Short airway telomeres are associated with primary graft dysfunction and chronic lung allograft dysfunction.

Primary graft dysfunction (PGD) is a major risk factor for chronic lung allograft dysfunction (CLAD) following lung transplantation, but the mechanisms linking these pathologies are poorly understood. We hypothesized that the replicative stress induced by PGD would lead to erosion of telomeres, and that this telomere dysfunction could potentiate CLAD. In a longitudinal cohort of 72 lung transplant recipients with >6 years median follow-up time, we assessed tissue telomere length, PGD grade, and freedom from CLAD. Epithelial telomere length and fibrosis-associated gene expression were assessed on endobronchial biopsies taken at 2 to 4 weeks post-transplant by TeloFISH assay and nanoString digital RNA counting. Negative-binomial mixed-effects and Cox-proportional hazards models accounted for TeloFISH staining batch effects and subject characteristics including donor age. Increasing grade of PGD severity was associated with shorter airway epithelial telomere lengths (p = 0.01). Transcriptomic analysis of fibrosis-associated genes showed alteration in fibrotic pathways in airway tissue recovering from PGD, while telomere dysfunction was associated with inflammation and impaired remodeling. Shorter tissue telomere length was in turn associated with increased CLAD risk, with a hazard ratio of 1.89 (95% CI 1.16-3.06) per standard deviation decrease in airway telomere length, after adjusting for subject characteristics. PGD may accelerate telomere dysfunction, potentiating immune responses and dysregulated repair. Epithelial cell telomere dysfunction may represent one of several mechanisms linking PGD to CLAD.

IL-1ß-dependent extravasation of preexisting lung-restricted autoantibodies during lung transplantation activates complement and mediates primary graft dysfunction.

Preexisting lung-restricted autoantibodies (LRAs) are associated with a higher incidence of primary graft dysfunction (PGD), although it remains unclear whether LRAs can drive its pathogenesis. In syngeneic murine left lung transplant recipients, preexisting LRAs worsened graft dysfunction, which was evident by impaired gas exchange, increased pulmonary edema, and activation of damage-associated pathways in lung epithelial cells. LRA-mediated injury was distinct from ischemia-reperfusion injury since deletion of donor nonclassical monocytes and host neutrophils could not prevent graft dysfunction in LRA-pretreated recipients. Whole LRA IgG molecules were necessary for lung injury, which was mediated by the classical and alternative complement pathways and reversed by complement inhibition. However, deletion of Fc receptors in donor macrophages or mannose-binding lectin in recipient mice failed to rescue lung function. LRA-mediated injury was localized to the transplanted lung and dependent on IL-1ß-mediated permeabilization of pulmonary vascular endothelium, which allowed extravasation of antibodies. Genetic deletion or pharmacological inhibition of IL-1R in the donor lungs prevented LRA-induced graft injury. In humans, preexisting LRAs were an independent risk factor for severe PGD and could be treated with plasmapheresis and complement blockade. We conclude that preexisting LRAs can compound ischemia-reperfusion injury to worsen PGD for which complement inhibition may be effective.

CD11b suppresses TLR activation of nonclassical monocytes to reduce primary graft dysfunction after lung transplantation.

Primary graft dysfunction (PGD) is the leading cause of postoperative mortality in lung transplant recipients and the most important risk factor for development of chronic lung allograft dysfunction. The mechanistic basis for the variability in the incidence and severity of PGD between lung transplant recipients is not known. Using a murine orthotopic vascularized lung transplant model, we found that redundant activation of Toll-like receptors 2 and 4 (TLR2 and -4) on nonclassical monocytes activates MyD88, inducing the release of the neutrophil attractant chemokine CXCL2. Deletion of Itgam (encodes CD11b) in nonclassical monocytes enhanced their production of CXCL2 and worsened PGD, while a CD11b agonist, leukadherin-1, administered only to the donor lung prior to lung transplantation, abrogated CXCL2 production and PGD. The damage-associated molecular pattern molecule HMGB1 was increased in peripheral blood samples from patients undergoing lung transplantation after reperfusion and induced CXCL2 production in nonclassical monocytes via TLR4/MyD88. An inhibitor of HMGB1 administered to the donor and recipient prior to lung transplantation attenuated PGD. Our findings suggest that CD11b acts as a molecular brake to prevent neutrophil recruitment by nonclassical monocytes following lung transplantation, revealing an attractive therapeutic target in the donor lung to prevent PGD in lung transplant recipients.

Genetic effect of ischemia-reperfusion injury upon primary graft dysfunction and chronic lung allograft dysfunction in lung transplantation: evidence based on transcriptome data.

The unclear mechanism that ischemia-reperfusion injury (IRI) contributes to the development of primary graft dysfunction (PGD) and chronic lung allograft dysfunction (CLAD) remains a major issue in lung transplantation. Differentially expressed PGD-related genes and CLAD-related genes during IRI (IRI-PGD common genes and IRI-CLAD common genes) were identified using GEO datasets (GSE127003, GSE8021, GSE9102) and GeneCards datasets. Enrichment analysis and four network analyses, namely, protein-protein interaction, microRNA (miRNA)-gene, transcription factor (TF)-gene, and drug-gene networks, were then performed. Moreover, GSE161520 was analyzed to identify the differentially expressed core miRNAs during IRI in rats. Finally, Pearson correlation analysis and ROC analysis were performed. Eight IRI-PGD common genes (IL6, TNF, IL1A, IL1B, CSF3, CXCL8, SERPINE1, and PADI4) and 10 IRI-CLAD common genes (IL1A, ICAM1, CCL20, CCL2, IL1B, TNF, PADI4, CXCL8, GZMB, and IL6) were identified. Enrichment analysis showed that both IRI-PGD and IRI-CLAD common genes were significantly enriched in "AGE-RAGE signaling pathway in diabetic complication" and "IL-17 signaling pathway". Among the core miRNAs, miR-1-3p and miR-335 were differentially expressed in IRI rats. Among core TFs, CEBPB expression had a significant negative correlation with P/F ratio (r = -0.33, P = 0.021). In the reperfused lung allografts, the strongest positive correlation was exhibited between PADI4 expression and neutrophil proportion (r = 0.76, P < 0.001), and the strongest negative correlation was between PADI4 expression and M2 macrophage proportion (r = -0.74, P < 0.001). In lung allografts of PGD recipients, IL6 expression correlated with activated dendritic cells proportion (r = 0.86, P < 0.01), and IL1B expression correlated with the neutrophils proportion(r = 0.84, P < 0.01). In whole blood of CLAD recipients, GZMB expression correlated with activated CD4+ memory T cells proportion (r = 0.76, P < 0.001).Our study provides the novel insights into the molecular mechanisms by which IRI contributes to PGD and CLAD and potential targets for therapeutic intervention.

Resolving the graft ischemia-reperfusion injury during liver transplantation at the single cell resolution.

Ischemia-reperfusion injury (IRI) remains the major reason for impaired donor graft function and increased mortality post-liver transplantation. The mechanism of IRI involves multiple pathophysiological processes and numerous types of cells. However, a systematic and comprehensive single-cell transcriptional profile of intrahepatic cells during liver transplantation is still unclear. We performed a single-cell transcriptome analysis of 14,313 cells from liver tissues collected from pre-procurement, at the end of preservation and 2 h post-reperfusion. We made detailed annotations of mononuclear phagocyte, endothelial cell, NK/T, B and plasma cell clusters, and we described the dynamic changes of the transcriptome of these clusters during IRI and the interaction between mononuclear phagocyte clusters and other cell clusters. In addition, we found that TNFAIP3 interacting protein 3 (TNIP3), specifically and highly expressed in Kupffer cell clusters post-reperfusion, may have a protective effect on IRI. In summary, our study provides the first dynamic transcriptome map of intrahepatic cell clusters during liver transplantation at single-cell resolution.

N-myc-interactor mediates microbiome induced epithelial to mesenchymal transition and is associated with chronic lung allograft dysfunction.

BACKGROUND: Recent evidence suggests a role for lung microbiome in occurrence of chronic lung allograft dysfunction (CLAD). However, the mechanisms linking the microbiome to CLAD are poorly delineated. We investigated a possible mechanism involved in microbial modulation of mucosal response leading to CLAD with the hypothesis that a Proteobacteria dominant lung microbiome would inhibit N-myc-interactor (NMI) expression and induce epithelial to mesenchymal transition (EMT). METHODS: Explant CLAD, non-CLAD, and healthy nontransplant lung tissue were collected, as well as bronchoalveolar lavage from 14 CLAD and matched non-CLAD subjects, which were followed by 16S rRNA amplicon sequencing and quantitative polymerase chain reaction (PCR) analysis. Pseudomonas aeruginosa (PsA) or PsA-lipopolysaccharide was cocultured with primary human bronchial epithelial cells (PBEC). Western blot analysis and quantitative reverse transcription (qRT) PCR was performed to evaluate NMI expression and EMT in explants and in PsA-exposed PBECs. These experiments were repeated after siRNA silencing and upregulation (plasmid vector) of EMT regulator NMI. RESULTS: 16S rRNA amplicon analyses revealed that CLAD patients have a higher abundance of phyla Proteobacteria and reduced abundance of the phyla Bacteroidetes. At the genera level, CLAD subjects had an increased abundance of genera Pseudomonas and reduced Prevotella. Human CLAD airway cells showed a downregulation of the N-myc-interactor gene and presence of EMT. Furthermore, exposure of human primary bronchial epithelial cells to PsA resulted in downregulation of NMI and induction of an EMT phenotype while NMI upregulation resulted in attenuation of this PsA-induced EMT response. CONCLUSIONS: CLAD is associated with increased bacterial biomass and a Proteobacteria enriched airway microbiome and EMT. Proteobacteria such as PsA induces EMT in human bronchial epithelial cells via NMI, demonstrating a newly uncovered mechanism by which the microbiome induces cellular metaplasia.

Identifying host microRNAs in bronchoalveolar lavage samples from lung transplant recipients infected with Aspergillus.

BACKGROUND: MicroRNAs (miRNAs) are small non-coding RNAs of ∼22 nucleotides that play a crucial role in post-transcriptional regulation of gene expression. Dysregulation of miRNA expression has been shown during microbial infections. We sought to identify miRNAs that distinguish invasive aspergillosis (IA) from non-IA in lung transplant recipients (LTRs). METHODS: We used NanoString nCounter Human miRNA, version 3, panel to measure miRNAs in bronchoalveolar lavage (BAL) samples from LTRs with Aspergillus colonization (ASP group) (n = 10), those with Aspergillus colonization and chronic lung allograft dysfunction (CLAD) (ASPCLAD group) (n = 7), those with IA without CLAD (IA group) (n = 10), those who developed IA with CLAD (IACLAD group) (n = 9), and control patients (controls) (n = 9). The miRNA profile was compared using the permutation test of 100,000 trials for each of the comparisons. We used mirDIP to obtain their gene targets and pathDIP to determine the pathway enrichment. RESULTS: We performed pairwise comparisons between patient groups to identify differentially expressed miRNAs. A total of 5 miRNAs were found to be specific to IA, including 4 (miR-145-5p, miR-424-5p, miR-99b-5p, and miR-4488) that were upregulated and the pair (miR-4454 + miR-7975) that was downregulated in IA group vs controls. The expression change for these miRNAs was specific to patients with IA; they were not significantly differentiated between IACLAD and IA groups. Signaling pathways associated with an immunologic response to IA were found to be significantly enriched. CONCLUSIONS: We report a set of 5 differentially expressed miRNAs in the BAL of LTRs with IA that might help in the development of diagnostic and prognostic tools for IA in LTRs. However, further investigation is needed in a larger cohort to validate the findings.

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

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.

"The Secret Life of Human Donor Hearts": An Examination of Transcriptomic Events During Cold Storage.

BACKGROUND: Ischemic tolerance of donor hearts has a major impact on the efficiency in utilization and clinical outcomes. Molecular events during storage may influence the severity of ischemic injury. METHODS: RNA sequencing was used to study the transcriptional profile of the human left ventricle (LV, n=4) and right ventricle (RV, n=4) after 0, 4, and 8 hours of cold storage in histidine-tryptophan-ketoglutarate preservation solution. Gene set enrichment analysis and gene ontology analysis was used to examine transcriptomic changes with cold storage. Terminal deoxynucleotidyl transferase 2´-Deoxyuridine, 5´-Triphosphate nick end labeling and p65 staining was used to examine for cell death and NFκB activation, respectively. RESULTS: The LV showed activation of genes related to inflammation and allograft rejection but downregulation of oxidative phosphorylation and fatty acid metabolism pathway genes. In contrast, inflammation-related genes were down-regulated in the RV and while oxidative phosphorylation genes were activated. These transcriptomic changes were most significant at the 8 hours with much lower differences observed between 0 and 4 hours. RNA velocity estimates corroborated the finding that immune-related genes were activated in the LV but not in the RV during storage. With increasing preservation duration, the LV showed an increase in nuclear translocation of NFκB (p65), whereas the RV showed increased cell death close to the endocardium especially at 8 hours. CONCLUSIONS: Our results demonstrated that the LV and RV of human donor hearts have distinct responses to cold ischemic storage. Transcriptomic changes related to inflammation, oxidative phosphorylation, and fatty acid metabolism pathways as well as cell death and NFκB activation were most pronounced after 8 hours of storage.

Circular RNA circTET3 mediates migration of rat vascular smooth muscle cells by targeting miR-351-5p.

Abnormal migration and proliferation of vascular smooth muscle cells (VSMCs) are the pathological basis of hyperplasia during vein graft disease. It remains unknown if circular RNAs (circRNAs) are involved in vein graft disease. In the present study, a rat vein graft model was constructed by the "cuff" technique, and whole transcriptome deep sequencing was applied to identify differential circRNAs in the grafted vein compared to the control. We identified a novel circRNA, named circTET3, whose structure was verified by Sanger sequencing and RNase R digestion. CircTET3 was increased in the grafted vein and stably located in the cytoplasm as detected by fluorescence in situ hybridization. Knockdown of circTET3 suppressed VSMC migration by acting as an endogenous miR-351-5p sponge detected by RNA pull-down and dual-luciferase reporter assays. PTPN1 was the targeted gene due to the competitive binding of circTET3 to miR-351-5p. This regulatory pathway may serve as a potential therapeutic avenue against intimal hyperplasia in vein graft disease.

Dectin-1 genetic deficiency predicts chronic lung allograft dysfunction and death.

BACKGROUNDInnate immune activation impacts lung transplant outcomes. Dectin-1 is an innate receptor important for pathogen recognition. We hypothesized that genotypes reducing dectin-1 activity would be associated with infection, graft dysfunction, and death in lung transplant recipients.METHODSWe assessed the rs16910526 CLEC7A gene polymorphism Y238X, which results in dectin-1 truncation, in 321 lung allograft recipients at a single institution and in 1,129 lung allograft recipients in the multicenter Lung Transplant Outcomes Group (LTOG) cohort. Differences in dectin-1 mRNA, cytokines, protein levels, immunophenotypes, and clinical factors were assessed.RESULTSY238X carriers had decreased dectin-1 mRNA expression (P = 0.0001), decreased soluble dectin-1 protein concentrations in bronchoalveolar lavage (P = 0.008) and plasma (P = 0.04), and decreased monocyte surface dectin-1 (P = 0.01) compared with wild-type subjects. Y238X carriers had an increased risk of fungal pathogens (HR 1.17, CI 1.0-1.4), an increased risk of graft dysfunction or death (HR 1.6, CI 1.0-2.6), as well increased mortality in the UCSF cohort (HR 1.8, CI 1.1-3.8) and in the LTOG cohort (HR 1.3, CI 1.1-1.6), compared with wild-type CLEC7A subjects.CONCLUSIONIncreased rates of graft dysfunction and death associated with this dectin-1 polymorphism may be amplified by immunosuppression that drives higher fungal burden from compromised pathogen recognition.FUNDINGThe UCSF Nina Ireland Program for Lung Health Innovative Grant program, the Clinical Sciences Research & Development Service of the VA Office of Research and Development, and the Joel D. Cooper Career Development Award from the International Society for Heart and Lung Transplantation.

Lung Transplant Outcomes in Patients With Pulmonary Fibrosis With Telomere-Related Gene Variants.

BACKGROUND: Pulmonary fibrosis (PF) is the most common disease indication for lung transplantation. Our recent work implicated an excess of rare genetic variants in the telomere-related genes TERT, RTEL1, and PARN in PF disease risk. The impact of such variants on posttransplant outcomes is uncertain. The objective of this study was to determine if patients with these PF-associated variants have altered rates of posttransplant acute rejection (AR), chronic lung allograft dysfunction (CLAD), and survival. METHODS: The study cohort consisted of 262 PF lung transplant recipients previously genetically characterized by whole exome sequencing. Thirty-one patients (11.8%) had variants in TERT, RTEL1, or PARN, whereas 231 (88.2%) did not. Multivariate Cox proportional hazards models adjusted for relevant clinical variables were used to assess the outcomes of death and CLAD. The AR burden was quantified and compared over the first posttransplant year. RESULTS: Patients with PF with disease-associated variants in TERT, RTEL1, or PARN had a significantly higher risk of death (adjusted hazard ratio [HR], 1.82; 95% CI, 1.07-3.08; P = .03) and CLAD (adjusted HR, 2.88; 95% CI, 1.42-5.87; P = .004) than patients without these variants. There was no difference in AR burden or rates of grade 3 primary graft dysfunction between the two groups. CONCLUSIONS: Rare variants in the telomere-related genes TERT, RTEL1, or PARN are associated with poor posttransplant outcomes among PF lung transplant recipients. Further research is needed to understand the biological mechanisms by which telomere-related variants increase the risk for death and CLAD.

A single-nucleotide polymorphism in a gene modulating glucocorticoid sensitivity is associated with the decline in total lung capacity after lung transplantation.

PURPOSE: Glucocorticoids are used to prevent chronic lung allograft dysfunction (CLAD) after lung transplantation (LT). Our study was aimed at assessing the association between the glucocorticoid-induced transcript 1 gene (GLCCI1) variant, which modulates glucocorticoid sensitivity, and the postoperative lung function and development of CLAD after LT. METHODS: A total of 71 recipients of LT were genotyped for the GLCCI1 variant (rs37972) and divided into three groups: the homozygous mutant allele (TT) group, the heterozygous mutant allele (CT) group, and the wild-type allele (CC) group. The results of pulmonary function tests were compared with the postoperative baseline values. RESULTS: The total lung capacity (TLC) in the TT group was significantly lower than that in the CC group at 3 years after LT (P = 0.029). In the recipients of cadaveric LT, the TLC and forced expiratory volume in 1 s in the TT group were significantly lower than those in the CC groups, resulting in a significant worse CLAD-free survival at 3 years after LT (P = 0.016). CONCLUSION: The GLCCI1 variant was associated with a significant decrease of the TLC at 3 years after LT and the development of CLAD at 3 years, especially in patients undergoing cadaveric LT.

Renal ischemia/reperfusion-induced mitophagy protects against renal dysfunction via Drp1-dependent-pathway.

Autophagy is upregulated under stress conditions to degrade superfluous proteins and recycle damaged organelles including damaged mitochondria. However, the occurrence of mitochondrial autophagy and its contribution remain to be elucidated during renal ischemia/reperfusion injury (IRI). In this study, mitophagosomes and engulfed mitochondria were frequently observed by electron microscopy after renal IRI vs. control. Meanwhile, the increase of lipidated microtubule associated protein light chain 3 (LC3-II) and decrease of mitochondrial proteins were detected by western blot, suggesting the presence of mitophagy. Drp1 translocated to mitochondria and was phosphorylated at S616 in response to IRI. Interestingly, we found that inhibiting drp1 phosphorylation with mdivi-1 significantly suppressed IRI-induced mitophagy without affecting general autophagy. Furthermore, our results showed that downregulation of mitophagy significantly exacerbated cell apoptosis and markedly aggravated kidney dysfunction induced by IRI. Taken together, these data indicate that mitophagy was activated via Drp1-dependent pathway and such mitophagic clearance of damaged mitochondria protects cells from IRI-induced apoptosis.

Donor Club Cell Secretory Protein G38A Polymorphism Is Associated With a Decreased Risk of Primary Graft Dysfunction in the French Cohort in Lung Transplantation.

BACKGROUND: Club Cell Secretory Protein (CCSP) G38A polymorphism has recently been involved in lung epithelial susceptibility to external injuries. Lung transplantation (LT) is currently limited by ischemia-reperfusion injury leading to primary graft dysfunction (PGD). We thus hypothesized that donor CCSP G38A polymorphism might impact the risk of PGD after LT. METHODS: We focused on LT included in the French multicentric Cohort in Lung Transplantation (COLT), performed between January 2009 and December 2014, and associated with preoperative blood samples from the donor and the recipient. Characteristics of the donors, recipients, procedures, early and late outcomes were prospectively recorded in COLT. The CCSP serum concentration and CCSP gene G38A polymorphism were retrospectively determined in a blind manner. Their association with grade 3 PGD was studied in univariate and multivariate analysis. RESULTS: The study group included 104 LT donors and recipients, 84 with grade 0 to 2 PGD and 20 with grade 3 PGD. Preoperative CCSP serum concentration was significantly higher in the donors (median, 22.54 ng/mL; interquartile range, 9.6-43.9) than in the recipients (median, 7.03 ng/mL; interquartile range, 0.89-19.2; P < 0.001) but none impacted the risk of grade 3 PGD (P = 0.93 and P = 0.69, respectively). Donor CCSP G38A polymorphism was associated with a decreased risk of grade 3 PGD in univariate (AG + AA 3/21 = 14.2% vs GG 10/26 = 38.4%, P = 0.044) and multivariate analysis (odds ratio associated with AG + AA, 0.22; 95% confidence interval, 0.041-0.88; P = 0.045), but recipient CCSP G38A polymorphism was not. CONCLUSIONS: Donor CCSP G38A polymorphism is associated with a decreased risk of severe PGD after LT in the COLT study. These findings should be confirmed in the frame of a prospective study.

Role of tumor necrosis factor-α in epithelial-to-mesenchymal transition in transplanted kidney cells in recipients with chronic allograft dysfunction.

BACKGROUND: Chronic allograft dysfunction (CAD) is characterized by allograft kidney interstitial fibrosis, the underlying mechanism of which is unclear. Our aim was to elucidate the role and mechanism of TNF-α-induced epithelial-to-mesenchymal transition (EMT) in transplant kidney tubular interstitial fibrosis. METHODS: Human kidney tissues from normal volunteers and CAD patients were assessed using periodic acid-Schiff, Masson trichrome and immunohistochemical staining. mRNA and protein expression of E-cadherin, α-smooth muscle actin (SMA) and fibronectin(FN) in renal proximal tubule epithelial (HK-2) cells after treatment with TNF-α under different conditions were assessed using western blot and qRT-PCR analysis. Cell motility and migration were assessed using wound healing and transwell assays. Expression of Smurf2 and TNF-α-signaling pathway-related proteins in HK-2 cells treated with TNF-α was detected by western blotting. E-cadherin and α-SMA expression was also assessed in Smurf2 plasmid-transfected or Smurf2 siRNA-treated HK-2 cells. RESULTS: The expression of TNF-α, Smurf2, α-SMA, and fibronectin was significantly upregulated, while the expression of E-cad was downregulated in the CAD group compared with the normal group. The in vitro results showed that TNF-α remarkably upregulated the expression of Smurf2, α-SMA and fibronectin and downregulated the expression of E-cadherin in HK-2 cells and enhanced motility and migration in HK-2 cells. Overexpression of Smurf2 could promote the expression of α-SMA and inhibit the expression of E-cad, whereas knockdown of Smurf2 expression reversed TNF-α-induced upregulation of α-SMA and prohibited the reduction of E-cad expression. Furthermore, TNF-α-induced Smurf2 expression promoted EMT through the Akt signaling pathway. CONCLUSIONS: TNF-α induced EMT via the TNF-α/Akt/Smurf2 signaling pathways, and it may play a role in aggravating allograft kidney interstitial fibrosis in CAD patients.

Influence of Donor Lung Surfactant-A and -B Protein Expression on the Development of Primary Graft Dysfunction After Lung Transplantation: A Pilot Study.

BACKGROUND Primary graft dysfunction (PGD) is responsible of high early mortality in lung transplanted patients. We measured the rate of surfactant proteins in the organ donor, and we observed the occurrence of lung PGD in the recipient. The co-relation between these two parameters was evaluated. MATERIAL AND METHODS In this pilot study, we prospectively collected blood samples and lung biopsies in thirteen donors at the time of recovery of organs before preservation. Gene expression of SP-A, SP-B, SP-D, and CC16 was evaluated by real-time quantitative PCR. Surfactant proteins plasma levels were evaluated by ELISA. Post-transplant assessments included hemodynamic, arterial blood gas measurements, and radiographic evaluation to determine PGD and lung biopsies. RESULTS Nine of the thirteen recipients (69%) developed lung infiltrates and four (31%) developed PGD at either stages 2 or 3. SP-A and SP-B expressions were dramatically reduced in lung allografts of these patients, while lung expression of SP-D and CC16 remained unchanged. Plasma levels of SP-A, SP-B, SP-D, and CC16 did not differ. CONCLUSIONS Primary graft dysfunction may be initiated in the donor. Lung allografts with low lung SP-A and SP-B gene expression prior to implantation are associated with increased incidence of lung infiltrates after transplantation.

Telomere length in patients with pulmonary fibrosis associated with chronic lung allograft dysfunction and post-lung transplantation survival.

BACKGROUND: Prior studies have shown that patients with pulmonary fibrosis with mutations in the telomerase genes have a high rate of certain complications after lung transplantation. However, few studies have investigated clinical outcomes based on leukocyte telomere length. METHODS: We conducted an observational cohort study of all patients with pulmonary fibrosis who underwent lung transplantation at a single center between January 1, 2007, and December 31, 2014. Leukocyte telomere length was measured from a blood sample collected before lung transplantation, and subjects were stratified into 2 groups (telomere length <10th percentile vs ≥10th percentile). Primary outcome was post-lung transplant survival. Secondary outcomes included incidence of allograft dysfunction, non-pulmonary organ dysfunction, and infection. RESULTS: Approximately 32% of subjects had a telomere length <10th percentile. Telomere length <10th percentile was independently associated with worse survival (hazard ratio 10.9, 95% confidence interval 2.7-44.8, p = 0.001). Telomere length <10th percentile was also independently associated with a shorter time to onset of chronic lung allograft dysfunction (hazard ratio 6.3, 95% confidence interval 2.0-20.0, p = 0.002). Grade 3 primary graft dysfunction occurred more frequently in the <10th percentile group compared with the ≥10th percentile group (28% vs 7%; p = 0.034). There was no difference between the 2 groups in incidence of acute cellular rejection, cytopenias, infection, or renal dysfunction. CONCLUSIONS: Telomere length <10th percentile was associated with worse survival and shorter time to onset of chronic lung allograft dysfunction and thus represents a biomarker that may aid in risk stratification of patients with pulmonary fibrosis before lung transplantation.