This study aimed to investigate the effectiveness of carrageenan (CGN) as an oral immune adjuvant. During the initial research, the inadvertent shallow insertion of an oral gavage needle confirmed CGN’s effect as an adjuvant for esophageal immunization. However, in oral immunization, antibody formation was not observed regardless of CGN’s presence or absence as an adjuvant. Conversely, when bovine serum albumin (used as an antigen) was introduced into the esophagus along with CGN, it resulted in the production of antigen-specific IgG. An exploration was conducted to ascertain whether CGN’s adjuvant effects were associated with prolonging the antigen’s residence time in the esophagus. Upon introducing the antigen into the esophagus without CGN, it was undetectable at two minutes post-introduction. Conversely, when administered with CGN, the antigen remained detectable in the esophagus for up to five minutes post-introduction. To investigate whether this immune response was elicited through mucosal immune mechanisms in the esophagus, the production of IgA, a representative immunoglobulin of mucosal immunity, was assessed. Following esophageal immunization with CGN as an adjuvant, total IgG, IgG1, and IgG2a were detected in serum, while IgA was not detected. These findings suggest that under specific conditions, the esophagus may serve as a site for initiating a novel immune response.

Aurora A kinase is a mitotic serine/threonine kinase whose proposed functions include the maturation of centrosomes, G2/M transition, alignment of chromosomes at metaphase, and cytokinesis. In this study, we investigated the effect of MLN8237, an aurora A kinase inhibitor, on the postovulatory aging of oocytes based on the frequency of oocyte fragmentation, cdk1 kinase activity, and cyclin B degradation. The fragmentation of ovulated oocytes during prolonged culture was inhibited by treatment with MLN8237 in a concentration-dependent manner. The frequency of fragmented oocytes was significantly lower in oocytes treated with 2 μM MLN8237 (13%) than in control oocytes (64%) after two days of culture. Most of the control (non-fragmented) oocytes (91%) were activated after two days of culture. In comparison, only 22% of the MLN8237-treated oocytes were activated; the rest of the oocytes (78%) were still in metaphase with an abnormal spindle and dispersed chromosomes. Next, cdk1 activity and the level of cyclin B were examined. The level of cyclin B and cdk1 activity in MLN8237-treated oocytes were nearly equal to those in control oocytes. Our results indicate that MLN8237 inhibited the fragmentation of ovulated oocytes during prolonged culture, although it blocked the spontaneous decrease in activity of cdk1 and degradation of cyclin B. This mechanism of inhibition is different from that in oocytes treated with nocodazole, which have high levels of cdk1 activity and cyclin B.

Oocyte enucleation is essential for somatic cell nuclear transfer (SCNT) in the production of cloned animals or embryonic stem cells from adult somatic cells. Most studies of oocyte enucleation have been performed using micromanipulator-based techniques, which are technically demanding, time-consuming, and expensive. Several recent studies have used chemical-induced oocyte enucleation; however, each has been plagued by low efficiency and toxicity. In this study, I found that the co-treatment of murine oocytes with demecolcine and BMI-1026, a potent cdk1 inhibitor, resulted in a high enucleation rate (97%). This method is entirely independent of a micromanipulator and is suitable for the large-scale production of enucleated oocytes. This new method of enucleation will be useful in SCNT and in the development of handmade cloning techniques.