Genomic analysis identifies candidate pathogenic variants in 9 of 18 patients with unexplained West syndrome

West syndrome, which is narrowly defined as infantile spasms that occur in clusters and hypsarrhythmia on EEG, is the most common early-onset epileptic encephalopathy (EOEE). Patients with West syndrome may have clear etiologies, including perinatal events, infections, gross chromosomal abnormalities, or cases followed by other EOEEs. However, the genetic etiology of most cases of West syndrome remains unexplained. DNA from 18 patients with unexplained West syndrome was subjected to microarray-based comparative genomic hybridization (array CGH), followed by trio-based whole-exome sequencing in 14 unsolved families. We identified candidate pathogenic variants in 50 % of the patients (n = 9/18). The array CGH revealed candidate pathogenic copy number variations in four cases (22 %, 4/18), including an Xq28 duplication, a 16p11.2 deletion, a 16p13.1 deletion and a 19p13.2 deletion disrupting CACNA1A. Whole-exome sequencing identified candidate mutations in known epilepsy genes in five cases (36 %, 5/14). Three candidate de novo mutations were identified in three cases, with two mutations occurring in two new candidate genes (NR2F1 and CACNA2D1) (21 %, 3/14). Hemizygous candidate mutations in ALG13 and BRWD3 were identified in the other two cases (14 %, 2/14). Evaluating a panel of 67 known EOEE genes failed to identify significant mutations. Despite the heterogeneity of unexplained West syndrome, the combination of array CGH and whole-exome sequencing is an effective means of evaluating the genetic background in unexplained West syndrome. We provide additional evidence for NR2F1 as a causative gene and for CACNA2D1 and BRWD3 as candidate genes for West syndrome.     

Acupuncture and moxibustion for stress-related disorders

Acupuncture and moxibustion, which medical doctors are licensed by the government of Japan to perform, can improve the psychological relationship between doctors and patients, especially when it is disturbed by a “game”, a dysfunctional interpersonal interaction that is repeated unintentionally. This advantage is due to the essential properties of acupuncture and moxibustion. Acupuncture and moxibustion are helpful in treating somatoform disorders, especially musculoskeletal symptoms. In Japan, a holistic acupuncture and moxibustion therapy called Sawada-style has been developed. This is based on fundamental meridian points that are considered to have effects on central, autonomic nervous, immune, metabolic, and endocrine systems to regulate the whole body balance. In addition, some of the fundamental points have effects on Qi, blood, and water patterns associated with major depression, generalized anxiety disorder, eating disorders, and somatoform disorders. The fixed protocol of Sawada-style would be suitable for large-scale, randomized, controlled studies in the future. Recent systematic reviews indicate that electroacupuncture would be a useful addition to antidepressant therapy for some symptoms accompanying fibromyalgia. Acupuncture and moxibustion are also recommended for irritable bowel syndrome, instead of Western drug therapy. Surprisingly, the dorsal prefrontal cerebral cortex, which is associated with a method of scalp acupuncture applied for gastrointestinal disorders, has been found to be activated in patients with irritable bowel syndrome. It is quite possible that regulation of this cortical area is related to the effect of scalp acupuncture. This acupuncture method can be effective not only for irritable bowel syndrome but also for other stress-related gastrointestinal disorders.     

Rosa26-GFP Direct Repeat (RaDR-GFP) Mice Reveal Tissue- and Age-Dependence of Homologous Recombination in Mammals In Vivo

Homologous recombination (HR) is critical for the repair of double strand breaks and broken replication forks. Although HR is mostly error free, inherent or environmental conditions that either suppress or induce HR cause genomic instability. Despite its importance in carcinogenesis, due to limitations in our ability to detect HR in vivo, little is known about HR in mammalian tissues. Here, we describe a mouse model in which a direct repeat HR substrate is targeted to the ubiquitously expressed Rosa26 locus. In the Rosa26 Direct Repeat-GFP (RaDR-GFP) mice, HR between two truncated EGFP expression cassettes can yield a fluorescent signal. In-house image analysis software provides a rapid method for quantifying recombination events within intact tissues, and the frequency of recombinant cells can be evaluated by flow cytometry. A comparison among 11 tissues shows that the frequency of recombinant cells varies by more than two orders of magnitude among tissues, wherein HR in the brain is the lowest. Additionally, de novo recombination events accumulate with age in the colon, showing that this mouse model can be used to study the impact of chronic exposures on genomic stability. Exposure to N-methyl-N-nitrosourea, an alkylating agent similar to the cancer chemotherapeutic temozolomide, shows that the colon, liver and pancreas are susceptible to DNA damage-induced HR. Finally, histological analysis of the underlying cell types reveals that pancreatic acinar cells and liver hepatocytes undergo HR and also that HR can be specifically detected in colonic somatic stem cells. Taken together, the RaDR-GFP mouse model provides new understanding of how tissue and age impact susceptibility to HR, and enables future studies of genetic, environmental and physiological factors that modulate HR in mammals.     

Double Columnar Structure with a Nanogradient Composite for Increased Oxygen Diffusivity and Reduction Activity

The cathodic performances that can be achieved in solid oxide fuel cells (SOFCs), particularly in terms of oxygen diffusion, need to be improved so that high power densities can be produced at intermediate temperatures. Here, to improve the cathodic performance, a double columnar functional interlayer (DCFL) consisting of Sm_0.2Ce_0.8O_2?δ (SDC) and Sm_0.5Sr_0.5CoO_3?δ (SSC) is fabricated between a La_0.9Sr_0.1Ga_0.8Mg_0.2O_3?δ electrolyte film and a SSC cathode film with pulsed laser deposition. The DCFL has a rough surface morphology consisting of nanosized grains (with diameters less than 5 nm), and it is formed of small columns that grow at an angle of ca. 45° from the substrate. Inserting the DCFL causes the electrical conductivity of the cathode to increase significantly, and the power density obtained by using it in a metal‐supported SOFC is increased. Atomic resolution scanning transmission electron microscopy (TEM) images and density functional theory calculations confirm that the samarium atoms in the SDC columns and cobalt atoms in the SSC columns are located at the interfaces between SDC and SSC columns. Therefore, it is possible a SmCoO_3?δ nanogradient is formed, which may cause lattice distortions. The ¹⁸O₂ concentration is actually much higher in the DCFL than in either of SSC or SDC films.     

Adaptation of a Cyanobacterium to a Biochemically Rich Environment in Experimental Evolution as an Initial Step toward a Chloroplast-Like State

Chloroplasts originated from cyanobacteria through endosymbiosis. The original cyanobacterial endosymbiont evolved to adapt to the biochemically rich intracellular environment of the host cell while maintaining its photosynthetic function; however, no such process has been experimentally demonstrated. Here, we show the adaptation of a model cyanobacterium, Synechocystis sp. PCC 6803, to a biochemically rich environment by experimental evolution. Synechocystis sp. PCC 6803 does not grow in a biochemically rich, chemically defined medium because several amino acids are toxic to the cells at approximately 1 mM. We cultured the cyanobacteria in media with the toxic amino acids at 0.1 mM, then serially transferred the culture, gradually increasing the concentration of the toxic amino acids. The cells evolved to show approximately the same specific growth rate in media with 0 and 1 mM of the toxic amino acid in approximately 84 generations and evolved to grow faster in the media with 1 mM than in the media with 0 mM in approximately 181 generations. We did not detect a statistically significant decrease in the autotrophic growth of the evolved strain in an inorganic medium, indicating the maintenance of the photosynthetic function. Whole-genome resequencing revealed changes in the genes related to the cell membrane and the carboxysome. Moreover, we quantitatively analyzed the evolutionary changes by using simple mathematical models, which evaluated the evolution as an increase in the half-maximal inhibitory concentration (IC₅₀) and estimated quantitative characteristics of the evolutionary process. Our results clearly demonstrate not only the potential of a model cyanobacterium to adapt to a biochemically rich environment without a significant decrease in photosynthetic function but also the properties of its evolutionary process, which sheds light of the evolution of chloroplasts at the initial stage.     

Simultaneous Copy Number Losses within Multiple Subtelomeric Regions in Early-Onset Type2 Diabetes Mellitus

Genetic factors play very important roles in the onset and progression of type 2 diabetes mellitus (T2DM). However, the genetic factors correlating with T2DM onset have not as yet been fully clarified. We previously found that copy number losses in the subtelomeric region on chromosome 4p16.3 were detected in early-onset Japanese T2DM patients (onset age <35 years) at a high frequency. Herein, we additionally found two novel copy number losses within the subtelomeric regions on chromosomes 16q24.2-3 and 22q13.31-33, which have significant associations with early-onset Japanese T2DM. The associations were statistically significant by Fisher''s exact tests with P values of 5.19×10⁻³ and 1.81×10⁻³ and odds ratios of 5.7 and 4.4 for 16q24.2-3 and 22q13.31-33, respectively. Furthermore, copy number variation (CNV) analysis of the whole genome using the CNV BeadChip system verified simultaneous copy number losses in all three subtelomeric regions in 11 of our 100 T2DM subjects, while none of 100 non-diabetic controls showed the copy number losses in all three regions. Our results suggest that the mechanism underlying induction of CNVs is involved in the pathogenesis of early-onset T2DM. Thus, copy number losses within multiple subtelomeric regions are strongly associated with early-onset T2DM and examination of simultaneous CNVs in these three regions may lead to the development of an accurate and selective procedure for detecting genetic susceptibility to T2DM.     

Spatial conservation planning under climate change: Using species distribution modeling to assess priority for adaptive management of Fagus crenata in Japan

Protected areas are the basis of modern conservation systems, but current climate change causes gaps between protected areas and the species distribution ranges. To mitigate the impact of climate change on species distribution ranges, revision of protected areas are necessary. Alternatively, active management such as excluding competitive species or transplanting target species would be effective. In this study, we assessed optimal actions (revision of protected areas or active management) in each geographical region to establish an effective spatial conservation plan in Japan. Gaps between the protected areas and future potential habitats were assessed using species distribution models and 20 future climate simulations. Fagus crenata, an endemic and dominant species in Japan, was used as a target species. Potential habitats within the protected areas were predicted to decrease from 22,122 km² at present to 12,309 km² under future climate conditions. Sustainable potential habitats (consistent potential habitats both at present and in future) without the protected areas extended to 13,208 km², and were mainly found in northeast Japan. These results suggest that, in northeast Japan, revisions to protected areas would be effective in preserving sustainable potential habitats under future climate change. However, the potential habitats of southwestern Japan, in which populations were genetically different from northeastern populations, were predicted to virtually disappear both within and outside of protected areas. Active management is thus necessary in southwestern Japan to ensure intraspecific genetic diversity under future climate change.