Age-related Changes in Humanin Expression in the Ovarian Tissue of Rat.

Abstract

This study examined humanin expression in rat ovarian tissue, its cellular localization, and its correlation with rat age under physiological conditions. A total of 40 Sprague-Dawley rats of various ages (2, 12, 30, and 60 days old and 1 year old) were grouped by age. Immunofluorescence and immunohistochemical techniques were used to observe humanin expression and cellular location in the ovarian tissues of rats from each age group. In addition, Western blotting and Real-time quantitative reverse transcription PCR (qRT-PCR) techniques were used to measure humanin expression level in the ovarian tissues of rats from each age group. The immunofluorescence and immunohistochemical results confirmed that humanin was expressed in rat ovarian tissues. In addition, cellular localization analysis showed that humanin was expressed in the cytoplasm of oocytes, interstitial cells, granulosa cells and theca cells in all levels of follicles after the primary follicles, and in the corpus luteum. The qRT-PCR results revealed that the level of humanin expression in the ovarian tissues of 12-day-old rats was non-significantly higher than that in the ovarian tissues of 2-day-old rats (P>0.05), whereas the levels of humanin expression in the ovarian tissues of 30-day-old, 60-day-old, and 1-year-old rats were significantly lower than that in the ovarian tissues of 2-day-old rats (P<0.05). The Western blotting results demonstrated that the levels of humanin protein expression in the ovarian tissues of 60-day-old and 1-year-old rats were significantly lower than those of 2-day-old rats (P<0.01), whereas there was no significant difference in the level of humanin protein expression between the ovarian tissues of 12-day-old and 30-day-old rats. This study confirmed that humanin is expressed in the cytoplasm of various cells in rat ovarian tissues. Moreover, the level of humanin expression was highest in the ovarian tissues of 12-day-old rats, and it subsequently decreased with age. The changes in the expression of humanin in the ovary of rats at different ages will lay the foundation for the role of humanin in ovarian aging. The effect of humanin on ovarian function is worthy of further study in the future.