Inhibition of RANKL-Induced Osteoclastogenesis by Novel Mutant RANKL

Background: Recently, it was reported that leucine-rich repeat-containing G-protein-coupled receptor 4 (LGR4, also called GPR48) is another receptor for RANKL and was shown to compete with RANK to bind RANKL and suppress canonical RANK signaling during osteoclast differentiation. The critical role of the protein triad RANK–RANKL in osteoclastogenesis has made their binding an important target for the development of drugs against osteoporosis. In this study, point-mutations were introduced in the RANKL protein based on the crystal structure of the RANKL complex and its counterpart receptor RANK, and we investigated whether LGR4 signaling in the absence of the RANK signal could lead to the inhibition of osteoclastogenesis.; Methods: The effects of point-mutated RANKL (mRANKL-MT) on osteoclastogenesis were assessed by tartrate-resistant acid phosphatase (TRAP), resorption pit formation, quantitative real-time polymerase chain reaction (qPCR), western blot, NFATc1 nuclear translocation, micro-CT and histomorphological assay in wild type RANKL (mRANKL-WT)-induced in vitro and in vivo experimental mice model. Results: As a proof of concept, treatment with the mutant RANKL led to the stimulation of GSK-3β phosphorylation, as well as the inhibition of NFATc1 translocation, mRNA expression of TRAP and OSCAR, TRAP activity, and bone resorption, in RANKL-induced mouse models; and Conclusions: The results of our study demonstrate that the mutant RANKL can be used as a therapeutic agent for osteoporosis by inhibiting RANKL-induced osteoclastogenesis via comparative inhibition of RANKL. Moreover, the mutant RANKL was found to lack the toxic side effects of most osteoporosis treatments.     

Diphenyl-Methane Based Thyromimetic Inhibitors for Transthyretin Amyloidosis

Thyromimetics, whose physicochemical characteristics are analog to thyroid hormones (THs) and their derivatives, are promising candidates as novel therapeutics for neurodegenerative and metabolic pathologies. In particular, sobetirome (GC-1), one of the initial halogen-free thyromimetics, and newly synthesized IS25 and TG68, with optimized ADME-Tox profile, have recently attracted attention owing to their superior therapeutic benefits, selectivity, and enhanced permeability. Here, we further explored the functional capabilities of these thyromimetics to inhibit transthyretin (TTR) amyloidosis. TTR is a homotetrameric transporter protein for THs, yet it is also responsible for severe amyloid fibril formation, which is facilitated by tetramer dissociation into non-native monomers. By combining nuclear magnetic resonance (NMR) spectroscopy, computational simulation, and biochemical assays, we found that GC-1 and newly designed diphenyl-methane-based thyromimetics, namely IS25 and TG68, are TTR stabilizers and efficient suppressors of TTR aggregation. Based on these observations, we propose the novel potential of thyromimetics as a multi-functional therapeutic molecule for TTR-related pathologies, including neurodegenerative diseases.     

Contribution of Invariant Natural Killer T Cells to the Clearance of Pseudomonas aeruginosa from Skin Wounds

Chronic infections are considered one of the most severe problems in skin wounds, and bacteria are present in over 90% of chronic wounds. Pseudomonas aeruginosa is frequently isolated from chronic wounds and is thought to be a cause of delayed wound healing. Invariant natural killer T (iNKT) cells, unique lymphocytes with a potent regulatory ability in various inflammatory responses, accelerate the wound healing process. In the present study, we investigated the contribution of iNKT cells in the host defense against P. aeruginosa inoculation at the wound sites. We analyzed the re-epithelialization, bacterial load, accumulation of leukocytes, and production of cytokines and antimicrobial peptides. In iNKT cell–deficient (Jα18KO) mice, re-epithelialization was significantly decreased, and the number of live colonies was significantly increased, when compared with those in wild-type (WT) mice on day 7. IL-17A, and IL-22 production was significantly lower in Jα18KO mice than in WT mice on day 5. Furthermore, the administration of α-galactosylceramide (α-GalCer), a specific activator of iNKT cells, led to enhanced host protection, as shown by reduced bacterial load, and to increased production of IL-22, IL-23, and S100A9 compared that of with WT mice. These results suggest that iNKT cells promote P. aeruginosa clearance during skin wound healing.     

Effect of milling ball size on the densification and optical properties of transparent Y2O3 ceramics

In this study, we report highly transparent Y₂O₃ ceramics fabricated by hot-pressing only at 1500 °C without a HIP treatment, featuring in-line transmittance levels of 77% and 84% at a wavelength of 400 and 1100 nm, respectively with the grain size suppressed to 710 nm. The effect of the ball size during the grinding of Y₂O₃ powders on the correlation between the thus-prepared Y₂O₃ powders and the optical properties of the hot-pressed samples is demonstrated for the first time. With a decrease in the diameter of the ZrO₂ balls from 5 mm to 1 mm, the milling efficiency was enhanced and admirable transparency of Y₂O₃ was attained at a short milling time. However, several micron-sized pores remained in the transparent specimens prepared with 1 mm balls, originating from the inhomogeneously packed region of the green body. Finally, the 2 mm was found to be optimum for obtaining a fine-grained and pore-free microstructure with the best in-line transmittance of Y₂O₃ ceramics.     

Characteristics and Cryopreservation of Semen of Sex-Reversed Females of Salmonid Fish

Sex reversal has been used as a breeding strategy by salmonid fish to produce genetically and phenotypically single sex populations. Production of all-female fish has great importance for the creation of monosex female triploids of salmonid fish, which are valued for their sterility, lack of female maturation, and larger commercial size. Among salmonids, the majority of rainbow trout (Oncorhynchus mykiss) production is based on all-female production with a high proportion of all-female triploid production in Europe. The main aim of this review is to present the recent knowledge regarding sex-reversed females (SRFs) of salmonid fish. We discuss the methods of sex reversal as well as their effects on the morphology and histology of the reproductive tract. We focus on the characteristics of SRF semen as well as the factors determining semen quality. The lower quality of SRF sperm compared to that of normal males has resulted in the need for the artificial maturation of semen. Most importantly, methods of semen storage—both short-term and long-term (cryopreservation)—that can improve hatchery operations are presented with the special emphasis on recent progress in development of efficient cryopreservation procedures and use of cryopreserved semen in hatchery practice. Moreover, we also address the emerging knowledge concerning the proteomic investigations of salmonid sperm, focusing primarily on the proteomic comparison of normal male and SRF testicular semen and presenting changes in SRF rainbow trout sperm proteome after in vitro incubation in artificial seminal plasma.     

Prelabeling Expansion Single-Molecule Localization Microscopy with Minimal Linkage Error

Expansion microscopy combined with single-molecule localization microscopy (ExSMLM) has a potential for approaching molecular resolution. However, ExSMLM faces multiple challenges such as loss of fluorophores and proteins during polymerization, digestion or denaturation, and an increase in linkage error arising from the distance between the fluorophore and the target molecule. Here, we introduce a trifunctional streptavidin to link the target, fluorophore and gel matrix via a biotinylizable peptide tag. The resultant ExSMLM images of vimentin filaments demonstrated high labeling efficiency and a minimal linkage error of ∼5 nm. Our ExSMLM provides a simple and practical means for fluorescence imaging with molecular resolution.     

基于改进 abcd 模型的黄河源区径流变化与归因

为改进模型对高寒地区融雪径流模拟不足的缺陷，将融雪模块耦合到传统 abcd 模型。利用 1980—2018 年 逐月实测的径流数据和通过 AnuSpline 方法插值的格网气象要素，驱动改进后的abcd 模型，分析三江源生态保护 措施实施前后（1980—1999 年和 2000—2018 年）黄河源区径流的动态变化，并量化关键气象因素与人类活动对 径流变化的影响程度，即相对贡献。结果表明：耦合融雪模块的 abcd-snow 模型完善了高寒地区水文过程的模拟， 提高对径流的模拟性能，在黄河源区表现出较好的适用性；整个研究时段黄河源区的实测径流呈不显著减少趋势 （?0.80?mm/a，p>0.05），但 2000 年前径流则呈现显著下降趋势（?4.12?mm/a，p<0.05），2000 年后径流则呈显著增加 趋势（3.16?mm/a，p<0.05）；?归因分析表明气候变化是源区径流变化的主导因素。2000 年前，气候变化对径流减少 的相对贡献率为 62.8%，人类活动对径流的贡献为 37.2%；2000 年后，气候变化对径流增加的贡献率达到 120.0?%， 人类活动对径流的贡献为?20.0%。其中：降水的变化是决定径流变化主导因素；其他气候因素的相对贡献较小； 以人类活动为主的生态恢复可显著降低河川径流。本研究有助于理解气候变化和下垫面变化对黄河源区水资源 变化的系统驱动机理，并为流域水资源合理配置提供科学参考依据。     

The Role of CTLA4 and Its Polymorphisms in Solid Organ and Haematopoietic Stem Cell Transplantation

HLA matching, transplantation technique, or underlying disease greatly influences the probability of long-term transplantation success. It has been hypothesised that genetic variation affecting antigen presentation also contributes to the outcomes of both solid organ transplantation and allogeneic haematopoietic stem cell transplantation (AHSCT). Those genes, along with those responsible for innate and adaptive immunity, have become targets of investigation. In this review, we focus on the role of CTLA4 in the process of acute graft rejection and summarise the progress in our understanding of its role in predicting the outcome. We present the results of the latest studies investigating the link between CTLA4 gene variability and AHSCT, as well as organ transplantation outcomes. While some studies found a link between +49 A/G and −318 C/T and transplantation outcomes, comprehensive meta-analyses have failed to present any association. The most recent field reviews suggest that the −1772 T/C (rs733618) CC genotype is weakly associated with a lower risk of acute graft rejection, while +49 A/G might be clinically meaningful when investigated in the context of combinations with other polymorphisms. Studies verifying associations between 12 CTLA4 gene SNPs and AHSCT outcomes present inexplicit results. Some of the most commonly studied polymorphisms in this context include +49 A/G (rs231775) and CT60 A/G (rs3087243). The results signify that, in order to understand the role of CTLA4 and its gene polymorphisms in transplantology, further studies must be conducted.     

Sensitization of MCF7 Cells with High Notch1 Activity by Cisplatin and Histone Deacetylase Inhibitors Applied Together

Histone deacetylase inhibitors (HDIs) are promising anti-cancer agents that inhibit proliferation of many types of cancer cells including breast carcinoma (BC) cells. In the present study, we investigated the influence of the Notch1 activity level on the pharmacological interaction between cisplatin (CDDP) and two HDIs, valproic acid (VPA) and suberoylanilide hydroxamic acid (SAHA, vorinostat), in luminal-like BC cells. The type of drug–drug interaction between CDDP and HDIs was determined by isobolographic analysis. MCF7 cells were genetically modified to express differential levels of Notch1 activity. The cytotoxic effect of SAHA or VPA was higher on cells with decreased Notch1 activity and lower for cells with increased Notch1 activity than native BC cells. The isobolographic analysis demonstrated that combinations of CDDP with SAHA or VPA at a fixed ratio of 1:1 exerted additive or additive with tendency toward synergism interactions. Therefore, treatment of CDDP with HDIs could be used to optimize a combined therapy based on CDDP against Notch1-altered luminal BC. In conclusion, the combined therapy of HDIs and CDDP may be a promising therapeutic tool in the treatment of luminal-type BC with altered Notch1 activity.