[Show abstract][Hide abstract] ABSTRACT: Poly(ADP-ribose) glycohydrolase (PARG) is the primary enzyme responsible for the degradation of poly(ADP-ribose). PARG dysfunction sensitizes cells to alkylating agents and induces cell death; however, the details of this effect have not been fully elucidated. Here, we investigated the mechanism by which PARG deficiency leads to cell death in different cell types using methylmethanesulfonate (MMS), an alkylating agent, and Parg(-/-) mouse ES cells and human cancer cell lines. Parg(-/-) mouse ES cells showed increased levels of γ-H2AX, a marker of DNA double strand breaks (DSBs), accumulation of poly(ADP-ribose), p53 network activation, and S-phase arrest. Early apoptosis was enhanced in Parg(-/-) mouse ES cells. Parg(-/-) ES cells predominantly underwent caspase-dependent apoptosis. PARG was then knocked down in a p53-defective cell line, MIAPaCa2 cells, a human pancreatic cancer cell line. MIAPaCa2 cells were sensitized to MMS by PARG knockdown. Enhanced necrotic cell death was induced in MIAPaCa2 cells after augmenting γ-H2AX levels and S-phase arrest. Taken together, these data suggest that DSB repair defect causing S-phase arrest, but p53 status was not important for sensitization to alkylation DNA damage by PARG dysfunction, whereas the cell death pathway is dependent on the cell type. This study demonstrates that functional inhibition of PARG may be useful for sensitizing at least particular cancer cells to alkylating agents.
[Show abstract][Hide abstract] ABSTRACT: Poly(ADP-ribose) glycohydrolase (Parg) is the main enzyme involved in poly(ADP-ribose) degradation. Here, the effects of Parg deficiency on sensitivity to low and high linear-energy-transfer (LET) radiation were investigated in mouse embryonic stem (ES) cells. Mouse Parg(-/-) and poly(ADP-ribose) polymerase-1 deficient (Parp-1(-/-)) ES cells were used and responses to low and high LET radiation were assessed by clonogenic survival and biochemical and biological analysis methods. Parg(-/-) cells were more sensitive to γ-irradiation than Parp-1(-/-) cells. Transient accumulation of poly(ADP-ribose) was enhanced in Parg(-/-) cells. Augmented levels of phosphorylated H2AX (γ-H2AX) from early phase were observed in Parg(-/-) ES cells. The induction level of p53 phophorylation at ser18 was similar in wild-type and Parp-1(-/-) cells and apoptotic cell death process was mainly observed in the both genotypes. These results suggested that the enhanced sensitivity of Parg(-/-) ES cells to γ-irradiation involved defective repair of DNA double strand breaks. The effects of Parg and Parp-1 deficiency on the ES cell response to carbon-ion irradiation (LET13 keV/μm and 70 keV/μm) and Fe-ion irradiation (200 keV/μm) were also examined. Parg(-/-) cells were more sensitive to LET 70 keV/μm carbon-ion irradiation than Parp-1(-/-) cells. Enhanced apoptotic cell death also accompanied augmented levels of γ-H2AX in a biphasic manner peaked at 1 and 24 hrs. The induction level of p53 phophorylation at ser18 was not different between wild-type and Parg(-/-) cells. The augmented level of poly(ADP-ribose) accumulation was noted after carbon-ion irradiation compared to γ-irradiation even in the wild-type cells. An enhanced poly(ADP-ribose) accumulation was further observed in Parg(-/-) cells. Both Parg(-/-) cells and Parp-1(-/-) cells did not show sensitization to Fe-ion irradiation. Parg deficiency sensitizes mouse ES cells to a wide therapeutic range of LET radiation through the effects on DNA double strand break repair responses and enhanced cell death.
Biochemical and Biophysical Research Communications 04/2013; 435(1). DOI:10.1016/j.bbrc.2013.04.048 · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Elucidation of the relationship between poly-ADP-ribosylation and carcinogenesis has markedly progressed by the recent development
of knockout or transgenic mice models of poly(ADP-ribose) polymerase (Parp)-1, Parp-2, and poly(ADP-ribose) glycohydrolase
(Parg). Parp-1 is involved in base excision repair (BER), single- and |double-strand break repair, and chromosomal stability.
These multiple functions explain why Parp-1 deficiency enhances carcinogenesis induced by alkylating agents and that in aged animals. Parp-1 is also involved in transcriptional
regulation through protein-protein interaction as a coactivator and/or poly-ADP-ribosylation reaction and is possibly involved
in epigenetic alteration during carcinogenesis and modulation of tumor phenotypes. Parp-1-dependent cell-death accompanying
NAD depletion may be another important issue in carcinogenesis because this process could lead to the selection of Parp-1 deficient cells due to their survival advantage during cancer growth. The relationship of Parp-2, Parp-3, tankyrase and Parg
with carcinogenesis is also discussed.
[Show abstract][Hide abstract] ABSTRACT: Many lines of evidence suggest that poly(ADP-ribose) polymerase-1 (Parp-1) is involved in transcriptional regulation of various genes as a coactivator or a corepressor by modulating chromatin structure. However, the impact of Parp-1-deficiency on the regulation of genome-wide gene expression has not been fully studied yet.
We employed a microarray analysis covering 12,488 genes and ESTs using mouse Parp-1-deficient (Parp-1-/-) embryonic stem (ES) cell lines and the livers of Parp-1-/- mice and their wild-type (Parp-1+/+) counterparts. Here, we demonstrate that of the 9,907 genes analyzed, in Parp-1-/- ES cells, 9.6% showed altered gene expression. Of these, 6.3% and 3.3% of the genes were down- or up-regulated by 2-fold or greater, respectively, compared with Parp-1+/+ ES cells (p < 0.05). In the livers of Parp-1-/- mice, of the 12,353 genes that were analyzed, 2.0% or 1.3% were down- and up-regulated, respectively (p < 0.05). Notably, the number of down-regulated genes was higher in both ES cells and livers, than that of the up-regulated genes. The genes that showed altered expression in ES cells or in the livers are ascribed to various cellular processes, including metabolism, signal transduction, cell cycle control and transcription. We also observed expression of the genes involved in the pathway of extraembryonic tissue development is augmented in Parp-1-/- ES cells, including H19. After withdrawal of leukemia inhibitory factor, expression of H19 as well as other trophoblast marker genes were further up-regulated in Parp-1-/- ES cells compared to Parp-1+/+ ES cells.
These results suggest that Parp-1 is required to maintain transcriptional regulation of a wide variety of genes on a genome-wide scale. The gene expression profiles in Parp-1-deficient cells may be useful to delineate the functional role of Parp-1 in epigenetic regulation of the genomes involved in various biological phenomena.
[Show abstract][Hide abstract] ABSTRACT: The impact of poly(ADP-ribose) polymerase-1 (Parp-1)-deficiency on 4-nitroquinoline 1-oxide (4NQO)-induced carcinogenesis was studied in mice with an ICR/129Sv mixed genetic background. Parp-1(+/+), Parp-1(+/-) and Parp-1(-/-) animals given 4NQO for thirty-two weeks at 0.001% in their drinking water developed papillomas and squamous cell carcinomas of the tongue, palate and esophagus, but with no statistically significant variation with the Parp-1 genotype. Thus Parp-1 deficiency does not elevate susceptibility to carcinogenesis induced by a carcinogen which gives rise to bulky DNA lesions. This study also indicated that the ICR/129Sv mixed genetic background is associated with high yield induction of esophageal tumors by 4NQO.
Cancer Letters 10/2006; 241(1):87-92. DOI:10.1016/j.canlet.2005.10.003 · 5.62 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Poly(ADP-ribose) polymerase-1 (Parp-1) localizes mainly in the nucleus and functions in DNA repair, genome stability and cell death regulation. Meanwhile, it also localizes in centrosomes and is involved in the regulation of centrosome duplication. An abnormal increase in centrosome numbers is frequently observed in Parp-1-deficient (Parp-1-/-) mouse embryonic fibroblasts (MEFs) (Kanai et al. (2003) Mol. Cell. Biol. 23, 2451-2462). However, there are no studies on whether the centrosome abnormality occurs also in other cell types under Parp-1 deficiency. In this study, we report that Parp-1-/- mouse embryonic stem (ES) cell lines did not show an abnormally increased number of centrosomes compared to wildtype ES cells. Recently, poly(ADP-ribose) glycohydrolase (Parg) has also been shown to localize in centrosomes (Ohashi et al. (2003) Biochem. Biophys. Res. Commun. 307, 915-921). The number of centrosomes of Pary-deficient (Parg-/-) ES cells was also analyzed in this study and was found to be stable under Parg deficiency. We also examined centrosome numbers in wild-type, Parp-1-/- and Parg-/- ES cell lines after treatment with methylmethanesulfonate (MMS) or γ-irradiation. Although a slight increase in the number of centrosomes is observed in each genotype twenty-four hours after treatment with MMS at 50 μM or with γ-irradiation at 1.4 Gy, there was no difference among the genotypes. These results suggest that loss of Parp-1 and Parg is insufficient to induce abnormality in centrosome numbers in ES cells and that ES cells possibly possess a strict mechanism for the maintenance of a normal number of centrosomes.
Proceedings of the Japan Academy Ser B Physical and Biological Sciences 06/2004; 80(6):290-294. DOI:10.2183/pjab.80.290 · 2.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: This report is of a 32-year-old man with central odontgenic frbroma arising in the maxilla. The clinical, radiographic, and histological features are described, and the relevant literature reviewed.
Asian Journal of Oral and Maxillofacial Surgery 12/2003; 15(4):288–291. DOI:10.1016/S0915-6992(03)80011-0
[Show abstract][Hide abstract] ABSTRACT: Poly(ADP-ribose) glycohydrolase plays a central role in poly(ADP-ribose) degradation, cleaving α(ribose-ribose) bonds to produce ADP-ribose. We previously generated Parg-deficient (Parg+/- and Parg-/-) mouse embryonic stem (ES) cells by disrupting exon 1 of the Parg gene, and showed the Parg-/- ES cells to be hypersensitive to methylmethanesulfonate, an alkylating agent. To clarify the role of Parg in the maintenance of genomic stability, we examined the frequency of sister-chromatid exchanges (SCEs) in Parg+/+ and Parg-/- ES cells with and without methylmethanesulfonate treatment. No difference was observed in the spontaneous frequency of SCEs between the Parg genotypes, treatment with methyl-methanesulfonate at 50 μM causing approximately 2-fold elevation in both cases.
Proceedings of the Japan Academy Ser B Physical and Biological Sciences 12/2003; 79(10):305-307. DOI:10.2183/pjab.79B.305 · 2.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Bone morphogenetic protein (BMP) induces bone formation in young rodents, but aging causes a reduction in the bone-forming ability of BMP. Most patients who require bone reconstruction are relatively old. Accordingly, we examined whether anabolic hormones could restore the bone inductive activity of rhBMP-2 in aged rats. rhBMP-2 in a carrier pellet was implanted subcutaneously in both 4- and 50-week-old female Wistar rats. PTH, PGE2, or 1,25(OH)2D3 was injected every day during the period of BMP implantation. The pellets were harvested, and were examined both histologically and biochemically 2 weeks after implantation. Bone-forming ability was measured by alkaline phosphatase (ALP) activity and calcium (Ca) content. Pellets in 50-week-old rats showed a significant reduction in bone formation compared to pellets in 4-week-old rats. However, daily injections of PTH into 50-week-old rats restored both ALP activity (103 +/- 4.6%) and Ca content (105 +/- 2.6%). 1,25(OH)2D3 and PGE2 also restored Ca content (103 +/- 4.5% and 98 +/- 3.8%, respectively) and stimulated ALP activity (142 +/- 2.3% and 133 +/- 3.6%). These results show that the administration of these hormones restores bone-forming ability in aged rats. A combination treatment of these hormones with rhBMP-2 might be applicable to the reconstruction of bone defects in elderly patients.
DNA and Cell Biology 09/2003; 22(9):541-6. DOI:10.1089/104454903322405428 · 2.06 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The purpose of this study was to evaluate the effectiveness of simultaneous cortex bone plate (CBP) graft with particulate marrow and cancellous bone (PMCB) graft for reliable closure of palatal fistulae associated with alveolar clefts.
Following standard secondary bone graft preparation of the cleft site, CBP harvested from the medial iliac crest was inserted into the palatal deficiency. This was followed by suturing the palatal mucosa. PMCB was then packed between the cortical bone and the reconstructed nasal floor.
Ten consecutive patients with palatal fistula were operated on at Tokyo Medical and Dental University Hospital from 1998 to 2000. Primary palatal repair was performed in 7 out of 10 patients at our center and in 3 out of 10 patients at other hospitals.
Ten patients (6 boys and men, 4 girls and women) with a palatal fistula associated with an alveolar cleft were studied. Ages ranged from 12 to 26 years.
All patients underwent simultaneous CBP graft with PMCB graft for closure of palatal fistula under general anesthesia.
Complete closure of palatal fistulae were obtained in 8 out of 10 cases. A very small asymptomatic fistula remained in one patient. Total necrosis of the labial flap with a residual palatal fistula occurred in one patient.
Simultaneous CBP graft with PMCB graft could be more reliable than PMCB alone for closure of a cleft associated palatal fistula.