Idiopathic pulmonary fibrosis is a progressive lung disease characterized by the rapid scarring of the lung parenchyma resulting in impaired gas exchange and early mortality. Current treatment options are limited; lung transplantation remains the only definitive treatment. The adaptive immune system has been increasingly evoked as a potential contributor to disease initiation or progression. Using spectral flow cytometry, immunofluorescence imaging, and in-vitro functional assays we studied T cells obtained from explanted lungs and lung draining lymph nodes (HLN) from patients with IPF and non-diseased controls. We found the accumulation of granzyme K producing, hypofunctional CD8+ T cells in the lungs and HLN from IPF compared to controls. We also showed the accumulation of regulatory T cells in both lung and HLN in patients with IPF. CD4+ and CD8+ T cells were found to accumulate around areas of active fibrosis in IPF lung sections. Finally, when exposed to extracellular granzyme K, epithelial cells from human lungs showed increased expression of genes related to fibrosis, proliferation, and inflammation. Together, these show that IPF lungs and lymph nodes are characterized by the accumulation of granzyme K producing T cells and that granzyme K can promote pro-fibrotic effects of lung epithelial cells, providing a potential means whereby T cells might contribute to lung fibrosis.

To evaluate and summarize the current landscape of peripheral blood monitoring and immunophenotyping of patients following heart and lung transplantation, highlighting recent advances in monitoring which may help to predict and improve clinical outcomes. Immunological contributions to primary graft dysfunction, acute allograft dysfunction, and chronic allograft dysfunction are reviewed. In heart transplantation, novel diagnostic approaches like donor-derived cell-free DNA (dd-cfDNA) and gene expression profiling (GEP) show promise as alternatives to invasive biopsies for detecting rejection. Following lung transplantation, various immune cell populations—including NK cells, T cells, B cells, and regulatory T cells—demonstrate distinct dynamics between peripheral blood and the allograft compartment. Key findings include associations between circulating immune cell phenotypes and chronic lung allograft dysfunction (CLAD), donor-specific antibodies correlating with graft survival, and telomere length serving as a potential biomarker. Immune cells in circulation do not always phenotypically reflect immune cells in the allograft. While some promising markers for allograft dysfunction exist, further studies are needed to correlate them with disease severity and/or progression.

Motivated by growing evidence that the presence of critically shortened telomeres influences interstitial lung disease (ILD) trajectories and is associated with extrapulmonary conditions relevant to lung transplant candidacy and post-transplant complications, this Consensus Statement aims to address gaps in the evaluation and management of patients with short telomere syndrome (STS). These considerations reflect the work of an international Writing Committee with expertise in STS and are grounded in current literature and expert consensus. The need for this document arises from the recognition that STS is an underdiagnosed contributor to ILD, and that its presence introduces complexities that require dedicated, multidisciplinary attention in the transplant setting. J Heart Lung Transplant 2026;45:e83-e103 (c) 2026 International Society for Heart and Lung Transplantation. All rights are reserved, including those for text and data mining, AI training, and similar technologies.

Chronic lung allograft dysfunction (CLAD) is the major barrier for long-term survival in lung transplant recipients (LTRs). CLAD remains a diagnosis of exclusion with poor responses to therapies. A molecular diagnostic for CLAD is needed to risk-stratify LTRs for prognosis and identify new targets to mitigate CLAD progression. We used weighted gene correlation network analysis on the airway brush-derived airway transcriptome to identify immune pathways and markers relevant to CLAD. Weighted gene correlation network analysis was performed on RNA sequencing from airway brushings of 37 LTRs with CLAD compared with 37 stable LTRs. We analyzed gene coexpression networks (modules) for their biological significance and association with CLAD. Three gene modules were positively correlated with CLAD, its severity, allograft dysfunction, and survival. These enriched components of the acute phase response, type 1 adaptive immunity, and innate immunity, respectively. A fourth module correlated with protection and was inversely correlated with the other modules. We validated our findings by identification of downstream protein and eicosanoid levels in the bronchoalveolar lavage, and an external validation cohort where module expression differentiated LTRs with CLAD and correlated with worse survival. The CLAD airway transcriptome enriches for coexpression networks associated with network modules that correlate with allograft dysfunction and survival.

Rationale CD8+ T cells canonically activated by the presentation of antigen by MHC class 1 leading to downstream proliferation and cytotoxicity. In certain instances, T cells may activate without exposure to antigen, but by cytokines alone, a process known as bystander activation. We examined and compared the potential effects of bystander activation in T cells from the lung tissue of patients with idiopathic pulmonary fibrosis (IPF). Methods T cells were obtained from explanted lung tissues from patients with IPF undergoing transplantation and non-diseased lungs. These cells underwent multi-parameter spectral flow cytometry to assess the functionality. Single cell suspensions were obtained after mechanical and enzymatic digestion. Three separate conditions were created: unstimulated, T cell receptor stimulation, and bystander activation. Four cytokines (IL-2, IL-12, IL-15, IL-18) were added to the bystander activation group. All cells were stimulated for five days. We studied proliferation by CFSE dilution and cytokine production using cytometric bead assay. Two-way ANOVA was used for statistical significance. Results Overall, 16 samples were obtained (8 IPF and 8 control). Only CD8+ T cells (CD45+) were assessed during this experiment. In comparison to control, lung tissue from IPF patients had overall increased % of NKG2D CD8+ cells present in the bystander activation and unstimulated group, although not statistically significant. In addition, all IPF groups demonstrated increased proliferation of NKG2D CD8+ T cells (p=ns). The total percentage of CD8 CD103+ T cells was increased in all groups (no stimulation p=0.05, stimulation p=ns, bystander p=0.05) however it was only elevated in the bystander proliferation group (p=ns). The supernatant from this experiment was assessed for the presence of other cytokines. Pro-inflammatory cytokine, IL-6, was increased in all groups (p=0.0001). In comparison to the IPF stimulation group, there was less secretion of Granzyme B (p=0.0001) from bystander supernatant. Both stimulation and bystander group had increase in IFN-γ (p=0.01 and p=ns, respectively) (Figure 1a-b). Conclusion Despite majority of the results not achieving statistical significance, a proportion of the CD8+ T cells from lung tissue of IPF patients demonstrated increased enrichment for NKG2D+ through bystander activation. In the presence of other pro-inflammatory cytokines, this pathway could potentiate further fibrosis development in the absence of repeated antigen exposure.

OBJECTIVES:Soluble ST2 (sST2), a decoy receptor for the alarmin interleukin-33 (IL-33), has been implicated in adverse clinical outcomes in acute respiratory failure (ARF). We evaluated sST2 distribution across diverse cohorts of patients with different etiologies of ARF, compared plasma and lower respiratory tract (LRT) concentrations, and examined associations with individual organ dysfunction, biological subphenotypes, and outcomes. DESIGN:Observational study. SETTING:Multicenter cohorts of ARF patients. PATIENTS:A total of 1432 ARF patients, including 863 non-COVID and 569 COVID-19 cases, from five cohorts. INTERVENTIONS:None. MEASUREMENTS AND MAIN RESULTS:sST2 levels were measured in plasma and LRT specimens (when available) and analyzed for associations with ARF etiology, severity, organ dysfunction, systemic host response, subphenotypes, and 30-day mortality. Plasma sST2 levels were higher in non-COVID ARF patients compared with COVID-19 patients ( p < 0.05) and were markedly elevated compared with LRT levels (> 19-fold), with weak intercompartmental correlation. Elevated plasma sST2 levels were associated with extrapulmonary organ dysfunction and a hyperinflammatory ARF subphenotype but not with respiratory indices, including hypoxemia. Plasma sST2 independently predicted 30-day mortality in pooled cohort data, adjusted for age, sex, and illness severity. In longitudinal measurements, nonsurvivors had persistently elevated plasma sST2 levels in the first 2 weeks of critical illness compared with survivors. CONCLUSIONS:Plasma sST2 levels independently predict outcomes in ARF and are strongly associated with extrapulmonary organ dysfunction. The weak correlation between plasma and LRT sST2 levels suggests a predominantly systemic source. These findings highlight the potential of the IL-33/ST2 axis as a therapeutic target and warrant further investigation into its role in multiple organ dysfunction in ARF.

PURPOSE OF REVIEW:Median survival after lung transplantation is 5.7 years, which lags behind other solid organ transplants, such as heart, liver, and kidney. The major barrier to long-term survival in lung transplant recipients is chronic lung allograft dysfunction (CLAD). This review discusses the challenge of CLAD as a barrier to tolerance and identifies key areas in the field that require further development. RECENT FINDINGS:CLAD is a heterogenous disease in its kinetics of onset and severity and remains a clinical diagnosis of exclusion, based on a decline in allograft function. While acute cellular rejection and antibody-mediated rejection are major risk-factors for CLAD, other barriers to long-term allograft acceptance are aspiration and primary graft dysfunction. However infections, particularly respiratory viral infections and Cytomegalovirus (CMV) remain the most significant risks for CLAD. Additionally, the lung transplant field is limited by a lack of molecular diagnostic assays for CLAD. Further, new targets are needed for precision immunosuppression, and more studies are needed to develop novel interventions to extend allograft acceptance. SUMMARY:This review discusses new lines of study to address important unmet needs necessary to extend lung allograft acceptance. Other studies, such as tandem lung transplant and bone marrow transplant in select patients with primary immunodeficiency may provide additional lessons on how to potentially establish tolerance. However, tolerance in lung transplant is extremely rare, and further studies are needed to pursue this ultimate goal.

Influenza infection predisposes individuals to secondary pneumonia caused by a range of pathogens, including both bacterial and fungal organisms. Neutrophils are critical effector cells during infection. In this study, we analyzed the transcriptional pathways of lung neutrophils isolated from mouse models of influenza-associated pulmonary aspergillosis (IAPA) and post-influenza methicillin-resistant Staphylococcus aureus (MRSA) pneumonia to examine the immunopathological mechanisms underlying post-influenza super-infection. Pathways associated with neutrophil chemotaxis and degranulation were inhibited in IAPA compared to singular A. fumigatus infection and pathways associated with neutrophil recruitment and phagocytosis were inhibited in IAPA compared to singular influenza infection. Pathways associated with neutrophil recruitment and degranulation were inhibited in post-influenza MRSA pneumonia compared to singular MRSA infection and pathways associated with cytokine signaling were inhibited in post-influenza MRSA pneumonia compared to singular influenza infection. When the 2 types of super-infection were directly compared, pathways related to cytokine induction and neutrophil function were inhibited in IAPA neutrophils compared to post-influenza MRSA pneumonia. These data demonstrate that influenza causes neutrophil dysfunction, predisposing to secondary fungal and bacterial infections.

Inhaled therapeutics have high potential for the treatment of chronic respiratory diseases of high unmet medical need, such as idiopathic pulmonary fibrosis (IPF). Preclinical and early clinical evidence show that cellular communication network factor 2 (CCN2), previously called connective tissue growth factor (CTGF), is a promising target for the treatment of IPF. In recent phase 3 clinical trials, however, systemic CCN2 inhibition failed to demonstrate a clinically meaningful benefit. Here, we present the preclinical profile of the inhaled anti-CCN2 Anticalin® protein PRS-220. Our study demonstrates that efficient pulmonary delivery directly translates into superior efficacy in relevant models of pulmonary fibrosis when compared to systemic CCN2 inhibition. Moreover, we present a holistic approach for the preclinical characterization of inhaled PRS-220 from state-of-the art in vitro and in vivo models to novel human ex vivo and in silico models, highlighting the advantage of inhaled drug delivery for treatment of respiratory disease.

Chronic lung allograft dysfunction (CLAD) substantially limits long-term survival following lung transplantation. To identify potential targets for CLAD prevention, T cells from explanted CLAD lungs and lung-draining lymph nodes, as well as diseased and nondiseased controls were isolated and single-cell RNA sequencing and TCR sequencing were performed. TCR sequencing revealed a clonally expanded population of CD8+ tissue-resident memory T cells (TRMs) with high cytotoxic potential, including upregulation of KLRK1, encoding the co-receptor NKG2D. These cytotoxic CD8+ TRMs accumulated around the CLAD airways and had a 100-fold increase in clonal overlap with lung-draining lymph nodes when compared with non-CLAD lungs. Using a murine model of orthotopic lung transplantation, we confirmed that cytotoxic CD8+ TRM accumulation was due to chronic rejection and not transplantation alone. Furthermore, blocking NKG2D in vivo attenuated the airway remodeling following transplantation and diminished airway accumulation of CD8+ T cells. Our findings support NKG2D as a potential therapeutic target for CLAD, affecting cytotoxic CD8+ TRM accumulation.

Rationale: Tissue-resident memory T cells (TRM) comprise the majority of T cells in the human lung; they are generated after an infection or antigen exposure and remain in the lung so they can be re-activated quickly in the case of recurrent infection. More recently, TRMs have been identified as important players in both physiologic and disease states. While surface markers such as ITGAE (CD103) and CD69 help to define the identity of TRMs, the gene expression signature of lung-specific TRM in humans is still not well defined. This project utilizes a novel approach to specifically isolate and characterize true tissue-resident T cells in human lungs ex vivo. Better knowledge of differential gene regulation in this population will allow for improved analysis of their role in various disease states, as well as identify potential therapeutic targets. Methods: We utilized an ex vivo lung perfusion (EVLP) model using human lungs declined for organ donation. Lungs were ventilated at 6 cc/kg of ideal body weight and perfused between 2-4 hours. A biotinylated CD45 antibody was added to the perfusate 20 minutes prior to tissue harvesting. Single cell suspensions were generated from lung biopsies with enzymatic and mechanical digestion. CITE-seq with combined single cell RNA and T cell receptor sequencing was then performed, including a streptavidin-oligonucleotide, a technique to identify those immune cells in communication with systemic circulation (“labeled”) versus those not exposed to the antibody (“protected”). TRM were defined as those “protected” cells. Results: UMAP analysis of single cell RNA-seq data identified two main clusters of protected T cells. One of these clusters contained a significant proportion of clonally expanded T cells. As expected, these protected T cells express canonical markers of tissue residency including ITGAE and CD69. In comparison to circulating T cells, TRMs in the clonally expanded cluster downregulate genes involved in T cell migration including CCR7 and S1PR1. Interestingly, MMP25, a matrix metalloproteinase implicated in regulation of innate immunity as well as T cell infiltration in solid tumors, is upregulated in TRMs. Conclusions: The EVLP model provides a unique platform to identify and analyze TRMs in the human lung. Preliminary data obtained using this model suggests that TRMs differentially downregulate migration-associated genes and upregulate MMP25. This suggests that TRMs may prioritize Future experimental directions include validation of these differentially regulated genes using a variety of in vitro and in vivo techniques.

Lung transplant remains the primary therapeutic option for patients with end-stage lung disease, but long-term survival rates remain suboptimal compared with other solid organ transplants. Acute cellular rejection (ACR) is a significant challenge in lung transplant recipients, with T cell- mediated mechanisms playing a major role. IL-10 is known for its immunoregulatory function, although its specific role in lung allograft rejection remains unclear. Using the mouse orthotopic lung transplant model, we investigated the role of IL-10 in regulating alloeffector T cell responses. Unexpectedly, we found that IL-10 was not required for early costimulation blockade-induced allograft acceptance. However, IL-10 deficiency or blockade resulted in increased CD4+ + T cell numbers, proliferation, graft infiltration, and alloeffector responses. In the absence of IL-10, CD4+ + T cell responses predominated over CD8 responses during ACR in contrast to wild-type mice. Type 1 immunity (IFN-g) g ) responses along with elevated CD4+NKG7+ + NKG7 + and CD4+CD107a+ + CD107a + responses predominated during ACR, highlighting a critical regulatory role for IL-10 in modulating CD4+ + T cell alloimmune responses. We further demonstrated increased colocalization of NKG7 and CD107a in CD4+ + T cells from IL-10-deficient allografts, suggesting coordination in cytotoxic activity. Together, our findings highlight a critical role for IL-10 in regulation of cytotoxic CD4+NKG7+ + NKG7 + T cells, an effector population that needs further investigation to elucidate their role in lung allograft rejection. The Journal of Immunology, , 2024, 213: 898-905.- 905.

BackgroundCirculating donor-derived cell-free DNA (dd-cfDNA) levels have been proposed as a potential tool for the diagnosis of graft injury. In this study, we prospectively investigated dd-cfDNA plasma levels and their association with severe primary graft dysfunction (PGD) and graft rejection after lung transplant.MethodsA total of 40 subjects undergoing de-novo lung transplants at our institution were recruited in this study. Blood samples were collected at various time points before and after lung transplant for 1 year. Dd-cfDNA in samples was determined using AlloSure assay (CareDx Inc.). The correlation of the value of %dd-cfDNA was investigated with the incidence of PGD, acute cellular rejection (ACR), and donor-specific antibody.ResultsWe observed a rapid increase of %dd-cfDNA in the blood of recipients after lung transplantation compared to baseline. The levels of dd-cfDNA decreased during the first two weeks. The peak was observed within 72 h after transplantation. The peak values of %dd-cfDNA varied among subjects and did not correlate with severe PGD incidence. We observed an association between levels of %dd-cfDNA from blood collected at the time of transbronchial biopsy and the histological diagnosis of ACR at 3 weeks.ConclusionOur data show that circulating dd-cfDNA levels are associated with ACR early after transplantation but not with severe PGD. Plasma levels of dd-cfDNA may be a less invasive tool to estimate graft rejection after lung transplantation however larger studies are still necessary to better identify thresholds.

Purpose: Lung transplantation remains the only therapeutic option for select patients with end-stage lung diseases. Acute cellular rejection (ACR) is the major risk factor for chronic lung allograft dysfunction (CLAD), the predominant barrier for long-term survival in lung transplant recipients (LTRs). However, little is known about the role of donor specific CD4+T cell responses in ACR and the potential impact on the development of CLAD in LTRs.

"Lung Transplant Outcomes Keep BUGging Us: Acute Cellular Rejection and the Lung Microbiome." American Journal of Respiratory and Critical Care Medicine, 0(ja), pp.

Purpose: Long term survival after lung transplantation is greatly limited by the high incidence of chronic lung allograft dysfunction (CLAD), however mechanisms are incompletely characterized and effective therapy is limited. NKG2D is a cell surface protein associated with lymphocyte cytoxicity that may be involved in CLAD pathogenesis.

GDF15 (growth differentiation factor 15) is a stress cytokine with several proposed roles, including support of stress erythropoiesis. Higher circulating GDF15 levels are prognostic of mortality during acute respiratory distress syndrome, but the cellular sources and downstream effects of GDF15 during pathogen-mediated lung injury are unclear. We quantified GDF15 in lower respiratory tract biospecimens and plasma from patients with acute respiratory failure. Publicly available data from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection were reanalyzed. We used mouse models of hemorrhagic acute lung injury mediated by Pseudomonas aeruginosa exoproducts in wild-type mice and mice genetically deficient for Gdf15 or its putative receptor, Gfral. In critically ill humans, plasma levels of GDF15 correlated with lower respiratory tract levels and were higher in nonsurvivors. SARS-CoV-2 infection induced GDF15 expression in human lung epithelium, and lower respiratory tract GDF15 levels were higher in coronavirus disease (COVID-19) nonsurvivors. Inmice, intratracheal P. aeruginosa type II secretion systemexoproducts were sufficient to induce airspace and plasma release of GDF15, which was attenuated with epithelial-specific deletion of Gdf15. Mice with global Gdf15 deficiency had decreased airspace hemorrhage, an attenuated cytokine profile, and an altered lung transcriptional profile during injury induced by P. aeruginosa type II secretion systemexoproducts, which was not recapitulated in mice deficient for Gfral. AirspaceGDF15 reconstitution did not significantlymodulate key lung cytokine levels but increased circulating erythrocyte counts. Lung epithelium releasesGDF15 during pathogen injury, which is associated with plasma levels in humans and mice and can increase erythrocyte counts in mice, suggesting a novel lung-blood communication pathway.

AbstractThe endothelial glycocalyx (eGC) is a carbohydrate‐rich layer on the vascular endothelium, and its damage can lead to endothelial and organ dysfunction. Heparanase (HPSE) degrades the eGC in response to cellular stress, but its role in organ dysfunction remains unclear. This study investigates HPSE's role in lung ischemia–reperfusion (I/R) injury. A left lung hilar occlusion model was used in B6 wildtype (WT) and HPSE genetic knockout (−/−) mice to induce I/R injury in vivo. The left lungs were ischemic for 1 h followed by reperfusion for 4 h prior to investigations of lung function and eGC status. Data were compared between uninjured lungs and I/R‐injured lungs in WT and HPSE−/− mice. WT lungs showed significant functional impairment after I/R injury, whereas HPSE−/− lungs did not. Inhibition or knockout of HPSE prevented eGC damage, inflammation, and cellular migration after I/R injury by reducing matrix metalloproteinase activities. HPSE−/− mice exhibited compensatory regulation of related gene expressions. HPSE facilitates eGC degradation leading to inflammation and impaired lung function after I/R injury. HPSE may be a therapeutic target to attenuate graft damage in lung transplantation.