Contributing factors in the development of acute lung injury in a murine double hit model

  • Philipp StörmannEmail author
  • Nils Becker
  • Leander Künnemeyer
  • Sebastian Wutzler
  • Jan Tilmann Vollrath
  • Thomas Lustenberger
  • Frank Hildebrand
  • Ingo Marzi
  • Borna Relja
Original Article



Blunt chest (thoracic) trauma (TxT) is known to contribute to the development of secondary pulmonary complications. Of these, acute lung injury (ALI) is common especially in multiply injured patients and might not only be due to the direct trauma itself, but seems to be caused by ongoing and multifactorial inflammatory changes. Nevertheless, the exact mechanisms and contributing factors of the development of ALI following blunt chest trauma are still elusive.


60 CL57BL/6N mice sustained either blunt chest trauma combined with laparotomy without further interventions or a double hit (DH) including TxT and cecal ligation puncture (CLP) after 24 h to induce ALI. Animals were killed either 6 or 24 h after the second procedure. Pulmonary expression of inflammatory mediators cxcl1, cxcl5, IL-1β and IL-6, neutrophil infiltration and lung tissue damage using the Lung Injury Score (LIS) were determined.


Next to a moderate increase in other inflammatory mediators, a significant increase in CXCL1, neutrophil infiltration and lung injury was observed early after TxT, which returned to baseline levels after 24 h. DH induced significantly increased gene expression of cxcl1, cxcl5, IL-1β and IL-6 after 6 h, which was followed by the postponed significant increase in the protein expression after 24 h compared to controls. Neutrophil infiltration was significantly enhanced 24 h after DH compared to all other groups, and exerted a slight decline after 24 h. LIS has shown a significant increase after both 6 and 24 h compared to both control groups as well the late TxT group.


Early observed lung injury with moderate inflammatory changes after blunt chest trauma recovered quickly, and therefore, may be caused by mechanical lung injury. In contrast, lung injury in the ALI group did not undergo recovery and is closely associated with significant changes of inflammatory mediators. This model may be used for further examinations of contributing factors and therapeutic strategies to prevent ALI.


Thoracic trauma Lung injury CLP ALI Neutrophils cxcl Cytokines 



We thank Alexander Schaible, Katrin Jurida, and Kerstin Kontradowitz for outstanding technical assistance. PS was supported by the “Frankfurter Forschungsförderung” of the Goethe University Frankfurt within the program “Nachwuchsforscher”. Grant support: The work was supported by grants from the DFG WU 820/2-1, HI 820/5-1, and RE 3304/8-1.

Compliance with ethical standards

Conflict of interest

The authors state that they have no conflict of interest.


  1. 1.
    Störmann P, Marzi I, Wutzler S. Rotational therapy in thoracic injuries: what is the evidence? Curr Opin Crit Care. 2017;23:527–32.CrossRefGoogle Scholar
  2. 2.
    Leenen LPH. Focus on chest trauma. Eur J Trauma Emerg Surg. 2017;43:153–4.CrossRefGoogle Scholar
  3. 3.
    Miller PR, Croce MA, Bee TK, Qaisi WG, Smith CP, Collins GL, et al. ARDS after pulmonary contusion: accurate measurement of contusion volume identifies high-risk patients. J Trauma Injury Infect Crit Care. 2001;51:223–8 (discussion 229–30).Google Scholar
  4. 4.
    Keel M, Meier C. Chest injuries—what is new? Curr Opin Crit Care. 2007;13:674–9.CrossRefGoogle Scholar
  5. 5.
    Battle CE, Hutchings H, James K, Evans PA. The risk factors for the development of complications during the recovery phase following blunt chest wall trauma: a retrospective study. Injury. 2013;44:1171–6.CrossRefGoogle Scholar
  6. 6.
    Wutzler S, Wafaisade A, Maegele M, Laurer H, Geiger EV, Walcher F, et al. Lung Organ Failure Score (LOFS): probability of severe pulmonary organ failure after multiple injuries including chest trauma. Injury. 2012;43:1507–12.CrossRefGoogle Scholar
  7. 7.
    Hoth JJ, Wells JD, Hiltbold EM, McCall CE, Yoza BK. Mechanism of neutrophil recruitment to the lung after pulmonary contusion. Shock. 2011;35:604–9.CrossRefGoogle Scholar
  8. 8.
    Rose S. Marzi I [Pathophysiology of polytrauma]. Zentralbl Chir. 1996;121:896–913.Google Scholar
  9. 9.
    Keel M, Trentz O. Pathophysiology of polytrauma. Injury. 2005;36:691–709.CrossRefGoogle Scholar
  10. 10.
    Raymond SL, Holden DC, Mira JC, Stortz JA, Loftus TJ, Mohr AM, et al. Microbial recognition and danger signals in sepsis and trauma. Biochim Biophys Acta. 2017;1863:2564–73.CrossRefGoogle Scholar
  11. 11.
    Bruns B, Hönle T, Kellermann P, Ayala A, Perl M. divergent effects of neutrophils on fas-induced pulmonary inflammation, apoptosis, and lung damage. Shock. 2017;47:225–35.CrossRefGoogle Scholar
  12. 12.
    Weckbach S, Hohmann C, Denk S, Kellermann P, Huber-Lang MS, Baumann B, et al. Apoptotic and inflammatory signaling via Fas and tumor necrosis factor receptor I contribute to the development of chest trauma-induced septic acute lung injury. J Trauma Acute Care Surg. 2013;74:792–800.CrossRefGoogle Scholar
  13. 13.
    Ware LB, Matthay MA. The acute respiratory distress syndrome. N Engl J Med Massachusetts Medical Society; 2000;342:1334–49.Google Scholar
  14. 14.
    Störmann P, Lustenberger T, Relja B, Marzi I, Wutzler S. Role of biomarkers in acute traumatic lung injury. Injury. New York: Elsevier; 2017.CrossRefGoogle Scholar
  15. 15.
    Parsons PE, Eisner MD, Thompson BT, Matthay MA, Ancukiewicz M, Bernard GR, et al. Lower tidal volume ventilation and plasma cytokine markers of inflammation in patients with acute lung injury. Crit Care Med. 2005;33:1–6.CrossRefGoogle Scholar
  16. 16.
    Toscano MG, Ganea D, Gamero AM. Cecal ligation puncture procedure. J Vis Exp. 2011;e2860.Google Scholar
  17. 17.
    Zou Y, Bao S, Wang F, Guo L, Zhu J, Wang J, et al. FN14 Blockade on pulmonary microvascular endothelial cells improves the outcome of sepsis-induced acute lung injury. Shock. 2017 (Publish Ahead of Print: 1).Google Scholar
  18. 18.
    Wagner N, Dieteren S, Franz N, Köhler K, Mörs K, Nicin L, et al. Ethyl pyruvate ameliorates hepatic injury following blunt chest trauma and hemorrhagic shock by reducing local inflammation, NF-kappaB activation and HMGB1 release. PLoS One; 2018;13:2171.Google Scholar
  19. 19.
    Ulger H, Deniz T, Comu F, Agalar C, Kisa U, Agalar F. Protective effect of hypothermia in a blunt thoracic trauma and hemorrhagic shock model. Thorac Cardiovasc Surg. 2014;62:716–21.CrossRefGoogle Scholar
  20. 20.
    Kaiser K, Prystaz K, Vikman A, Haffner-Luntzer M, Bergdolt S, Strauss G, et al. Pharmacological inhibition of IL-6 trans-signaling improves compromised fracture healing after severe trauma. Arch Pharmacol. 2018;391:523–36.CrossRefGoogle Scholar
  21. 21.
    Kilkenny C, Browne WJ, Cuthill IC, Emerson M, Altman DG. Improving bioscience research reporting: the ARRIVE guidelines for reporting animal research. PLoS Biol. 2010;e1000412.Google Scholar
  22. 22.
    Knöferl MW, Liener UC, Seitz DH, Perl M, Brückner UB, Kinzl L, et al. Cardiopulmonary, histological, and inflammatory alterations after lung contusion in a novel mouse model of blunt chest trauma. Shock. 2003;19:519–25.CrossRefGoogle Scholar
  23. 23.
    Knöferl MW, Liener UC, Perl M, Brückner UB, Kinzl L, Gebhard F. Blunt chest trauma induces delayed splenic immunosuppression. Shock. 2004;22:51–6.CrossRefGoogle Scholar
  24. 24.
    Medina E. Murine model of polymicrobial septic peritonitis using cecal ligation and puncture (CLP). Methods Mol Biol. 2010;602:411–5.CrossRefGoogle Scholar
  25. 25.
    Vaschetto R, Kuiper JW, Chiang SR, Haitsma JJ, Juco JW, Uhlig S, et al. Inhibition of poly(adenosine diphosphate-ribose) polymerase attenuates ventilator-induced lung injury. Anesthesiology. 2008;108:261–8.CrossRefGoogle Scholar
  26. 26.
    Relja B, Horstmann JP, Kontradowitz K, Jurida K, Schaible A, Neunaber C, et al. Nlrp1 inflammasome is downregulated in trauma patients. J Mol Med. 2015;93:1391–400.CrossRefGoogle Scholar
  27. 27.
    Relja B, Henrich D, Wetzel G, Sander AL, Jakob H, Maraslioglu M, et al. Effects of acute ethanol gavage on intestinal integrity after hemorrhage/resuscitation. Scand J Gastroenterol. 2013;48:448–58.CrossRefGoogle Scholar
  28. 28.
    Perl M, Hohmann C, Denk S, Kellermann P, Lu D, Braumüller S, et al. Role of activated neutrophils in chest trauma-induced septic acute lung injury. Shock. 2012;38:98–106.CrossRefGoogle Scholar
  29. 29.
    Kalbitz M, Karbach M, Braumueller S, Kellermann P, Gebhard F, Huber-Lang M, et al. Role of complement C5 in experimental blunt chest trauma-induced septic acute lung injury (ALI). PLoS One. 2016;11:e0159417.CrossRefGoogle Scholar
  30. 30.
    Horst K, Simon TP, Pfeifer R, Teuben M, Almahmoud K, Zhi Q, et al. Characterization of blunt chest trauma in a long-term porcine model of severe multiple trauma. Sci Rep. 2016;6:39659.CrossRefGoogle Scholar
  31. 31.
    Hoth JJ, Martin RS, Yoza BK, Wells JD, Meredith JW, McCall CE. Pulmonary contusion primes systemic innate immunity responses. J Trauma Injury Infect Crit Care. 2009;67:14–22.CrossRefGoogle Scholar
  32. 32.
    Relja B, Mörs K, Marzi I. Danger signals in trauma. Eur J Trauma Emerg Surg. 2018;44:301–16.CrossRefGoogle Scholar
  33. 33.
    Moore FA, Sauaia A, Moore EE, Haenel JB, Burch JM, Lezotte DC. Postinjury multiple organ failure: a bimodal phenomenon. J Trauma Injury Infect Crit Care. 1996;40:501–10 (discussion 510–512).CrossRefGoogle Scholar
  34. 34.
    Hafner S, Wagner K, Wepler M, Matallo J, Gröger M, McCook O, et al. Physiological and immune-biological characterization of a long-term murine model of blunt chest trauma. Shock. 2015;43:140–7.CrossRefGoogle Scholar
  35. 35.
    Bhatia RK, Pallister I, Dent C, Jones SA, Topley N. Enhanced neutrophil migratory activity following major blunt trauma. Injury. 2005;36:956–62.CrossRefGoogle Scholar
  36. 36.
    Cohn SM, Dubose JJ. Pulmonary contusion: an update on recent advances in clinical management. World J Surg. 2010;34:1959–70.CrossRefGoogle Scholar
  37. 37.
    Gill SE, Rohan M, Mehta S. Role of pulmonary microvascular endothelial cell apoptosis in murine sepsis-induced lung injury in vivo. Respir Res. 2015;16:109.CrossRefGoogle Scholar
  38. 38.
    Kozan A, Kilic N, Alacam H, Guzel A, Guvenc T, Acikgoz M. The effects of dexamethasone and L-NAME on acute lung injury in rats with lung contusion. Inflammation. 2016;39:1747–56.CrossRefGoogle Scholar
  39. 39.
    Gao H, Neff T, Ward PA. Regulation of lung inflammation in the model of IgG immune-complex injury. Annu Rev Pathol. 2006;1:215–42.CrossRefGoogle Scholar
  40. 40.
    Wagner N, Franz N, Dieteren S, Perl M, Mörs K, Marzi I, et al. Acute alcohol binge deteriorates metabolic and respiratory compensation capability after blunt chest trauma followed by hemorrhagic shock—a new research model. Alcohol Clin Exp Res. 2017;41:1559–67.CrossRefGoogle Scholar
  41. 41.
    Liener UC, Knöferl MW, Sträter J, Barth TFE, Pauser E-M, Nüssler AK, et al. Induction of apoptosis following blunt chest trauma. Shock. 2003;20:511–6.CrossRefGoogle Scholar
  42. 42.
    Guo R-F, Ward PA. Mediators and regulation of neutrophil accumulation in inflammatory responses in lung: insights from the IgG immune complex model. Free Radic Biol Med. 2002;33:303–10.CrossRefGoogle Scholar
  43. 43.
    Mehta D, Ravindran K, Kuebler WM. Novel regulators of endothelial barrier function. Am J Physiol Lung Cell Mol Physiol. 2014;307:L924–35.CrossRefGoogle Scholar
  44. 44.
    Seok J, Warren HS, Cuenca AG, Mindrinos MN, Baker HV, Xu W, et al. Genomic responses in mouse models poorly mimic human inflammatory diseases. Proc Natl Acad Sci USA. 2013;110:3507–12.CrossRefGoogle Scholar
  45. 45.
    Tompkins RG. Genomics of injury: the Glue Grant experience. J Trauma Acute Care Surg. 2015;78:671–86.CrossRefGoogle Scholar
  46. 46.
    Opal SM, Laterre P-F, Francois B, LaRosa SP, Angus DC, Mira J-P, et al. Effect of eritoran, an antagonist of MD2-TLR4, on mortality in patients with severe sepsis: the ACCESS randomized trial. JAMA. 2013;309:1154–62.CrossRefGoogle Scholar
  47. 47.
    Shirey KA, Lai W, Scott AJ, Lipsky M, Mistry P, Pletneva LM, et al. The TLR4 antagonist Eritoran protects mice from lethal influenza infection. Nature. 2013;497:498–502.CrossRefGoogle Scholar
  48. 48.
    Perl M, Kieninger M, Huber-Lang MS, Gross H-J, Bachem MG, Braumüller S, et al. Divergent effects of activated neutrophils on inflammation, Kupffer cell/splenocyte activation, and lung injury following blunt chest trauma. Shock. 2012;37:210–8.CrossRefGoogle Scholar
  49. 49.
    Fujita M, Kuwano K, Kunitake R, Hagimoto N, Miyazaki H, Kaneko Y, et al. Endothelial cell apoptosis in lipopolysaccharide-induced lung injury in mice. Int Arch Allergy Immunol. 1998;117:202–8.CrossRefGoogle Scholar
  50. 50.
    Wiener-Kronish JP, Albertine KH, Matthay MA. Differential responses of the endothelial and epithelial barriers of the lung in sheep to Escherichia coli endotoxin. J Clin Invest. 1991;88:864–75.CrossRefGoogle Scholar
  51. 51.
    Matute-Bello G, Frevert CW, Martin TR. Animal models of acute lung injury. Am J Physiol Lung Cell Mol Physiol. 2008;295:L379–99.CrossRefGoogle Scholar
  52. 52.
    White TO, Jenkins PJ, Smith RD, Cartlidge CWJ, Robinson CM. The epidemiology of posttraumatic adult respiratory distress syndrome. J Bone Joint Surg Am. 2004;86-A:2366–76.CrossRefGoogle Scholar
  53. 53.
    Ehrnthaller C, Flierl M, Perl M, Denk S, Unnewehr H, Ward PA, et al. The molecular fingerprint of lung inflammation after blunt chest trauma. Eur J Med Res. 2015;20:70.CrossRefGoogle Scholar
  54. 54.
    Neunaber C, Oestern S, Andruszkow H, Zeckey C, Mommsen P, Kutter D, et al. Cytokine productive capacity of alveolar macrophages and Kupffer cells after femoral fracture and blunt chest trauma in a murine trauma model. Immunol Lett. 2013;152:159–66.CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Philipp Störmann
    • 1
    Email author
  • Nils Becker
    • 1
  • Leander Künnemeyer
    • 1
  • Sebastian Wutzler
    • 1
    • 2
  • Jan Tilmann Vollrath
    • 1
  • Thomas Lustenberger
    • 1
  • Frank Hildebrand
    • 3
  • Ingo Marzi
    • 1
  • Borna Relja
    • 1
  1. 1.Department of Trauma, Hand and Reconstructive SurgeryHospital of the Goethe University Frankfurt/MainFrankfurt/MainGermany
  2. 2.Department of Trauma, Hand and Orthopedic SurgeryHelios Horst Schmidt ClinicWiesbadenGermany
  3. 3.Department of Trauma SurgeryRWTH UniversityAachenGermany

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