硅肺病理过程中肺泡巨噬细胞内cAMP结合蛋白的筛选与鉴定

doi:10.16352/j.issn.1001-6325.2026.08.1042

基础医学与临床 ›› 2026, Vol. 46 ›› Issue (8): 1042-1050.doi: 10.16352/j.issn.1001-6325.2026.08.1042

硅肺病理过程中肺泡巨噬细胞内cAMP结合蛋白的筛选与鉴定

周欢, 王逸阳, 张田甜^*, 王婧^*

中国医学科学院北京协和医学院基础医学研究所,呼吸和共病全国重点实验室,北京 100005

收稿日期:2026-04-14 修回日期:2026-05-25 发布日期:2026-07-22
通讯作者: ^*tiantianzhang@ibms.pumc.edu.cn; wangjing@ibms.pumc.edu.cn
基金资助:
国家科技重大专项(2024ZD0528904);国家自然科学基金(82300098)

Screening and identification of cAMP-binding proteins in alveolar macrophages during the pathological process of silicosis

ZHOU Huan, WANG Yiyang, ZHANG Tiantian^*, WANG Jing^*

State Key Laboratory of Respiratory Health and Multimorbidity, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, China

Received:2026-04-14 Revised:2026-05-25 Published:2026-07-22
Contact: ^*tiantianzhang@ibms.pumc.edu.cn; wangjing@ibms.pumc.edu.cn

摘要/Abstract

摘要： 目的通过筛选与鉴定二氧化硅(silica)刺激小鼠肺泡巨噬细胞系(MH-S)中与环磷酸腺苷(cAMP)相互结合的新型蛋白,探究硅肺病理过程中cAMP信号通路的调控机制。方法将MH-S细胞分为对照组和实验组,用酶联免疫吸附测定法(ELISA)检测细胞内cAMP水平;用偶联cAMP的琼脂糖珠进行pull-down实验,捕获与cAMP结合的蛋白;通过SDS-PAGE和银染进行可视化分析;用液相色谱-串联质谱(LC-MS/MS)蛋白质组学技术筛选潜在的cAMP结合蛋白;利用AlphaFold对部分候选蛋白进行结构预测以及结合能力评估。结果与对照组比较,实验组细胞内cAMP水平显著降低(P＜0.01),cAMP结合蛋白谱发生变化,共鉴定到5个差异上调蛋白和413个差异下调蛋白;AlphaFold预测上调的蛋白为潜在cAMP结合蛋白的可信度较高。结论 Silica刺激可改变肺泡巨噬细胞内cAMP水平及结合蛋白谱,这些新型cAMP结合蛋白可能通过增加表达水平或增强与cAMP的结合,从而参与硅肺的发生发展,为发现硅肺的潜在治疗靶点提供新的方向。

关键词: 硅肺, 肺泡巨噬细胞, cAMP结合蛋白, 蛋白质组学

Abstract: Objective To investigate the regulatory mechanisms of the cyclic adenosine monophosphate (cAMP) signaling pathway in the pathogenesis of silicosis by screening and identifying novel proteins that interact with cAMP in mouse alveolar macrophage cells (MH-S) stimulated with silica. Methods MH-S cells were divided into a control group and an experimental group. Intracellular cAMP levels were measured using enzyme-linked immunosorbent assay (ELISA). Pull-down assays were performed with cAMP-conjugated agarose beads to capture cAMP-binding proteins. These proteins were separated by SDS-PAGE and visualized by silver staining. Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics was used to screen for potential cAMP-binding proteins. AlphaFold was used to predict the structures and assess the binding capacities of selected candidate proteins. Results Compared with the control group, the intracellular cAMP level in the experimental group was significantly decreased (P＜0.01) and the profile of cAMP-binding proteins was altered. A total of 5 upregulated and 413 down- regulated proteins were identified. AlphaFold predictions indicated that the upregulated proteins might be potential cAMP-binding proteins with high confidence. Conclusions Silica stimulation can alter the intracellular cAMP levels and profile of cAMP-binding proteins in alveolar macrophages. These novel cAMP-binding proteins may participate in the occurrence and progression of silicosis by increasing expression levels or enhancing binding affinity to cAMP, providing a new avenue for the discovery of potential therapeutic targets for silicosis.

Key words: silicosis, alveolar macrophage, cAMP-binding proteins, proteomics

中图分类号:

R363.2

周欢, 王逸阳, 张田甜, 王婧. 硅肺病理过程中肺泡巨噬细胞内cAMP结合蛋白的筛选与鉴定[J]. 基础医学与临床, 2026, 46(8): 1042-1050.

ZHOU Huan, WANG Yiyang, ZHANG Tiantian, WANG Jing. Screening and identification of cAMP-binding proteins in alveolar macrophages during the pathological process of silicosis[J]. Basic & Clinical Medicine, 2026, 46(8): 1042-1050.

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硅肺病理过程中肺泡巨噬细胞内cAMP结合蛋白的筛选与鉴定

Screening and identification of cAMP-binding proteins in alveolar macrophages during the pathological process of silicosis

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