Residual endotoxin induces primary graft dysfunction through ischemia/reperfusion-primed alveolar macrophages
Mahzad Akbarpour, … , G.R. Scott Budinger, Ankit Bharat
Mahzad Akbarpour, … , G.R. Scott Budinger, Ankit Bharat
Published May 19, 2020
Citation Information: J Clin Invest. 2020;130(8):4456-4469. https://doi.org/10.1172/JCI135838.
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Research Article Immunology Transplantation

Residual endotoxin induces primary graft dysfunction through ischemia/reperfusion-primed alveolar macrophages

Abstract

Despite the widespread use of antibiotics, bacterial pneumonias in donors strongly predispose to the fatal syndrome of primary graft dysfunction (PGD) following lung transplantation. We report that bacterial endotoxin persists in human donor lungs after pathogen is cleared with antibiotics and is associated with neutrophil infiltration and PGD. In mouse models, depletion of tissue-resident alveolar macrophages (TRAMs) attenuated neutrophil recruitment in response to endotoxin as shown by compartmental staining and intravital imaging. Bone marrow chimeric mice revealed that neutrophils were recruited by TRAM through activation of TLR4 in a MyD88-dependent manner. Intriguingly, low levels of endotoxin, insufficient to cause donor lung injury, promoted TRAM-dependent production of CXCL2, increased neutrophil recruitment, and led to PGD, which was independent of donor NCMs. Reactive oxygen species (ROS) increased in human donor lungs starting from the warm-ischemia phase and were associated with increased transcription and translocation to the plasma membrane of TLR4 in donor TRAMs. Consistently, scavenging ROS or inhibiting their production to prevent TLR4 transcription/translocation or blockade of TLR4 or coreceptor CD14 on donor TRAMs prevented neutrophil recruitment in response to endotoxin and ameliorated PGD. Our studies demonstrate that residual endotoxin after successful treatment of donor bacterial pneumonia promotes PGD through ischemia/reperfusion-primed donor TRAMs.

Authors

Mahzad Akbarpour, Emilia Lecuona, Stephen F. Chiu, Qiang Wu, Melissa Querrey, Ramiro Fernandez, Félix L. Núñez-Santana, Haiying Sun, Sowmya Ravi, Chitaru Kurihara, James M. Walter, Nikita Joshi, Ziyou Ren, Scott C. Roberts, Alan Hauser, Daniel Kreisel, Wenjun Li, Navdeep S. Chandel, Alexander V. Misharin, Thalachallour Mohanakumar, G.R. Scott Budinger, Ankit Bharat

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Figure 2

TRAMs upregulate CXCL2 in response to endotoxin and pseudomonas.

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TRAMs upregulate CXCL2 in response to endotoxin and pseudomonas. Transcr...
Transcriptomic profiling of human TRAMs during Gram-negative bacterial pneumonia revealed upregulation of CXCL2, a known neutrophil chemoattractant. TRAMs were identified as CD15–HLA–DR+CD169+CD206++, and flow sorted for RNA isolation, library preparation, and sequencing (culture-negative, n = 4; P. aeruginosa–positive, n = 6). (A) Heatmap of top 1000 most significantly differentially expressed genes ranked by FDR q value. Both samples and genes are grouped by hierarchical clustering. (B) Functional enrichment analysis with GO biological processes was performed with GOrilla with the top 1000 genes upregulated in P. aeruginosa–positive samples for all clusters. In total, 443 significantly upregulated genes were used to identify the top 5 biological processes (–log10 [P value]). (C) Heatmap of pathway genes (750 genes) based on the top GO processes identified in B (GO:0002376,GO:0019221, GO:0002237,GO:0006954,GO:0006950). Hierarchical clustering shows the expression of these genes in naive, low, and high virulence samples for MISTRG mice. Selective genes are highlighted (n = 3–4). (D) Normalized CPM counts of CXCL2 in both human and MISTRG mouse following infection with P. aeruginosa (n = 3–6). Graph show mean ± SD and was analyzed by unpaired t test.**P < 0.01.