Dr. Chao’s Research

Extracellular RNA, inflammation, and sepsis

Sepsis is life-threatening organ dysfunction caused by a dysregulated host response to infection. Cardiovascular collapse induced by cardiac dysfunction and profound vasodilatation represents a main feature of septic shock and contributes to its high mortality. Dysregulated innate immune response to pathogen components and endogenous host danger molecules proves to be deleterious in sepsis. We have discovered that host cellular RNAs are released into the blood during sepsis and that circulating host RNA is closely associated with sepsis severity in animals. miRNA array identifies six miRNAs that are elevated during sepsis; four of them (miR-34a, -122, -145, -146a) induce marked innate immune activation in immune cells, cardiomyocytes, and in intact animals. The over goal of this proposal is to determine the role of ex-miRNAs in host innate immune activation and in cardiac bioenergetic dysfunction in sepsis. The proposal is based on the following 3 hypotheses: 1) that host miRNAs are released from injured cells and play an important role in innate immune activation and cardiac dysfunction in sepsis, 2) that ex-miRNAs act through endosomal TLR7 signaling, 3) that pharmacological blockade of miRNAs after polymicrobial infection will attenuate innate immune activation and offer cardiac and survival benefit in sepsis. In Aim 1, we will determine the function and mechanism of plasma exosomes and ex-miRNAs in mediating innate immune responses in sepsis. In Aim 2, we will define the impact of ex-miRNAs to sepsis-induced cardiac bioenergetic dysfunction. In Aim 3, through a series of proof-of-concept pre-clinical studies, we will determine the therapeutic efficacy of synthetic anti-miR oligonucleotide inhibitors to modulate organ inflammation, organ injury, and mortality in sepsis. This proposal addresses a unique function of endogenous ex-miRNAs in host innate immunity and organ dysfunction with a significant implication to sepsis pathogenesis, treatment, and miRNA biology.

Related Publications

Sepsis-Induced Coagulopathy: A Comprehensive Narrative Review of Pathophysiology, Clinical Presentation, Diagnosis, and Management Strategies
Williams, Brittney MD, Zou, Lin MD, PhD, Pittet, Jean-Francois MD, Chao, Wei MD, PhD

Physiological hemostasis is a balance between pro- and anticoagulant pathways, and in sepsis, this equilibrium is disturbed, resulting in systemic thrombin generation, impaired anticoagulant activity, and suppression of fibrinolysis, a condition termed sepsis-induced coagulopathy (SIC). SIC is a common complication, being present in 24% of patients with sepsis and 66% of patients with septic shock, and is often associated with poor clinical outcomes and high mortality.  Recent preclinical and clinical studies have generated new insights into the molecular pathogenesis of SIC. In this article, the authors analyze the complex pathophysiology of SIC with a focus on the role of procoagulant innate immune signaling in hemostatic activation––tissue factor production, thrombin generation, endotheliopathy, and impaired antithrombotic functions. Clinical presentations of SIC are also reviewed, the diagnostic scoring system and laboratory tests, the current standard of care, and clinical trials evaluating the efficacies of anticoagulant therapies.

Emerging Role of Extracellular RNA in Innate Immunity, Sepsis, and Trauma
Brittney Williams, Rosemary Kozar, Wei Chao

Sepsis and trauma remain the leading causes of morbidity and mortality. The understanding of the molecular pathogenesis in development of multiple organ dysfunction in sepsis and trauma has evolved as more focus is on secondary injury from innate immunity, inflammation, and the potential role of endogenous danger molecules. Studies have generated evidence for extracellular RNAs (exRNAs) as biologically active mediators in health and disease. The authors review studies on plasma exRNA profiling in mice and humans with sepsis and trauma, the role and mode of action by exRNAs, such as ex-micro(mi)RNAs, in host innate immune response, and their potential implications in various organ injury during sepsis and trauma.

Innate immune TLR7 signaling mediates platelet activation and platelet-leukocyte aggregate formation in murine bacterial sepsis.
Williams B, Zhu J, Zou L, Chao W.

Thrombocytopenia is a common complication in sepsis and is associated with higher mortality. Activated platelets express CD62P, which facilitates platelet-leukocyte aggregate (PLA) formation and contributes to thrombocytopenia in sepsis. The authors have reported that thrombocytopenia in murine sepsis is partly attributable to TLR7 signaling, but the underlying mechanism is unclear. In the current study, they tested the hypothesis that TLR7 mediates platelet activation and PLA formation during sepsis.

TLR7 Mediates Acute Respiratory Distress Syndrome in Sepsis by Sensing Extracellular miR-146a. 
Huang H, Zhu J, Gu L, Hu J, Feng X, Huang W, Wang S, Yang Y, Cui P, Lin SH, Suen A, Shimada BK, Williams B, Kane MA, Ke Y, Zhang CO, Birukova AA, Birukov KG, Chao W, Zou L.

Toll-like receptor 7 (TLR7), the sensor for single-stranded RNA, contributes to systemic inflammation and mortality in murine polymicrobial sepsis. Recent studies show that extracellular miR-146a-5p serves as a TLR7 ligand and plays an important role in regulating host innate immunity. However, the role of miR-146a-5p and TLR7 signaling in pulmonary inflammation, endothelial activation, and sepsis-associated acute respiratory distress syndrome remains unclear. Here, the authors show that intratracheal administration of exogenous miR-146a-5p in mice evokes lung inflammation, activates endothelium, and increases endothelial permeability via TLR7-dependent mechanisms.

Pyruvate-Driven Oxidative Phosphorylation is Downregulated in Sepsis-Induced Cardiomyopathy: A Study of Mitochondrial Proteome. 
Shimada BK, Boyman L, Huang W, Zhu J, Yang Y, Chen F, Kane MA, Yadava N, Zou L, Lederer WJ, Polster BM, Chao W.

Evidence suggests that cardiac mitochondrial oxidative phosphorylation is attenuated in sepsis, but the underlying molecular mechanisms remain incompletely understood. Data from this study, utilizing adult male mice of 9-12 weeks old, demonstrates that broad mitochondrial protein remodeling, pyruvate dehydrogenase inactivation and impaired pyruvate-fueled oxidative phosphorylation during SIC, provide a molecular framework for further exploration.

Role of Extracellular MicroRNA-146a-5p in Host Innate Immunity and Bacterial Sepsis.
Wang S, Yang Y, Suen A, Zhu J, Williams B, Hu J, Chen F, Kozar R, Shen S, Li Z, Jeyaram A, Jay MS, Zou L, Chao W.

Data from this study supports a critical role for extracellular miR-146a-5p in innate immune inflammation and sepsis pathogenesis and offers a rationale for selective targeting extracellular miRNAs as a novel strategy for sepsis intervention.

Brain Innate Immune Response via miRNA-TLR7 Sensing in Polymicrobial Sepsis
Zou L, He J, Gu L, Shahror RA, Li Y, Cao T, Wang S, Zhu J, Huang H, Chen F, Fan X, Wu J, Chao W.

Taking a loss-of-function approach, the data from this study found a contributory role of endogenous miR-146a and TLR7 sensing in animal SAE. Hence, suggesting extracellular miRNA sensing via TLR7 as a possible link of neuroinflammation within the brain microenvironment during polymicrobial sepsis.

Therapeutic Potential of Extracellular Vesicles for Sepsis Treatment
Kronstadt SM, Pottash AE, Levy D, Wang S, Chao W, Jay SM

Sepsis is a deadly condition lacking a specific treatment despite decades of research. This has prompted the exploration of new approaches, with extracellular vesicles (EVs) emerging as a focal area. EVs are nanosized, cell-derived particles that transport bioactive components (i.e., proteins, DNA, and RNA) between cells, enabling both normal physiological functions and disease progression depending on context.

miR-19b targets pulmonary endothelial syndecan-1 following hemorrhagic shock
Wu F, Wang JY, Chao W, Sims C, Kozar RA.

This study presents evidence that miR-19b targets syndecan-1 mRNA to downregulate its expression. The results demonstrate that miR-19b was increased in hemorrhagic shock patients and in-vitro specifically bound to syndecan-1 mRNA and caused its degradation.

Extracellular miR-146a-5p Induces Cardiac Innate Immune Response and Cardiomyocyte Dysfunction
Shimada BK, Yang Y, Zhu J, Wang S, Suen A, Kronstadt SM, Jeyaram A, Jay SM, Zou L, Chao W.

Previous studies have demonstrated that transient myocardial ischemia leads to release of cellular nucleic acids such as RNA. Extracellular RNA reportedly plays a pivotal role in myocardial inflammation and ischemic injury in animals.

Hypobaria Exposure Worsens Cardiac Function and Endothelial Injury in an Animal Model of Polytrauma: Implications for Aeromedical Evacuation
Lopez K, Suen A, Yang Y, Wang S, Williams B, Zhu J, Hu J, Fiskum G, Cross A, Kozar R, Miller C, Zou L, Chao W.

Aeromedical evacuation can expose traumatically injured patients to low pressure (hypobaria) and hypoxia. This study sought to assess the impact of hypobaria on inflammation, organ injury, and mortality in a mouse model of polytrauma.

Targeting Toll-Like Receptors in Sepsis – From Bench to Clinical Trials
Chen F, Zou L, Williams B, Chao W.

This study reviews various TLRs and the specific molecules these TLRs sense – both the pathogen-associated and host-derived stress molecules, and their converging signaling pathways. We critically analyze preclinical investigations into the role of TLRs in animal sepsis, the complexity of targeting TLRs for sepsis intervention, and the disappointing clinical trials of the TLR4 antagonist eritoran.

Enhanced loading of functional miRNA cargo via pH-gradient modification of extracellular vesicles.
Jeyaram A, Lamichhane TN, Wang S, Dahal E, Kronstadt SM, Levy D, Parajuli B, Knudsen D, Chao W, Jay SM.

Based on their identification as physiological nucleic acid carriers in humans and other organisms, extracellular vesicles (EVs) have been explored as therapeutic delivery vehicles for DNA, RNA, and other cargo.

Toll-like receptor 2 and 7 mediate coagulation activation and coagulopathy in murine sepsis.
Williams B, Neder J, Cui P, Suen A, Tanaka K, Zou L, Chao W.

Sepsis is a life-threatening condition often manifested as marked inflammation and severe coagulopathy. Toll-like receptors (TLRs) play a pivotal role in inflammation, organ dysfunction, and mortality in animal sepsis…

Toll-like Receptor 7 Contributes to Inflammation, Organ Injury, and Mortality in Murine Sepsis.
Jian W, Gu L, Williams B, Feng Y, Chao W, Zou L.

Sepsis remains a critical illness with high mortality. The authors have recently reported that mouse plasma RNA concentrations are markedly increased during sepsis and closely associated with its severity. Toll-like receptor 7, originally identified as the sensor for single-stranded RNA virus, also mediates host extracellular RNA-induced innate immune responses in vitro and in vivo. Here, the authors hypothesize that innate immune signaling via Toll-like receptor 7 contributes to inflammatory response, organ injury, and mortality during polymicrobial sepsis…

Circulating Plasma Extracellular Vesicles from Septic Mice Induce Inflammation via MicroRNA- and TLR7-Dependent Mechanisms.
Xu J, Feng Y, Jeyaram A, Jay SM, Zou L, Chao W.

We have previously reported that a group of host cellular microRNAs (miRNAs; miR-34a-5p, miR-122-5p, miR-145-5p, miR-146a-5p, miR-210-3p) are released into the blood during sepsis, some of which are capable of inducing complement activation, cytokine production, and leukocyte migration. Extracellular vesicles (EVs) have been proposed as vehicles for extracellular miRNA-mediated intercellular communication. However, the biological function of plasma EVs and the associated miRNAs in sepsis are largely unknown…

Complement factor B is the downstream effector of Toll-like receptors and plays an important role in a mouse model of severe sepsis.
Zou L, Feng Y, Li Y, Zhang M, Chen C, Cai JY, Gong Y, Wang L, Thurman J, Wu X, Atkinson JP, Chao W.

Severe sepsis involves massive activation of the innate immune system and leads to high mortality. Previous studies have demonstrated that various types of Toll-like receptors (TLRs) mediate a systemic inflammatory response and contribute to organ injury and mortality in animal models of severe sepsis…

Myocardial ischemia induces a rapid activation of innate immune signaling via cardiac heat-shock protein 60 and Toll-like receptor 4.
Li Y, Feng Y, Chen H, Zou L, Si R, Wang E, Zhang M, Warren S, Sosnovik D, Chao W.

Innate immune response after transient ischemia is the most common cause of myocardial inflammation and may contribute to injury, yet the detailed signaling mechanisms leading to such a response are not well understood. Herein we tested the hypothesis that myocardial ischemia activates interleukin receptor-associated kinase-1 (IRAK-1), a kinase critical for the innate immune signaling such as that of Toll-like receptors (TLRs), via a mechanism that involves heat shock proteins (HSPs) and TLRs…

Role of extracellular RNA and TLR3-Trif signaling in myocardial ischemia-reperfusion injury.
Chen C, Feng Y, Zou L, Chen HH, Cai JY, Xu JM, Sosnovik DE, Chao W.

Toll-like receptor 3 (TLR3) was originally identified as the receptor for viral RNA and represents a major host antiviral defense mechanism. TLR3 may also recognize extracellular RNA (exRNA) released from injured tissues under certain stress conditions. However, a role for exRNA and TLR3 in the pathogenesis of myocardial ischemic injury has not been tested. This study examined the role of exRNA and TLR3 signaling in myocardial infarction (MI), apoptosis, inflammation, and cardiac dysfunction during ischemia-reperfusion (I/R) injury.

Cardiac RNA induces inflammatory responses in cardiomyocytes and immune cells via Toll-like receptor 7 signaling.
Feng Y, Chen H, Cai J, Zou L, Yan D, Xu G, Li D, Chao W.

We have recently reported that extracellular RNA (exRNA) released from necrotic cells induces cytokine production in cardiomyocytes and immune cells and contributes to myocardial ischemia/reperfusion injury. However, the signaling mechanism by which exRNA exhibits its pro-inflammatory effect is unknown. Here we hypothesize that exRNA directly induces inflammation through specific Toll-like receptors (TLRs)…

Splenic RNA and microRNA mimics promote complement factor B production and alternative pathway activation via innate immune signaling.
Zou L, Feng Y, Xu G, Jian W, Chao W.

Complement factor B (cfB) is an essential component of the alternative pathway (AP) and plays an important role in the pathogenesis of polymicrobial sepsis. However, the mechanism leading to cfB production and AP activation during sepsis remains poorly understood. In this study, we found that plasma cell-free RNA was significantly increased following cecal ligation and puncture (CLP), an animal model of polymicrobial sepsis, and was closely associated with sepsis severity…

Extracellular microRNAs induce potent innate immune responses via TLR7/MyD88-dependent mechanisms.
Feng Y, Zou L, Yan D, Chen H, Xu G, Jian W, Cui P, Chao W.

Tissue ischemia, such as transient myocardial ischemia, leads to release of cellular RNA including microRNA(miRNA) into the circulation and extracellular (ex-) space, but the biological function of the ex-RNA is poorly understood. We recently reported that cardiac RNA of both human and rodent origins induced cytokine production and immune cell activation…