Establishment of mitochondrial DNA depleted A549 cell line and the change of its radiosensitivity / 中华放射医学与防护杂志

Objective To establish the mitochondrial DNA depleted cell line (ρ0 cells) of lung cancer cell A549 and to observe the radiosensitivity difference between ρ0 cells and normal A549 cells (ρ + cells).Methods The ρ0 cells were depleted of mitochondrial DNA by culturing chronically in the presence of low concentration of ethidium bromide (EB),and then the cell model was confined.Radiosensitivity of both ρ0 cells and ρ + cells was detected using the cologenic formation assay.After 6 MV X-ray irradiation in vitro,cell cycle distribution and reactive oxygen species (ROS) level were analyzed by flow cytometry and fluorescence microplate reader,respectively.Results A ρ0 cell line was successfully established and had a lower radiosensitivity than ρ + cells (t =12.57,P ＜ 0.01).After irradiation with a dose of 8 Gy,compared to ρ+ cells,ρ0 cells showed prolonged G2 arrest with less cells in G2 (t =6.82,P ＜ 0.01) and had lower increase of ROS level (t =14.51,P ＜ 0.01).Conclusions ρ0 cells have a lower radiosensitivity than ρ + cells,in which the reduction of ROS production and the prolongation of G2 arrest post-irradiation may be involved.

Prolonged expression of senescence markers in mice exposed to gamma-irradiation

Although ionizing radiation is known to induce cellular senescence in vitro and in vivo, its long-term in vivo effects are not well defined. In this study, we examined the prolonged expression of senescence markers in mice irradiated with single or fractionated doses. C57BL/6 female mice were exposed to 5 Gy of gamma-rays in single or 5, 10, 25 fractions. At 2, 4, and 6 months after irradiation, senescence markers including mitochondrial DNA (mtDNA) common deletion, p21, and senescence-associated beta-galactosidase (SA beta-gal) were monitored in the lung, liver, and kidney. Increases of mtDNA deletion were detected in the lung, liver, and kidney of irradiated groups. p21 expression and SA beta-gal staining were also increased in the irradiated groups compared to the non-irradiated control group. Increases of senescence markers persisted up to 6 months after irradiation. Additionally, the extent of mtDNA deletion and the numbers of SA beta-gal positive cells were greater as the number of radiation fractions increased. In conclusion, our results showed that ionizing radiation, especially that delivered in fractions, can cause the persistent upregulation of senescence marker expression in vivo. This should be considered when dealing with chronic normal tissue injuries caused by radiation therapy or radiation accidents.

Interrelationship of aging and mitochondrial DNA deletion in luteinized granulosa cells / 대한산부인과학회지

OBJECTIVE: Many clinical trials have proven the close relationship between the loss of human mitochondrial DNA and aging process. The purpose of this study was to evaluate the different types of mitochondrial DNA deletion and its frequency in luteinized granulosa cells in different aged groups of women undergoing in vitro fertilization (IVF). METHODS: The ovum pick up was done in 51 women who participated in Konyang University IVF program, and mitochondrial DNAs extracted from luteinized granulosa cells, were screened to search for different types of deletion and its frequency. The deleted mitochondrial DNA were analyzed by polymerase chain reaction method. DNA sequencing was performed to reveal exact deletion point. RESULTS: Three different types of deletions (4,977 bp, 7,150 bp, and 5,777 bp) were confirmed. To find the difference between the aged groups, we have divided women into groups younger than 32 years, between 32 to 37 years, and older than 37 years. The deletion of 4,977 bp was 60.9% (14/23) in younger than 32 years, 46.2% (6/13) in 32 to 37 years, 46.7% (7/15) in older than 37 years. There was no statistical significance between aged groups and the incidence of the deletion. The deletion of 7150 bp was 34.8% (8/23), in younger than 32 years, 30.8% (4/13) in 32 to 37 years, 40% (6/15) in older than 37 years. We investigated relationship between the frequency of deletion and the aging, but there was no statistical significance. In case of 5,777 bp, the deletion was 43.5% (10/23) in younger than 32 years, 30.8% (4/13) in 32 to 37 years, 53.3% (8/15) in older than 37 years. It showed no statistical significance as well as other types. CONCLUSION: In this study we have found three different types of deletion of mitochondrial DNA obtained from luteinized granulosa cells in women with infertility. There were no significant differnces of each type of deletion in 3 different aged groups of infantile women. The limitation of this study is that the analyze were done qualitatively. If we could provide the quantitative analyze it could be applied clinically.

Effect of Sijunzi Decoction on Mitochondrial DNA Missing of the Aging Model Mice / 中药新药与临床药理

Objective To observe the effect of Sijunzi Decoction (SD) on the mitochondrial DNA deletion of the aging model mice, and to explore its anti-aging biomolecular mechanism. Methods Kunming mice were divided into 6 groups according to the body weight: normal control group, .model control group, high-, middle-and low-dose SD groups (at the dose of 20, 10, 5 g·kg-1 respectively), and vitamin E group (250 mg·kg-1), 20 mice in each group.The mice models of aging were induced by D-galactose. Water maze test was used to evaluate the learning and memory ability of the aging model mice. The effect of SD on the mitochondrial DNA deletion of the aging model mice was ob-served. Results In the hippocampal mtDNA of model control group and low-dose SD group, there were about 3.87 kb fragment missing. But there was no missing pieces in high-and middle-dose SD groups. Conclusion The anti-aging biomolecular mechanism of SD is probably related to the preventive effect on the mutation and deletion of the mitochondrial DNA in the aging model mice.