Overaccumulation of Fat Caused Rapid Reproductive Senescence but not Loss of Ovarian Reserve in ob/ob Mice.

Ovarian reserve and fertility are reduced by aging and a poor energy balance. To date, the relationships of high energy accumulation and aging with the ovarian reserve have not been elucidated. Here, the effects of obesity on the aging ovarian reserve were evaluated in a leptin-deficient (ob/ob) mouse model. Abnormal estrous cyclicity appeared as early as 6 weeks and worsened with aging. The blood level patterns of 17ß-estradiol (E2), testosterone (T), and progesterone (P4) with aging were similar between lean and ob/ob mice. The blood level of E2 but not P4 or T was similar at 24 weeks. Many more atretic follicles but fewer corpora lutea were observed in ob/ob mice than in lean mice within all age groups. Anti-Müllerian hormone (Amh) mRNA levels were similar between genotypes. Dazl, Stra8, and ZP3 mRNAs were highly expressed in ob/ob mice after 12 weeks. Sohlh1 and Ybx2 mRNAs were highly expressed at 24 weeks in ob/ob compared with lean mice. In addition, SOHLH1-positive primordial follicle counts were significantly increased in ob/ob mice at 24 weeks. The proportions of AMH-positive secondary and small antral follicles were similar between genotypes. Together, these results show that the ovarian reserve lasts longer in ob/ob mice than in lean mice, suggesting that the loss of normal physiological or physical status causes decreased fertility at a young age in ob/ob mice and that an increase in adipocytes without leptin, as in ob/ob mice, can improve the ovarian reserve. Such knowledge can be applied to understanding reproductive dysfunction.

All-in-one platform for salivary cotinine detection integrated with a microfluidic channel and an electrochemical biosensor.

Medical disorders caused by second-hand smoke are a major public health concern worldwide. To estimate the level of second-hand smoke exposure, salivary diagnostics for cotinine analysis is a compelling alternative in conventional diagnostics using bio-fluids, such as blood and urine, owing to its simple and non-invasive collection method. However, there are several critical issues, such as tedious multisteps, demand for expertise, and field unavailability to collect and transport the purified saliva for further analysis. Here, an all-in-one platform is presented to simply collect real human saliva and directly deliver it onto the biosensing surface. The platform consists of a commercial cotton-swab-type collector, 3D-printed housing, and microfluidic channel integrated with an electrochemical competitive immunosensor to evaluate the level of salivary cotinine. The immunosensor is based on a competitive binding assay between cotinine-conjugated horseradish peroxidase (C-HRP) and cotinine for anti-cotinine binding sites. The current responses obtained from the HRP-thionine-H2O2 system decreased proportionally to the cotinine concentration. This immunosensor successfully detected its target over a range of 1 × 10-1 to 1 × 104 pg ml-1 with a low limit of detection of 6 × 10-2 pg ml-1 and a limit of quantification of 1 × 10-1 pg ml-1. In addition, the platform is applicable to various commercial cotton-swab-type saliva collectors and can successfully transfer the saliva in wide flow rates ranging from 0.1 to 30 ml min-1 without leakage or damage to the sensing surface. Furthermore, the practicality of the proposed platform was evaluated by measuring cotinine in real human saliva from eight non-smokers. The concentration of cotinine was from 45.7 to 890.8 pg ml-1, which was in good agreement with that measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The introduced all-in-one platform represented a reliable performance delivering simple and practical steps in salivary diagnostics.

Development of Polymeric Micelles of Oleanolic Acid and Evaluation of Their Clinical Efficacy.

Oleanolic acid has been used only as a subsidiary agent in cosmetic products. The aim of the study is to show the effect of oleanolic acid as an active ingredient for the alleviation of wrinkles in humans and to develop a polymeric micelle formulation that enables poorly soluble oleanolic acid to be used as a main ingredient in cosmetic products for reducing wrinkles. The solubility of oleanolic acid was evaluated in solubilizers, surfactants, and polymers. The particle sizes and shapes of polymeric micelles containing oleanolic acid were evaluated by electrophoretic light scattering spectrophotometer and scanning electron cryomicroscopy. Encapsulation efficiency and skin permeation were measured by HPLC. Stability of the polymeric micelles stored at 40 °C for 3 months was evaluated by visual observation, particle size measurement, and oleanolic acid content measurement. Polymeric micelles in final product ampoule form were applied around the eyes of 23 female subjects for 8 weeks. Five skin parameters were evaluated by optical profilometry every 4 weeks for 8 weeks. In addition, professionals made visual observations of the skin and a human skin irritation study was conducted. Polymeric micelles of oleanolic acid with a particle size of less than 100 nm were prepared using Capryol 90® and poloxamer. The skin permeation rate of the oleanolic acid in the polymeric micelles was higher than that in the other solutions made of oleanolic acid dispersed in 2 different surfactants. No significant changes in particle size, color, or oleanolic acid content were observed, and the polymeric micelles stored at 40 °C for 3 months did not undergo phase separation. After 8 weeks of application, skin irritation had not developed and all five parameters evaluated by optical profilometry as well as the visual evaluation scores were significantly improved. This study showed that the polymeric micelles of oleanolic acid prepared in this study were stable and effective at alleviating wrinkles in humans as the principal active ingredient. Based on these findings, it is expected that polymeric micelles of oleanolic acid can be widely used in cosmetic applications.