Diagnosis Of Acute Cellular Rejection Research Articles

Flexible bronchoscopy in pediatric lung transplantation.

Pediatric lung transplantation represents a treatment option for children with advanced lung disease or pulmonary vascular disorders who are deemed an appropriate candidate. Pediatric flexible bronchoscopy is an important and evolving field that is highly relevant in the pediatric lung transplant population. It is thus important to advance our knowledge to better understand how care for children after lung transplant can be maximally optimized using pediatric bronchoscopy. Our goals are to continually improve procedural skills when performing bronchoscopy and to decrease the complication rate while acquiring adequate samples for diagnostic evaluation. Attainment of these goals is critical since allograft assessment by bronchoscopic biopsy is required for histological diagnosis of acute cellular rejection and is an important contributor to establishing chronic lung allograft dysfunction, a common complication after lung transplant. Flexible bronchoscopy with bronchoalveolar lavage and transbronchial lung biopsy plays a key role in lung transplant graft assessment. In this article, we discuss the application of bronchoscopy in pediatric lung transplant evaluation including historical approaches, our experience, and future directions not only in bronchoscopy but also in the evolving pediatric lung transplantation field. Pediatric flexible bronchoscopy has become a vital modality for diagnosing lung transplant complications in children as well as assessing therapeutic responses. Herein, we review the value of flexible bronchoscopy in the management of children after lung transplant and discuss the application of novel techniques to improve care for this complex pediatric patient population and we provide a brief update about new diagnostic techniques applied in the growing lung transplantation field.

Acute rejection post lung transplant.

To review what is currently known about the pathogenesis, diagnosis, treatment, and prevention of acute rejection (AR) in lung transplantation. Epigenomic and transcriptomic methods are gaining traction as tools for earlier detection of AR, which still remains primarily a histopathologic diagnosis. Acute rejection is a common cause of early posttransplant lung graft dysfunction and increases the risk of chronic rejection. Detection and diagnosis of AR is primarily based on histopathology, but noninvasive molecular methods are undergoing investigation. Two subtypes of AR exist: acute cellular rejection (ACR) and antibody-mediated rejection (AMR). Both can have varied clinical presentation, ranging from asymptomatic to fulminant ARDS, and can present simultaneously. Diagnosis of ACR requires transbronchial biopsy; AMR requires the additional measuring of circulating donor-specific antibody (DSA) levels. First-line treatment in ACR is increased immunosuppression (pulse-dose or tapered dose glucocorticoids); refractory cases may need antibody-based lymphodepletion therapy. First line treatment in AMR focuses on circulating DSA removal with B and plasma cell depletion; plasmapheresis, intravenous human immunoglobulin (IVIG), bortezomib, and rituximab are often employed.

Longitudinal Lower Airway Microbial Signatures of Acute Cellular Rejection in Lung Transplantation.

Rationale: Acute cellular rejection (ACR) after lung transplant is a leading risk factor for chronic lung allograft dysfunction. Prior studies have demonstrated dynamic microbial changes occurring within the allograft and gut that influence local adaptive and innate immune responses. However, the lung microbiome's overall impact on ACR risk remains poorly understood. Objectives: To evaluate whether temporal changes in microbial signatures were associated with the development of ACR. Methods: We performed cross-sectional and longitudinal analyses (joint modeling of longitudinal and time-to-event data and trajectory comparisons) of 16S rRNA gene sequencing results derived from lung transplant recipient lower airway samples collected at multiple time points. Measurements and Main Results: Among 103 lung transplant recipients, 25 (24.3%) developed ACR. In comparing samples acquired 1 month after transplant, subjects who never developed ACR demonstrated lower airway enrichment with several oral commensals (e.g., Prevotella and Veillonella spp.) than those with current or future (beyond 1 mo) ACR. However, a subgroup analysis of those who developed ACR beyond 1 month revealed delayed enrichment with oral commensals occurring at the time of ACR diagnosis compared with baseline, when enrichment with more traditionally pathogenic taxa was present. In longitudinal models, dynamic changes in α-diversity (characterized by an initial decrease and a subsequent increase) and in the taxonomic trajectories of numerous oral commensals were more commonly observed in subjects with ACR. Conclusions: Dynamic changes in the lower airway microbiota are associated with the development of ACR, supporting its potential role as a useful biomarker or in ACR pathogenesis.

Single center study investigating the clinical association of donor-derived cell-free DNA with acute outcomes in lung transplantation.

Circulating 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. A 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. We 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. Our 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.

Higher Molecular Injury at Diagnosis of Acute Cellular Rejection Increases the Risk of Lung Allograft Failure: A Clinical Trial.

Rationale: The association of acute cellular rejection (ACR) with chronic lung allograft dysfunction (CLAD) in lung transplant recipients has primarily been described before consensus recommendations incorporating restrictive phenotypes. Furthermore, the association of the degree of molecular allograft injury during ACR with CLAD or death remains undefined. Objectives: To investigate the association of ACR with the risk of CLAD or death and to further investigate if this risk depends on the degree of molecular allograft injury. Methods: This multicenter, prospective cohort study included 188 lung transplant recipients. Subjects underwent serial plasma collections for donor-derived cell-free DNA (dd-cfDNA) at prespecified time points and bronchoscopy. Multivariable Cox proportional-hazards analysis was conducted to analyze the association of ACR with subsequent CLAD or death as well as the association of dd-cfDNA during ACR with risk of CLAD or death. Additional outcomes analyses were performed with episodes of ACR categorized as "high risk" (dd-cfDNA ⩾ 1%) and "low risk" (dd-cfDNA < 1%). Measurements and Main Results: In multivariable analysis, ACR was associated with the composite outcome of CLAD or death (hazard ratio [HR], 2.07 [95% confidence interval (CI), 1.05-4.10]; P = 0.036). Elevated dd-cfDNA ⩾ 1% at ACR diagnosis was independently associated with increased risk of CLAD or death (HR, 3.32; 95% CI, 1.31-8.40; P = 0.012). Patients with high-risk ACR were at increased risk of CLAD or death (HR, 3.13; 95% CI, 1.41-6.93; P = 0.005), whereas patients with low-risk status ACR were not. Conclusions: Patients with ACR are at higher risk of CLAD or death, but this may depend on the degree of underlying allograft injury at the molecular level. Clinical trial registered with www.clinicaltrials.gov (NCT02423070).

Role of NRF2 and reactive aldehydes in acute cellular rejection in liver transplant recipients

ObjectiveTo evaluate the level of oxidative stress and antioxidative response in the transplanted liver and its role in acute cellular rejection (ACR). Particular attention was paid to ACR diagnosis in patients with hepatitis C (HCV), as histopathological features of ACR and viral disease recurrence overlap. MethodsThis retrospective study included 40 liver transplant patients who underwent liver transplantation with two consecutive liver biopsies performed during one hospitalization period: 1.) initial biopsy of the donor liver (before implantation) and 2.) indication biopsy (after suspected ACR). Based on the etiology, patients were divided into two groups: 22 patients with alcoholic liver cirrhosis (EtOH group) and 18 patients with hepatitis C cirrhosis (HCV group). We analyzed the presence of acrolein, HNE (4-hydroxynonenal), and the major antioxidant transcription factor NRF2 (nuclear factor erythroid 2-related factor 2) in both biopsies. ResultsThe presence of acrolein and HNE in both biopsies indicates increased oxidative stress, while the decrease in these aldehydes in the indication biopsies indicates a decrease in oxidative stress over time, reflecting liver graft recovery. The absence of NRF2 in both biopsies reflects significantly reduced antioxidant protection in patients undergoing liver transplantation. ConclusionThe results support the role of oxidative stress in the pathogenesis of ACR. The presence of acrolein and the absence of HNE in the indication biopsy in patients with ACR could contribute to the diagnosis of ACR in clinical practice when functional antibodies are tested in the clinical setting.

Application of Intestinal Barrier Molecules in the Diagnosis of Acute Cellular Rejection After Intestinal Transplantation.

Diagnosing acute rejection after intestinal transplantation currently heavily relies on histopathological analysis of graft biopsies. However, the invasive risks associated with ileoscopic examination and the inaccessibility for biopsy after ileostomy closure hinder real-time detection of rejection responses. Molecules comprising the intestinal barrier have been identified as physiological and molecular biomarkers for various bowel conditions and systemic diseases. To investigate the potential of barrier function-related molecules in diagnosing rejection after intestinal transplantation, plasma samples were collected longitudinally from transplant recipients. The samples were categorized into "indeterminate for rejection (IND)" and "acute rejection (AR)" groups based on clinical diagnoses at each time point. The longitudinal association between plasma levels of these barrier function-related molecules and acute rejection was analyzed using the generalized estimating equations (GEE) method. Logistic GEE models revealed that plasma levels of claudin-3, occludin, sIgA, and zonulin were independent variables correlated with the clinical diagnosis of acute rejection. The subsequent prediction model demonstrated moderate ability in discriminating between IND and AR samples, with a sensitivity of 76.0%, specificity of 89.2%, and accuracy of 84.6%. In conclusion, monitoring plasma levels of claudin-3, occludin, sIgA, and zonulin shows great potential in aiding the diagnosis of acute rejection after intestinal transplantation.

Value of Doppler ultrasonography in predicting clinical outcomes for patients with acute cellular rejection after liver transplantation.

This study investigated the value of Doppler ultrasonography in predicting clinical outcomes after antirejection treatment for patients with acute cellular rejection (ACR) following liver transplantation (LT). This retrospective study included 84 patients who were pathologically diagnosed with ACR and received antirejection treatment within 90 days following LT. Two radiologists searched for abnormal Doppler parameters at ACR diagnosis and within 7 days after antirejection treatment initiation, including portal blood velocity (PBV) <20 cm/s, hepatic artery resistive index <0.5, and a monophasic hepatic vein flow pattern. Interval PBV changes were also evaluated. The frequencies of abnormal Doppler parameters and PBV changes were compared by treatment outcome. The frequency of abnormal PBV in the early post-treatment phase (PBVearly post-treatment) was significantly higher among poor responders (50.0% [10/20]) than among good responders (7.8% [5/64]) (P<0.001). The sensitivity, specificity, and accuracy of abnormal PBVearly post-treatment as a predictor of poor response to antirejection treatment were 50.0% (10/20), 92.2% (59/64), and 82.1% (69/84), respectively. A decrease (>10%) from the PBV at event (PBVevent) to PBVearly post-treatment was significantly more common among poor responders (50.0% [10/20]) than among good responders (20.3% [13/64]) (P=0.019). The sensitivity, specificity, and accuracy of this PBV decrease in predicting poor treatment response were 50.0% (10/20), 79.7% (51/64), and 72.6% (61/84), respectively. Abnormal PBVearly post-treatment and a decrease between PBVevent and PBVearly post-treatment were significantly associated with poor treatment response in patients with ACR after LT. Consequently, Doppler ultrasonography may be useful for predicting clinical outcomes in these patients.

Regulatory T Cell Biomarkers Identify Patients at Risk of Developing Acute Cellular Rejection in the First Year Following Heart Transplantation.

Acute cellular rejection (ACR), an alloimmune response involving CD4+ and CD8+ T cells, occurs in up to 20% of patients within the first year following heart transplantation. The balance between a conventional versus regulatory CD4+ T cell alloimmune response is believed to contribute to developing ACR. Therefore, tracking these cells may elucidate whether changes in these cell populations could signal ACR risk. We used a CD4+ T cell gene signature (TGS) panel that tracks CD4+ conventional T cells (Tconv) and regulatory T cells (Treg) on longitudinal samples from 94 adult heart transplant recipients. We evaluated combined diagnostic performance of the TGS panel with a previously developed biomarker panel for ACR diagnosis, HEARTBiT, while also investigating TGS' prognostic utility. Compared with nonrejection samples, rejection samples showed decreased Treg- and increased Tconv-gene expression. The TGS panel was able to discriminate between ACR and nonrejection samples and, when combined with HEARTBiT, showed improved specificity compared with either model alone. Furthermore, the increased risk of ACR in the TGS model was associated with lower expression of Treg genes in patients who later developed ACR. Reduced Treg gene expression was positively associated with younger recipient age and higher intrapatient tacrolimus variability. We demonstrated that expression of genes associated with CD4+ Tconv and Treg could identify patients at risk of ACR. In our post hoc analysis, complementing HEARTBiT with TGS resulted in an improved classification of ACR. Our study suggests that HEARTBiT and TGS may serve as useful tools for further research and test development.

Histologic features of allograft livers in patients treated for rejection before biopsy

Liver biopsy is essential for management in liver transplant patients with clinical features suspicious for acute cellular rejection (ACR). As more patients are transplanted for noninfectious indications, it has become increasingly common for them to receive treatment for presumed ACR before biopsy. The effect of pretreatment on the classic histologic triad of ACR's mixed portal inflammation, endothelialitis, and bile duct damage is not well described. Here we report a retrospective study of 70 liver transplant biopsies performed on 53 patients for suspected ACR between 2018 and 2021. Thirty-seven biopsies had a clinical diagnosis of ACR after biopsy. Pretreatment with steroids, antithymocyte globulin, or other increased immunosuppression was given before biopsy in 17 of 37 cases; 20 not-pretreated cases acted as controls. A representative hematoxylin and eosin-stained slide from each biopsy was reviewed independently in a blinded fashion by 3 hepatic pathologists, graded according to the Banff system, assigned a Rejection Activity Index (RAI), and assessed for other histologic features. We found that pretreated biopsies had significantly less portal inflammation (P<.001), less endothelialitis (P<.001), lower RAI (P<.001), and less prominent eosinophils (P=.048) compared to not-pretreated biopsies. There was no significant difference for the other examined variables, including bile duct inflammation/damage (P=.32). Our findings suggest that portal inflammation and endothelialitis become less prominent with pretreatment, whereas bile duct inflammation/damage may take longer to resolve. When evaluating biopsies for suspected ACR, the finding of bile duct inflammation/damage should raise the possibility of partially treated ACR, even in the absence of endothelialitis and portal inflammation.

Circulating microRNAs in cellular and antibody-mediated heart transplant rejection.

Noninvasive monitoring of heart allograft health is important to improve clinical outcomes. MicroRNAs (miRs) are promising biomarkers of cardiovascular disease and limited studies suggest they can be used to noninvasively diagnose acute heart transplant rejection. The Genomic Research Alliance for Transplantation (GRAfT) is a multicenter prospective cohort study that phenotyped heart transplant patients from 5 mid-Atlantic centers. Patients who had no history of rejection after transplant were compared to patients with acute cellular rejection (ACR) or antibody-mediated rejection (AMR). Small RNA sequencing was performed on plasma samples collected at the time of an endomyocardial biopsy. Differential miR expression was performed with adjustment for clinical covariates. Regression was used to develop miR panels with high diagnostic accuracy for ACR and AMR. These panels were then validated in independent samples from GRAfT and Stanford University. Receiver operating characteristic curves were generated and area under the curve (AUC) statistics calculated. Distinct ACR and AMR clinical scores were developed to translate miR expression data for clinical use. The GRAfT cohort had a median age of 52 years, with 35% females and 45% Black patients. Between GRAfT and Stanford, we included 157 heart transplant patients: 108 controls and 49 with rejection (50 ACR and 38 AMR episodes). After differential miR expression and regression analysis, we identified 12 miRs that accurately discriminate ACR and 17 miRs in AMR. Independent validation of the miR panels within GRAfT led to an ACR AUC 0.92 (95% confidence interval [CI]: 0.86-0.98) and AMR AUC 0.82 (95% CI: 0.74-0.90). The externally validated ACR AUC was 0.72 (95% CI: 0.59-0.82). We developed distinct ACR and AMR miR clinical scores (range 0-100), a score ≥ 65, identified ACR with 86% sensitivity, 76% specificity, and 98% negative predictive value, for AMR score performance was 82%, 84% and 97%, respectively. We identified novel miRs that had excellent performance to noninvasively diagnose acute rejection after heart transplantation. Once rigorously validated, the unique clinical ACR and AMR scores usher in an era whereby genomic biomarkers can be used to screen and diagnose the subtype of rejection. These novel biomarkers may potentially alleviate the need for an endomyocardial biopsy while facilitating the initiation of targeted therapy based on the noninvasive diagnosis of ACR or AMR.

216.6: Regulatory T Cell Biomarkers Identify Patients at Risk of Developing Acute Cellular Rejection in the First Year Following Heart Transplantation

Background: Acute cellular rejection (ACR), an alloimmune response involving CD4+ and CD8+ T cells, is a leading cause of mortality and morbidity following heart transplantation. While immunosuppressive drugs such as corticosteroids and calcineurin inhibitors (e.g., cyclosporine, tacrolimus) have largely reduced ACR events immediately following transplantation, ACR remains a risk factor in the long-term, with outcomes including graft loss and death. The balance between a conventional versus regulatory CD4+ T cell alloimmune response is believed to play a major role in the development of ACR. Therefore tracking these cells following transplantation would elucidate if changes in these cell populations could signal ACR risk. Understanding this link could be an important step towards developing tools that can detect and/or prevent ACR. Methods: To describe changes in conventional versus regulatory CD4+ T cells during the course of heart transplantation, we used a CD4+ T cell gene signature panel (TGS) that tracks CD4+ T conventional (Tconv) and regulatory cells (Treg) on longitudinal samples from 94 adult heart transplant recipients. We investigated Tconv and Treg levels pre-transplant vs. post-transplant, as well as between non-rejection and ACR samples. Also, we compared TGS with HEARTBiT assay, a gene expression assay for exclusionary diagnosis of ACR in heart transplant patients in the first year following transplantation, on ACR diagnosis performance while also investigating its prognostic utility. Results: Compared to pre-transplant samples, non-rejection samples showed increased Treg- and decreased Tconv-gene expression while such differences were reduced in ACR samples. Within the post-transplant samples, we saw that the TGS panel could discriminate between ACR and non-rejection samples with comparable performance to the HEARTBiT assay. When HEARTBiT and TGS were combined, they showed improved specificity to HEARTBiT alone (up to 10% increase in specificity at the same sensitivity level). Furthermore, within the first two months following transplantation, Treg genes showed reduced expression in patients who had developed one or more ACR events. Reduced Treg gene expression was positively associated with younger recipient age and higher intra-patient tacrolimus variability. Conclusion: We demonstrated that expression of genes associated with CD4+ Tconvs and Tregs could identify patients at greater risk of ACR. Also, complementing HEARTBiT with TGS improved the diagnostic performance of HEARTBiT. Taken together, these biomarker tools may have clinical utility in heart transplant care. This work was supported by the Canadian Institute for Health Research, the Canadian Foundation for Innovation, Genome Canada and British Columbia, the Mathematics of Information Technology and Complex Systems (Mitacs), Michael Smith Health Research BC, the PROOF Centre of Excellence, IBM, Novartis, and Astellas.

The Role of Left Atrial Longitudinal Strain in the Diagnosis of Acute Cellular Rejection in Heart Transplant Recipients.

Our aim was to investigate the role of left atrial longitudinal strain (LALS) in the non-invasive diagnosis of acute cellular rejection (ACR) episodes in heart transplant (HTx) recipients. Methods: We performed successive echocardiographic exams in 18 consecutive adult HTx recipients in their first year after HTx within 3 h of the routine surveillance endomyocardial biopsies (EMB) in a single center. LALS parameters were analyzed with two different software. We investigated LALS association with ACR presence, as well as inter-vendor variability in comparable LALS values. Results: A total of 147 pairs of EMB and echo exams were carried out. Lower values of LALS were significantly associated with any grade of ACR presence. Peak atrial longitudinal strain (PALS) offered the best diagnostic value for any grade of ACR, with a C statistic of 0.77 using one software (95% CI 0.68–0.84, p < 0.0005) and 0.64 with the other (95% CI 0.54–0.73, p = 0.013) (p = 0.02 for comparison between both curves). Reproducibility between comparable LALS parameters was poor (intraclass correlation coefficients were 0.60 for PALS, 95% CI 0.42–0.73, p < 0.0005; and 0.42 for PALS rate, 95% CI −0.13–0.68, p < 0.0005). Conclusions: LALS variables might be a sensitive marker of ACR in HTx recipients, principally discriminating between those studies without rejection and those with any grade of ACR. Inter-vendor variability was significant.

Peripheral blood and bronchoalveolar leukocyte profile in lung transplant recipients and their changes according to immunosuppressive regimen: A single-center experience.

BackgroundAfter lung transplantation (LuTX), lower respiratory tract infections (LRTI) and acute cellular rejection (ACR) are associated with changes in peripheral blood and bronchoalveolar lavage fluid mononuclear cell profile (PBMC and BALIC). PBMC is also influenced by immunosuppressive regimen and its changes with postoperative time. First‐year PBMC and BALIC changes were evaluated in this study with rabbit anti‐thymocyte globulin (ATG) and alemtuzumab (AL) induction therapy.MethodsIn total, 64 LuTX recipients were included, 53 of them received AL and 11 ATG as induction therapy. PBMC and BALIC were examined routinely and in cases suspicious of infection and/or rejection. A PBMC‐ and BALIC‐based algorithm for infection and rejection prediction was also tested.ResultsIn the AL group, peripheral blood lymphocyte and basophil cell numbers were significantly reduced, while the neutrophil cell number elevation during LRTI was significantly higher compared to the control. Early postoperative measurements showed a lower BALIC lymphocyte count. The algorithm had 17% sensitivity and 94% specificity for ACR in all patients and 33% sensitivity and 95% specificity for ACR with coexisting LRTI.ConclusionBALIC is not significantly influenced by the immunosuppressive regimen. PBMC‐ and BALIC‐based algorithm may improve the differential diagnosis of ACR.

Transbronchial Cryobiopsy Compared to Forceps Biopsy for Diagnosis of Acute Cellular Rejection in Lung Transplants: Analysis of 63 Consecutive Procedures.

Background: Acute cellular rejection (ACR) is a complication after lung transplantation (LTx). The diagnosis of ACR is based on histologic findings using transbronchial forceps biopsy (FB). However, its diagnostic accuracy is limited because of the small biopsy size and crush artifacts. Transbronchial cryobiopsy (CB) provides a larger tissue size compared with FB. Methods: FB and CB were obtained consecutively during the same bronchoscopy (February 2020–April 2021). All biopsies were scored according to the ISHLT criteria by three pathologists. Interobserver agreement was scored by the kappa index. We assessed the severity of bleeding and the presence of pneumothorax. Results: In total, 35 lung transplant recipients were included, and 126 CBs and 315 FBs were performed in 63 consecutive bronchoscopies. ACR (A1–A3, minimal–moderate) was detected in 18 cases (28.6%) by CB, whereas ACR was detected in 3 cases (4.8%) by FB. Moderate and severe bleeding complicated FB and CB procedures in 23 cases (36.5%) and 1 case (1.6%), respectively. Pneumothorax occurred in 6.3% of patients. The interobserver agreement was comparable for both CB and FB. Conclusions: CB provided an improved diagnostic yield for ACR diagnosis, leading to reclassification and changes in treatment strategies in 28.6% of cases. Prospective studies should better define the role of CB after LTx.

Cardiac Allograft Rejection Induces Changes in Nucleocytoplasmic Transport: RANGAP1 as a Potential Non-Invasive Biomarker.

The non-invasive diagnosis of acute cellular rejection (ACR) is a major challenge. We performed a molecular study analyzing the predictive capacity of serum RanGTPase AP1 (RANGAP1) for diagnosing ACR during the first year after heart transplantation (HT). We included the serum samples of 75 consecutive HT patients, extracted after clinical stability, to determine the RANGAP1 levels through ELISA. In addition, various clinical, analytical, and echocardiographic variables, as well as endomyocardial biopsy results, were collected. RANGAP1 levels were higher in patients who developed ACR (median 63.15 ng/mL; (inter-quartile range (IQR), 36.61–105.69) vs. 35.33 ng/mL (IQR, 19.18–64.59); p = 0.02). Receiver operating characteristic (ROC) curve analysis confirmed that RANGAP1 differentiated between patients with and without ACR (area under curve (AUC), 0.70; p = 0.02), and a RANGAP1 level exceeding the cut-off point (≥90 ng/mL) was identified as a risk factor for the development of ACR (OR, 6.8; p = 0.006). Two independent predictors of ACR identified in this study were higher RANGAP1 and N-terminal pro-brain natriuretic peptide levels. The analysis of the ROC curve of the model showed a significant AUC of 0.77, p = 0.001. Our findings suggest that RANGAP1 quantification facilitates risk prediction for the occurrence of ACR and could be considered as a novel non-invasive biomarker of ACR.

Transbronchial Cryobiopsy vs Conventional Forceps Biopsy in Lung Transplant Recipients Multicenter Study

<h3>Purpose</h3> Transbronchial biopsy is considered the gold standard in the diagnosis of acute cellular rejection after lung transplantation. There still remains the discussion in conventional forceps biopsy comparing to cryobiopsy in lung transplant recipients. This study aims to compare the diagnostic yield and potential complications in both procedures. <h3>Methods</h3> In retrospective multicenter study all transbronchial biopsies (TBBs) were evaluated since 2010 till 2019. Surveillance TBBs were performed in 1, 3, 6 and 12 months after lung transplantation. Additional diagnostic bronchoscopies were performed if clinically indicated. Pneumothorax was categorized as none, partial recovered without treatment and pneumothorax required inserting of chest drainage. Bleeding was predefined as none, mild bleeding requiring suction and instillation of cold saline solution and severe bleeding requiring additional instillation of adrenaline. TBBs were graded according to the International Society of Heart and Lung Transplantation (ISHLT) consensus guidelines for the diagnosis of lung allograft rejection. <h3>Results</h3> In 309 patients a total of 448 forceps biopsies and 573 cryobiopsies were performed. The incidence of pneumothorax was significantly (p<0,001) higher after forceps biopsies (9,2 % vs 3,3 %). Bleeding was significantly greater in the cryobiopsy group (cryobiopsy/forceps biopsy: no bleeding 80,4%/96,4%; mild 15,4%/2,4%; severe 4,2%/1.1%; p < 0.001). The mean size of cryo-specimen was five times larger than forceps-group samples. The diagnostic rate of acute rejection using cryobiopsy and forceps biopsy was similar A0 grade 36% vs 39%, A1 grade 46% vs 35%, A2 grade 14% vs 17,5%, A3-A4 grade 4% vs 8,5%. But evidently larger specimens from cryobiopsy catched the presence of bronchiolus in 51% compared with 8% from forceps samples which shows the major significance (p<0,001). <h3>Conclusion</h3> Transbronchial cryobiopsies compared to traditional forceps biopsies provided superior quality of the specimens with significantly higher capture of bronchiolus and lower rate of non-diagnostic samples which also enable potential possibility for futher evaluation of humoral and chronic rejection. Cryobiopsies reduce the number of specimens with acceptable complications.

Circulating microRNA Biomarkers in Cellular and Antibody-Mediated Heart Transplant Rejection

<h3>Purpose</h3> Non-invasive monitoring of heart allograft health is an important clinical goal and circulating microRNAs (miRs) have been associated with acute rejection in limited studies. <h3>Methods</h3> The Genomic Research Alliance for Transplantation (GRAfT) is a multicenter prospective study. Patients with no history of rejection after transplant were selected as controls. Cases were patients with acute cellular rejection (ACR) or antibody-mediated rejection (AMR). Small RNA sequencing was performed on plasma samples collected at the time of an endomyocardial biopsy. Differential miR expression analysis and LASSO regression was used to develop ACR and AMR miR panels while adjusting for clinical covariates. The panels were validated in an independent sample from GRAfT and Stanford University. Receiver operating characteristic curves were generated and area under the curve (AUC) statistics are reported. Distinct ACR and AMR clinical scores were developed to translate gene expression data for clinical use. <h3>Results</h3> The GRAfT cohort had a median age of 52 years, with 35% females and 45% Blacks. We included 85 controls and 34 patients with rejection (20 ACR, 14 AMR). In ACR and AMR, using differential gene expression and regression analysis, we identified 11 and 15 miRs that accurately discriminated ACR and AMR (p<0.05). Independent validation of the miR panels within GRAfT led to excellent diagnostic accuracy for ACR AUC 0.92 (95% CI: 0.86-0.98) and AMR AUC 0.82 (95% CI: 0.74-0.90). The externally validated ACR AUC was 0.91 (95% CI: 0.82-0.99). Distinct ACR and AMR miR clinical scores were developed ranging from 0-100; using a threshold of 65 identified ACR with a sensitivity of 86%, specificity of 78% and negative predictive value of 98%, for AMR the respective performance was 82%, 84% and 97% (Figure). <h3>Conclusion</h3> The unique clinical ACR and AMR scores usher in an era whereby genomic biomarkers are used not only for rejection surveillance but permit accurate, non-invasive diagnosis of ACR and AMR.

Impact of Liver Biopsy Size on Histopathologic Evaluation of Liver Allograft Rejection.

Allograft liver biopsy is the gold standard in assessing transplant recipients for graft dysfunction. The impact of biopsy sample size on the diagnosis of acute cellular rejection (ACR) has not been studied. To assess the relationship of biopsy sample length with the diagnosis and determine optimal biopsy sample size in the transplant setting. We retrospectively reviewed 68 core biopsies from patients with a history of liver transplant. Each biopsy sample was read, on 5 different occasions with differing lengths, to assess for ACR per Banff criteria. Categorical agreement was calculated from rejection severity. The length of biopsy sample strongly correlated with the number of portal tracts. ACR rates increased from 73.5% to 79.4% with increase in length from 1 cm to 2 cm, and moderate rejection increased from 27.9% to 33.82%. At 1.0 and 1.5 cm, no cases of severe rejection were detected; at 2.0 cm, 1 case was detected; and at 3.0 cm, 2 cases were detected. The major error rate was reduced to less than 10% with a length of 2.0 cm, at which length the average number of complete and partial portal triads was 10 and 13, respectively. The likelihood of diagnosing ACR and rejection grade increased substantially with increase in biopsy sample length. This study suggests that a minimum length of 2 cm, 10 complete portal triads, or 13 partial/complete portal triads should be obtained for confident exclusion and grading of ACR.

Plasma Exchange Is an Effective Adjunct in the Management of Hyperbilirubinemia in Early Allograft Dysfunction and Small for Size Syndrome after Liver Transplant

Background: Management of early allograft dysfunction (EAD) and small for size syndrome (SFSS) after liver transplant remains challenging and care is largely based around supportive treatment. The role of plasmapheresis/plasma exchange in SFSS is not well established. However, it can potentially serve an important role as a salvage option by reducing the metabolic load of toxins including bilirubin and supplying important coagulation proteins to maintain coagulation homeostasis. Methods: We present three cases. A 19-year-old female underwent living-donor liver transplant (LDLT) and received a healthy right lobe graft. Graft-to-recipient weight ratio (GRWR) was 0.85. She developed SFSS and underwent one cycle of plasma exchange on post-operative day (POD) 8 (peak bilirubin 414umol/l) that was prematurely terminated as she developed an allergic response. A 58-year-old female underwent deceased-donor liver transplant and developed acute worsening hyperbilirubinemia on POD8-10 (from 116umol/l to 858 umol/l). She underwent one cycle of plasma exchange that was sufficient to serve as a bridge to treat the hyperbilirubinemia until liver biopsy diagnosis of acute cellular rejection was obtained and definitive steroid treatment was initiated. A 42-year-old female underwent LDLT and received a left lobe graft (GRWR 0.75). She developed SFSS despite portal inflow modulation performed during transplant. She underwent three sessions of plasma exchange (POD25,27,30). Results: In the three cases bilirubin levels improved significantly following plasma exchange, and graft recovery was evident. Conclusion: Our cases suggest the efficacy of plasma exchange as a potential adjunct for the supportive treatment of hyperbilirubinemia in EAD and SFSS to avoid re-transplantation.