Yuko Noda

Rag-dependent and Rag-independent mechanisms of Notch1 rearrangement in thymic lymphomas of Atm(-/-) and scid mice

The pathways of thymic lymphomagenesis are classified as Rag-dependent or -independent according to their dependence on recombination-activating gene (Rag1/2) proteins. The role of the two-lymphoma pathways in oncogene rearrangements and the connection between lymphoma pathways and rearrangement mechanisms, however, remain obscure. We compared the incidence and latency of thymic lymphomas, and associated rearrangements of the representative oncogene Notch1 among Rag2(-/-), ataxia telangiectasia mutated (Atm)(-/-), and severe combined immune deficiency (scid) mice combined with Rag2 deficiency. Contrary to expectations, Rag2(-/-) mice were prone to thymic lymphoma development, suggesting the existence of a Rag2-independent lymphoma pathway in Rag2(-/-) mice. The lymphoma incidence in Rag2(-/-)Atm(-/-) mice was lower than that in Atm(-/-) mice, but higher than that in Rag2(-/-) mice, indicating that Atm(-/-) mice develop lymphomas through both pathways. Scid mice developed lymphomas with an incidence and latency similar to Rag2(-/-)scid mice, suggesting that Rag2-mediated V(D)J recombination-driven events are not necessarily required for lymphomagenesis in scid mice. Notch1 rearrangement mechanisms were classified as Rag2-dependent or Rag2-independent based on the presence of recombination signal-like sequences at rearranged sites. In Rag2(-/-) lymphomas, Notch1 must be rearranged independently of Rag2 function, implying that Rag2(-/-) mice are susceptible to lymphomagenesis due to the presence of other rearrangement mechanisms. The results in Atm(-/-) mice suggest that Notch1 was rearranged through both lymphoma pathways. In scid mice, the frequency of Rag2-mediated rearrangements was relatively low compared with that in wild-type mice, suggesting that the Rag2-independent lymphoma pathway prevails in the development of thymic lymphomas in scid mice. Thus, two rearrangement mechanisms underlie the lymphoma pathways and constitute the mechanistic bases for lymphomagenesis, thereby providing the molecular criteria for distinguishing between Rag2-dependent and Rag2-independent lymphoma pathways.

Selective down‐regulation of Th2 cell‐mediated airway inflammation in mice by pharmacological intervention of CCR4

The chemokine receptor CCR4 has been implicated in Th2 cell‐mediated immune responses. However, other T cell subsets are also known to participate in allergic inflammation.

Effect of Atm Disruption on Spontaneously Arising and Radiation-Induced Deletion Mutations in Mouse Liver

Abstract Furuno-Fukushi, I., Masumura, K., Furuse, T., Noda, Y., Takahagi, M., Saito, T., Hoki, Y., Suzuki, H., Wynshaw-Boris, A., Nohmi, T. and Tatsumi, K. Effect of Atm Disruption on Spontaneously Arising and Radiation-Induced Deletion Mutations in Mouse Liver. Radiat. Res. 160, 549–558 (2003). Deletion mutations were efficiently recovered in mouse liver after total-body irradiation with X rays by using a transgenic mouse “gpt-delta” system that harbored a lambda EG10 shuttle vector with the red and gam genes for Spi− (sensitive to P2 lysogen interference) selection. We incorporated this system into homozygous Atm-knockout mice as a model of the radiosensitive hereditary disease ataxia telangiectasia (AT). Lambda phages recovered from the livers of X-irradiated mice with the Atm+/+ genotype showed a dose-dependent increase in the Spi− mutant frequency up to sixfold at 50 Gy over the unirradiated control of 2.8 × 10−6. The livers from Atm−/− mice yielded a virtually identical dose–response curve for X rays with a background fraction of 2.4 × 10−6. Structural analyses revealed no significant difference in the proportion of −1 frameshifts and larger deletions between Atm+/+ and Atm−/− mice, although larger deletions prevailed in X-ray-induced Spi− mutants irrespective of Atm status. While a possible defect in DNA repair after irradiation has been strongly indicated in the literature for nondividing cultured cells in vitro from AT patients, the Atm disruption does not significantly affect radiation mutagenesis in the stationary mouse liver in vivo.

Comparison of dose-dependent enhancing effects of γ-ray irradiation on urethan-induced lung tumorigenesis in athymic nude (nu/nu) mice and euthymic (nu/+) littermates

The role of immunological surveillance in carcinogenesis is still controversial. In our previous experiments, urethan-induced lung tumorigenesis in athymic (nu/nu) mice and euthymic (nu/+) littermates was examined, and it was concluded that immunosurveillance mediated by T cells could not be demonstrated. However, the reported enhancement of development of various tumors following ionizing radiation might be achieved through modulating the host immunological conditions. In the present experiment,nu/nu and littermatenu/+ mice were treated with 1–4 Gy γ-rays alone at 6 weeks of age or treated with urethan at 0.5 mg/g body weight when aged 14 days followed by 1–4 Gy γ-rays 4 weeks later. Lung tumors were assessed at 6.5 months of age. Ionizing radiation itself caused a very low incidence of these lesions. On the other hand, multiplicities and incidences of lung tumors after urethan treatment at 0.5 mg/g body weight were similar between the two phenotypically different groups of mice (1.66 and 1.84 tumors/mouse, 73% and 80% incidences, fornu/nu andnu/+ cases respectively). This urethan-induced lung tumorigenesis was significantly enhanced by γ-rays in bothnu/nu andnu/+ mice, and the magnitude of tumor enhancement was somewhat higher innu/+ mice than innu/nu mice, especially with a 2-Gy dose. In conclusion, it may be said that lung tumorigenicity of γ-ray irradiation itself and the enhancing effect of radiation on urethan-induced tumorigenesis are scarcely influenced by immunosurveillance mechanisms mediated by T cells

Mutant mouse p53 transgene elevates the chemical induction of tumors that respond to gene silencing with siRNA

To study the role of mutant p53 in the induction and cure of tumors, we generated transgenic mice carrying mutant p53 (mp53) containing a 9 bp deletion in exon 6 in addition to wild-type p53, expressing both p53 and mp53. The mp53 cDNA was cloned from a radiation-induced mouse tumor and ligated to the chicken β-actin promoter/CMV-IE enhancer in the expression vector. The presence of mp53 suppressed p21 expression in primary fibroblasts after ionizing irradiation, indicating the dominant-negative activity of mp53 in the mice. These mice developed fibrosarcomas after the subcutaneous injection of 3-methylcholanthrene with an incidence 1.7-fold higher than that of wild-type mice (42% excess). The tumors were then treated via a potent atelocollagen delivery system with small interfering RNA (siRNA), that targeted the promoter/enhancer of the expression vector, resulting in the suppression of tumor growth in 30% of 44 autochthonous tumors, including four cures, and their transplants, the total fraction corresponding to the tumor excess. This suppressive effect involved the induction of apoptosis. These results indicate that mp53 activity causes tumors that can be suppressed by subsequent silencing of mp53 in the presence of wild-type p53 alleles.

Induction of myeloid leukemias in C3H/He mice with low-dose rate irradiation

Abstract C3H/He male mice were exposed to whole body gamma-ray irradiation at 8 weeks of age. Radiation at a high-dose rate of 88.2 cGyears/min with doses of 0.125–5.0 Gyears, a medium dose rate of 9.56 cGyears/min with doses of 1.0–5.0 Gyears, and low-dose rates of 0.0298 cGyear/min with doses of 1.0–10 Gyears, 0.0067 cGyear/min with doses of 1.0–10.0 Gyears or 0.0016 cGyear/min with doses of 1.0–4.0 Gyears were delivered from 137 Cs sources. The mice in the low-dose rate groups were irradiated continuously for 22 h daily during a period of 3 to 200 days. All the mice were maintained for their entire life span and were pathologically examined after their death. Myeloid leukemia developed significantly more frequently in the irradiated groups with doses over 1 Gyear than in the unirradiated groups. The maximum values were 23.5% in the high-dose rate group, 11% in the medium dose rate group, 7.3%, 7.2%, 6.3% in the low-dose rate groups, and 0.99% in the unirradiated control group. These dose–effect curves had their highest values on each curve at about 3 Gyears. We obtained the dose and dose rate effectiveness factor (DDREF) values of 2.96 by linear fittings for their dose–response curves of the dose ranges in which leukemia incidences were increasing.