Xuebijing Ameliorates Cytokine Storm in Sepsis by Regulating the ACTN4/STAT1 Pathway in Monocytes

Monday, March 23, 2026

9:00 AM - 10:00 AM Central Time

Location: Connections Central - RST 16

Dingji Hu, MD

CHN

Disclosure(s): No relevant financial relationship(s) to disclose.
YZ

Yike Zhu, MD

Resident Physician
Jiangsu Provincial Key Laboratory of Critical Care Medicine, Department of Critical Care Medicine, Zhongda Hospital, School of Medicine, Southeast University, Nanjing 210009, Jiangsu, China

Disclosure information not submitted.
JX

Jianfeng Xie, MD, PhD

Professor of ICU & Chief Physician
Department of Critical Care Medicine, Zhongda Hospital, School of Medicine, Southeast University, Nanjing 210009, China.

Disclosure information not submitted.
HQ

Haibo Qiu, MD, PhD

Professor of ICU & Chief Physician
Department of Critical Care Medicine, Zhongda Hospital, School of Medicine, Southeast University, Nanjing 210009, China.

Disclosure information not submitted.
SL

Songqiao Liu, MD, PhD

Chief Physician
Jiangsu Provincial Key Laboratory of Critical Care Medicine, Department of Critical Care Medicine, Zhongda Hospital, School of Medicine, Southeast University, Nanjing 210009, Jiangsu, China

Disclosure information not submitted.

Introduction: Monocytes play a crucial role in infection and immune responses by secreting various cytokines, which initiate and regulate inflammatory reactions. While moderate cytokine release is essential for immune defense, excessive cytokine production can lead to a cytokine storm, causing tissue damage and multiple organ dysfunction, severely impacting clinical outcomes in sepsis patients. Previous studies suggested that Xuebijing injection (XBJ) exerts therapeutic effects in sepsis by modulating interferon signaling in monocytes, but the specific molecular targets and mechanisms require experimental validation.

Methods: Human monocytes (THP-1 cells) were stimulated with LPS to establish an inflammatory model, with PBS as control. Cell viability assays determined the safe and optimal therapeutic concentrations of XBJ. Four groups were designed: PBS+Veh, PBS+XBJ, LPS+Veh, and LPS+XBJ. After LPS stimulation, cells were treated with the optimal concentration of XBJ or vehicle (Veh). Interferons, interferon-stimulated genes, and inflammatory cytokines were quantified via RT-qPCR and ELISA. Drug affinity responsive target stability (DARTS) combined with ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-HRMS) identified direct protein targets of XBJ. AlphaFold 3 predicted the target protein structure, followed by virtual screening and molecular docking to pinpoint key bioactive components. WB and Co-IP assays validated target expression and pathway interactions.

Results: LPS significantly upregulated type I interferons, interferon-stimulated genes, and pro-inflammatory cytokines. XBJ treatment reversed these effects, while no significant changes were observed for type II interferon. DARTS-MS identified α-actinin-4 (ACTN4) as a direct target of XBJ. Molecular docking revealed strong binding affinity between ACTN4 and two XBJ components: Miltionone II and stigmasterol. WB demonstrated LPS-induced downregulation of ACTN4 and upregulation of STAT1, both reversed by XBJ. Co-IP confirmed interaction between STAT1 and ACTN4.

Conclusions: Xuebijing injection mitigates sepsis-associated cytokine storm by modulating the ACTN4/STAT1 pathway in monocytes. ACTN4 is a direct target, with Miltionone II and stigmasterol as potential key active constituents.