NOTE: End date is now 9/30/2015 per NSSC information (Ed., 7/11/14)

NOTE: End date is now 9/30/2014 per PI and NSSC information (Ed., 8/3/2013)

NOTE: End date is now 9/30/2013 per NSSC information (Ed., 3/12/2013)

NOTE: Extended to 3/31/2013 per NSSC information (Ed., 12/18/12)

ED. NOTE (7/7/15): Project continues as "Molecular Basis of DNA Repair and Protection from Apoptosis in Neuronal Progenitors Exposed to Space Radiation" grant NNX14AT39G, due to PI move to Prairie View A&M University in September 2014.

NOTE: End date is now 9/30/2014 per PI and NSSC information (Ed., 8/3/2013)

NOTE: End date is now 9/30/2013 per NSSC information (Ed., 3/12/2013)

NOTE: Extended to 3/31/2013 per NSSC information (Ed., 12/18/12)

Task progress: In this year, we performed the following studies:

1. Live imaging DNA damage process in neuronal cells following exposure to 0.1- 1Gy of 56Fe(1 GeV) and proton (1 GeV). We found that HZE particle induced persistent DNA damage in neuronal cells. This DNA damage signaling may transit to progeny cells.

2. Detection of effect of poly(ADP-ribose) polymerase 1 in DNA damage response in neuronal cells. We found that PARP-1 dependent DNA repair pathway is involved in clustered DNA damage repair in neuronal cells following exposure HZE particles.

3. Detection of the effect of glycogen synthase kinase 3 on radiation response in neuronal cells. We found that inhibition of GSK3 protect neuronal cells from space radiation.

4. Study on the effect of HZE on the processing of Amyloid precusor proteins in neuronal cells. 56Fe(1GeV) induces cleavage of APP c-terminal and translocation to nuclear in neuronal cells.

5. Study on High LET radiation produces sustained DNA damage signaling and changes cellular homeostasis in hippocampal neuronal cells. 1 Gy 56Fe(1GeV) induced persistent increased p53 expression and camp response element binding protein phosphorylation.

The results are described in published papers, and presented as abstracts in the following list and in several manuscripts.

Manuscripts supported by NNX08BA08G

1. Wang, T., Wang, M., Tian, L., Wang, H. PARP-1 is involved in clustered DNA damage in neuronal cells. Pols one revised

2. Ramachandiran, S. and Wang, H., Bernal-Mizrachi L. The noncanonical NF-kB pathway suppresses DNA damage, centrosome amplification and chromosomal aberrations in diffuse large cell lymphomas, Leukemia, Clinical Cancer Research, revising.

3. Wang, M., Tian L., Wang, T., Tang X. and Wang H. Function of GSK3 in radiation induced apopotosis in neural cells, in preparation.

4. Li Z, Hudson FZ, Wang H, Wang Y, Bian Z, Murnane JP, Dynan WS "Increased mutagenic joining of enzymatically-induced DNA double-strand breaks in high-charge and energy particle irradiated human cells" Radiat Res. 2013 Jul;180(1):17-24 PMID: 23692479 , May-2013

Abstracts in professional meeting

1. Wang T Kandimalla R, , Tang X, Wang H APP intracellular domain increases DNA damage response in hippocampal neuronal cells following exposure to high LET radiation Heavy ion in therapy and space radiation symposium , 24th Annual NASA Space Radiation Investigators' Workshop, Chiba, Japan, May 15-18 2013

2. Wang T, Kandimalla R, Tang X, Wang H High LET radiation produces sustained DNA damage signaling and changes cellular homeostasis in hippocampal neuronal cells Heavy ion in therapy and space radiation symposium , 24th Annual NASA Space Radiation Investigators' Workshop, Chiba, Japan, May 15-18 2013

3. Werner E, Kandimalla R, Wang H, and Doetsch PW, A role for reactive oxgen species in the resolution of persistent genomic instability after exposure to radiation Heavy ion in therapy and space radiation symposium , 24th Annual NASA Space Radiation Investigators' Workshop, Chiba, Japan, May 15-18 2013

4. Werner E, Kandimalla R, Wang H, and Doetsch PW. Early and persistent effect of PARP inhibition after exposure to radiation Heavy ion in therapy and space radiation symposium, 24th Annual NASA Space Radiation Investigators' Workshop, Chiba, Japan, May 15-18 2013

5. Li Z, Hudson FZ, Wang H, Murane JP, Dynan Ws. Secretoty protein phenotypes accompanied by mutagenic joining of enzymatically-induced DNA double-strand breaks in a population of HZE-exposed human cells. Heavy ion in therapy and space radiation symposium, 24th Annual NASA Space Radiation Investigators' Workshop, Chiba, Japan, May 15-18 2013

NOTE: Extended to 3/31/2013 per NSSC information (Ed., 12/18/12)

Current NASA Grant: NNX08BA08G PI: Dr. Huichen Wang Project Title: Funding period: 10/1/09 to 9/30/12

The health and performance risks to astronauts exposed to space radiation from galactic cosmic rays (GCR) and solar particle events (SPEs) during space missions are still uncertain. Acute and late radiation damage to the central nervous system (CNS) may lead to changes in motor function and behavior, or neurological disorders. Evidence of space radiation risk to the CNS has been accumulated and reported. However, the pathogenesis of space radiation-induced cognitive dysfunction remains largely uncharacterized. The neurodegenerative effects of space radiation are likely to be derived from DNA damage in the central nervous system (CNS). In this project we used an in vitro system based on cultures of mouse embryo neuronal progenitor cells (neurospheres) to study the function of PARP-1 in the induction of clustered DNA damage after low LET an high LET radiation as well as molecular basis of DNA repair. Space radiation induced clustered DNA damage including oxidized base damage, DNA single-strand breaks and double-strand breaks were detected by a modified single cell electrophoresis, and immunodetection of 8-oxo-dG, in neurospheres and PARP-1 proficient and deficient cells. We found that Poly(ADP-ribose) Polymerase 1 (PARP-1) inhibitor induce more DNA damage in neuronal progenitor cells following 56Fe particle and proton irradiation. PARP-1 inhibition also delayed the decay of gamma H2AX foci and the residual foci associated with new replicated DNA. The level of ATM (Ataxia Telangiectasia Mutated) phosphorylations was increased in neurospheres after treatment with PARP-1 inhibitor and following irradiation. Mutation of PARP-1 in MEF cells dramatically decrease DNA damage repair. We observed that PARP-1 and PAR formed a foci track and colocalized with phosphorylated ATM and MRE11. Cyclin-dependent kinase 5 (CDK5) is a proline-directed serine/threonine cyclin-dependent kinase in post-mitotic neurons and plays a critical role in neurogenesis. We found that the interaction of PARP-1 and CDK5 in radiation-induced DNA damage response in an in vitro system based on cultures of mouse embryo neuronal progenitor cells (neurospheres) from PARP-1 knockout mice and mouse hippocampal neuronal cells. CDK5 inhibitor (roscovitine) reversed the radiosensitivity of the PARP inhibitor in neural cells exposed to X-ray, proton and heavy ion radiation using a clonogenic survival assay. The PARP inhibitor increased radiation-induced DNA damage and apoptosis in neural cells. These effects were abrogated when the PARP inhibitor was combined with the CDK5 inhibitor. CDK5 PARP inhibitors increased radiation induced ATM expression and ATM phosphorylation in both cytoplasm and nucleus of neural cells. The GSK3 beta inhibitor also reversed radiosentitivity of neural cells. These suggest that the DNA repair pathway and the neuronal survival factors play a critical role in protecting neuronal cells following space radiation. The details of this mechanism and biological effectiveness will be discussed.

Conclusion

1. We described DNA repair pathways (PARP-1 and ATM) in neural cell survival following exposure to high LET radiation. High energy particles induced larger RBE in cell survival and DNA damage of hippocampal neurons compared to X-ray radiation. PARP and ATM deficiency increased the RBE of cell survival and DNA damage induced by high charge and energy particles.

Related publications:

Publications/submitted manuscripts supported by NNX08BA08G

1. Wang H., Liu S., et al. (2009). "S-phase cells are more sensitive to high-linear energy transfer radiation." Int J Radiat Oncol Biol Phys 74(4): 1236-1241.

2. Cucinotta F, Wang H, Huff JL. "Risk of Acute or Late Central Nervous System Effects from Radiation Exposure." Human Health and Performance Risks of Space Exploration Missions. ed. by J.C. McPhee and J.B. Charles. Houston, Texas. NASA, 2010. p. 191-212, NASA SP 2009

3. Shi Y. Zhang X., Tang X., Wang P., Wang H., and Wang Y.MiR-21 is Continually Elevated Long-Term in the Brain after Exposure to Ionizing Radiation, Radiation Research, 177(1):124-8, 2012

4. Zhang X., Wooi-Loon N., Wang P. Tian L., Werner E., Wang H. Doetsch PW., and Wang Y. MicroRNA-21 modulates reactive oxygen species levels via targeting SOD3 and TNFa, Cancer Research,

5. Ramachandiran, S. and Wang, H., Bernal-Mizrachi L. The noncanonical NF-gammaB pathway suppresses DNA damage, centrosome amplification and chromosomal aberrations in diffuse large cell lymphomas, Leukemia, Clinical Cancer Research, revising.

Manuscripts in preparation and in progression supported by NNX08BA08G

1. Wang, M., Tian, L., Wang, H. PARP-1 is involved in clustered DNA damage in neuronal cells. In preparation.

2. Wang, M., Tian L., Wang, T., Tang X. and Wang H. Function of GSK3 in radiation induced apopotosis in neural cells, in preparation.

3. Wang Y., Tang X. Wang M. and Wang H. Amentoflavone-mediated radioprotection of hippocampal neuronal cells through reduction of ROS and DNA damage, in preparation.

4. Homologous recombination mediates persistent clustered DNA damage induced by high LET radiation, in preparation.

PARP and ATM deficiency increased the RBE of cell survival and DNA damage induced by high charge and energy particles.

We found that Inhibition of cyclin dependent kinase 5 and glycogen synthase kinase 3 beta could reverse radiation sensitivity in neuronal cells and neurospheres, but enhance radiation sensitivity in glioblastoma cells.

A Summary of the results:

The health risks to astronauts exposed to space radiation include cognitive deficits and possibly accelerated aging. However, the pathogenesis of radiation-induced cognitive dysfunction remains largely uncharacterized. The neurodegenerative effects of space radiation are likely to derive from DNA damage in the central nervous system (CNS). PARP-1 catalyzes poly(ADP-ribosyl)ation on DNA and proteins as immediate response to DNA damage induced by ionizing radiation or following oxidative stress. . In this study, we used an in vitro system based on cultures of mouse embryo neuronal progenitor cells (neurospheres) to study the function of PARP-1 in the induction of clustered DNA damage after low LET an high LET radiation as well as molecular basis of DNA repair. Space radiation induced clustered DNA damage including oxidized base damage, DNA single-strand breaks and double-strand breaks were detected by a modified single cell electrophoresis, and immunodetection of 8-oxo-dG, in neurospheres and PARP-1 proficient and deficient cells. We found that Poly(ADP-ribose) Polymerase 1 (PARP-1) inhibitor induce more DNA damage in neuronal progenitor cells following 56Fe particle and proton irradiation. PARP-1 inhibition also delayed the decay of gamma-H2AX foci and the residual foci associated with new replicated DNA. The level of ATM (Ataxia Telangiectasia Mutated) phosphorylations was increased in neurospheres after treatment with PARP-1 inhibitor and following irradiation. Mutation of PARP-1 in MEF cells dramatically decrease DNA damage repair. These suggest that PARP-1 may play a role in protecting neuronal cells following space radiation. The detail of mechanism and biological effectiveness are under study.