased clonogenic
survival in vitro, demonstrating that HIF-1α promotes radiation resistance in a cell autonomous manner.
In order to determine changes in HIF-1α-regulated gene expression that contribute to cell-autonomous
radiation resistance, we performed sequencing on ribosomal-depleted RNA from primary sarcoma cell
lines with or without HIF-1α at baseline, under hypoxia, and at 8h, 24h, 48h, and 96 hours after 6Gy
irradiation. Although hypoxia induced classical HIF-1α targets, radiation induces other HIF-1α dependent
pathways involving cellular inflammation and immunity. These results implicate HIF-1α as a master
regulator of the cellular immune response to radiation that leads to radiation resistance.
1
(PS2-62) Transgenic mouse model for reducing oxidative damage in bone. Ann-Sofie Schreurs ;
1
1
1
1
1
2
Samantha Torres ; Tiffany Truong ; Akhilesh Kumar ; Josh Alwood ; Charlie Limoli ; and Ruth Globus
2
1
NASA ARC, Mountain View, CA and UC Irvine, Irvine, CA
Exposure to musculoskeletal disuse and radiation result in bone loss; we hypothesized that these
catabolic treatments cause excess reactive oxygen species (ROS), and thereby alter the tight balance
between bone resorption by osteoclasts and bone formation by osteoblasts, culminating in bone loss. To
test this, we used transgenic mice which over-express the human gene for catalase, targeted to
mitochondria (MCAT). Catalase is an anti-oxidant that converts the ROS hydrogen peroxide into water and
oxygen. MCAT mice were shown previously to display reduced mitochondrial oxidative stress and
radiosensitivity of the CNS compared to wild type controls (WT). As expected, MCAT mice expressed the
transgene in skeletal tissue, and in marrow-derived osteoblasts and osteoclast precursors cultured ex vivo,
and also showed greater catalase activity compared to wildtype (WT) mice (3-6 fold). Colony expansion in
marrow cells cultured under osteoblastogenic conditions was 2-fold greater in the MCAT mice compared
to WT mice, while the extent of mineralization was unaffected. MCAT mice had slightly longer tibiae than
WT mice (2%, P<0.01), although cortical bone area was slightly lower in MCAT mice than WT mice (10%,
p=0.09). To challenge the skeletal system, mice were treated by exposure to combined disuse (2 wk
Hindlimb Unloading) and total body irradiation 137 Cs (2 Gy, 0.8 Gy/min), then bone parameters were
analyzed by 2-factor ANOVA to detect possible interaction effects. Treatment caused a 2-fold increase
(p=0.015) in malondialdehyde levels of bone tissue (ELISA) in WT mice, but had no effect in MCAT mice.
These findings indicate that the transgene conferred protection from oxidative damage caused by
treatment. Unexpected differences between WT and MCAT mice emerged in skeletal responses to
treatment. In WT mice, treatment did not alter osteoblastogenesis, cortical bone area, moment of inertia,
or bone perimeter, whereas in MCAT mice, treatment increased these parameters. Taken together, this
typically catabolic treatment (disuse and irradiation) appeared to stimulate cortical expansion in MCAT
mice but not WT mice. In conclusion, these results reveal the importance of mitochondrial ROS generation
in skeletal remodeling and show that MCAT mice provide a useful animal model for bone studies.
169 | P a g e
