Spatiotemporal Role of the PKD1-NLRP3 axis in Sepsis-induced Cardiomyopathy

M. Ghait (Greifswald)¹, L. Lang (Greifswald)¹, N. Dörmann (Greifswald)², J. Rüdebusch (Greifswald)¹, B. Brinschwitz (Greifswald)³, S. Schwanz (Greifswald)², E. Martin (Greifswald)¹, P. Giri (Greifswald)¹, K. Bartels (Greifswald)⁴, B. Fielitz (Greifswald)⁴, E. Hammer (Greifswald)⁵, U. Völker (Greifswald)⁵, J. Fielitz (Greifswald)^2
1University Medicine Greifswald, Clinic for Internal Medicine B The partner site Greifswald of the German Center of Cardiovascular Research (DZHK) Greifswald, Deutschland; ²Universitätsmedizin Greifswald Klinik und Poliklinik für Innere Medizin B Greifswald, Deutschland; ³Universitätsmedizin Greifswald Klinik für Innere Medizin B Greifswald, Deutschland; ⁴University Medicine Greifswald Department of Internal Medicine B, Cardiology, DZHK (German Center for Cardiovascular Research), partner site Greifswald Greifswald, Deutschland; ⁵Interfaculty Institute for Genetics and Functional Genomics Greifswald, Deutschland

Sepsis is a major cause of death in critically ill patients and sepsis-induced cardiomyopathy (SICM) contributes to this unfavorable outcome. We reported that the NLRP3 inflammasome, which mediates the production of interleukin-1β (IL-1β) and induces pyroptosis during sepsis, is activated and contributes to SICM. NLRP3 undergoes phosphorylation by protein kinase D1 (PKD1) at the trans-Golgi network to mediate proximity-induced caspase-1 autoactivation. Active caspase 1 mediates IL-1β maturation and promotes activation of the membrane pore-forming protein gasdermin-D to facilitate the release of IL-1β and execute pyroptosis.
We investigated whether the PKD1-NLRP3 signalling axis is contained in cardiomyocytes and if cardiomyocyte-specific deletion of PKD1 (Prkd1) preserves cardiac function in sepsis.  Male cardiomyocyte-specific Prkd1 knockout (cKO; Prkd1^{loxP/loxP; αMHC-CRE}) mice and wild type littermate controls (WT; Prkd1^loxP/loxP) were exposed to polymicrobial sepsis by cecal ligation and puncture (CLP) surgery (cKO, n = 8; WT, n = 9) to induce SICM (permit nr. 7221.3-1.1-14/18). Sham‐treated mice served as controls (cKO, n = 4; WT, n = 4). SICM was assessed by echocardiography 24 hours after surgery. Neonatal rat ventricular cardiomyocytes (NRCM) were primed with lipopolysaccharide (LPS) followed by NLRP3 inflammasome activation with nigericin. PKD1 was inhibited with CRT0066101. Changes in protein contents in different subcellular compartments and differences in RNA expression were assessed by immunoblotting and RT-qPCR, respectively.
cKO mice showed a preserved cardiac function and less cardiac atrophy in response to sepsis as indicated by improved left ventricular ejection fraction and fractional shortening and a diminished cardiac stress response. NRCM underwent an intensive NLRP3 activation and induction of pyroptosis that was PKD1 dependent. Mechanistically, we found that PKD1 in cardiomyocytes is post-transcriptionally required for the spatial arrangement of NLRP3 from mitochondria to endomembrane fractions, a necessary step for assembly of the mature NLRP3 inflammasome. Our results indicate that PKD1 functions as a key mediator of SICM and regulates the spatial and temporal organization of the NLRP3 inflammasome in cardiomyocytes.
Despite therapeutic advances, SICM worsens the prognosis of critically ill patients. Clinical data suggest that IL-1β, which is activated by the NLRP3 inflammasome downstream of PKD1, impairs cardiac function. Thus, inhibition of the PKD1-NLRP3 axis in critically ill patients may help to control the immune response and prevent cardiac dysfunction in sepsis.