Hepatocyte nuclear factor (HNF) 4␣ is a key transcription factor regulating endo/xenobiotic-metabolizing enzymes and transporters. We investigated whether microRNAs are involved in the regulation of human HNF4␣. Potential recognition elements for miR-24 (MRE24) were identified in the coding region and the 3-untranslated region (3-UTR), and those for miR-34a (MRE34a) were identified in the 3-UTR in HNF4␣ mRNA. The HNF4␣ protein level in HepG2 cells was markedly decreased by the overexpression of miR-24 and miR-34a. The HNF4␣ mRNA level was significantly decreased by the overexpression of miR-24 but not by miR-34a. In luciferase analyses in HEK293 cells, the reporter activity of plasmid containing the 3-UTR of HNF4␣ was significantly decreased by miR-34a. The reporter activity of plasmid containing the HNF4␣ coding region downstream of the luciferase gene was significantly decreased by miR-24. These results suggest that the MRE24 in the coding region and MRE34a in the 3-UTR are functional in the negative regulation by mRNA degradation and translational repression, respectively. The down-regulation of HNF4␣ by these microRNAs resulted in the decrease of various target genes such as cytochrome P450 7A1 and 8B1 as well as morphological changes and the decrease of the S phase population in HepG2 cells. We also clarified that the expressions of miR-24 and miR-34a were regulated by protein kinase C/mitogen-activated protein kinase and reactive oxygen species pathways, respectively. In conclusion, we found that human HNF4␣ was down-regulated by miR-24 and miR-34a, the expression of which are regulated by cellular stress, affecting the metabolism and cellular biology.Human hepatocyte nuclear factor 4␣ (HNF4␣, NR2A1), 3 which belongs to the nuclear hormone receptor superfamily, is highly expressed in liver and regulates the expression of various genes involved in the synthesis/metabolism of fatty acid, cholesterol, glucose, and urea (1). It is well recognized that endo/xenobiotic-metabolizing enzymes such as cytochrome P450s (CYPs), UDP-glucuronosyltransferases, sulfotransferase as well as ATP-binding cassette transporters, organic anion transporters and organic cation transporters are under the control of HNF4␣ (2). HNF4␣ transactivates the expression of target genes not only via direct binding to their regulatory sequences but also through the regulation of other transcriptional factors such as pregnane X receptor and constitutive androstane receptor, which regulate these target genes. HNF4␣ forms large transcriptional regulatory networks in the liver. Therefore, it is believed that the change of HNF4␣ expression has a great impact upon the function of the liver.Bile acids are important regulatory molecules mediating cholesterol synthesis and glucose metabolism as well as their own synthesis (3). It is well known that HNF4␣ positively regulates the expression of bile acid-synthesizing enzymes such as CYP7A1 and CYP8B1. When bile acids are accumulated, the HNF4␣-mediated transactivation is inhibited by short heterodimer p...
A new water-soluble thermoresponsive pillar[6]arene with triethylene oxide groups was synthesized. The pillar[6]arene showed lower critical solution temperature behavior in aqueous solution. Its clouding point was photoreversibly switched based on a photoresponsive host-guest system. The trans form of an azobenzene guest formed a stable 1:1 complex with the pillar[6]arene. Complexation increased the clouding point. Irradiation with UV light induced a conformation change for the azobenzene guest from the trans to cis form, and dethreading occurred because of a size mismatch between the cis form and the pillar[6]arene cavity. This dethreading decreased the clouding point. The photoresponsive host-guest system was reversible, and the clouding point could be switched by alternating irradiation with UV or visible light. We demonstrated photoresponsive reversible clear-to-turbid and turbid-to-clear transitions for the solution based on the reversible switching of the clouding point using the photosensitive host-guest system.
Extracellular vesicles (EVs) are secreted from most cells and play important roles in cell–cell communication by transporting proteins, lipids, and nucleic acids. As the involvement of EVs in diseases has become apparent, druggable regulators of EV secretion are required. However, the lack of a highly sensitive EV detection system has made the development of EV regulators difficult. We developed an ELISA system using a high-affinity phosphatidylserine-binder TIM4 to capture EVs and screened a 1567-compound library. Consequently, we identified one inhibitor and three activators of EV secretion in a variety of cells. The inhibitor, apoptosis activator 2, suppressed EV secretion via a different mechanism and had a broader cellular specificity than GW4869. Moreover, the three activators, namely cucurbitacin B, gossypol, and obatoclax, had broad cellular specificity, including HEK293T cells and human mesenchymal stem cells (hMSCs). In vitro bioactivity assays revealed that some regulators control EV secretion from glioblastoma and hMSCs, which induces angiogenesis and protects cardiomyocytes against apoptosis, respectively. In conclusion, we developed a high-throughput method to detect EVs with high sensitivity and versatility, and identified four compounds that can regulate the bioactivity of EVs.
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