C-type natriuretic peptide (CNP) is a recently identified meiotic inhibitor in mice. However, it has not been investigated in porcine oocytes to date. This study aimed to demonstrate the inhibitory effect of CNP against germinal vesicle breakdown (GVBD) in porcine oocyte meiotic resumption. Immunohistochemical analysis revealed intense natriuretic peptide receptor 2 (NPR2) immunoreactivity in the oocyte surrounded cumulus cells in the follicles. Furthermore, reverse transcription polymerase chain reaction (RT-PCR) analysis showed the expression of npr2 mRNA only in cumulus cells but not in oocytes, suggesting that cumulus cells are the targets of CNP. When cumulus-oocyte complexes (COCs) or denuded oocytes (DOs) were cultured with various concentrations of CNP (10, 50, 100, 500, and 1,000 nM), inhibitory effect was observed in the COC group, but not in the DO group, confirming that CNP indirectly inhibits GVBD via cumulus cells. This evidence is the first indication that the CNP-NPR2 pathway is involved in meiotic arrest in porcine oocytes. Furthermore, we investigated the effect of oocyte-derived paracrine factor (ODPF) on npr2 mRNA expression level in cumulus cells by evaluating changes in mRNA expression in oocytectomised COCs (OXCs) by real-time PCR. A significant decrease in npr2 mRNA expression level was observed in OXCs, whereas mRNA expression level was restored in OXCs with DOs, indicating that ODPF participates in the regulation of npr2 expression in porcine cumulus cells.

A novel SAAF was isolated from the title ascidian. The structure was elucidated using the entire sample of 4 nmol, suggesting that the position of the OH group confers genus-specificity to sperm chemotaxis in ascidians. This study not only provides insight into the chemical tactics in sperm chemotaxis but demonstrates that the innovative techniques allow structure determination of natural products in trace amounts.

Historically, vaccine safety assessments have been conducted by animal testing (e.g., quality control tests and adjuvant development). However, classical evaluation methods do not provide sufficient information to make treatment decisions. We previously identified biomarker genes as novel safety markers. Here, we developed a practical safety assessment system used to evaluate the intramuscular, intraperitoneal, and nasal inoculation routes to provide robust and comprehensive safety data. Influenza vaccines were used as model vaccines. A toxicity reference vaccine (RE) and poly I:C-adjuvanted hemagglutinin split vaccine were used as toxicity controls, while a non-adjuvanted hemagglutinin split vaccine and AddaVax (squalene-based oil-in-water nano-emulsion with a formulation similar to MF59)-adjuvanted hemagglutinin split vaccine were used as safety controls. Body weight changes, number of white blood cells, and lung biomarker gene expression profiles were determined in mice. In addition, vaccines were inoculated into mice by three different administration routes. Logistic regression analyses were carried out to determine the expression changes of each biomarker. The results showed that the regression equations clearly classified each vaccine according to its toxic potential and inoculation amount by biomarker expression levels. Interestingly, lung biomarker expression was nearly equivalent for the various inoculation routes. The results of the present safety evaluation were confirmed by the approximation rate for the toxicity control. This method may contribute to toxicity evaluation such as quality control tests and adjuvant development.