Protocol to detect neutral lipids with BODIPY staining in myeloid-derived suppressor cells in mouse mammary tumors.

Neutral lipids affect the immunosuppressive function of myeloid-derived suppressor cells (MDSCs). Here, we present a protocol for measuring neutral lipids in MDSCs using BODIPY from mouse mammary tumor derived from triple-negative breast cancer cells, 4T1, which is applicable to other mammary tumors of interest. We describe steps for 4T1 cell culture, single-cell isolation from tumors, staining of cells with antibodies and BODIPY, and flow cytometry.

ΔNp63 regulates MDSC survival and metabolism in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) contributes greatly to mortality of breast cancer, demanding new targetable options. We have shown that TNBC patients have high expression in tumors. However, the function of ΔNp63 in established TNBC is yet to be explored.

The triazole fungicide metconazole inhibits the homodimerization of human androgen receptors to suppress androgen-induced transcriptional activation.

We assessed the mechanism of human androgen receptor-mediated endocrine-disrupting effect by a triazole fungicide, metconazole in this study. The internationally validated stably transfected transactivation (STTA) in vitro assay, which was established for determination of a human androgen receptor (AR) agonist/antagonist by using 22Rv1/MMTV_GR-KO cell line, alongside an in vitro reporter-gene assay to confirm AR homodimerization was used. The STTA in vitro assay results showed that metconazole is a true AR antagonist.

Chlorpropham, a carbamate ester herbicide, has an endocrine-disrupting potential by inhibiting the homodimerization of human androgen receptor.

This study was carried out to provide the evidence with respect to the adverse potential of chlorpropham, a representative carbamate ester herbicide product, on the endocrine system by using in vitro testing methods in accordance with the Organization for Economic Cooperation and Development Test Guideline No. 458 (22Rv1/MMTV_GR-KO human androgen receptor [AR] transcriptional activation assay) and a bioluminescence resonance energy transfer-based AR homodimerization assay. Results revealed that chlorpropham had no AR agonistic effects, but it was determined to be a true AR antagonist without intrinsic toxicity against the applied cell lines.

Hyperacetylation of the C-terminal domain of p53 inhibits the formation of the p53/p21 complex.

Given our previous finding that certain tumor-suppressing functions of p53 are exerted by the p53/p21 complex, rather than p53 alone, cells may have a system to regulate the p53/p21 interaction. As p53 binds to p21 via its C-terminal domain, which contains acetylable lysine residues, we investigated whether the C-terminal acetylation of p53 influences the p53/p21 interaction. Indeed, the p53/p21 interaction was reduced when various types of cells (HCT116 colon cancer, A549 lung cancer, and MCF7 breast cancer cells) were treated with MS-275, an inhibitor of SIRT1 (a p53 deacetylase), or with SIRT1-targeting small interfering RNAs.

Involvement of the p53/p21 complex in p53-dependent gene expression.

The p53 tumor suppressor regulates cell functions either by acting as a transcription factor or by interacting with other proteins. Previously, we reported that the non-transcriptional actions of p53 can be facilitated by the binding of p53 to p21. Herein, we investigated whether p53/p21 interaction influences the transcriptional activity of p53.

Next-Generation Intestinal Toxicity Model of Human Embryonic Stem Cell-Derived Enterocyte-Like Cells.

The gastrointestinal tract is the most common exposure route of xenobiotics, and intestinal toxicity can result in systemic toxicity in most cases. It is important to develop intestinal toxicity assays mimicking the human system; thus, stem cells are rapidly being developed as new paradigms of toxicity assessment. In this study, we established human embryonic stem cell (hESC)-derived enterocyte-like cells (ELCs) and compared them to existing and models.

Slug promotes p53 and p21 protein degradation by inducing Mdm2 expression in HCT116 colon cancer cells.

Our previous study revealed that the tumor suppressor/transcription factor p53 directly binds to its transcriptional target, p21, and that the p53/p21 complex binds to zinc finger protein SNAI2 (Slug), a tumor promoter/transcription factor; thereby promoting the degradation of Slug by Mdm2, an E3 ligase. The present study demonstrated that Slug reduced the cellular expression levels of p53 and p21 in HCT116 colon cancer by decreasing their protein stability. In parallel, Slug increased the mRNA and protein expression levels of Mdm2 in these cells.

Toxicity Assessment of Transfluthrin, Benzyl Butyl Phthalate, and 17β-Estradiol on the Primary Fibroblast of the Striped Field Mouse, Apodemus agrarius.

Environmental pollution (EP) is a well-known threat to wild animals, but its toxicological impact is poorly understood. In vitro toxicity evaluation using cells of lower predators could be a promising way to assess and monitor the effects of EPs on whole wildlife populations that are related in the food web. Here, we describe EPs' toxic effect and mechanism in the primary fibroblast derived from the embryo of the striped field mouse, Apodemus agrarius.

Albendazole exerts antiproliferative effects on prostate cancer cells by inducing reactive oxygen species generation.

Benzimidazole derivatives are used for their antihelmintic properties, but have also been reported to exert anticancer effects. In the present study, the anticancer effects of albendazole on prostate cancer cells were assessed using proliferation, clonogenic and migration assays. To investigate the anticancer mechanisms of albendazole, reactive oxygen species (ROS) levels were measured, and the expression of genes associated with oxidative stress and Wnt/β-catenin signaling was confirmed by reverse transcription-quantitative PCR and western blotting.

p21 promotes p53 protein degradation by facilitating p53-Wip1 and p53-Mdm2 interaction.

Downregulation of the p53 tumor suppressor in cancers is frequently accompanied by the upregulation of Wip1 (a phosphatase) and Mdm2 (an E3 ubiquitin ligase). Mdm2 binds and ubiquitinates p53, promoting its degradation by the proteasome. As the p53/Mdm2 interaction is alleviated by the phosphorylation of the serine-15 (S15) residue of p53, Wip1, which can directly dephosphorylate phospho-S15, facilitates the Mdm2-mediated degradation of p53.

TALEN-mediated generation of Nkx3.1 knockout rat model.

Background: Recent developments in gene editing, using transcriptional activator-like effector nucleases (TALENs), have greatly helped the generation of genetically engineered animal models. The NK3 homeobox 1 (NKX3.1) protein plays important roles in prostate development and protein production, and functions as a tumor suppressor.

Vitrification for cryopreservation of 2D and 3D stem cells culture using high concentration of cryoprotective agents.

Background: Vitrification is the most promising technology for successful cryopreservation of living organisms without ice crystal formation. However, high concentrations (up to ~ 6-8 M) of cryoprotective agents (CPAs) used in stem cell induce osmotic and metabolic injuries. Moreover, the application of conventional slow-freezing methods to cultures of 3-D organoids of stem cells in various studies, is limited by their size.

Loss of glutathione peroxidase 3 induces ROS and contributes to prostatic hyperplasia in Nkx3.1 knockout mice.

Background: Glutathione peroxidase 3 (Gpx3) protects cells from oxidative stress, and its reduced expression in human prostate cancer has been reported.

Objectives: We hypothesized that Gpx3 might play an important role in the development of prostatic intraepithelial neoplasia (PIN), a pre-cancerous state of the prostate, and aimed to highlight the underlying molecular mechanism.

Materials And Methods: The following double-knockout mice Nkx3.

Development and evaluation of next-generation cardiotoxicity assay based on embryonic stem cell-derived cardiomyocytes.

In accordance with requirements of the ICH S7B safety pharmacology guidelines, numerous next-generation cardiotoxicity studies using human stem cell-derived cardiomyocytes (CMs) are being conducted globally. Although several stem cell-derived CMs are being developed for commercialization, there is insufficient research to verify if these CMs can replace animal experiments. In this study, in vitro high-efficiency CMs derived from human embryonic stem cells (hESC-CMs) were compared with Sprague-Dawley rats as in vivo experimental animals, and primary cultured in vitro rat-CMs for cardiotoxicity tests.

Improved human hematopoietic reconstitution in HepaRG co-transplanted humanized NSG mice.

Several humanized mouse models are being used to study humanspecific immune responses and diseases. However, the pivotal needs of fetal tissues for the humanized mice model have been huddled because of the demand for ethical and medical approval. Thus, we have verified the hematopoietic and immunomodulatory function of HepaRG and developed a new and easy humanized mouse model to replace the use of fetal liver tissue.

Pimozide Inhibits the Human Prostate Cancer Cells Through the Generation of Reactive Oxygen Species.

The United States Food and Drug Administration-approved antipsychotic drug, pimozide, has anticancer activities. However, the role of reactive oxygen species (ROS) in its effect on prostate cancer is not well-known. We examined cell proliferation, colony formation, migration, ROS production, and the expression of antioxidant-related genes after treatment of human prostate cancer PC3 and DU145 cells with pimozide.

Correction to: Phloretin Inhibits the Human Prostate Cancer Cells through the Generation of Reactive Oxygen Species.

AbstractThe original version of this article unfortunately contained an error in Figs. 1, 5 and 6. The asterisks and bars indicating statistical significance were missing in the figures.

Phenotyping analysis of p53 knockout mice produced by gene editing and comparison with conventional p53 knockout mice.

Background: Knockout (KO) mice developed by homologous recombination (HR) have become useful tools to elucidate gene function. However, HR has low KO efficiency and is time-consuming, labor-intensive, and expensive. 'Gene editing' has received much attention for efficient genetic manipulation.

Phloretin Inhibits the Human Prostate Cancer Cells Through the Generation of Reactive Oxygen Species.

Phloretin is a flavonoid with known anticancer activities. However, we do not fully understand how phloretin mitigates prostate cancer on the molecular level. In the present study, we examined changes in proliferation, colony formation, and migration after phloretin treatment in human prostate cancer cells PC3 and DU145.

Trimethyltin chloride induces reactive oxygen species-mediated apoptosis in retinal cells during zebrafish eye development.

Trimethyltin chloride (TMT), one of the most widely used organotin compounds in industrial and agricultural fields, is widespread in soil, aquatic systems, foodstuffs and household items. TMT reportedly has toxic effects on the nervous system; however, there is limited information about its effects on eye development and no clear associated mechanisms have been identified. Therefore, in the present study, we investigated eye morphology, vison-related behavior, reactive oxygen species (ROS) production, apoptosis, histopathology, and gene expression to evaluate the toxicity of TMT during ocular development in zebrafish embryos.

Establishment, characterization, and toxicological application of a spontaneous immortalized cell line from the striped field mouse, Apodemus agrarius.

It is important to secure various biological resources in situations of diminishing wildlife genetic diversity. Cultured cells are useful bioresources because they can stably store genetic information for a long time and can be expanded efficiently. Here, we established fibroblast cell lines from Apodemus agrarius as a new living resource.

Troglitazone inhibits the migration and invasion of PC-3 human prostate cancer cells by upregulating E-cadherin and glutathione peroxidase 3.

Troglitazone (TGZ) is a synthetic peroxisome proliferator-activated receptor γ (PPARγ) ligand that exhibits potential antitumor effects on a number of cancer subtypes, including prostate cancer. However, little is known about the effect of TGZ on metastasis in prostate cancer. The aim of the present study was to determine the inhibitory effect and mechanism underlying TGZ on cell growth, migration and invasion using the prostate cancer PC-3 cell line.

Respiratory Toxicity of Polyhexamethylene Guanidine Phosphate Exposure in Zebrafish.

Humidifier disinfectants containing polyhexamethylene guanidine phosphate (PHMG-P) can induce pulmonary toxicity and has caused human casualties in South Korea since 2006. Thereby, the safety evaluation of household chemicals such as PHMG-P has garnered increased importance. However, many limitations, such as the lack of specialized facilities and animal welfare concerns associated with the use of murine models, persist.

A case of active incomplete biliary cirrhosis in an aged female Japanese macaque (Macaca fuscata).

We describe the first case of biliary cirrhosis in Japanese macaque. Clinical signs had not been detected. The liver was nodular.