一条新型母乳来源肽ANECP-1防治NEC的功能与机制研究

The ANECP-1, identified by peptidomics techniques, is an peptide derived from casein protein of breast milk without any function report. Previous study showed that ANECP-1 alleviated LPS caused enterocytes apoptosis and inflammation status, simultaneously restored the proliferation ability. The study of in vivo also exhibited a protective role of ANECP-1 in intestine injury and inflammation. Besides, pull down assay revealed ANECP-1 interacted with COX-2, which had been reported to be involved in TLR4 signaling that regulated enterocytes apoptosis, proliferation and inflammation, suggesting COX-2 could be the target of ANECP-1. In future work, we aim to determine the content of ANECP-1 in different breast milk and evaluate its safety. Meanwhile, more NEC cell models and animal models would be employed to confirm ANECP-1’s function. At last, physical methods and rescue approach would also be conducted to elucidate the mechanism that ANECP-1 works. Our work may provide new insights into the prevention and treatment of NEC.

ANECP-1是我们借助多肽组学发现的，一个功能未知的母乳酪蛋白来源多肽。前期发现，ANECP-1可降低LPS诱导的肠上皮细胞凋亡和炎症因子释放，恢复细胞增殖能力；在体实验显示，ANECP-1可显著缓解NEC大鼠模型的肠道损伤和炎症水平；Pull down实验提示COX-2为ANECP-1的可能结合靶标，有报道COX-2介导TLR4信号引起的肠细胞凋亡、增殖和炎症反应，提示ANECP-1可能通过调控COX-2发挥其生物学功能。研究拟通过检测不同母乳中ANECP-1含量的变化、评估ANECP-1的毒副作用，论证其防治NEC的可行性；通过细胞与动物实验，论证其保护肠上皮细胞损伤的作用及防治NEC的可能性；借助物理学方法和挽救实验策略，深入阐明COX-2介导ANECP-1发挥功能的分子机制。研究可为NEC防治提供新手段。

坏死性小肠结肠炎（Necrotizing enterocolitis, NEC）是早产儿最常见且致命的肠道疾病之一，严重威胁新生儿健康，目前针对NEC的干预措施十分有限，积极寻找更安全、有效的NEC防治方法已十分迫切。临床与基础研究表明，母乳含有多种生物活性分子，且对NEC具有良好的防治效果。借助多肽组学分析，我们从母乳蛋白水解产物中寻找到一条多肽ANECP-1具有良好的理化性质和潜在的生物学活性。临床相关性分析显示，ANECP-1在早产儿母乳及初乳中具有相对高丰度。体外实验证明ANECP-1抑制LPS诱导的人、大鼠肠上皮细胞的凋亡且不影响肠上皮细胞增殖，ANECP-1还可抑制巨噬细胞M1型极化并促进M2型相关抗炎相关标志物的表达。体内实验显示，ANECP-1显著改善NEC动物模型导致的肠损伤，减轻炎症反应。进一步通过免疫共沉淀及挽救实验对ANECP-1的作用机制进行了探索，发现ANECP-1通过靶向FHL2并调控NF-κB信号发挥抑制肠上皮细胞凋亡和巨噬细胞炎症反应的功能。综上，本研究系统阐明了ANECP-1在体外与体内对NEC肠损伤及炎症的抑制功能，并对机制进行了探索，不仅为母乳喂养防治NEC提供了理论基础，也为NEC干预药物的研发提供新线索。