Silver nanocolloid, a dense suspension of ligand-encapsulated silver nanoparticles, is an important material for printing-based device production technologies. However, printed conductive patterns of sufficiently high quality and resolution for industrial products have not yet been achieved, as the use of conventional printing techniques is severely limiting. Here we report a printing technique to manufacture ultrafine conductive patterns utilizing the exclusive chemisorption phenomenon of weakly encapsulated silver nanoparticles on a photoactivated surface. The process includes masked irradiation of vacuum ultraviolet light on an amorphous perfluorinated polymer layer to photoactivate the surface with pendant carboxylate groups, and subsequent coating of alkylamine-encapsulated silver nanocolloids, which causes amine–carboxylate conversion to trigger the spontaneous formation of a self-fused solid silver layer. The technique can produce silver patterns of submicron fineness adhered strongly to substrates, thus enabling manufacture of flexible transparent conductive sheets. This printing technique could replace conventional vacuum- and photolithography-based device processing.

Epigenetic modifications, such as DNA methylation and histone modifications, are dynamically altered predominantly in paternal pronuclei soon after fertilization. To identify which histone modifications are required for early embryonic development, we utilized histone K-M mutants, which prevent endogenous histone methylation at the mutated site. We prepared four single K-M mutants for histone H3.3, K4M, K9M, K27M, and K36M, and demonstrate that overexpression of H3.3 K4M in embryos before fertilization results in developmental arrest, whereas overexpression after fertilization does not affect the development. Furthermore, loss of H3K4 methylation decreases the level of minor zygotic gene activation (ZGA) predominantly in the paternal pronucleus, and we obtained similar results from knockdown of the H3K4 methyltransferase Mll3/4. We therefore conclude that H3K4 methylation, likely established by Mll3/4 at the early pronuclear stage, is essential for the onset of minor ZGA in the paternal pronucleus, which is necessary for subsequent preimplantation development in mice.

Since spermatogonial stem cells (SSCs) are capable of both self-renewal and differentiation to daughter cells for subsequent spermatogenesis, the development of an efficient in vitro culture system is essential for studies related to spermatogenesis. Although the currently available system is serum-free and contains only chemically-defined components, it highly relies upon bovine serum albumin (BSA), a component with batch-to-batch quality variations similar to those of fetal bovine serum. Thus, we searched for an alternative BSA-free culture system that preserved the properties of SSCs. In this study, we utilized Knockout Serum Replacement (KSR) in the SSC culture medium, as a substitute for BSA. The results demonstrated that KSR supported the continuous growth of SSCs in vitro and the SSC activity in vivo without BSA, in a feeder-cell combination with mouse embryonic fibroblasts. The addition of BSA to KSR further facilitated cell cycle progression, whereas a transplantation assay revealed that the addition of BSA did not affect the number of SSCs in vivo. The combination of KSR with BSA also allowed the elimination of GFRA1 and FGF2, and the reduction of the GDNF concentration from 20 ng/ml to 5 ng/ml, while maintaining the growth rate and the expression of SSC markers. Furthermore, KSR was also useful with SSCs from non-DBA/2 strains, such as C57BL/6 and ICR. These results suggested that KSR is an effective substitute for BSA for long-term in vitro cultures of SSCs. Therefore, this method is practical for various studies related to SSCs, including spermatogenesis and germ stem cell biology.

Metastasis is the major cause of cancer-related deaths due to the lack of effective therapies. Emerging evidence suggests that certain epigenetic and transcriptional regulators drive cancer metastasis and could be targeted for metastasis treatment. To identify epigenetic regulators of breast cancer metastasis, we profiled the transcriptomes of matched pairs of primary breast tumors and metastases from human patients. We found that distant metastases are more immune inert with increased M2 macrophages compared to their matched primary tumors. The acetyl-lysine reader, cat eye syndrome chromosome region candidate 2 (CECR2), was the top up-regulated epigenetic regulator in metastases associated with an increased abundance of M2 macrophages and worse metastasis-free survival. CECR2 was required for breast cancer metastasis in multiple mouse models, with more profound effect in the immunocompetent setting. Mechanistically, the nuclear factor κB (NF-κB) family member v-rel avian reticuloendotheliosis viral oncogene homolog A (RELA) recruits CECR2 to increase chromatin accessibility and activate the expression of their target genes. These target genes include multiple metastasis-promoting genes, such as TNC , MMP2 , and VEGFA , and cytokine genes CSF1 and CXCL1 , which are critical for immunosuppression at metastatic sites. Consistent with these results, pharmacological inhibition of CECR2 bromodomain impeded NF-κB–mediated immune suppression by macrophages and inhibited breast cancer metastasis. These results reveal that targeting CECR2 may be a strategy to treat metastatic breast cancer.

Cadmium (Cd) from a zinc mine polluted Jinzu River basin, Toyama prefecture and produced a disease with severe bone pain (Itai-itai disease), from the early part of the 20th century. Cd caused renal tubular dysfunction and osteomalacia accompanied by osteoporosis. The main symptoms of the disease were bone pain, pseud fracture (Looser's zone), fracture, and renal tubular dysfunction. The pollution of paddy-field soil by Cd still remains and our epidemio1ogica1 studies of the inhabitants living in the Cd-polluted areas show the presence of renal tubular dysfunction and decrease of bone mass; characteristic of Itai-itai disease.

Epigenetic regulators have been implicated in tumorigenesis of many types of cancer; however, their roles in endothelial cell cancers such as canine hemangiosarcoma (HSA) have not been studied. In this study, we found that lysine-specific demethylase 2B (Kdm2b) was highly expressed in HSA cell lines compared to normal canine endothelial cells. Silencing of Kdm2b in HSA cells resulted to increased cell death in vitro compared to the scramble control by inducing apoptosis through the inactivation of the DNA repair pathways and accumulation of DNA damage. Similarly, doxycycline-induced Kdm2b silencing in tumor xenografts resulted to decreased tumor sizes compared to the scramble control. Furthermore, Kdm2b was also highly expressed in clinical cases of HSA, and its expression levels was higher than in hemangioma, a benign counterpart of HSA. Based on these results, we hypothesized that pharmacological Kdm2b inhibition can also induce HSA cell death and can be used as an alternative treatment for HSA. We treated HSA cells with GSK-J4, a histone demethylase inhibitor, and found that GSK-J4 treatment also induced apoptosis and cell death. On top of that, GSK-J4 treatment in HSA tumor-bearing mice decreased tumor sizes without any obvious side-effects. In this study, we demonstrated that Kdm2b acts as an oncogene in HSA by enhancing DNA damage response and can be used as a biomarker to differentiate HSA from hemangioma. Moreover, we indicated that histone demethylase inhibitor GSK-J4 can be used as a therapeutic alternative to doxorubicin for HSA treatment.

Canine hemangiosarcoma (HSA) is one of the most common mesenchymal tumors in dogs. Its high metastatic and growth rates are usually associated with poor prognosis. Neoplastic cells of HSA can show various levels of cellular atypia in the same mass and may consist of various populations at different differentiated stages. Up to present, however, there is no report analyzing their differentiation states by comparing cellular atypia with differentiation-related protein expressions. To evaluate whether cellular atypia can be used as a differentiation marker in HSA, we analyzed correlation between cellular atypia and intensities of CD31 and von Willebrand Factor (vWF) staining in HSA cases. We also compared cellular atypia and expression levels of CD31 and vWF in each growth patterns. Our results show that cellular atypia was negatively correlated to CD31 and vWF expression levels but no significant correlation was found between growth patterns and cellular atypia or CD31 and vWF expression levels. Our study suggests that cellular atypia is useful for identifying differentiation levels in HSA cases. This study also provides useful information to determine differentiation levels of cell populations within HSA based only on morphological analysis, which will aid further HSA research such as identifying undifferentiation markers of endothelial cells or finding undifferentiated cell population in tissue sections.

In vivo fluorescent imaging technique is a strong tool to visualize the various cellular events such as the proliferation, differentiation, migration, and a lineage tracing in living cells requiring no further experimental procedure such as immunostaining. During spermatogenesis, unique and dynamic histone exchanges occur. Since the timing and types of histone exchanges defines the particular stages of spermatogenesis, visualizing certain types of histones in testes is useful not only for researching specific histone dynamics, but also for monitoring the stages of spermatogenesis in vivo. In this study, we report the establishment of two transgenic (Tg) mouse lines expressing histone H4-Venus (H4V) and histone H3.3-mCherry (H33C) fusion proteins in testicular germ cells, and demonstrated their utility for monitoring germ cell development in vivo. Because of the choice of promoter as well as the nature of these histones, H4V and H33C were exclusively expressed in the germ cells of the distinct stages, which allowed the determination of spermatogenic stages in real time. In addition, disappearance of H4V and H33C at particular stages of differentiation/fertilization also represented dynamic histone removal. Collectively, these Tg mice are a valuable resource not only for monitoring differentiation stages, but also for studying the chromatin dynamics of post-natal testicular germ cell development in vivo.