Protease activated receptor 4 limits bacterial growth and lung pathology during late stage Streptococcus pneumoniae induced pneumonia in mice

Summary

Streptococcus pneumoniae is a common causative pathogen of pneumonia and sepsis. Pneumonia and sepsis are associated with enhanced activation of coagulation, resulting in the production of several host-derived proteases at the primary site of infection and in the circulation. Serine proteases cleave protease activated receptors (PARs), which form a molecular link between coagulation and inflammation. PAR4 is one of four subtypes of PARs and is widely expressed by multiple cell types in the respiratory tract implicated in pulmonary inflammation, by immune cells and by platelets. In mice, mouse (m)PAR4 is the only thrombin receptor expressed by platelets. We here sought to determine the contribution of mPAR4 to the host response during pneumococcal pneumonia. Pneumonia was induced by intranasal inoculation with S. pneumoniae in mPAR4-deficient (par4^-/- ) and wild-type mice. Mice were sacrificed after 6, 24 or 48 hours (h). Blood, lungs, liver and spleen were collected for analyses. Ex vivo stimulation assays were performed with S. pneumoniae and mPAR4 activating peptides. At 48 h after infection, higher bacterial loads were found in the lungs and blood of par4^-/- mice (p < 0.05), accompanied by higher histopathology scores and increased cytokine levels (p < 0.05) in the lungs. Ex vivo, co-stimulation with mPAR4 activating peptide enhanced the whole blood cytokine response to S. pneumoniae. Thrombin inhibition resulted in decreased cytokine release after S. pneumoniae stimulation in human whole blood. Our findings suggest that mPAR4 contributes to antibacterial defence during murine pneumococcal pneumonia.

Keywords

Protease activated receptors - Streptococcus pneumoniae - thrombin - platelet immunology - infectious diseases

• References

• 1 van der Poll T, Opal SM. Pathogenesis, treatment, and prevention of pneumococcal pneumonia. Lancet 2009; 374: 1543-1556.
• 2 Dockrell DH, Whyte MK, Mitchell TJ. Pneumococcal pneumonia: mechanisms of infection and resolution. Chest 2012; 142: 482-491.
• 3 Garau J. Treatment of drug-resistant pneumococcal pneumonia. Lancet Infect Dis 2002; 02: 404-415.
• 4 Levi M, Schultz MJ, Rijneveld AW. et al. Bronchoalveolar coagulation and fibrinolysis in endotoxemia and pneumonia. Crit Care Med 2003; 31 (Suppl. 04) S238-S242.
• 5 Shpacovitch V, Feld M, Bunnett NW. et al. Protease-activated receptors: novel PARtners in innate immunity. Trends Immunol 2007; 28: 541-550.
• 6 Peters T, Henry PJ. Protease-activated receptors and prostaglandins in inflammatory lung disease. Br J Pharmacol 2009; 158: 1017-1033.
• 7 Sokolova E, Reiser G. A novel therapeutic target in various lung diseases: airway proteases and protease-activated receptors. Pharmacol Ther 2007; 115: 70-83.
• 8 Ando S, Otani H, Yagi Y. et al. Protease-activated receptor 4-mediated Ca2+ signaling in mouse lung alveolar epithelial cells. Life Sci 2007; 81: 794-802.
• 9 Ando S, Otani H, Yagi Y. et al. Proteinase-activated receptor 4 stimulation-induced epithelial-mesenchymal transition in alveolar epithelial cells. Respir Res 2007; 08: 31
• 10 Vergnolle N, Derian CK, D’Andrea MR. et al. Characterization of thrombin-induced leukocyte rolling and adherence: a potential proinflammatory role for proteinase-activated receptor-4. J Immunol 2002; 169: 1467-1473.
• 11 Moffatt JD, Lever R, Page CP. Effects of inhaled thrombin receptor agonists in mice. Br J Pharmacol 2004; 143: 269-275.
• 12 Kahn ML, Zheng YW, Huang W. et al. A dual thrombin receptor system for platelet activation. Nature 1998; 394: 690-694.
• 13 Kataoka H, Hamilton JR, McKemy DD. et al. Protease-activated receptors 1 and 4 mediate thrombin signaling in endothelial cells. Blood 2003; 102: 3224-3231.
• 14 Fujiwara M, Jin E, Ghazizadeh M. et al. Activation of PAR4 induces a distinct actin fiber formation via p38 MAPK in human lung endothelial cells. J Histochem Cytochem 2005; 53: 1121-1129.
• 15 Soh UJ, Dores MR, Chen B. et al. Signal transduction by protease-activated receptors. Br J Pharmacol 2010; 160: 191-203.
• 16 Rijneveld AW, Weijer S, Bresser P. et al. Local activation of the tissue factor-factor VIIa pathway in patients with pneumonia and the effect of inhibition of this pathway in murine pneumococcal pneumonia. Crit Care Med 2006; 34: 1725-1730.
• 17 Van Den Boogaard FE, Brands X, Schultz MJ. et al. Recombinant human tissue factor pathway inhibitor exerts anticoagulant, anti-inflammatory and antimicrobial effects in murine pneumococcal pneumonia. J Thromb Haemost 2011; 09: 122-132.
• 18 Rijneveld AW, Florquin S, Bresser P. et al. Plasminogen activator inhibitor type-1 deficiency does not influence the outcome of murine pneumococcal pneumonia. Blood 2003; 102: 934-939.
• 19 Hahn I, Klaus A, Janze AK. et al. Cathepsin G and neutrophil elastase play critical and nonredundant roles in lung-protective immunity against Streptococcus pneumoniae in mice. Infect Immun 2011; 79: 4893-4901.
• 20 Sambrano GR, Weiss EJ, Zheng YW. et al. Role of thrombin signalling in platelets in haemostasis and thrombosis. Nature 2001; 413: 74-78.
• 21 Duitman J, Hoogendijk AJ, Groot AP. et al. CCAAT-enhancer binding protein delta (C/EBPdelta) protects against Klebsiella pneumoniae-induced pulmonary infection: potential role for macrophage migration. J Infect Dis 2012; 206: 1826-1835.
• 22 Lammers AJ, de Porto AP, Florquin S. et al. Enhanced vulnerability for Streptococcus pneumoniae sepsis during asplenia is determined by the bacterial capsule. Immunobiology 2011; 216: 863-870.
• 23 Dessing MC, Florquin S, Paton JC. et al. Toll-like receptor 2 contributes to antibacterial defence against pneumolysin-deficient pneumococci. Cell Microbiol 2008; 10: 237-246.
• 24 Paterson GK, Orihuela CJ. Pneumococci: immunology of the innate host response. Respirology 2010; 15: 1057-1063.
• 25 Schouten M, Wiersinga WJ, Levi M. et al. Inflammation, endothelium, and coagulation in sepsis. J Leukoc Biol 2008; 83: 536-545.
• 26 Connolly AJ, Ishihara H, Kahn ML. et al. Role of the thrombin receptor in development and evidence for a second receptor. Nature 1996; 381: 516-519.
• 27 Connolly TM, Condra C, Feng DM. et al. Species variability in platelet and other cellular responsiveness to thrombin receptor-derived peptides. Thromb Haemost 1994; 72: 627-633.
• 28 Asokananthan N, Graham PT, Fink J. et al. Activation of protease-activated receptor (PAR)-1, PAR-2, and PAR-4 stimulates IL-6, IL-8, and prostaglandin E2 release from human respiratory epithelial cells. J Immunol 2002; 168: 3577-3585.
• 29 Slofstra SH, Bijlsma MF, Groot AP. et al. Protease-activated receptor-4 inhibition protects from multiorgan failure in a murine model of systemic inflammation. Blood 2007; 110: 3176-3182.
• 30 Lan RS, Knight DA, Stewart GA. et al. Role of PGE(2) in protease-activated receptor-1, -2 and -4 mediated relaxation in the mouse isolated trachea. Br J Pharmacol 2001; 132: 93-100.
• 31 Semple JW, Italiano Jr. JE, Freedman J. Platelets and the immune continuum. Nat Rev Immunol 2011; 11: 264-274.
• 32 Yeaman MR. Platelets in defense against bacterial pathogens. Cell Mol Life Sci 2010; 67: 525-544.
• 33 Tang YQ, Yeaman MR, Selsted ME. Antimicrobial peptides from human platelets. Infect Immun 2002; 70: 6524-6533.
• 34 Krauel K, Potschke C, Weber C. et al. Platelet factor 4 binds to bacteria, [corrected] inducing antibodies cross-reacting with the major antigen in heparin-induced thrombocytopenia. Blood 2011; 117: 1370-1378.
• 35 Monroe DM, Hoffman M, Roberts HR. Platelets and thrombin generation. Arterioscler Thromb Vasc Biol 2002; 22: 1381-1389.
• 36 Su X, Camerer E, Hamilton JR. et al. Protease-activated receptor-2 activation induces acute lung inflammation by neuropeptide-dependent mechanisms. J Immunol 2005; 175: 2598-2605.
• 37 Borensztajn K, Duitman J, Bruggemann LW. et al. Protease-activated receptor-4 deficiency does not protect against bleomycin-induced pulmonary fibrosis in mice. Eur Respir J 2012; 40: 1056-1057.
• 38 Schouten M, Van’t Veer C, Roelofs JJ. et al. Protease-activated receptor-1 impairs host defense in murine pneumococcal pneumonia: a controlled laboratory study. Crit Care 2012; 16: R238.