肠道微生物组对人类健康和疾病的作用不断取得进展，需要明确的肠道细胞模型来研究和快速评估宿主、微生物组和药物相互作用。分化的 Caco-2 细胞系通常用作药物渗透性研究的上皮模型，最近已用于研究宿主-微生物组相互作用。然而，其研究这种相互作用的适用性仍有待确定。在这里，我们采用多层次蛋白质组学来证明自发和丁酸诱导的 Caco-2 分化表现出相似的蛋白质和途径变化，包括与翻译和增殖相关的蛋白质的下调以及与宿主-微生物组相互作用有关的功能的上调，例如细胞粘附、紧密连接、细胞外囊泡和对刺激的反应。赖氨酸乙酰组学显示，组蛋白乙酰化水平随着细胞分化而降低，而与线粒体功能相关的蛋白质的乙酰化水平则增加。这项研究还表明，与自发分化方法相比，含丁酸盐培养基可加速 Caco-2 分化，并较早上调与宿主-微生物组相互作用相关的蛋白质，表明其在使用该肠道模型进行检测开发时具有优越性。总而言之，这项多组学研究强调了 Caco-2 向专门的肠上皮样细胞分化的受控进展，并确立了其研究宿主-微生物组相互作用的适用性。

Emergent advancements on the role of the intestinal microbiome for human health and disease necessitate well-defined intestinal cellular models to study and rapidly assess host, microbiome, and drug interactions. Differentiated Caco-2 cell line is commonly utilized as an epithelial model for drug permeability studies and has more recently been utilized for investigating host-microbiome interactions. However, its suitability to study such interactions remains to be characterized. Here, we employed multilevel proteomics to demonstrate that both spontaneous and butyrate-induced Caco-2 differentiations displayed similar protein and pathway changes, including the downregulation of proteins related to translation and proliferation and upregulation of functions implicated in host-microbiome interactions, such as cell adhesion, tight junction, extracellular vesicles, and responses to stimuli. Lysine acetylomics revealed that histone protein acetylation levels were decreased along with cell differentiation, while the acetylation in proteins associated with mitochondrial functions was increased. This study also demonstrates that, compared to spontaneous differentiation methods, butyrate-containing medium accelerates Caco-2 differentiation, with earlier upregulation of proteins related to host-microbiome interactions, suggesting its superiority for assay development using this intestinal model. Altogether, this multiomics study emphasizes the controlled progression of Caco-2 differentiation toward a specialized intestinal epithelial-like cell and establishes its suitability for investigating the host-microbiome interactions.