Article Text

GW24-e3668 TXNIP mediates NLRP3 inflammasome activation in cardiac microvascular endothelial cells as a novel mechanism in myocardial ischaemia/reperfusion injury
  1. Liu Yi,
  2. Ling Tao
  1. Department of Cardiology, Xijing Hospital, The Fourth Military Medical University


Objectives The present study sought to determine the role of the NLRP3 inflammasome in MI/R injury and to investigate further the signalling pathways that lead to the activation of NLRP3 inflammasome by MI/R injury

Methods Two methods were utilised for blocking NLRP3 signalling in mice: Firstly, small-interfering RNA (siRNA) gene silencing technique was used to knockdown NLRP3 expression by three separate intramyocardial injections. Hearts were subjected myocardial ischaemia/reperfusion (MI/R) 48 hours after siRNA injection. Secondly, BAY 11-7028, an inflammasome inhibitor was administered via intraperitoneal (IP) injection 10 min prior to MI/R. Murine neonatal cardiomyocytes and cardiac microvascular endothelial cells (CMECs) were subjected to hypoxia/reoxygenation (H/R) injury. Before H/R injury, the CMECs were treated with TXNIP-specific siRNA for 48 hours. The cardiac function and myocardial infarct size were determined by echocardiography and TTC, respectively. The macrophage and neutrophil infiltration were assessed by immunohistochemistry. The IL-1β and IL-18 levels were assessed using ELISA kit. LDH release and caspase-3 activity were determined using an enzyme activity assay kit.

Results The ischaemic heart exhibited enhanced inflammasome activation as evidenced by increased NLRP3 and caspase-1 expression and increased IL-1β production. Cardiac NLRP3 was knocked down using either an intramyocardial siRNA injection or an intraperitoneal injection of BAY 11-7028, an inflammasome inhibitor. Knocking down cardiac NLRP3 attenuated macrophage and neutrophil infiltration and decreased MI/R injury, as measured by cardiomyocyte apoptosis and infarct size. In vitro experiments revealed that NLRP3 was expressed in cardiac microvascular endothelial cells (CMECs) but was hardly expressed in cardiomyocytes. Hypoxia/reoxygenation (H/R) stimulated NLRP3 inflammasome activation in CMECs but not in cardiomyocytes. Moreover, CMECs subjected to H/R injury increased both Txnip expression and interactions between Txnip and NLRP3, the latter of which has been shown to be a mechanism for activating NLRP3. Txnip siRNA diminished NLRP3 inflammasome activation and hypoxia/reoxygenation induced injury, as measured by LDH release and apoptosis in CMECs. ROS scavenger dissociated TXNIP from NLRP3 and inhibited the activation of NLRP3 inflammasome in the CMECs.

Conclusions Several novel observations were found in this study. First, NLRP3 inflammasomes in myocardial resident cells played an essential role in the pathophysiology of MI/R injury. Second, NLRP3 inflammasomes activation occured in CMECs but not in cardiomyocytes, indicating that CMECs may mediate inflammation in response to I/R. Third, Txnip activated NLRP3 inflammasomes in MI/R injury in a ROS dependent fashion. Finally, blocking Txnip/NLRP3 signalling inhibited the activation of NLRP3 inflammasomes and may provide insight into novel therapies for mitigating MI/R injury.

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