Ischemia-Reperfusion Injury in a Simulated Lung Transplant Setting Differentially Regulates Transcriptomic Profiles between Human Lung Endothelial and Epithelial Cells.

Current understanding of mechanisms of ischemia-reperfusion-induced lung injury during lung preservation and transplantation is mainly based on clinical observations and animal studies. Herein, we used cell and systems biology approaches to explore these mechanisms at transcriptomics levels, especially by focusing on the differences between human lung endothelial and epithelial cells, which are crucial for maintaining essential lung structure and function. Human pulmonary microvascular endothelial cells and human lung epithelial cells were cultured to confluent, subjected to different cold ischemic times (CIT) to mimic static cold storage with preservation solution, and then subjected to warm reperfusion with a serum containing culture medium to simulate lung transplantation.

Use of metabolomics to identify strategies to improve and prolong ex vivo lung perfusion for lung transplants.

Background: Normothermic ex vivo lung perfusion (EVLP) allows for functional assessment of donor lungs; thus has increased the use of marginal lungs for transplantation. To extend EVLP for advanced organ reconditioning and regenerative interventions, cellular metabolic changes need to be understood. We sought to comprehensively characterize the dynamic metabolic changes of the lungs during EVLP, and to identify strategies to improve EVLP.

Transcriptomic investigation reveals donor-specific gene signatures in human lung transplants.

Introduction: Transplantation of lungs from donation after circulatory death (DCD) in addition to donation after brain death (DBD) became routine worldwide to address the global organ shortage. The development of lung perfusion (EVLP) for donor lung assessment and repair contributed to the increased use of DCD lungs. We hypothesise that a better understanding of the differences between lungs from DBD and DCD donors, and between EVLP and directly transplanted (non-EVLP) lungs, will lead to the discovery of the injury-specific targets for donor lung repair and reconditioning.

Eosinophils in transbronchial biopsies: a predictor of chronic lung allograft dysfunction and reduced survival after lung transplantation - a retrospective single-center cohort study.

Long-term outcomes after lung transplantation remain inferior to those of other solid organ groups. The significance of eosinophils detected on transbronchial biopsies (TBBx) after lung transplantation and their relationship to long-term outcomes remain unknown. A retrospective single-center cohort study was performed of patients transplanted between January 01, 2001, and July 31, 2018, who had at least 1 TBBx with evaluable parenchymal tissue.

Strategies to prolong homeostasis of ex vivo perfused lungs.

Objectives: Ex vivo lung perfusion provides an innovative method to assess and repair donor lungs. The current Toronto ex vivo lung perfusion protocol can reliably and reproducibly preserve lungs for 12 hours. A longer ex vivo lung perfusion preservation time could enable the application of more advanced repair therapies and the rescue of more donor lungs for lung transplant.

Bronchoalveolar bile acid and inflammatory markers to identify high-risk lung transplant recipients with reflux and microaspiration.

Background: Gastroesophageal reflux disease (GERD) is a risk factor for chronic lung allograft dysfunction. Bile acids-putative markers of gastric microaspiration-and inflammatory proteins in the bronchoalveolar lavage (BAL) have been associated with chronic lung allograft dysfunction, but their relationship with GERD remains unclear. Although GERD is thought to drive chronic microaspiration, the selection of patients for anti-reflux surgery lacks precision.

Potential therapeutic targets for lung repair during human lung perfusion.

Introduction: The lung perfusion (EVLP) technique has been developed to assess the function of marginal donor lungs and has significantly increased donor lung utilisation. EVLP has also been explored as a platform for donor lung repair through injury-specific treatments such as antibiotics or fibrinolytics. We hypothesised that actively expressed pathways shared between transplantation and EVLP may reveal common mechanisms of injury and potential therapeutic targets for lung repair prior to transplantation.

Variation at and Asthma on the Island of Tristan da Cunha.

A high prevalence of asthma has been documented among the inhabitants of Tristan da Cunha, an isolated island in the South Atlantic. The population derives from just 28 founders. We performed lung function testing, including methacholine inhalation challenge, allergen skin prick testing, and collected DNA from essentially all of the current island population (269 individuals), and genotyped a panel of 43 single-nucleotide polymorphisms (SNPs) reported as associated with asthma and atopy.

Normothermic ex vivo lung perfusion: Does the indication impact organ utilization and patient outcomes after transplantation?

Background: Ex vivo lung perfusion (EVLP) is being increasingly applied as a method to evaluate and treat donor lungs for transplantation. However, with the previous limited worldwide experience, no studies have been able to evaluate the impact of indication for EVLP on organ utilization rates and recipient outcomes after lung transplantation (LTx). We examined these outcomes in a large-cohort, single-center series of clinical EVLP cases.

Ischemia-reperfusion induces death receptor-independent necroptosis via calpain-STAT3 activation in a lung transplant setting.

Ischemia-reperfusion (I/R)-induced lung injury undermines lung transplantation (LTx) outcomes by predisposing lung grafts to primary graft dysfunction (PGD). Necrosis is a feature of I/R lung injury. However, regulated necrosis (RN) with specific signaling pathways has not been explored in an LTx setting.

Towards donor lung recovery-gene expression changes during ex vivo lung perfusion of human lungs.

We and others have demonstrated that acellular normothermic ex vivo lung perfusion of high-risk donor lungs can result in posttransplant outcomes equivalent to that of contemporaneous lung transplantation using standard donor lungs. However, the mechanism of this effect remains unclear. Given the restoration of cellular metabolic activity during normothermic perfusion, one possibility is that of lung healing via natural innate recovery mechanisms.

Metabolic Profile of Ex Vivo Lung Perfusate Yields Biomarkers for Lung Transplant Outcomes.

Objective: To identify potential biomarkers during ex vivo lung perfusion (EVLP) using metabolomics approach.

Summary Background Data: EVLP increases the number of usable donor lungs for lung transplantation (LTx) by physiologic assessment of explanted marginal lungs. The underlying paradigm of EVLP is the normothermic perfusion of cadaveric lungs previously flushed and stored in hypothermic preservation fluid, which allows the resumption of active cellular metabolism and respiratory function.

Higher M30 and high mobility group box 1 protein levels in ex vivo lung perfusate are associated with primary graft dysfunction after human lung transplantation.

Background: Ex vivo lung perfusion (EVLP) enables assessment of marginal donor lungs for transplantation, with similar clinical outcomes to conventional lung transplantation. We investigated whether cell death-related proteins in the EVLP perfusate could predict primary graft dysfunction (PGD) after transplantation.

Methods: M30 (indicating epithelial apoptosis), M65 (indicating total epithelial cell death), and high mobility group box 1 (HMGB-1, related to cell death and inflammation) protein levels in EVLP perfusate were measured by enzyme-linked immunosorbent assay and correlated with clinical outcomes.

Importance of left atrial pressure during ex vivo lung perfusion.

Background: Ex vivo lung perfusion (EVLP) allows for the evaluation and treatment of donor lungs before transplant. Different EVLP strategies have been described using either an open left atrium (LA) (pressure of 0 mm Hg) or closed LA (pressure of 5 mm Hg). We hypothesized that maintaining a physiologic positive LA pressure during EVLP is protective to the lung.

Circulating Cell Death Biomarkers May Predict Survival in Human Lung Transplantation.

Rationale: Immediate graft performance after lung transplantation is associated with short- and long-term clinical outcomes. However, the biologic mechanism that determines outcomes is not fully understood.

Objectives: To investigate the impact of cell death signals at 24 and 48 hours after lung transplantation on short- and long-term clinical outcomes.

Fractal circuit sensors enable rapid quantification of biomarkers for donor lung assessment for transplantation.

Biomarker profiling is being rapidly incorporated in many areas of modern medical practice to improve the precision of clinical decision-making. This potential improvement, however, has not been transferred to the practice of organ assessment and transplantation because previously developed gene-profiling techniques require an extended period of time to perform, making them unsuitable in the time-sensitive organ assessment process. We sought to develop a novel class of chip-based sensors that would enable rapid analysis of tissue levels of preimplantation mRNA markers that correlate with the development of primary graft dysfunction (PGD) in recipients after transplant.

XB130 promotes bronchioalveolar stem cell and Club cell proliferation in airway epithelial repair and regeneration.

Proliferation of bronchioalveolar stem cells (BASCs) is essential for epithelial repair. XB130 is a novel adaptor protein involved in the regulation of epithelial cell survival, proliferation and migration through the PI3K/Akt pathway. To determine the role of XB130 in airway epithelial injury repair and regeneration, a naphthalene-induced airway epithelial injury model was used with XB130 knockout (KO) mice and their wild type (WT) littermates.

The role of the endothelin-1 pathway as a biomarker for donor lung assessment in clinical ex vivo lung perfusion.

Background: Normothermic ex vivo lung perfusion (EVLP) is a preservation technique that allows reassessment of donor lungs before transplantation. We hypothesized that the endothelin-1 (ET-1) axis would be associated with donor lung performance during EVLP and recipient outcomes after transplantation.

Methods: ET-1, Big ET-1, endothelin-converting enzyme (ECE), and nitric oxide (NO) metabolites were quantified in the perfusates of donor lungs enrolled in a clinical EVLP trial.

Potential role of CC chemokine receptor 6 in prediction of late-onset cytomegalovirus infection following solid organ transplant.

CCR6 is a chemokine receptor involved in homing memory T cells, particularly Th17 cells, to sites of mucosal inflammation. Despite the critical role of memory T cells in long-term protective immunity against cytomegalovirus (CMV), a virus that reactivates at multiple mucosal sites, the ability of CCR6 or other Th17 marker expression to predict CMV reactivation following transplantation is not clear. Using 11-color flow cytometry, in this prospective single-center pilot study, we measured the expression of CCR6 and other markers of T-cell function in peripheral blood samples obtained from 21 SOT recipients at the time of discontinuation of anti-CMV prophylaxis.

Protein expression profiling predicts graft performance in clinical ex vivo lung perfusion.

Objectives: To study the impact of ex vivo lung perfusion (EVLP) on cytokines, chemokines, and growth factors and their correlation with graft performance either during perfusion or after transplantation.

Background: EVLP is a modern technique that preserves lungs on normothermia in a metabolically active state. The identification of biomarkers during clinical EVLP can contribute to the safe expansion of the donor pool.

Metabolomic heterogeneity of pulmonary arterial hypertension.

Although multiple gene and protein expression have been extensively profiled in human pulmonary arterial hypertension (PAH), the mechanism for the development and progression of pulmonary hypertension remains elusive. Analysis of the global metabolomic heterogeneity within the pulmonary vascular system leads to a better understanding of disease progression. Using a combination of high-throughput liquid-and-gas-chromatography-based mass spectrometry, we showed unbiased metabolomic profiles of disrupted glycolysis, increased TCA cycle, and fatty acid metabolites with altered oxidation pathways in the human PAH lung.

Elevated CXCL10 (IP-10) in bronchoalveolar lavage fluid is associated with acute cellular rejection after human lung transplantation.

Background: CXCL10 (IP-10) is a potent chemoattractant for T cells that has been postulated to play a role in infection and acute cellular rejection (ACR) in animal models. We measured CXCL10 (IP-10) (and other cytokines previously implicated in the pathogenesis of ACR) in the bronchoalveolar lavage (BAL) of lung transplant recipients (LTRs) to determine the association between CXCL10 (IP-10) and ACR in LTRs.

Methods: In a prospective study of 85 LTRs, expression of cytokines (tumor necrosis factor, interferon-γ, interleukin [IL]-6, IL-8, IL-15, IL-16, IL-17, CXCL10 [IP-10], and MCP-1 [CCL2]) in BAL samples (n=233) from patients with episodes of ACR (n=44), infection ("Infect"; n=25), concomitant "Infect+ACR" (n=10), and "No Infect and No ACR" (n=154) were analyzed.

PKC activation induces inflammatory response and cell death in human bronchial epithelial cells.

A variety of airborne pathogens can induce inflammatory responses in airway epithelial cells, which is a crucial component of host defence. However, excessive inflammatory responses and chronic inflammation also contribute to different diseases of the respiratory system. We hypothesized that the activation of protein kinase C (PKC) is one of the essential mechanisms of inflammatory response in airway epithelial cells.

Ribonucleic acid microarray analysis from lymph node samples obtained by endobronchial ultrasonography-guided transbronchial needle aspiration.

Purpose: The aim of this study was to evaluate the feasibility of messenger RNA (mRNA) and microRNA (miRNA) expression analysis by microarray using samples obtained by endobronchial ultrasonography-guided transbronchial needle aspiration (EBUS-TBNA).

Description: We isolated total RNA from 24 archived clinical EBUS-TBNA samples. The purified RNA that met the quality threshold was used for two different kinds of microarray analyses: whole transcript-based (WT) array for mRNA expression and miRNA expression array.