衰老表现为生物体随着年龄增加从分子、细胞到组织和器官等多个层次发生的 系统性的退行性变化。长期以来，衰老研究偏重衰老进程的中晚期，此时各种宏观 衰老表型充分显现，便于研究者观察和检验干预效果。对衰老起始阶段的研究一直 未得到重视。关于衰老如何发生、从什么时候开始这一基本问题的研究尚未起步。

本文以秀丽隐杆线虫（以下简称线虫）作为模式生物，通过分析衰老早期密集 时间点采样获得的野生型和长寿突变体样品的蛋白质组数据，寻找在衰老早期发生 改变且影响寿命的关键生物学过程、以及这些生物过程的作用。据文献报道，线虫 成年 24 小时内蛋白质稳态维持能力骤降，这是目前已知最早的衰老相关表型。本 文研究发现，线虫成年 24 小时内发生剧烈的分子水平变化，其中最显著的特征是 蛋白质合成代谢的改变。SUnSET 实验揭示了蛋白质合成速率在幼虫发育末期达到 顶峰，在成年早期迅速下降。蛋白质组数据提示幼虫发育末期蛋白质合成代谢旺盛 可能与成虫阶段寿命反向关联。与之呼应，从发育末期开始抑制蛋白质合成，可提 高成年早期的蛋白质稳态维持能力，并延长线虫成虫寿命。通过在不同时间窗口内 施加 RNA 干扰下调蛋白质合成，并测量其对蛋白质稳态和线虫寿命的影响，本文 发现发育末期到成年早期是关键阶段。进一步实验发现，虽然长时间抑制蛋白质合 成或改善蛋白质稳态均能延长线虫寿命，但最有效的时间窗口分别在发育末期和成 年早期。结合文献报道和本文其它实验证据，我们提出发育末期高速率的蛋白质合 成可能导致成年早期蛋白质稳态失衡，进而引发衰老。

鞘脂作为第二大类膜脂，具有广泛的生物学功能。秀丽线虫鞘脂有独特的结构。 目前关于线虫鞘脂的研究不够深入，分析方法也不够成熟。深入解析鞘脂功能需要 考虑到鞘脂种类和结构的多样性。针对线虫鞘脂分析，本文建立了一个高效简便的 液相色谱-串联质谱方法，成功鉴定到三类（神经酰胺、鞘磷脂、葡萄糖神经酰胺） 共 54 个鞘脂，并对其中 47 个进行了定量。本文逐个表征了 12 个鞘脂合成代谢酶， 对其基因突变或沉默后鞘脂分子的变化进行了系统性分析，揭示了同工酶在底物选 择性和活性上的差异。此外，通过检测衰老过程中的鞘脂变化，发现鞘磷脂和一类 结构上具有共性的神经酰胺（脂肪酰基链不含羟基）在衰老早期明显积累。本文还 发现抑制鞘脂合成代谢也能提高成年早期的蛋白质稳态维持能力，降低鞘磷脂合成 代谢酶的表达能延长线虫寿命。

综上，本论文重点关注衰老早期分子水平的改变，发现线虫发育末期高速率的 蛋白质合成以及成虫早期特定鞘脂分子的积累皆有促进衰老的作用，抑制蛋白质合 成代谢或鞘脂合成代谢能改善成年早期的蛋白质稳态维持能力，并延长线虫寿命。

Aging is characterized by systematic age-related degenerative changes occurred at multiple levels, from the molecular and cellular level to the tissue and organ level. For a long time, research on aging has focused only on the middle or the late stage of aging, when various macroscopic aging phenotypes are fully manifested, which facilitates observation and assessment of aging phenotypes or anti-aging effects. However, research on the early stage of aging has not received sufficient attention. Explorations on how and when aging occurs in adulthood are highly limited. This study uses the nematode Caenorhabditis elegans (C. elegans) as a model organism and aims to identify key biological processes that have changed early and affect lifespan by analyzing the proteomic data of wild-type and long-lived daf-2 mutant at dense time intervals starting from the beginning of adulthood. According to the literature, protein homeostasis (proteostasis) collapsed within 24 hours of adulthood, and this is the earliest aging-related phenotype. In this study, we uncovered a dramatic molecular-level changes within 24 hours of adulthood, with the most significant feature being the alterations in protein synthesis metabolism. SUnSET assay revealed that the rate of protein synthesis peaks at the end of larval development and rapidly declines in early adulthood. Proteomic data suggest that protein synthesis activity at the end of larval development may be inversely correlated with adult lifespan. In accordance, inhibition of protein synthesis from the end of larval development stage enhances proteostasis in early adulthood and extend adult lifespan. By knocking-down protein synthesis using RNA interference from different life stages and measuring the effects on proteostasis and lifespan, it is revealed that the end of larval development and early adulthood is a critical time window. Further studies found that although life-long inhibition of protein synthesis or enhancement of proteostasis could extend lifespan, the most effective treatment time is the end of larval development or early adulthood, respectively. In combination with literature reports and other evidence in this study, we propose that very fast rate of protein synthesis at the end of larval development may lead to proteostasis collapse and aging in adulthood. Sphingolipids, as the second most abundant class of membrane lipids, exert a plethora of biological functions. Sphingolipids in C. elegans have a peculiar chemical structure. Current researches on the function of worm sphingolipids are very limited, and the analysis method are not mature enough. An in-depth analysis of sphingolipid functions requires consideration of the heterogeneity in subclasses and fatty acyl chain compositions. This study has established an effective and simple liquid chromatography-tandem mass spectrometry workflow for the analysis of worm sphingolipids, identified a total of 54 sphingolipids, including ceramide (Cer), sphingomyelins (SM), and glucosylceramides (GlcCer), and quantified 47 of them. We systematically characterized the sphingolipid profiles of 12 sphingolipid synthetic enzymes after functional depletion by gene mutation or silencing, and revealed the differential functions of isozymes in terms of the substrate selectivity and enzymatic activity. In addition, by profiling changes in sphingolipids during the aging process, we found that sphingomyelins and a subset of ceramides (without hydroxyl groups on the N-acyl chain) accumulate in the early phase of aging. Inhibition of sphingolipid biosynthesis metabolism could also improve the proteostasis maintenance capacity in early adulthood. Moreover, knock-down the expression of sphingomyelin synthases contributes to the lifespan extension of C. elegans. In summary, this thesis focuses on identifying molecular-level changes in the early phase of aging, uncovering that high rates of protein synthesis at the end of larval development and the accumulation of specific sphingolipids play a role in promoting aging. Inhibition of protein synthesis metabolism or sphingolipid synthesis metabolism can improve the capacity to maintain proteostasis in early adulthood and extend worm lifespan.