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Inducing Acute Lung Injury in Mice by Direct Intratracheal Lipopolysaccharide Instillation
Published on: July 6, 2019
Poldip2 knockdown protects against lipopolysaccharide-induced acute lung injury via Nox4/Nrf2/NF-κB signaling pathway
Yueguo Wang1, Wenwen Wang1,2, Shusheng Zhou1
1Department of Emergency Medicine, First Affiliated Hospital of USTC, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, China.
Abstract:
Polymerase δ-interacting protein 2 (Poldip2) has been reported to mediate acute lung injury (ALI); however, the underlying mechanism is not fully explored. Male C57BL/6 mice and A549 cells were used to establish the lipopolysaccharide (LPS)-induced ALI model, then the expression of Poldip2 and its effect on oxidative stress and the resulting inflammation were detected. Adeno-associated virus serotype 6 (AAV6) mediated Poldip2 knockdown was transfected into mice via intratracheal atomization. And A549 cells stimulated with LPS was used to further confirm our hypothesis in vitro. ML385, specifically inhibited the activation of the Nrf2 signaling pathway. Our data suggested that LPS stimulation remarkably increased protein levels of Nox4 and p-P65, activities of NADPH and MPO, and generation of ROS, TNF-α, and IL-1β while decreased protein levels of Nrf2 and HO-1 compared with those in NC shRNA + Saline group, which were obviously reversed by Poldip2 knockdown. Concomitantly, Poldip2 knockdown dramatically reduced contents of MDA and enhanced activities of SOD and GSH-Px compared to NC shRNA + LPS group. In vitro, we found that knockdown of Poldip2 significantly reversed LPS-induced increase protein levels of Nox4 and p-P65, activity of NADPH, and generation of ROS, TNF-α, and IL-1β, and decrease protein levels of Nrf2 and HO-1, ML385 pretreatment reversed the effects of Poldip2 knockdown mentioned above. Our study indicated that Poldip2 knockdown alleviates LPS-induced ALI via inhibiting Nox4/Nrf2/NF-κB signaling pathway.
Insights
Polymerase δ-interacting protein 2 (Poldip2) knockdown alleviates lipopolysaccharide (LPS)-induced acute lung injury (ALI) by inhibiting the Nox4/Nrf2/NF-κB pathway, reducing oxidative stress and inflammation.
Area of Science:
- Cell Biology
- Molecular Biology
- Immunology
Background:
- Acute lung injury (ALI) is a critical condition with limited therapeutic options.
- Polymerase δ-interacting protein 2 (Poldip2) is implicated in ALI pathogenesis, but its precise mechanism remains unclear.
- Understanding Poldip2's role is crucial for developing novel ALI treatments.
Purpose of the Study:
- To investigate the role of Poldip2 in lipopolysaccharide (LPS)-induced ALI.
- To elucidate the underlying molecular mechanisms involving oxidative stress and inflammation.
- To evaluate Poldip2 knockdown as a potential therapeutic strategy for ALI.
Main Methods:
- Establishment of LPS-induced ALI models in C57BL/6 mice and A549 cells.
- Adeno-associated virus serotype 6 (AAV6)-mediated Poldip2 knockdown via intratracheal atomization.
- Assessment of oxidative stress markers (ROS, MDA, SOD, GSH-Px) and inflammatory cytokines (TNF-α, IL-1β).
- Analysis of key signaling pathway proteins (Nox4, Nrf2, HO-1, p-P65) and pathway inhibition using ML385.
Main Results:
- LPS stimulation increased Nox4, p-P65, ROS, TNF-α, IL-1β, MDA, and MPO activity, while decreasing Nrf2 and HO-1.
- Poldip2 knockdown reversed these LPS-induced changes, reducing oxidative stress and inflammation.
- ML385 pretreatment counteracted the protective effects of Poldip2 knockdown, confirming pathway involvement.
Conclusions:
- Poldip2 plays a significant role in exacerbating LPS-induced ALI.
- Poldip2 knockdown alleviates ALI by inhibiting the Nox4/Nrf2/NF-κB signaling pathway.
- Targeting Poldip2 presents a promising therapeutic avenue for ALI treatment.
