The 'ataxia telangiectasia mutated' (Atm) gene maintains genomic stability by activating a key cell-cycle checkpoint in response to DNA damage, telomeric instability or oxidative stress. Mutational inactivation of the gene causes an autosomal recessive disorder, ataxia-telangiectasia, characterized by immunodeficiency, progressive cerebellar ataxia, oculocutaneous telangiectasia, defective spermatogenesis, premature ageing and a high incidence of lymphoma. Here we show that ATM has an essential function in the reconstitutive capacity of haematopoietic stem cells (HSCs) but is not as important for the proliferation or differentiation of progenitors, in a telomere-independent manner. Atm-/- mice older than 24 weeks showed progressive bone marrow failure resulting from a defect in HSC function that was associated with elevated reactive oxygen species. Treatment with anti-oxidative agents restored the reconstitutive capacity of Atm-/- HSCs, resulting in the prevention of bone marrow failure. Activation of the p16(INK4a)-retinoblastoma (Rb) gene product pathway in response to elevated reactive oxygen species led to the failure of Atm-/- HSCs. These results show that the self-renewal capacity of HSCs depends on ATM-mediated inhibition of oxidative stress.

Induction of pluripotent stem cells from human fibroblasts has been achieved by the ectopic expression of two different sets of four genes. However, the mechanism of the pluripotent stem cell induction has not been elucidated. Here we identified a marked heterogeneity in colonies generated by the four-gene (Oct3/4, Sox2, c-Myc, and Klf4) transduction method in human neonatal skin-derived cells. The four-gene transduction gave a higher probability of induction for archetypal pluripotent stem cell marker genes (Nanog, TDGF, and Dnmt3b) than for marker genes that are less specific for pluripotent stem cells (CYP26A1 and TERT) in primary induction culture. This tendency may reflect the molecular mechanism underlying the induction of human skin-derived cells into pluripotent stem cells. Among the colonies induced by the four-gene transduction, small cells with a high nucleus-to-cytoplasm ratio could be established by repeated cloning. Subsequently established cell lines were similar to human embryonic stem cells as well as human induced pluripotent stem (iPS) cells derived from adult tissue in morphology, gene expression, long-term self-renewal ability, and teratoma formation. Genome-wide single-nucleotide polymorphism array analysis of the human iPS cell line indicates that the induction process did not induce DNA mutation.

We have cloned a gene (aphA) encoding acetylpolyamine amidohydrolase from Mycoplana ramosa ATCC 49678 (previously named Mycoplana bullata). A genomic library of M. ramosa was screened with an oligonucleotide probe designed from a N-terminal amino acid sequence of the enzyme purified from M. ramosa. Nucleotide sequence analysis revealed an open reading frame of 1,023 bp which encodes a polypeptide with a molecular mass of 36,337 Da. This is the first report of the structure of acetylpolyamine amidohydrolase. The aphA gene was subcloned under the control of the trc promoter and was expressed in Escherichia coli MM294. The recombinant enzyme was purified, and the enzymatic properties were characterized. Substrate specificities, K m values, and V max values were identical to those of the native enzyme purified from M. ramosa. In the analysis of the metal-substituted enzymes, we found that the acid limb of pH rate profiles shifts from 7.2 for the original zinc enzyme to 6.6 for the cobalt enzyme. This change suggests that the zinc atom is essential for the catalytic activity of the enzyme similarly to the zinc atom in carboxypeptidase A.Polyamines (putrescine, cadaverine, spermidine, and spermine) are bioactive amines widely distributed among both eukaryotic and prokaryotic cells. Polyamines and their metabolites are essential for normal cell growth, although details of their functions have not been fully elucidated (22). In higher eukaryotes, polyamines modulate channel activities of N-methyl-D-aspartate receptors, which play an important role in glutamate-mediated neuronal plasticity and neurotoxicity in the central nervous system (17). Spermine also functions to stimulate the protein kinase casein kinase 2 (12).Biosynthetic pathways for polyamines are well established (14,22). Four key enzymes make up the pathway of polyamine synthesis. L-Ornithine decarboxylase forms putrescine from Lornithine. S-Adenosylmethionine decarboxylase forms decarboxylated S-adenosylmethionine, which acts as an aminopropyl donor. Spermidine synthase and spermine synthase transfer the aminopropyl group from decarboxylated S-adenosylmethionine to putrescine and spermidine, respectively. Specific inhibitors of these enzymes are now available and are used as therapeutic agents for African sleeping sickness and cancer.In eukaryotic cells, the retroconversion of spermine to putrescine can be accomplished by sequential actions of spermidine-spermine N 1 -acetyltransferase and polyamine oxidase. In microorganisms, however, polyamine oxidase and acetylpolyamine amidohydrolase are involved in degradative pathways of acetylpolyamines (14). It is uncertain whether some types of acetylpolyamines are catabolized via deacetylation by acetylpolyamine amidohydrolases while the others are catabolized by polyamine oxidases. The acetylpolyamine amidohydrolases found in Streptomyces avellaneus (18), Arthrobacter sp. (10), and Micrococcus rubens (15) act only on acetylputrescine. A novel acetylpolyamine amidohydrolase which acts upon all types of acetylpolya...

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Objective The purpose of this research was to develop a deep-learning model to assess radiographic finger joint destruction in RA. Methods The model comprises two steps: a joint-detection step and a joint-evaluation step. Among 216 radiographs of 108 patients with RA, 186 radiographs were assigned to the training/validation dataset and 30 to the test dataset. In the training/validation dataset, images of PIP joints, the IP joint of the thumb or MCP joints were manually clipped and scored for joint space narrowing (JSN) and bone erosion by clinicians, and then these images were augmented. As a result, 11 160 images were used to train and validate a deep convolutional neural network for joint evaluation. Three thousand seven hundred and twenty selected images were used to train machine learning for joint detection. These steps were combined as the assessment model for radiographic finger joint destruction. Performance of the model was examined using the test dataset, which was not included in the training/validation process, by comparing the scores assigned by the model and clinicians. Results The model detected PIP joints, the IP joint of the thumb and MCP joints with a sensitivity of 95.3% and assigned scores for JSN and erosion. Accuracy (percentage of exact agreement) reached 49.3–65.4% for JSN and 70.6–74.1% for erosion. The correlation coefficient between scores by the model and clinicians per image was 0.72–0.88 for JSN and 0.54–0.75 for erosion. Conclusion Image processing with the trained convolutional neural network model is promising to assess radiographs in RA.