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探討PP4在敗血症時的細胞焦亡機制
Other Title
To Explore the Mechanism of PP4 in Pyroptosis during Sepsis
Type
thesis
Date Issued
2026-07-23
Author(s)
江為睿
Advisor
楊豐名; 吳友志
Subjects
系所名稱：呼吸治療學系胸腔醫學碩士班
Publisher
呼吸治療學系胸腔醫學碩士班
Description
學位別：碩士
語文別：英文
指導教授：楊豐名; 吳友志
口試委員：楊豐名; 吳友志; 蘇秉驊; 王紹安; 鄭文豪
授權範圍：網際網路,開放日期為2026-07-30
電子論文連結：https://handle.ncl.edu.tw/11296/k2scxh
語文別：英文
指導教授：楊豐名; 吳友志
口試委員：楊豐名; 吳友志; 蘇秉驊; 王紹安; 鄭文豪
授權範圍：網際網路,開放日期為2026-07-30
電子論文連結：https://handle.ncl.edu.tw/11296/k2scxh
Abstract
Background: Sepsis is a life-threatening syndrome characterized by a dysregulated host immune response to infection, often leading to multi-organ failure and high mortality. A critical driver of this systemic inflammation is pyroptosis, a gasdermin D (GSDMD)-mediated programmed inflammatory cell death. Protein phosphatase 4 (PP4), a serine/threonine phosphatase, has been identified as a negative modulator of Toll-like receptor 4 (TLR4)-mediated innate immunity. However, its specific role in governing pyroptosis during sepsis is still unknown.
Objective: In this study, we hypothesize that PP4 serves as a negative regulator of pyroptosis by modulating TLR4-mediated signaling and subsequent inflammasome assembly. Our goal is to elucidate the molecular mechanisms by which PP4 influences GSDMD cleavage and inflammatory cytokine release.
Methods: In vitro, PP4 protein knockdown by specific siRNA in THP-1-derived macrophages stimulated by Lipopolysaccharide (LPS) was performed to evaluate the expression of GSDMD cleavage, PP4, and canonical/non-canonical pathways via Western blotting (WB) and human phospho-kinase array. In vivo, different organs harvested from PP4-overexpressing and myeloid-specific PP4 knockout (PP4-KO) mice subjected to endotoxemia were used to assess pyroptosis by WB and immunohistochemical (IHC) staining.
Results: Through dose- and time-dependent experiments, stimulation of 0.25 μg/ml LPS for 2 hours was selected to establish the cellular pyroptosis model in THP-1derived macrophages. Loss of PP4 significantly increased LPS-induced GSDMD cleavage, suggesting that PP4 may act as a negative regulator of pyroptosis in vitro. Furthermore, increasing heat shock protein 27 (HSP27) phosphorylation was found in the siPP4+LPS group, compared to siNT+LPS group. In vivo, administration of PP4 significantly decreased GSDMD cleavage in lung tissue, but not in liver or kidney. In contrast, GSDMD cleavage was increased in lung tissue collected from cecal ligation and puncture (CLP)-challenged PP4-KO mice. Mechanistically, caspase-11 was significantly increased across endotoxemia models, ranging from intraperitoneal to intranasal LPS challenge. IHC analysis further revealed that GSDMD cleavage was predominantly localized in parenchyma region of lung tissues, indicating a potential tissue-specific role of PP4 in regulating pyroptosis.
Conclusion: Taken together, these findings suggest that PP4 functions as a negative regulator of pyroptosis via regulating HSP27 phosphorylation and caspase-11 signaling during sepsis. Further studies are required to elucidate more details of molecular mechanisms in PP4-mediated pyroptosis.
Key words: pyroptosis, gasdermin D, protein phosphatase 4, heat shock protein 27, caspase-11
Objective: In this study, we hypothesize that PP4 serves as a negative regulator of pyroptosis by modulating TLR4-mediated signaling and subsequent inflammasome assembly. Our goal is to elucidate the molecular mechanisms by which PP4 influences GSDMD cleavage and inflammatory cytokine release.
Methods: In vitro, PP4 protein knockdown by specific siRNA in THP-1-derived macrophages stimulated by Lipopolysaccharide (LPS) was performed to evaluate the expression of GSDMD cleavage, PP4, and canonical/non-canonical pathways via Western blotting (WB) and human phospho-kinase array. In vivo, different organs harvested from PP4-overexpressing and myeloid-specific PP4 knockout (PP4-KO) mice subjected to endotoxemia were used to assess pyroptosis by WB and immunohistochemical (IHC) staining.
Results: Through dose- and time-dependent experiments, stimulation of 0.25 μg/ml LPS for 2 hours was selected to establish the cellular pyroptosis model in THP-1derived macrophages. Loss of PP4 significantly increased LPS-induced GSDMD cleavage, suggesting that PP4 may act as a negative regulator of pyroptosis in vitro. Furthermore, increasing heat shock protein 27 (HSP27) phosphorylation was found in the siPP4+LPS group, compared to siNT+LPS group. In vivo, administration of PP4 significantly decreased GSDMD cleavage in lung tissue, but not in liver or kidney. In contrast, GSDMD cleavage was increased in lung tissue collected from cecal ligation and puncture (CLP)-challenged PP4-KO mice. Mechanistically, caspase-11 was significantly increased across endotoxemia models, ranging from intraperitoneal to intranasal LPS challenge. IHC analysis further revealed that GSDMD cleavage was predominantly localized in parenchyma region of lung tissues, indicating a potential tissue-specific role of PP4 in regulating pyroptosis.
Conclusion: Taken together, these findings suggest that PP4 functions as a negative regulator of pyroptosis via regulating HSP27 phosphorylation and caspase-11 signaling during sepsis. Further studies are required to elucidate more details of molecular mechanisms in PP4-mediated pyroptosis.
Key words: pyroptosis, gasdermin D, protein phosphatase 4, heat shock protein 27, caspase-11