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Extracellular CIRP dysregulates macrophage bacterial phagocytosis in sepsis
Mian Zhou1, Monowar Aziz1,2, Hao-Ting Yen1
1Center for Immunology and Inflammation, The Feinstein Institutes for Medical Research, Manhasset, NY, USA.
Abstract:
In sepsis, macrophage bacterial phagocytosis is impaired, but the mechanism is not well elucidated. Extracellular cold-inducible RNA-binding protein (eCIRP) is a damage-associated molecular pattern that causes inflammation. However, whether eCIRP regulates macrophage bacterial phagocytosis is unknown. Here, we reported that the bacterial loads in the blood and peritoneal fluid were decreased in CIRP-/- mice and anti-eCIRP Ab-treated mice after sepsis. Increased eCIRP levels were correlated with decreased bacterial clearance in septic mice. CIRP-/- mice showed a marked increase in survival after sepsis. Recombinant murine CIRP (rmCIRP) significantly decreased the phagocytosis of bacteria by macrophages in vivo and in vitro. rmCIRP decreased the protein expression of actin-binding proteins, ARP2, and p-cofilin in macrophages. rmCIRP significantly downregulated the protein expression of βPIX, a Rac1 activator. We further demonstrated that STAT3 and βPIX formed a complex following rmCIRP treatment, preventing βPIX from activating Rac1. We also found that eCIRP-induced STAT3 phosphorylation was required for eCIRP's action in actin remodeling. Inhibition of STAT3 phosphorylation prevented the formation of the STAT3-βPIX complex, restoring ARP2 and p-cofilin expression and membrane protrusion in rmCIRP-treated macrophages. The STAT3 inhibitor stattic rescued the macrophage phagocytic dysfunction induced by rmCIRP. Thus, we identified a novel mechanism of macrophage phagocytic dysfunction caused by eCIRP, which provides a new therapeutic target to ameliorate sepsis.
Insights
Extracellular cold-inducible RNA-binding protein (eCIRP) impairs macrophage bacterial phagocytosis during sepsis. Inhibiting eCIRP or its downstream signaling pathway improves bacterial clearance and survival.
Area of Science:
- Immunology
- Cellular Biology
- Molecular Medicine
Background:
- Sepsis is characterized by impaired macrophage bacterial phagocytosis, a critical immune defense mechanism.
- The precise molecular mechanisms underlying this phagocytic dysfunction remain incompletely understood.
- Extracellular cold-inducible RNA-binding protein (eCIRP), a damage-associated molecular pattern, is known to induce inflammation but its role in phagocytosis is unexplored.
Purpose of the Study:
- To investigate the role of extracellular cold-inducible RNA-binding protein (eCIRP) in regulating macrophage bacterial phagocytosis during sepsis.
- To elucidate the molecular pathways through which eCIRP affects phagocytic function.
- To identify eCIRP as a potential therapeutic target for sepsis treatment.
Main Methods:
- Utilized CIRP knockout (CIRP-/-) mice and anti-eCIRP antibody treatment in a sepsis model.
- Assessed bacterial loads, survival rates, and macrophage phagocytic activity in vivo and in vitro.
- Analyzed protein expression of key regulators of actin remodeling, including ARP2, p-cofilin, and βPIX, using Western blotting.
- Investigated the interaction between STAT3 and βPIX and the role of STAT3 phosphorylation via immunoprecipitation and Western blotting.
- Employed the STAT3 inhibitor stattic to assess its effect on macrophage phagocytosis.
Main Results:
- CIRP-/- mice and anti-eCIRP antibody-treated mice exhibited reduced bacterial loads and improved survival post-sepsis.
- Increased eCIRP levels correlated with decreased bacterial clearance in septic mice.
- Recombinant murine CIRP (rmCIRP) significantly impaired macrophage phagocytosis both in vivo and in vitro.
- rmCIRP treatment led to decreased expression of ARP2, p-cofilin, and βPIX, and inhibited Rac1 activation.
- eCIRP-induced STAT3 phosphorylation formed a complex with βPIX, hindering Rac1 activation and actin remodeling.
- Inhibition of STAT3 phosphorylation or targeting the STAT3-βPIX complex restored phagocytic function.
Conclusions:
- Extracellular cold-inducible RNA-binding protein (eCIRP) directly impairs macrophage bacterial phagocytosis during sepsis.
- The mechanism involves eCIRP-induced STAT3 phosphorylation, leading to the formation of a STAT3-βPIX complex that inhibits Rac1 activation and actin remodeling.
- Targeting the eCIRP-STAT3 signaling pathway presents a promising therapeutic strategy to enhance macrophage function and improve outcomes in sepsis.
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