Important roles of odontoblast membrane phospholipids in early dentin mineralization

The objective of this study was to first identify the timing and location of early mineralization of mouse first molar, and subsequently, to characterize the nucleation site for mineral formation in dentin from a materials science viewpoint and evaluate the effect of environmental cues (pH) affecting early dentin formation. Early dentin mineralization in mouse first molars began in the buccal central cusp on post-natal day 0 (P0), and was first hypothesized to involve collagen fibers. However, elemental mapping indicated the co-localization of phospholipids with collagen fibers in the early mineralization area. Co-localization of phosphatidylserine and annexin V, a functional protein that binds to plasma membrane phospholipids, indicated that phospholipids in the pre-dentin matrix were derived from the plasma membrane. A 3-dimensional in vitro biomimetic mineralization assay confirmed that phospholipids from the plasma membrane are critical factors initiating mineralization. Additionally, the direct measurement of the tooth germ pH, indicated it to be alkaline. The alkaline environment markedly enhanced the mineralization of cell membrane phospholipids. These results indicate that cell membrane phospholipids are nucleation sites for mineral formation, and could be important materials for bottom-up approaches aiming for rapid and more complex fabrication of dentin-like structures

Required Elements in tRNA for Methylation by the Eukaryotic tRNA (Guanine-N2-) Methyltransferase (Trm11-Trm112 Complex)

The Saccharomyces cerevisiae Trm11 and Trm112 complex (Trm11-Trm112) methylates the 2-amino group of guanosine at position 10 in tRNA and forms N2-methylguanosine. To determine the elements required in tRNA for methylation by Trm11-Trm112, we prepared 60 tRNA transcript variants and tested them for methylation by Trm11-Trm112. The results show that the precursor tRNA is not a substrate for Trm11-Trm112. Furthermore, the CCA terminus is essential for methylation by Trm11-Trm112, and Trm11-Trm112 also only methylates tRNAs with a regular-size variable region. In addition, the G10-C25 base pair is required for methylation by Trm11-Trm112. The data also demonstrated that Trm11-Trm112 recognizes the anticodon-loop and that U38 in tRNAAla acts negatively in terms of methylation. Likewise, the U32-A38 base pair in tRNACys negatively affects methylation. The only exception in our in vitro study was tRNAValAAC1. Our experiments showed that the tRNAValAAC1 transcript was slowly methylated by Trm11-Trm112. However, position 10 in this tRNA was reported to be unmodified G. We purified tRNAValAAC1 from wild-type and trm11 gene deletion strains and confirmed that a portion of tRNAValAAC1 is methylated by Trm11-Trm112 in S. cerevisiae. Thus, our study explains the m2G10 modification pattern of all S. cerevisiae class I tRNAs and elucidates the Trm11-Trm112 binding sites

Male mice, caged in the International Space Station for 35 days, sire healthy offspring

Matsumura, T., Noda, T., Muratani, M. et al. Male mice, caged in the International Space Station for 35 days, sire healthy offspring. Sci Rep 9, 13733 (2019). https://doi.org/10.1038/s41598-019-50128-

Male mice, caged in the International Space Station for 35 days, sire healthy offspring

The effect on the reproductive system and fertility of living in a space environment remains unclear. Here, we caged 12 male mice under artificial gravity (approximate to 1 gravity) (AG) or microgravity (MG) in the International Space Station (ISS) for 35 days, and characterized the male reproductive organs (testes, epididymides, and accessory glands) after their return to earth. Mice caged on earth during the 35 days served as a "ground" control (GC). Only a decrease in accessory gland weight was detected in AG and MG males; however, none of the reproductive organs showed any overt microscopic defects or changes in gene expression as determined by RNA-seq. The cauda epididymal spermatozoa from AG and MG mice could fertilize oocytes in vitro at comparable levels as GC males. When the fertilized eggs were transferred into pseudo-pregnant females, there was no significant difference in pups delivered (pups/transferred eggs) among GC, AG, and MG spermatozoa. In addition, the growth rates and fecundity of the obtained pups were comparable among all groups. We conclude that short-term stays in outer space do not cause overt defects in the physiological function of male reproductive organs, sperm function, and offspring viability

Postnatal lethality and chondrodysplasia in mice lacking both chondroitin sulfate N-acetylgalactosaminyltransferase-1 and -2

Chondroitin sulfate (CS) is a sulfated glycosaminoglycan (GAG) chain. In cartilage, CS plays important roles as the main component of the extracellular matrix (ECM), existing as side chains of the major cartilage proteoglycan, aggrecan. Six glycosyltransferases are known to coordinately synthesize the backbone structure of CS; however, their in vivo synthetic mechanism remains unknown. Previous studies have suggested that two glycosyltransferases, Csgalnact1 (t1) and Csgalnact2 (t2), are critical for initiation of CS synthesis in vitro. Indeed, t1 single knockout mice (t1 KO) exhibit slight dwarfism and a reduction in CS content in cartilage compared with wild-type (WT) mice. To reveal the synergetic roles of t1 and t2 in CS synthesis in vivo, we generated systemic single and double knockout (DKO) mice and cartilage-specific t1 and t2 double knockout (Col2-DKO) mice. DKO mice exhibited postnatal lethality, whereas t2 KO mice showed normal size and skeletal development. Col2-DKO mice survived to adulthood and showed severe dwarfism compared with t1 KO mice. Histological analysis of epiphyseal cartilage from Col2-DKO mice revealed disrupted endochondral ossification, characterized by drastic GAG reduction in the ECM. Moreover, DKO cartilage had reduced chondrocyte proliferation and an increased number of apoptotic chondrocytes compared with WT cartilage. Conversely, primary chondrocyte cultures from Col2-DKO knee cartilage had the same proliferation rate as WT chondrocytes and low GAG expression levels, indicating that the chondrocytes themselves had an intact proliferative ability. Quantitative RT-PCR analysis of E18.5 cartilage showed that the expression levels of Col2a1 and Ptch1 transcripts tended to decrease in DKO compared with those in WT mice. The CS content in DKO cartilage was decreased compared with that in t1 KO cartilage but was not completely absent. These results suggest that aberrant ECM caused by CS reduction disrupted endochondral ossification. Overall, we propose that both t1 and t2 are necessary for CS synthesis and normal chondrocyte differentiation but are not sufficient for all CS synthesis in cartilage

MAFB is dispensable for the fetal testis morphogenesis and the maintenance of spermatogenesis in adult mice

The transcription factor MAFB is an important regulator of the development and differentiation of various organs and tissues. Previous studies have shown that MAFB is expressed in embryonic and adult mouse testes and is expected to act as the downstream target of retinoic acid (RA) to initiate spermatogenesis. However, its exact localization and function remain unclear. Here, we localized MAFB expression in embryonic and adult testes and analyzed its gene function using Mafb-deficient mice. We found that MAFB and c-MAF are the only large MAF transcription factors expressed in testes, while MAFA and NRL are not. MAFB was localized in Leydig and Sertoli cells at embryonic day (E) 18.5 but in Leydig cells, Sertoli cells, and pachytene spermatocytes in adults. Mafb-deficient testes at E18.5 showed fully formed seminiferous tubules with no abnormal structure or differences in testicular somatic cell numbers compared with those of control wild-type mice. Additionally, the expression levels of genes related to development and function of testicular cells were unchanged between genotypes. In adults, the expression of MAFB in Sertoli cells was shown to be stage specific and induced by RA. By generating Mafbfl/fl CAG-CreER™ (Mafb-cKO) mice, in which Cre recombinase was activated upon tamoxifen treatment, we found that the neonatal cKO mice died shortly upon Mafb deletion, but adult cKO mice were alive upon deletion. Adult cKO mice were fertile, and spermatogenesis maintenance was normal, as indicated by histological analysis, hormone levels, and germ cell stage-specific markers. Moreover, there were no differences in the proportion of seminiferous stages between cKO mice and controls. However, RNA-Seq analysis of cKO Sertoli cells revealed that the down-regulated genes were related to immune function and phagocytosis activity but not spermatogenesis. In conclusion, we found that MAFB is dispensable for fetal testis morphogenesis and spermatogenesis maintenance in adult mice, despite the significant gene expression in different cell types, but MAFB might be critical for phagocytosis activity of Sertoli cells

Practice and Research on Media Literacy Workshop in Disaster-stricken Areas

本論文では，防災カードゲーム「クロスロード」のフォーマットを用いて，携帯電話やネット・コミュニケーションにおけるさまざまなトラブルなどの問題に応用したワークショップを，東日本大震災の被災地で実施した試みについて紹介する．これまでの研究から「クロスロード」を用いた実践は，メディアリテラシーの啓発と向上に効果をもたらしうることが確認されてきた．本研究においては，震災被災者を対象に実施することで，従来のインタヴュー調査などでは引き出しにくいと考えられる，深い内容のデータも収集しうるのではないかという目論見から調査実践をおこなった．結果としては，期待した成果を十分得られなかった部分もあったが，ワークショップのデザインを修正していくことにより，方法の発展の余地があることも確認された．This study discusses the proceedings of a workshop that examined the format of a disaster prevention card gaming called “Crossroad,” applied to resolve various problems related with mobile phone usage and Internet communication. Through subsequent studies, we have also confirmed that it is effective forpractices applied the Crossroad method to enlighten people and to improve media literacy. As this method could dig out the deeply bottomed content which had been seemed to be difficult to be picked up with great disaster victims, we were going to make workshops of gaming in this case. Though we could not derive the expected results, we also recognized possibilities for improvement with the design of the workshop

SARS-CoV-2 disrupts respiratory vascular barriers by suppressing Claudin-5 expression

臓器チップ技術を用いて新型コロナウイルスが血管へ侵入するメカニズムを解明 --Claudin-5発現抑制による呼吸器の血管内皮バリア破壊--. 京都大学プレスリリース. 2022-09-22.A study using an organ-on-a-chip reveals a mechanism of SARS-CoV-2 invasion into blood vessels --Disruption of vascular endothelial barrier in respiratory organs by decreasing Claudin-5 expression--. 京都大学プレスリリース. 2022-09-27.In the initial process of coronavirus disease 2019 (COVID-19), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infects respiratory epithelial cells and then transfers to other organs the blood vessels. It is believed that SARS-CoV-2 can pass the vascular wall by altering the endothelial barrier using an unknown mechanism. In this study, we investigated the effect of SARS-CoV-2 on the endothelial barrier using an airway-on-a-chip that mimics respiratory organs and found that SARS-CoV-2 produced from infected epithelial cells disrupts the barrier by decreasing Claudin-5 (CLDN5), a tight junction protein, and disrupting vascular endothelial cadherin–mediated adherens junctions. Consistently, the gene and protein expression levels of CLDN5 in the lungs of a patient with COVID-19 were decreased. CLDN5 overexpression or Fluvastatin treatment rescued the SARS-CoV-2–induced respiratory endothelial barrier disruption. We concluded that the down-regulation of CLDN5 expression is a pivotal mechanism for SARS-CoV-2–induced endothelial barrier disruption in respiratory organs and that inducing CLDN5 expression is a therapeutic strategy against COVID-19

2006ネン ニ トクシマシ シロヤマ ニ ショクジュ サレタ ホルトノキ ノ ナエギ ノ ゲンジョウ : セイイク ジョウキョウ ト ホルトノキ イオウビョウ カンセン ジョウキョウ ニツイテ

Elaeocarpus zollingeri (synonym: E. sylvestris) was one of dominant trees in Mt. Shiroyama in the Tokushima City until 1970s, however, most of adult trees have died by Elaeocarpus yellows. To prevent extinction 300 nursery trees were planted at the foot of the mountain by local volunteers in 2006. In this study we investigated their survival, growth, and infection status of the disease. We found 40 nursery trees have survived (survival rate was 13%). Survived trees were abundant in areas facing the south, and most trees disappeared in areas facing the north and the west. Growth of the trees was well also in areas facing the south. Among the survived 40 nursery trees, we found three trees infected with phytoplasma, the pathogen of Elaeocarpus yellows, based on PCR analysis. It is not clear when they were infected with phytoplasma, whether they have infected after the plantation or they already had phytoplasma in the seed. Anyway, the fact that only a few tree is infected must be gratifying in considering the conservation E. zollinger in Mt. Shiroyama

Indirect Adhesion of Hydrogels via the Radical Polymerization Mediated by <i>N</i>,<i>N</i>,<i>N</i>′,<i>N</i>′‑Tetramethylethylenediamine and Ammonium Persulfate

Recently, developing superior adhesive hydrogel systems is in demand because it is important when hydrogel materials are used for applications in engineering, medical, and other fields. Among the adhesive gel systems, the ones for which adhesion can be controlled indirectly have high potential for various applications in the medical and engineering fields because they have merits compared to the systems that need to be regulated directly at the adhesive interfaces when we need to make materials adhere at places that are difficult to touch. In this manuscript, we report the adhesive system which can be regulated indirectly by the reaction of N,N,N′,N′-tetramethylethylenediamine (TMEDA) and ammonium persulfate (APS) inside the gel networks. Applying this system, a step-by-step adhesion, in which adhesive timing can be controlled, and a multiple adhesion, which can adhere multiple gel pieces at once, are constructed. These indirect and continuous adhesions will help us use hydrogels for constructing artificial biomaterials and other various applications