Primary graft dysfunction (PGD) is the clinical syndrome of acute lung injury after lung transplantation (LTx). However, PGD is an umbrella term that encompasses the ongoing pathophysiological and -biological mechanisms occurring in the lung grafts. Therefore, we aim to provide a focused review on the clinical, physiological, radiological, histological and cellular level of PGD. PGD is graded based on hypoxemia and chest X-ray (CXR) infiltrates. High-grade PGD is associated with inferior outcome after LTx. Lung edema is the main characteristic of PGD and alters pulmonary compliance, gas exchange and circulation. A conventional CXR provides a rough estimate of lung edema, while a chest computed tomography (CT) results in a more in-depth analysis. Macroscopically, interstitial and alveolar edema can be distinguished below the visceral lung surface. On the histological level, PGD correlates to a pattern of diffuse alveolar damage (DAD). At the cellular level, ischemia-reperfusion injury (IRI) is the main trigger for the disruption of the endothelial-epithelial alveolar barrier and inflammatory cascade. The multilevel approach integrating all PGD-related aspects results in a better understanding of acute lung failure after LTx, providing novel insights for future therapies.

Objective: To describe our experience with lung transplantation (LTx) from donors ≥70 years and compare short and long-term outcomes to a propensity-matched cohort of donors <70 years. Background: Although extended-criteria donors have been widely used to enlarge the donor pool, the experience with LTx from older donors (≥70 years) remains limited. Methods: All single-center bilateral LTx between 2010 and 2020 were retrospectively analyzed. Matching (1:1) was performed for the donor (type, sex, smoking history, x-ray abnormalities, partial pressure of oxygen/fraction of inspired oxygen ratio, and time on ventilator) and recipient characteristics (age, sex, LTx indication, perioperative extracorporeal life support, and cytomegalovirus mismatch). Primary graft dysfunction grade-3, 5-year patient, and chronic lung allograft dysfunction-free survival were analyzed. Results: Out of 647 bilateral LTx, 69 were performed from donors ≥70 years. The mean age in the older donor cohort was 74 years (range: 70–84 years) versus 49 years (range: 12–69 years) in the matched younger group. No significant differences were observed in the length of ventilatory support, intensive care unit, or hospital stay. Primary graft dysfunction-3 was 26% in the older group versus 29% in younger donor recipients (P = 0.85). Reintervention rate was comparable (29% vs 16%; P = 0.10). Follow-up bronchoscopy revealed no difference in bronchial anastomotic complications (P = 1.00). Five-year patient and chronic lung allograft dysfunction-free survivals were 73.6% versus 73.1% (P = 0.72) and 51.5% versus 59.2% (P = 0.41), respectively. Conclusions: LTx from selected donors ≥70 years is feasible and safe, yielding comparable short and long-term outcomes in a propensity-matched analysis with younger donors (<70 years).

Primary graft dysfunction (PGD) is a major obstacle after lung transplantation (LTx), associated with increased early morbidity and mortality. Studies in liver and kidney transplantation revealed prolonged anastomosis time (AT) as independent risk factor for impaired short- and long-term outcomes. We investigated if AT during LTx is a risk factor for PGD. In this retrospective single-center cohort study we included all first double lung transplantations (DLTx) between 2008-2016. The association of AT with any PGD grade 3 (PGD3) within the first 72 hours post-transplant was analyzed by univariable and multivariable logistic regression analysis. Data on AT and PGD was available for 427 patients of which 130 (30.2%) developed PGD3. AT was independently associated with the development of any PGD3 ≤72 hours in uni- (OR per 10min 1.293, 95%CI (1.136-1.471), p<.0001) and multivariable (OR 1.205, 95%CI (1.022-1.421), p=0.03) logistic regression analysis. There was no evidence that the relation between AT and PGD3 differed between lung recipients from donation after brain death versus donation after circulatory death donors. This study identified AT as independent risk factor for development of PGD3 post-LTx. We suggest that the implantation time should be kept short and the lung cooled to decrease PGD-related morbidity and mortality post-LTx.

SARS-CoV-2 is a single-stranded positive-sense RNA virus. Several negative-sense SARS-CoV- 2 RNA species, both full-length genomic and subgenomic, are produced transiently during viral replication. Methodologies for rigorously characterising cell tropism and visualising ongoing viral replication at single-cell resolution in histological sections are needed to assess the virological and pathological phenotypes of future SARS-CoV-2 variants. We aimed to provide a robust methodology for examining the human lung, the major target organ of this RNA virus. A prospective cohort study took place at the University Hospitals Leuven in Leuven, Belgium. Lung samples were procured postmortem from 22 patients who died from or with COVID-19. Tissue sections were fluorescently stained with the ultrasensitive single-molecule RNA in situ hybridisation platform of RNAscope combined with immunohistochemistry followed by confocal imaging. We visualised perinuclear RNAscope signal for negative-sense SARS-CoV-2 RNA species in ciliated cells of the bronchiolar epithelium of a patient who died with COVID-19 in the hyperacute phase of the infection, and in ciliated cells of a primary culture of human airway epithelium that had been infected experimentally with SARS-CoV-2. In patients who died between 5 and 13 days after diagnosis of the infection, we detected RNAscope signal for positive-sense but not for negative-sense SARS-CoV-2 RNA species in pneumocytes, macrophages, and among debris in the alveoli. SARS-CoV-2 RNA levels decreased after a disease course of 2–3 weeks, concomitant with a histopathological change from exudative to fibroproliferative diffuse alveolar damage. Taken together, our confocal images illustrate the complexities stemming from traditional approaches in the literature to characterise cell tropism and visualise ongoing viral replication solely by the surrogate parameters of nucleocapsid-immunoreactive signal or in situ hybridisation for positive-sense SARS-CoV-2 RNA species. Confocal imaging of human lung sections stained fluorescently with commercially available RNAscope probes for negative-sense SARS-CoV-2 RNA species enables the visualisation of viral replication at single-cell resolution during the acute phase of the infection in COVID-19. This methodology will be valuable for research on future SARS-CoV-2 variants and other respiratory viruses. 10.13039/501100004189 Max Planck Society , Coronafonds 10.13039/501100004857 UZ / 10.13039/501100004040 KU Leuven , European Society for Organ Transplantation