Lueli Wang,1,* Rui Shi,1,* Zhaoliang Li,1 Ruoqiu Ma,1 Chongyu Wang,1,2 Changli Xu,1 Rong Guo,1 Chuang Xiao,1 Xiaohua Du,3 Weimin Yang1 

1Yunnan Key Laboratory of Pharmacology for Natural Products, Kunming Medical University, Kunming, People’s Republic of China; 2Pharmacy Department, The First People’s Hospital of Zhaotong, Zhaotong, People’s Republic of China; 3Geriatric Respiratory Department, First Affiliated Hospital of Kunming Medical University, Kunming, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Xiaohua Du, Geriatric Respiratory Department, First Affiliated Hospital of Kunming Medical University, Kunming, People’s Republic of China, Email dxiaohuabs@126.com Weimin Yang, Yunnan Key Laboratory of Pharmacology for Natural Products, Kunming Medical University, Chunrong West Road, Kunming, 650500, People’s Republic of China, Email yangweimin@kmmu.edu.cn

Objective: Chronic obstructive pulmonary disease (COPD) is a disease with a complex etiology. The secretion of inflammatory factors, such as IL-1β and oxidative stress, plays an important role in the pathogenesis of COPD. The aim of this paper is to investigate the changes in the redox protein thioredoxin (TRX) in COPD and the role TRX plays in IL-1β release.
Methods: We analyzed data from single-cell RNA sequencing (scRNA-seq) of COPD lung tissue in the GEO database. Changes in TRX expression levels and activity were assessed by protein activity analysis of alveolar macrophages(AM). Using Monocle2 and molecular dynamics(MD) to analyze which pathways through TRX achieves regulation of the inflammatory response. The analytical results were subsequently validated by constructing vivo and vitro models.
Results: AM that specifically synthesize IL-1β were identified by scRNA-seq. No change in TRX expression levels and decreased protein antioxidant activity in IL-1β+ AM with COPD. We confirmed an increase in oxidized TRX (oxTRX) and a decrease in reduced TRX (reTRX) in COPD mouse model and THP-1 cell model. The decrease in reTRX was accompanied by the upregulation of NLRP3 activity, which played a catalytic role in the synthesis of IL-1β in this subgroup. This was subsequently confirmed in a cigarette smoke-induced THP-1 cell model. A decrease in reTRX level accompanied by an upregulation of NLRP3 activity, which plays a facilitating role in the synthesis of IL-1β. We determined that reTRX reduction was followed by depolymerization of thioredoxin-interacting protein (TXNIP) through MD and immune co-precipitation (CO-IP). Then TNXIP interacted with NLRP3 and up-regulate NLRP3 activity, which in turn promoted IL-1β release.
Conclusion: Our study shows that the reTRX is abnormally altered in COPD and leads to the upregulation of NLRP3 activity in AM to enhance IL-1β production.

Keywords: chronic obstructive pulmonary disease, single-cell RNA sequencing, alveolar macrophages, thioredoxin, inflammations