Background: Mitochondrial DNA (MT-DNA) are intrinsically inflammatory nucleic acids released by damaged solid organs. Whether circulating cell-free MT-DNA quantitation could be used to predict the risk of poor COVID-19 outcomes remains undetermined. Methods: We measured circulating MT-DNA levels in prospectively collected, cell-free plasma samples from 97 subjects with COVID-19 at hospital presentation. Our primary outcome was mortality. Intensive care unit (ICU) admission, intubation, vasopressor, and renal replacement therapy requirements were secondary outcomes. Multivariate regression analysis determined whether MT-DNA levels were independent of other reported COVID-19 risk factors. Receiver operating characteristic and area under the curve assessments were used to compare MT-DNA levels with established and emerging inflammatory markers of COVID-19. Results: Circulating MT-DNA levels were highly elevated in patients who eventually died or required ICU admission, intubation, vasopressor use, or renal replacement therapy. Multivariate regression revealed that high circulating MT-DNA was an independent risk factor for these outcomes after adjusting for age, sex, and comorbidities. We also found that circulating MT-DNA levels had a similar or superior area under the curve when compared against clinically established measures of inflammation and emerging markers currently of interest as investigational targets for COVID-19 therapy. Conclusion: These results show that high circulating MT-DNA levels are a potential early indicator for poor COVID-19 outcomes. Funding: Washington University Institute of Clinical Translational Sciences COVID-19 Research Program and Washington University Institute of Clinical Translational Sciences (ICTS) NIH grant UL1TR002345.

The mechanisms by which innate immune signals regulate alloimmune responses remain poorly understood. In the present study, we show by intravital 2-photon microscopy direct interactions between graft-infiltrating neutrophils and donor CD11c⁺ dendritic cells (DCs) within orthotopic lung allografts immediately after reperfusion. Neutrophils isolated from the airways of lung transplantation recipients stimulate donor DCs in a contact-dependent fashion to augment their production of IL-12 and expand alloantigen-specific IFN-γ⁺ T cells. DC IL-12 expression is largely regulated by degranulation and induced by TNF-α associated with the neutrophil plasma membrane. Extended cold ischemic graft storage enhances G-CSF–mediated granulopoiesis and neutrophil graft infiltration, resulting in exacerbation of ischemia-reperfusion injury after lung transplantation. Ischemia reperfusion injury prevents immunosuppression-mediated acceptance of mouse lung allografts unless G-CSF–mediated granulopoiesis is inhibited. Our findings identify granulopoiesis-mediated augmentation of alloimmunity as a novel link between innate and adaptive immune responses after organ transplantation.

Neutrophils are often viewed as nonspecialized effector cells whose presence is a simple indicator of tissue inflammation. There is new evidence that neutrophils exist in subsets and have specialized effector functions that include extracellular trap generation and the stimulation of angiogenesis. The application of intravital imaging to transplanted organs has revealed novel requirements for neutrophil trafficking into graft tissue and has illuminated direct interactions between neutrophils and other leukocytes that promote alloimmunity. Paradoxically, retaining some neutrophilia may be important to induce or maintain tolerance. Neutrophils can stimulate anti‐inflammatory signals in other phagocytes and release molecules that inhibit T cell activation. In this article, we will review the available evidence of how neutrophils regulate acute and chronic inflammation in transplanted organs and discuss the possibility of targeting these cells to promote tolerance.

Primary graft dysfunction (PGD) is a major limitation to short and long‐term lung transplant survival. Recent work has shown mitochondrial damage associated molecular patterns (Mt‐DAMPs) can promote solid organ injury but whether they contribute to PGD severity remains unclear. We quantitated circulating plasma Mt‐DNA in 62 patients, prior to lung transplant and shortly after ICU arrival. Although all recipients released Mt‐DNA, high levels were associated with severe PGD development. In a mouse orthotopic lung transplant model of PGD we detected airway cell‐free damaged mitochondria and Mt‐DNA in the peripheral circulation. Pharmacological inhibition or genetic deletion of the formylated peptide receptor 1 (FPR1), a chemotaxis sensor for N‐formylated peptides released by damaged mitochondria, inhibited graft injury. An analysis of intragraft neutrophil trafficking patterns reveals that FPR1 enhances neutrophil transepithelial migration and retention within airways, but does not control extravasation. Using donor lungs that express a mitochondria‐targeted reporter protein, we also show that FPR1‐mediated neutrophil trafficking is coupled to the engulfment of damaged mitochondria, which in turn triggers reactive oxygen species (ROS)‐induced pulmonary edema. Therefore, our data demonstrate an association between Mt‐DAMP release and PGD development and suggest that neutrophil trafficking and effector responses to damaged mitochondria are drivers of graft damage.

Surgery is the first line of treatment for laryngotracheal stenosis; Montgomery tube or permanent tracheostomy have been so far the only alternatives. Nd-YAG laser resection and indwelling endotracheal stents have rarely been used in subglottic stenosis for anatomic and technical reasons. We have used the latter approach to optimize the timing of surgery or to achieve palliation without tracheostomy. Between 1991 and 2001 we have treated 18 patients with subglottic stenosis (10 males, 8 females; age range 14-78, mean 34). The upper margin of the stricture was 2mm to 1cm below the vocal cords; the stenotic segment extended from 1.5 to 5 cm. Three patients had tracheostomy done elsewhere. Four patients (Group I) had laser and stenting by a Dumon prosthesis as the only treatment; six had laser and stenting (#4) followed after 1-6 months by laryngotracheal resection (Group II); eight had surgery alone (Group III). In Group I, one patient required repositioning of the stent and in two the stent was removed; two patients died of their underlying disease; at a follow-up of 2-9 years all living patients did well but required permanent aerosolized therapy and periodical bronchoscopy. In Group II, we had two wound infections due to airway colonization by staphylococcus aureus. In Group III, two patients developed anastomotic postoperative stenosis, treated by laser (#2) and stenting (#1), and one patient with previous tracheostomy had a wound infection. Overall, in the 14 surgical patients (Groups II and III) stenosis occurred in 14.2% and infection in 21.3%. After a follow up of 15 months to 12 years, all surgical patients breathe and speak well. Laser resection and endoluminal stenting can be a viable alternative to surgery or optimize the timing of operation in patients with subglottic stenosis.