Direct Injection of CRISPR/Cas9-Related mRNA into Cytoplasm of Parthenogenetically Activated Porcine Oocytes Causes Frequent Mosaicism for Indel Mutations

Some reports demonstrated successful genome editing in pigs by one-step zygote microinjection of mRNA of CRISPR/Cas9-related components. Given the relatively long gestation periods and the high cost of housing, the establishment of a single blastocyst-based assay for rapid optimization of the above system is required. As a proof-of-concept, we attempted to disrupt a gene (GGTA1) encoding the α-1,3-galactosyltransferase that synthesizes the α-Gal epitope using parthenogenetically activated porcine oocytes. The lack of α-Gal epitope expression can be monitored by staining with fluorescently labeled isolectin BS-I-B₄ (IB4), which binds specifically to the α-Gal epitope. When oocytes were injected with guide RNA specific to GGTA1 together with enhanced green fluorescent protein (EGFP) and human Cas9 mRNAs, 65% (24/37) of the developing blastocysts exhibited green fluorescence, although almost all (96%, 23/24) showed a mosaic fluorescent pattern. Staining with IB4 revealed that the green fluorescent area often had a reduced binding activity to IB4. Of the 16 samples tested, six (five fluorescent and one non-fluorescent blastocysts) had indel mutations, suggesting a correlation between EGFP expression and mutation induction. Furthermore, it is suggested that zygote microinjection of mRNAs might lead to the production of piglets with cells harboring various mutation types.     

Structure and properties of poly(vinyl alcohol)/κ‐carrageenan blends

Blends of a commercial atactic poly(vinyl alcohol) (a‐PVA) derived from vinyl acetate and κ‐carrageenan were prepared by mixing the aqueous solutions of both samples. Blend films prepared by casting were transparent. In the DSC curves of the blend films, the endothermic peaks shifted to lower temperature with an increase of the content of κ‐carrageenan. The Young's modulus and the strength at break increased with an increase of the content of a‐PVA. As the standing temperature of the blend solutions decreased, the gelation region increased also at high content of carrageenan. In the amorphous regions of blend films, a‐PVA and κ‐carrageenan were miscible. © 2001 Society of Chemical Industry     

Highly anisotropic thermal conductivity of mesogenic epoxy resin film through orientation control

To develop insulating materials with a high thermally conductive anisotropy, planarly aligned mesogenic epoxy (ME) resin film was fabricated by uniaxial coating on a hydrophobic polyethylene terephthalate substrate. Grazing incidence small-angle X-ray scattering (GISAXS) and transmission SAXS measurements exhibited that the films spontaneously formed uniaxially aligned monodomain-like smectic structures by curing on the hydrophobic substrate. Then, an in- and out-of-plane thermal conductivity of 10 and 0.048 W m⁻¹ K⁻¹ and outstanding thermal conductivity anisotropy of 208 have been confirmed, respectively. The ME resin films with high thermal conductivity can be applied as insulating materials for multiple-layer electrical and electronic devices.     

Links between Immune Cells from the Periphery and the Brain in the Pathogenesis of Epilepsy: A Narrative Review

Accumulating evidence has demonstrated that the pathogenesis of epilepsy is linked to neuroinflammation and cerebrovascular dysfunction. Peripheral immune cell invasion into the brain, along with these responses, is implicitly involved in epilepsy. This review explored the current literature on the association between the peripheral and central nervous systems in the pathogenesis of epilepsy, and highlights novel research directions for therapeutic interventions targeting these reactions. Previous experimental and human studies have demonstrated the activation of the innate and adaptive immune responses in the brain. The time required for monocytes (responsible for innate immunity) and T cells (involved in acquired immunity) to invade the central nervous system after a seizure varies. Moreover, the time between the leakage associated with blood–brain barrier (BBB) failure and the infiltration of these cells varies. This suggests that cell infiltration is not merely a secondary disruptive event associated with BBB failure, but also a non-disruptive event facilitated by various mediators produced by the neurovascular unit consisting of neurons, perivascular astrocytes, microglia, pericytes, and endothelial cells. Moreover, genetic manipulation has enabled the differentiation between peripheral monocytes and resident microglia, which was previously considered difficult. Thus, the evidence suggests that peripheral monocytes may contribute to the pathogenesis of seizures.     

Pore structure of carbons coated on ceramic particles

Pore structure of carbon coated on ceramic particles by carbonization of precursor in a powder mixture at 900 °C was studied by focusing on the effects of substrate ceramics (MgO, TiO₂ and various phases of Al₂O₃) and of carbon precursor (poly(vinyl alcohol) (PVA), poly(ethylene terephthalate) (PET) and hydroxyl propyl cellulose (HPC)). By dissolving substrate MgO particles, carbon coated was found to have a high BET surface area, more than 1000 m²/g, which was almost the same as the value estimated from apparent surface area measured on carbon-coated MgO particles under the assumption of zero surface area of the substrate. The carbon separated was found to be rich in micropores from the analyses by DFT method and α_s plot. The dependence of the BET surface area on the amount of carbon coated on TiO₂ with a high surface area was the same for three carbon precursors, although the carbon yields from the precursors were slightly different. Porous Al₂O₃ substrates, γ-Al₂O₃as-received and that formed from Al(OH)₃ during carbonization, gave a high BET surface area, but dense Al₂O₃, α-Al₂O₃, gave a low surface area.     

Synthesis of stable and soluble one-handed helical poly(substituted acetylene)s without chiral pendant groups via polymer reaction in membrane state

A soluble and stable one-handed helical conjugated polymer without the coexistence of any other chiral moieties was successfully synthesized by asymmetric-induced polymerization of a chiral monomer having two hydroxyl groups followed by desubstitution of the chiral groups in a solid membrane state. The reason for the success was the polymer reaction was carried out in the membrane state. This is an alternative method to obtain such a unique chiral polymer which was obtained only by the helix-sense-selective polymerization (HSSP) we reported before. In addition the efficiency of the chiral induction was higher than that of the HSSP. It is interesting that the “Membrane state” acted like as if a protecting group.     

Validating the usefulness of examiners’ forward citations from the viewpoint of applicants’ self-selection during the patent application procedure

In this study, we validated the usefulness of examiners’ forward citations, especially from the viewpoint of the applicants’ self-selection (ASS) decisions during the patent application procedure. We believe that the ASS in an early stage would be decided by a potential-value comparison among patent applications. We focused on six self-selection decision points of the applicants: whether to file patent applications in foreign countries, request for examination, request for accelerated examination, reply to a notification of reasons for refusal, appeal after receiving a decision of refusal, and register after receiving a decision to grant a patent as patent value parameters. We found that application groups that selected “Yes” have a significantly larger number of examiners’ forward citations than groups that selected “No” at all decision points. In addition, we confirmed that applications that were finally granted and those that were renewed for a full term after grant have a significantly large number of examiners’ forward citations. We concluded that the number of examiners’ forward citations would be a useful indicator of the potential value of patent applications in macroscopic analysis.     

Room temperature crystallization by RF plasma

The crystallization of amorphous thin films was achieved by radiofrequency (RF) plasma treatment. Although various amorphous films are crystallized after 2 min or so, the sample temperature is lower than 150 °C without compulsory cooling even when the films are treated for 1 h. This treatment works on amorphous films of various materials, independently of the film preparation method and substrate materials. Sol-gel-derived TiO₂ films were densified and simultaneously crystallized to anatase structure by the plasma treatment and the obtained films indicate almost the same photocatalytic activities as that of thermally crystallized TiO₂ films. Plasma-crystallized sputtered indium tin oxide (ITO) films have a bixbite structure and the resistivity reached to 1.6 × 10^− 4 Ω cm while the crystallization condition was not optimized. Amorphous silicon films with a small mount of crystallites were deposited by sputtering method and were crystallized by the plasma treatment.     

Crystal growth and scintillation properties of Ce-doped sodium calcium lutetium complex fluoride

0.1%, 0.5%, 1% and 3% Ce doped scintillation crystals based on NaF-CaF₂-LuF₃ solid solutions were grown from the melt using Micro-Pulling-Down (μ-PD) method. The grown crystals were transparent and their transmittance was approximately 80% for the wavelengths longer than 320 nm. Concerning the scintillation properties, radio-luminescence peaks of the crystals were detected at approximately 330 nm. The light yield of the crystals was also measured using γ-ray (¹³⁷Cs and ²²Na) as the excitation source.     

Long-term observation of subcutaneous tissue reaction to synthetic auditory ossicle (Apaceram) in rats

The present study evaluates histological characteristics of the soft tissue response to long-term implantation of Apaceram discs composed of dense hydroxyapatite in rats. Discs were implanted into the subcutaneous tissue of 76 rats for six to 20 months. Decalcified histological sections stained with haematoxylin and eosin (H & E) and Mallory's azan were examined. Different cell types surrounding implants were counted. The greatest proportion of macrophages was found at six months (13.5 per cent). This proportion gradually decreased to four per cent at 20 months. Small numbers of lymphocytes and foreign body giant cells were observed in every group, but neither neutrophils nor osteogenesis were observed in any specimens. Results of the present study and previous related studies indicate that despite reappearance of a small number of macrophages six months after implantation, Apaceram is useful for reconstructive surgery.