HEP14 treatment improves ovarian function in aged mice through mitophagy enhancement and oxidative stress reducton

Ovarian senescence impacts reproductive health and accelerates the overall aging process. Despite extensive research in this area there is still a dearth of effective therapeutic strategies. We here report the rejuvenating effects of HEP14, a natural activator of PKC pathway, on aged ovarian function. Further, we detail a method to fabricate HEP14-loaded PLGA microspheres designed for controlled and sustained drug release in vivo. Transcriptomic analysis revealed a significant overlap between the transcriptional profiles of HEP14-treated aged ovaries and those of adult ovaries, suggesting molecular rejuvenation process. Histopathological evaluations following treatment demonstrated that HEP14 not only enhances mitophagy but also exhibits antioxidative properties and promotes follicular regeneration. As a result, ovarian endocrine function was ultimately restored in aged mice, evident in raised serum levels of E2, AMH, Inhibin A/B and lowered FSH levels. In vitro studies further showcased restorative effect of HEP14 on enhancement of mitophagy and mitochondrial function by the activation of the PKC-ERK1/2 pathway in senescent GCs, which are integral to the action mechanism of HEP14. These findings pave the way for new therapeutic approaches for developing therapeutic strategies aimed at improving reproductive health in aging individuals. To delve into the molecular mechanisms behind HEP14-driven ovarian function restoration in aged mice, we conducted a comparative transcriptomic analysis on ovaries from four groups of mice: young (7~8 weeks), adult (27~28 weeks), aged mice (73~74 weeks) treated with vehicle, and aged mice (73~74 weeks) treated with HEP14 (aged+HEP14 mice).

卵巢衰老（Ovarian senescence）会影响生殖健康并加速整体衰老进程。尽管该领域已开展大量研究，但目前仍缺乏有效的治疗策略。本研究报道了蛋白激酶C（PKC）通路天然激活剂HEP14对衰老卵巢功能的复壮作用。此外，本研究详细阐述了一种制备载HEP14聚乳酸-羟基乙酸共聚物（PLGA）微球的方法，该微球可实现体内药物的控释与缓释。转录组学分析显示，经HEP14处理的衰老卵巢与成年卵巢的转录谱存在显著重叠，提示其存在分子层面的复壮过程。治疗后的组织病理学评估结果表明，HEP14不仅可增强线粒体自噬（mitophagy），还具有抗氧化特性并能促进卵泡再生。最终，衰老小鼠的卵巢内分泌功能得以恢复，具体表现为血清雌二醇（E2）、抗缪勒管激素（AMH）、抑制素A/B（Inhibin A/B）水平升高，而卵泡刺激素（FSH）水平降低。体外实验进一步证实，HEP14可通过激活衰老颗粒细胞（GCs）中的PKC-ERK1/2通路，增强线粒体自噬与线粒体功能，这是HEP14发挥作用的核心机制。上述研究结果为开发针对衰老个体生殖健康改善的治疗策略提供了新的方向。为深入探究HEP14修复衰老小鼠卵巢功能的分子机制，本研究对四组小鼠的卵巢开展了对比转录组学分析：年轻组（7~8周龄）、成年组（27~28周龄）、溶剂对照衰老组（73~74周龄）以及HEP14处理衰老组（73~74周龄，简称aged+HEP14小鼠）。