Novel findings obtained with the colony formation assay of normal human cells

Hamada, Nobuyuki*; Sakashita, Tetsuya; Hara, Takamitsu*; Fujimichi, Yuki*

Hoshasen Seibutsu Kenkyu, 49(3), p.318 - 331, 2014/09

Over the past half century, a colony formation assay has been most commonly used to evaluate cellular radiosensitivity. Our colony formation assay of primary normal human diploid fibroblasts and the analysis of the size and cell numbers in abortive colonies comprising 50 cells and clonogenic colonies comprising 50 cells led to observations that can potentially answer a fundamental question of what is actually being evaluated in the colony formation assay, and a new phenomenon was also unveiled with the colony formation assay whereby the higher the dose the larger the size of colonies arising from irradiated primary normal human diploid ocular lens epithelial cells. This paper shall review these findings.

A Framework for analysis of abortive colony size distributions using a model of branching processes in irradiated normal human fibroblasts

Sakashita, Tetsuya; Hamada, Nobuyuki*; Kawaguchi, Isao*; Ouchi, Noriyuki; Hara, Takamitsu*; Kobayashi, Yasuhiko; Saito, Kimiaki

PLOS ONE (Internet), 8(7), p.e70291_1 - e70291_10, 2013/07

https://doi.org/10.1371/journal.pone.0070291

Clonogenicity gives important information about the cellular reproductive potential following ionizing irradiation. We firstly plotted the experimentally determined colony size distribution of abortive colonies in irradiated normal human fibroblasts, and found the linear relationship on the log-log plot. By applying the simple model of branching processes to the linear relationship, we found the persistent reproductive cell death (RCD) over several generations following irradiation. To verify the estimated probability of RCD, abortive colony size distribution (15 cells/colony) and the surviving fraction were simulated by the Monte Carlo computational approach. Radiation-induced RCD (i.e. excess probability) lasted over 16 generations and mainly consisted of two components in the early and late phases. We found that short-term RCD is critical to the abortive colony size distribution, and long-lasting RDC is important for the dose response of the surviving fraction.

The Small-molecule Bcl-2 inhibitor HA14-1 sensitizes cervical cancer cells, but not normal fibroblasts, to heavy-ion radiation

Hamada, Nobuyuki*; Kataoka, Keiko*; Sora, Sakura*; Hara, Takamitsu*; Omura, Motoko*; Funayama, Tomoo; Sakashita, Tetsuya; Nakano, Takashi*; Kobayashi, Yasuhiko

Radiotherapy and Oncology, 89(2), p.227 - 230, 2008/11

https://doi.org/10.1016/j.radonc.2008.08.006

Exposure of normal human fibroblasts to heavy-ion radiation promotes their morphological differentiation

Sora, Sakura*; Hamada, Nobuyuki*; Hara, Takamitsu*; Funayama, Tomoo; Sakashita, Tetsuya; Yokota, Yuichiro; Nakano, Takashi*; Kobayashi, Yasuhiko

Uchu Seibutsu Kagaku, 22(2), p.54 - 58, 2008/10

https://doi.org/10.2187/bss.22.54

Energetic heavy ions accelerate differentiation in the descendants of irradiated normal human diploid fibroblasts

Hamada, Nobuyuki*; Hara, Takamitsu*; Funayama, Tomoo; Sakashita, Tetsuya; Kobayashi, Yasuhiko

Mutation Research; Fundamental and Molecular Mechanisms of Mutagenesis, 637(1-2), p.190 - 196, 2008/01

https://doi.org/10.1016/j.mrfmmm.2007.07.002

Dependence of ionizing radiation-induced genomic instability on the linear energy transfer (LET) of the radiation is incompletely characterized; however, our previous work has shown that delayed reductions in clonogenicity can be most pronounced at LET of 108 keV/m. To gain insight into potential cellular mechanisms involved in LET-dependent delayed loss of clonogenicity, we investigated morphological changes in colonies arising from normal human diploid fibroblasts exposed to -rays or energetic carbon ions (108 keV/m). Exposure of confluent cultures to carbon ions was 4-fold more effective at inactivating cellular clonogenic potential and produced more abortive colonies containing reduced number of cells per colony than -rays. Second, colonies were assessed for clonal morphotypic heterogeneity. The yield of differentiated cells was elevated in a dose- and LET-dependent fashion in clonogenic colonies, whereas differentiated cells predominated to a comparable extent irrespective of radiation type or dose in abortive colonies.

Intercellular and intracellular signaling pathways mediating ionizing radiation-induced bystander effects

Hamada, Nobuyuki*; Matsumoto, Hideki*; Hara, Takamitsu*; Kobayashi, Yasuhiko

Journal of Radiation Research, 48(2), p.87 - 95, 2007/03

https://doi.org/10.1269/jrr.06084

Experimental evidence has accumulated demonstrating that biological effects of ionizing radiation arise in non-irradiated bystander cells that have received signals from near or distant irradiated cells. This phenomenon has been termed the radiation-induced bystander effect and challenges the long-held tenet that cell nucleus is the quintessential target for radiation damage. Bystander effects encompass a range of endpoints, such as genetic and epigenetic alterations, changes in gene expression, signal transduction pathway activation, and late-arising effects in their descendants. Proposed mechanisms involve gap junctional cell-to-cell communication, transmissible soluble factors, oxidative stress, lipid rafts, and calcium fluxes. In this article, we review proposed mechanisms underpinning the bystander effect and also briefly discuss the potential benefit of bystander effects in cancer radiotherapy.

LET dependence of radiation-induced delayed reproductive death in normal human diploid cells

Hamada, Nobuyuki*; Hara, Takamitsu*; Funayama, Tomoo; Sakashita, Tetsuya; Wada, Seiichi*; Kakizaki, Takehiko; Suzuki, Michiyo; Fukamoto, Kana; Kobayashi, Yasuhiko

no journal, , 

no abstracts in English

Bystander effect studies using heavy-ion microbeam

Kobayashi, Yasuhiko; Funayama, Tomoo; Sakashita, Tetsuya; Furusawa, Yoshiya*; Wada, Seiichi*; Hamada, Nobuyuki*; Yokota, Yuichiro; Kakizaki, Takehiko; Hara, Takamitsu*; Fukamoto, Kana; et al.

no journal, , 

no abstracts in English

Simulation of stress responses in by using a neuronal circuit model of chemotaxis

Suzuki, Michiyo; Sakashita, Tetsuya; Tsuji, Toshio*; Fukamoto, Kana; Hamada, Nobuyuki*; Wada, Seiichi*; Hara, Takamitsu*; Kakizaki, Takehiko; Kobayashi, Yasuhiko

no journal, , 

no abstracts in English

LET dependence of late-arising loss of clonogenicity

Hamada, Nobuyuki*; Hara, Takamitsu*; Sakashita, Tetsuya; Wada, Seiichi*; Kakizaki, Takehiko; Suzuki, Michiyo; Funayama, Tomoo; Kobayashi, Yasuhiko

no journal, , 

no abstracts in English