Extrathoracic multiple trauma dysregulates neutrophil function and exacerbates pneumonia-induced lung injury - PubMed (original) (raw)

Extrathoracic multiple trauma dysregulates neutrophil function and exacerbates pneumonia-induced lung injury

Jennifer M Leonard et al. J Trauma Acute Care Surg. 2021.

Abstract

Background: Forty percent of critically ill trauma patients will develop an infectious complication. Pneumonia is the most common cause of death of trauma patients surviving their initial insult. We previously demonstrated that polytrauma (PT), defined as two or more severe injuries in at least two areas of the body, induces emergency hematopoiesis characterized by accelerated myelopoiesis in the bone marrow and increased myeloid cell frequency in the peripheral tissues. We hypothesized that PT alone induces priming of neutrophils, resulting in hyperactivation upon secondary exposure to bacteria and causing acute lung injury and increased susceptibility to secondary exposure to Pseudomonas aeruginosa pneumonia.

Methods: C57BL/6 mice were subjected to PT consisting of a lower extremity pseudofracture, liver crush injury, and 15% blood-volume hemorrhage. Pneumonia was induced by intratracheal injection of 5 × 106 CFU live P. aeruginosa or 1 × 107 of heat-killed P. aeruginosa (HKPA). For reactive oxygen species (ROS), studies polymorphonuclear neutrophils (PMNs) were isolated by immunomagnetic bead negative selection and stimulated ex-vivo with HKPA. Reactive oxygen species production was measured by immunofluorescence. For histology, lung sections were stained by hematoxylin-eosin and analyzed by a blinded grader.

Results: Polytrauma induced persistent changes in immune function at baseline and to secondary infection. Pneumonia after injury resulted in increased mortality (60% vs. 5% p < 0.01). Blood neutrophils from PT mice had higher resting (unstimulated) ROS production than in naive animals (p < 0.02) demonstrating priming of the neutrophils following PT. After intratracheal HKPA injection, bronchoalveolar lavage PMNs from injured mice had higher ROS production compared with naive mice (p < 0.01), demonstrating an overexuberant immunopathologic response of neutrophils following PT.

Conclusion: Polytrauma primes neutrophils and causes immunopathologic PMN ROS production, increased lung injury and susceptibility to secondary bacterial pneumonia. These results suggest that trauma-induced immune dysfunction can cause immunopathologic response to secondary infection and suggests neutrophil-mediated pulmonary damage as a therapeutic target for posttrauma pneumonia.

PubMed Disclaimer

Conflict of interest statement

Disclosure

No conflicts of interest were associated with this publication. This study was funded by AAST Research and Education Fund Scholarship Reward.

Figures

Figure 1 –

(A) Polytrauma (PT) results in changes in absolute numbers of resident monocytes (p=0.022) and eosinophils (p=0.0002) in blood (PBL) while absolute counts of neutrophils remain the same, however, (B) CD101 expression assayed by flow cytometry shows the neutrophil population in blood is altered in PT mice resulting in a shift towards an immature (CD101 low) population.

Figure 2 –

Box and whisker plot of cytokine analysis of PBMC isolated from Naïve (n=10) and Polytrauma (PT) mice (n=6) in response to stimulation with LPS and heat-killed pseudomonas aeruginosa (HKPa) injection demonstrates exaggerated secretion of (A) TNF-alpha, (B) MCP-1 and (C) IL-6 by polytrauma (PT) mice compared to Naïve. Boxes are 25th to 75th percentiles and contain the median (bar), whiskers are 5–95th percentile range. (* p=<0.05, **p<0.005)

Figure 3 –

ROS production by neutrophils from blood isolated from Naïve (n=6) versus Polytrauma (PT) mice (n=6) with and without in vitro heat-killed pseudomonas aeruginosa (HKPA) stimulation shows blood neutrophils from PT mice produce less ROS after HKPA stimulation (p=0.01) consistent with findings of an immature population of neutrophils mobilized to the blood in PT. Area Under Curve (AUC) values were normalized to those from naïve mice.

Figure 4 –

Kaplan-Meier curve of survival from pseudomonas aeruginosa (Pa) lung infection of polytrauma (PT) and sham operated mice starting from 48 hours from surgery demonstrates significant mortality from combination of PT and Pa infection compared to PT alone, or Sham surgery with Pa infection. PT mice also demonstrate greatly increased bacterial counts assayed by colony-forming units (CFU) in both lung tissue (p=0.022) and blood (p=0.037) compared to sham controls.

Figure 5 –

Mice Naïve and Polytrauma (PT) mice were exposed to pseudomonas aeruginosa (PA) and assayed with cytokine bead array for (A) TNF-alpha, (B) IL-6 and (C) MCP-1 with PT (n=7 (0h), n=8 (6h), n= 6 (24h) consistently showing an altered and prolonged elevation in cytokines within 24 hours of PA exposure compared to naïve animals (N=7 at each time point). P values by Mann-Whitney test on 24 hour time point.

Figure 6 –

Histology graded by a blinded investigator showed (A) increased Alveolar Neutrophil score in polytrauma (PT) animals compared to naïve animals (p=0.002), and (B) increased Acute Lung Injury score in PT compared to naïve animals (p=0.002) 24 hours following live pseudomonas aeruginosa (PA) dosing. Consistent with (C) this reactive oxygen species (ROS) production from neutrophils isolated from bronchioalveolar lavage in naïve versus PT animals and stimulated in vitro with heat-killed pseudomonas aeruginosa (HKPA) showed elevated ROS in PT neutrophils from the lung (p=<0.01) and consistent with a mature population of neutrophils recruited to lungs in extrathoracic PT, in contrast to the neutrophils from blood. (D) Pulmonary histology of naïve and PT mice with and without induction of 2.5 × 108 CFU/ml live PA.

References

1. Hoover L, Bochicchio GV, Napolitano LM, Joshi M, Bochicchio K, Meyer W, Scalea TM. Systemic inflammatory response syndrome and nosocomial infection in trauma. J Trauma. 2006;61(2):310–6; discussion 6–7. -PubMed
1. Surgeons ACo. National Trauma Database Annual Report. Chicago, IL; 2014.
1. Magill SS, Edwards JR, Bamberg W, Beldavs ZG, Dumyati G, Kainer MA, Lynfield R, Maloney M, McAllister-Hollod L, Nadle J, et al. Multistate point-prevalence survey of health care-associated infections. N Engl J Med. 2014;370(13):1198–208. -PMC -PubMed
1. Meakins JL, Pietsch JB, Bubenick O, Kelly R, Rode H, Gordon J, MacLean LD. Delayed hypersensitivity: indicator of acquired failure of host defenses in sepsis and trauma. Ann Surg. 1977;186(3):241–50. -PMC -PubMed
1. MacLean LD, Meakins JL, Taguchi K, Duignan JP, Dhillon KS, Gordon J. Host resistance in sepsis and trauma. Ann Surg. 1975;182(3):207–17. -PMC -PubMed

Extrathoracic multiple trauma dysregulates neutrophil function and exacerbates pneumonia-induced lung injury - PubMed (original) (raw)