Marker Of Rejection Research Articles

Antithymocyte Globulin Induction is Associated with Complement Deposition in Pediatric Cardiac Transplant Biopsies

Purpose Antithymocyte globulin (ATG) has been increasingly used as a means of induction immunosuppression (I-I) in pediatric cardiac transplantation (PCT). ATG contains polyclonal antibodies directed primarily against human T-lymphocytes and may also contain other antibodies with affinity for endothelial surface antigens and other tissues in the transplanted organ, resulting in complement (C4d) deposition (C4dD). This phenomenon has been demonstrated in endomyocardial biopsies (EMB) of adult cardiac transplants though the effects of C4dD on rejection risk remain unclear. It has been thought that use of basiliximab for induction may avoid the risk of (C4dD) and decrease the risk of rejection. We examined C4dD and incidence of rejection the relationship among children treated with ATG I-I as well as those treated with basiliximab I-I. Methods A retrospective review of charts of patients with PCT performed at our center between June 2012 and April 2018 was performed. Results of C4d immunohistochemistry were available from all EMB of patients (n=56). Results The population was predominantly male (70%) with a median age of 10 years (IQR 2.63 -14.46 years). 28 patients received ATG and 28 patients received basiliximab as induction therapy. Sixty percent (60%) of the ATG patients had C4dD on the 1st biopsy post-transplant and 25% of patients had a persistent C4dD on the next 5 EMB. When C4dD was demonstrated in ATG patients, other markers of significant rejection were rare; only 4% had ACR grade 2 or higher by the revised 2004 ISHLT grading system (historically 3A, 3B and 4 rejection) and/or histologic evidence of AMR. Only 3% of patients receiving basiliximab for induction had C4dD on any EMB in the first year post-transplant. When C4dD was demonstrated in basiliximab patients, other markers of significant rejection were found frequently; 57% had ACR grade 2 or higher by the revised 2004 ISHLT grading system and/or histologic evidence of AMR. Conclusion C4d deposition was more common on EMB up to 1 year post-PCT in patients receiving ATG induction, compared to basiliximab. Despite less C4dD in basiliximab EMBs, these patients had more rejection episodes. The presence of C4dD may thus not be an accurate measure of rejection in PCT patients and should be taken into consideration when reviewing and determining treatment options. Antithymocyte globulin (ATG) has been increasingly used as a means of induction immunosuppression (I-I) in pediatric cardiac transplantation (PCT). ATG contains polyclonal antibodies directed primarily against human T-lymphocytes and may also contain other antibodies with affinity for endothelial surface antigens and other tissues in the transplanted organ, resulting in complement (C4d) deposition (C4dD). This phenomenon has been demonstrated in endomyocardial biopsies (EMB) of adult cardiac transplants though the effects of C4dD on rejection risk remain unclear. It has been thought that use of basiliximab for induction may avoid the risk of (C4dD) and decrease the risk of rejection. We examined C4dD and incidence of rejection the relationship among children treated with ATG I-I as well as those treated with basiliximab I-I. A retrospective review of charts of patients with PCT performed at our center between June 2012 and April 2018 was performed. Results of C4d immunohistochemistry were available from all EMB of patients (n=56). The population was predominantly male (70%) with a median age of 10 years (IQR 2.63 -14.46 years). 28 patients received ATG and 28 patients received basiliximab as induction therapy. Sixty percent (60%) of the ATG patients had C4dD on the 1st biopsy post-transplant and 25% of patients had a persistent C4dD on the next 5 EMB. When C4dD was demonstrated in ATG patients, other markers of significant rejection were rare; only 4% had ACR grade 2 or higher by the revised 2004 ISHLT grading system (historically 3A, 3B and 4 rejection) and/or histologic evidence of AMR. Only 3% of patients receiving basiliximab for induction had C4dD on any EMB in the first year post-transplant. When C4dD was demonstrated in basiliximab patients, other markers of significant rejection were found frequently; 57% had ACR grade 2 or higher by the revised 2004 ISHLT grading system and/or histologic evidence of AMR. C4d deposition was more common on EMB up to 1 year post-PCT in patients receiving ATG induction, compared to basiliximab. Despite less C4dD in basiliximab EMBs, these patients had more rejection episodes. The presence of C4dD may thus not be an accurate measure of rejection in PCT patients and should be taken into consideration when reviewing and determining treatment options.

Peripheral Blood Eosinophil Count as a Marker of Pulmonary Allograft Rejection

Purpose Non-invasive methods for early diagnosis of acute cellular rejection (ACR) are needed. Previous studies of other solid organ transplanted recipients have shown a relationship between the eosinophil count in peripheral blood (EOSp) and the presence of ACR. However, the relationship between this biomarker and ACR in lung transplant patients remains unclear.Objective: To asses the relationship between EOSp and the presence of allograft rejection in lung transplant recipients. Methods A retrospective clinical review of 363 transbronchial biopsies (BTB) performed in 194 lung transplant patients between 2014 and 2018. We analyzed: age, date of transplant, the presence and degree of ACR, and the simultaneous absolute number of EOSp. BTB indications were classified as: 1. Protocol BTB in the first month after transplant and after ACR treatment. 2. Decrease of lung function. 3. Other indications The relationship between the EOSp count and ACR was analyzed by the Chi-square test . We calculated the sensitivity, specificity of the EOSp count to diagnose ACR. The precision of the EOS count to predict ACR was analyzed using a receiver operating characteristic ROC curve. Results BTB showed ACR in 117 of 363 cases (32.2%). The most frequent ACR degree was A2 (53%). The EOSp count in patients with ACR was higher than non-ACR patients (197 vs. 92.7; p 197 EOSp was 91% and the area under the ROC curve was 0.724 with 95% confidence interval (CI) 0.636-0.813. Conclusion The monitoring of the EOSp count can be useful to increase the diagnostic yield for ACR of BTB performed by protocol or lung function deterioration.

Cell-Free DNA in Different Clinical Scenarios after Heart Transplantation: Shedding Light or Obscuring the Picture?

Purpose Donor-derived cell-free DNA (dd-cfDNA) as a highly sensitive marker of rejection after heart transplantation (HTx) has gained emerging interest. Recent studies are based on sequencing techniques and use fractional abundance (dd-cfDNA as a fraction of total cfDNA) as their outcome. Here we present patient examples of the BIODRAFT-study (NCT03477383) based on PCR-techniques. Methods Blood samples are taken prospectively in parallel with endomyocardial biopsies (EMB) during the first year after HTx. dd-cfDNA is analyzed using targeted preamplification of 35 single nucleotide polymorphisms followed by digital droplet-PCR. Outcome is fractional abundance as well as total number of DNA copies, both from the donor and the recipient. Results 71 patients (57 adults, 14 children) are so far included and more than 500 blood samples analyzed. In otherwise stable patients, both fractional abundance and total DNA copies seem to follow biopsy patterns with respect to rejection. However, we could also identify different scenarios in which the distribution of cfDNA seems more complicated than hitherto described: Patients with primary graft failure, clinically silent CMV-infection or mild right heart failure show elevated levels of dd-cfDNA, thus complicating the interpretation of results. Some patients present with markedly elevated levels of their own cfDNA without obvious clinical reasons. Conclusion Besides the well-known concept of fractional abundance, our approach gives results that allow following absolute DNA copy numbers of both donor and recipient. This widens the horizon of cfDNA as a marker of graft injury after HTx, and gives an outlook to clinical scenarios with possible false positive (mimicking rejection) and, even worse, false negative results: high levels of recipient-cfDNA could mask ongoing rejection if only fractional abundance can be reported. The biology of cell-free DNA is complex and seems not yet fully understood.

PLASMA CELL-FREE DNA AS A NOVEL MARKER OF DISEASE SEVERITY IN PULMONARY ARTERIAL HYPERTENSION

Cell-free DNA (cfDNA) has recently been utilized as a marker of transplant rejection and is also elevated in states associated with vascular endothelial damage, including septic shock and cardiopulmonary bypass. The present study is the first to investigate the potential for cfDNA to predict disease

Global Left Ventricular Relaxation: A Useful Echocardiographic Marker of Heart Transplant Rejection and Recovery in Children

Tissue Doppler velocities are impaired after heart transplantation and further diminished in acute rejection. Left ventricular relaxation index (LVRI) was calculated as the sum of E' of the left ventricular lateral, septal and posterior walls divided by left ventricular posterior wall (LVPW) thinning (LVRI=E' lateral+E' septal+E' posterior/[systolic LVPW-diastolic LVPW/systolic LVPW]). On the basis of a prior study, LVRI > 0.8 was considered normal after transplantation. Serial LVRI measurements (n=941) were analyzed in a total of 35 patients who underwent transplantation. The sensitivity and specificity of LVRI < 0.8 for detecting rejection were calculated. LVRI was compared at baseline, at diagnosis of rejection, and at recovery after rejection treatment for each patient. The potential role of ischemic graft time, pretransplantation waiting period, and pretransplantation diagnosis on LVRI recovery was also assessed. LVRI was low early after transplantation (mean, 0.69) normalizing (mean, 0.91) at a median of 39.6days (range, 5-115days) after transplantation. Fifteen episodes of rejection were seen in 11 patients. LVRI was lower at diagnosis of rejection compared with baseline (P=.0013). LVRI < 0.8 had 93.3% sensitivity (95% CI, 68%-99.8%) and 89.5% specificity (95% CI, 67%-99%) for detecting all rejection. LVRI recovered at a mean of 28.3days after onset of treatment. No correlation was found to ischemic graft time, to pretransplantation waiting period, or to pretransplantation diagnosis. After the early posttransplantation period, serial measurements of LVRI appear to be a useful echocardiographic marker of heart transplantation rejection in children and of the effectiveness of rejection treatment. As such, this method may be of value in the ongoing clinical management of these difficult patients.

Native T1 Mapping in the Diagnosis of Cardiac Allograft Rejection: A Prospective Histologically Validated Study

ObjectivesThis study aimed to determine the role of T1 mapping in identifying cardiac allograft rejection. BackgroundEndomyocardial biopsy (EMBx), the current gold standard to diagnose cardiac allograft rejection, is associated with potentially serious complications. Cardiac magnetic resonance (CMR)–based T1 mapping detects interstitial edema and fibrosis, which are important markers of acute and chronic rejection. Therefore, T1 mapping can potentially diagnose cardiac allograft rejection noninvasively. MethodsPatients underwent CMR within 24 h of EMBx. T1 maps were acquired at 1.5-T. EMBx-determined rejection was graded according to International Society of Heart and Lung Transplant (ISHLT) criteria. ResultsOf 112 biopsies with simultaneous CMR, 60 were classified as group 0 (ISHLT grade 0), 35 as group 1 (ISHLT grade 1R), and 17 as group 2 (2R, 3R, clinically diagnosed rejection, antibody-mediated rejection). Native T1 values in patients with grade 0 biopsies and left ventricular ejection fraction >60% (983 ± 42 ms; 95% confidence interval: 972 to 994 ms) were comparable to values in nontransplant healthy control subjects (974 ± 45 ms; 95% confidence interval: 962 to 987 ms). T1 values were significantly higher in group 2 (1,066 ± 78 ms) versus group 0 (984 ± 42 ms; p = 0.0001) and versus group 1 (1,001 ± 54 ms; p = 0.001). After excluding patients with an estimated glomerular filtration rate <50 ml/min/m2, there was a moderate correlation of log-transformed native T1 with high-sensitivity troponin T (r = 0.54, p < 0.0001) and pro–B-type natriuretic peptide (r = 0.67, p < 0.0001). Using a T1 cutoff value of 1,029 ms, the sensitivity, specificity, and negative predictive value were 93%, 79%, and 99%, respectively. ConclusionsMyocardial tissue characterization with T1 mapping displays excellent negative predictive capacity for the noninvasive detection of cardiac allograft rejection and holds promise to reduce substantially the EMBx requirement in cardiac transplant rejection surveillance.

Effects of esomeprazole and pantoprazole on renal function in stable kidney transplantation recipients: A randomized clinical trial.

Renal allograft survival requires the administration of multiple immunosuppressive drugs. This strategy may lead to gastric complications that necessitate gastro-protective medications, notably, proton pump inhibitors (PPIs). This study aimed to compare the effects of pantoprazole and esomeprazole on renal function in stable renal transplant recipients. A prospective, parallel, open-label clinical trial was performed with forty-seven adult renal transplant recipients receiving immunosuppressive therapy with cyclosporine (CSA) doses adjusted to attain trough concentrations of 100-150 µg/L, mycophenolate mofetil (MMF) at 750 mg q12 hr and prednisolone at 5 mg daily at Nasser Institute, Cairo, Egypt. The enrolled participants were randomized into two groups, which received either esomeprazole or pantoprazole at the same dose (40 mg once daily). Renal function was measured at baseline and monthly for 6 months. The study was conducted between January-September 2016. Main outcome measures clinical signs of rejection reflected by renal function decline, assessed by elevated levels of serum creatinine. The mean serum creatinine level was significantly lower in the sixth month than at baseline in esomeprazole group (p 0.004); interestingly there was a continuous decrease of serum creatinine levels in esomeprazole group and nearly constant values in pantoprazole group. There was no significant difference in serum creatinine levels between the two groups. From this study, it could be concluded that esomeprazole may be preferred over pantoprazole in renal transplant recipients because it decreased serum creatinine which is one of the markers of chronic allograft rejection in stable renal transplantation recipients.

Clinical use of an immune monitoring panel in liver transplant recipients: A prospective, observational study

Immunosuppressive therapy greatly contributed to making liver transplantation the standard treatment for end-stage liver diseases. However, it remains difficult to predict and measure the efficacy of pharmacological immunosuppression. Therefore, we used a panel of standardized, commonly available, biomarkers with the aim to describe their changes in the first 3 weeks after the transplant procedure and assess if they may help therapeutic drug monitoring in better tailoring the dose of the immunosuppressive drugs.We prospectively studied 72 consecutive patients from the day of liver transplant (post-operative day #0) until the post-operative day #21. Leukocytes, neutrophils, lymphocytes (CD4+, CD8+), natural killer cells, monocytes, immunoglobulins and tacrolimus serum levels were measured on peripheral blood (at day 0, 3, 7, 14, 21 after surgery). Patients who developed infections showed significantly higher CD64+ monocytes on post operative day #7. IgG levels were lower on post operative day #3 among patients who later developed infections. We also found that a sharp decrease in IgA from post operative day #0 to 3 (−226 mg/dL in the ROC curve analysis) strongly correlates with the onset of infections among HCV− patients. No specific markers of rejection emerged from the tested panel of markers. Our results show that some early changes in peripheral blood white cells and immunoglobulins may predict the onset of infections and may be useful in modulating the immunosuppressive therapy. However, a panel of commonly available, standardized biomarkers do not support in improving therapeutic drug monitoring ability to individualize immunosuppressive drugs dosing.

Increased levels of circulating MMP3 correlate with severe rejection in face transplantation

Face transplantation is a viable treatment option for carefully selected patients with devastating injuries to the face. However, acute rejection episodes occur in more than 80% of recipients in the first postoperative year. Unfortunately, neither a correlation between histological grades of rejection and anti-rejection treatment nor systemic surrogate markers of rejection in face transplantation are established in clinical routine. Therefore, we utilized next generation aptamer-based SOMAscan proteomics platform for non-invasive rejection biomarker discovery. Longitudinal serum samples from face transplant recipients with long-term follow-up were included in this study. From the 1,310 proteins analyzed by SOMAscan, a 5-protein signature (MMP3, ACY1, IL1R2, SERPINA4, CPB2) was able to discriminate severe rejection from both no-rejection and nonsevere rejection samples. Technical validation on ELISA platform showed high correlation with the SOMAscan data for the MMP3 protein (rs = 0.99). Additionally, MMP3 levels were significantly increased during severe rejection as compared to no-rejection (p = 0.0009) and nonsevere rejection (p = 0.0173) episodes. Pathway analyses revealed significant activation of the metallopeptidase activity during severe face transplant rejection. This pilot study demonstrates the feasibility of SOMAscan to identify non-invasive candidate biomarkers of rejection in face transplantation. Further validation in a larger independent patient cohort is needed.

Effector memory CD4+ T-cells and dendritic cells are noninvasive biomarkers of late cellular rejection after kidney transplantation

Introduction. Diagnosis of the kidney transplant cellular rejection in the long-term after transplantation remains a challenge. Usual surrogate markers are not enough sensitive and specific. Rejection is an immune reaction to donor alloantigens. The kidney transplant biopsy to diagnose a dysfunction is an invasive procedure with the incidence of complications about 12.6% and can lead to transplant loss. In this regard, the search of immunological biomarkers for early noninvasive and accurate diagnosis of kidney transplant rejection is an actual task.Material and methods. This is a report of the observational retrospective single-center, comparative case-control study in two groups involving 44 patients who underwent kidney transplantation. The first group (REJ) included the patients with the chronic graft dysfunction caused by a biopsy-confirmed late cellular rejection (22 patients). The second group (STA) included the recipients who had no dysfunction in the posttransplant period (22 patients). Flow cytometry of peripheral blood cells was performed to identify immunophenotyping markers of late cellular rejection after kidney transplantation (we determined subpopulations of T, B lymphocytes, and dendritic cells).Results. As a result of our work, we found significant differences in the absolute count of effector memory T cells making 0.147 (0.115–0.260) × 109 cell/L in REJ group, and 0.106 (0.067–0.136) × 109 cell/L in STA group (р = 0.0167). Relative and absolute counts of myeloid dendritic cells were also different between the groups: 0.65 (0.36–0.73) vs. 1.05 (0.67–1.4) % and 0.039 (0.028–0.056) vs. 0.063 (0.049–0.076) × 109 cell/L, respectively (р = 0.0009, р = 0.003). The numbers of plasmacytoid dendritic cells were also different between the study groups: 0.0038 (0.0021–0.0054) vs. 0.005 (0.0035–0.007) × 109 cell/L for an absolute count (р = 0.0414), and 0.055 (0.04–0.085) vs. 0.09 (0.05–0.12) % for a relative count (р = 0.0197).Conclusion. The obtained data showed that the blood level of dendritic cells, which are the main “professional” initiators of immune reaction, and the level of effector helper T memory cells, which constitute the main lymphocyte subpopulation posing a destructive impact on the kidney transplant, can be considered as diagnostic markers of kidney transplant cellular rejection in the long-term after surgery.

Elevated C3a level in aqueous humor is the most reliable predictive marker of the graft rejection in corneal xenotransplantation

Elevated C3a level in aqueous humor is the most reliable predictive marker of the graft rejection in corneal xenotransplantation

Selective Deletion of Heparan Sulfotransferase Enzyme, Ndst1, in Donor Endothelial and Myeloid Precursor Cells Significantly Decreases Acute Allograft Rejection

Early damage to transplanted organs initiates excess inflammation that can cause ongoing injury, a leading cause for late graft loss. The endothelial glycocalyx modulates immune reactions and chemokine-mediated haptotaxis, potentially driving graft loss. In prior work, conditional deficiency of the glycocalyx-modifying enzyme N-deacetylase-N-sulfotransferase-1 (Ndst1f/f TekCre+) reduced aortic allograft inflammation. Here we investigated modification of heparan sulfate (HS) and chemokine interactions in whole-organ renal allografts. Conditional donor allograft Ndst1 deficiency (Ndst1−/−; C57Bl/6 background) was compared to systemic treatment with M-T7, a broad-spectrum chemokine-glycosaminoglycan (GAG) inhibitor. Early rejection was significantly reduced in Ndst1−/− kidneys engrafted into wildtype BALB/c mice (Ndst1+/+) and comparable to M-T7 treatment in C57Bl/6 allografts (P < 0.0081). M-T7 lost activity in Ndst1−/− allografts, while M-T7 point mutants with modified GAG-chemokine binding displayed a range of anti-rejection activity. CD3+ T cells (P < 0.0001), HS (P < 0.005) and CXC chemokine staining (P < 0.012), gene expression in NFκB and JAK/STAT pathways, and HS and CS disaccharide content were significantly altered with reduced rejection. Transplant of donor allografts with conditional Ndst1 deficiency exhibit significantly reduced acute rejection, comparable to systemic chemokine-GAG inhibition. Modified disaccharides in engrafted organs correlate with reduced rejection. Altered disaccharides in engrafted organs provide markers for rejection with potential to guide new therapeutic approaches in allograft rejection.

Correlation between Invasive Hemodynamic Measurements and Echocardiographic Markers of LV Diastolic Function in Heart Transplant Recipients

Introduction Left ventricular diastolic dysfunction in orthotopic heart transplant patients has been thought to be a marker of rejection and predictive of increased mortality. However, standard echocardiographic measurements of diastolic function have not been validated in this population. Factors unique to transplant recipients including larger atria, younger donor hearts, and electrical dissociation between donor and recipient hearts may affect echo measurements. We evaluated the correlation between echo parameters and left-ventricular end diastolic pressure in transplant recipients. Hypothesis At one year post-transplant, standard echocardiographic markers of diastolic function do not correlate well with invasive hemodynamic measurements of diastolic function given the unique characteristics of cardiac transplantation. Methods A series of 115 consecutive heart transplant recipients at our institution between January 2010 and December 2015 who were assessed at the time of their one-year post-transplant follow-up visit with same-day echocardiograms and cardiac catheterizations were studied. Left ventricular end-diastolic pressure (LVEDP) was measured at cardiac catheterization. Echocardiographic parameters of diastolic function included E/A, septal E/e', lateral E/e', average E/e', and left atrial volume. Chart abstraction was performed for clinical data evaluation. Linear association among catheterization and echo parameters was determined by the Pearson product moment correlation coefficient. Results None of the standard echocardiographic parameters of diastolic function were well correlated with LVEDP ( Table 1 ). Only E/A had a statistically significant correlation with LVEDP, with a correlation coefficient of 0.27 (p = 0.007). Septal, lateral, and average E/e’ were predictors of death and re-hospitalization for rejection (p ≤ 0.001 for all). Conclusions Standard echocardiographic markers of diastolic function were not well-correlated with LVEDP, suggesting they are not reliable surrogates of LVEDP. However, elevated E/e’ ratio was associated with subsequent death and hospitalization for allograft rejection, implying that early mitral filling and tissue velocity may describe important allograft functional properties independent of resting LVEDP.

PRO – AND ANTI-INFLAMMATORY CYTOKINES ARE EARLY MARKERS OF ACUTE REJECTION OF THE TRANSPLANTED KIDNEY

Aim. Determination of diagnostic significance of pro − and anti-inflammatory cytokines in early prognosis of posttransplant renal failure in patients with chronic renal disease.Materials and methods. In the peripheral blood of patients with chronic kidney disease 6 hours before kidney transplantation, multiplex analysis using Simplex ProcartaPlex panel (Bioscience, USA) and xMAP technology (principle of flow cytometry) was used to estimate the content of 10 cytokines: 7 proinflammatory (IL-1b, IL-6, IL-12p70, IL-27, IL-17A, IL-18) and 3 antiinflammatory (IL-1RA, IL-4, IL-13). Identification of HLA-antibodies was carried out with the help of multiplex immunological analysis, using test systems (Gen-Prob, USA), flow analyzer Luminex 200 xMAP technology (bimolecular reactions on the surface of microspheres).Results. The limited diagnostic significance of HLA-antibodies is due to the fact that their detection in the posttransplantation period can be either in the development of acute graft rejection, or in the favorable course of the period after the operation. Meanwhile, the determination of a number of blood cytokines before kidney transplantation allows predicting post-transplantation rejection. In particular, certain criteria favorable course of the period after kidney transplantation by absence of HLA antibodies in patients with chronic renal failure can be considered as the initial (within 6 hours of transplantation) low levels of IL1β, IL6, IL17а. Prognostically the increase in the blood levels of proinflammatory cytokines − IL6, IL17a and anti-inflammatory IL1-RA is a significant marker of acute rejection of a transplanted kidney. Along with this, it is important to note that the appearance of HLA antibodies in patients with a favorable course of the post-transplantation period is associated with an initially elevated level of proinflammatory cytokines such as Il1ß and IL6.Conclusion. Diagnostic value of the evaluated cytokines at the pre-transplant kidney patients determines the feasibility of the inclusion of evaluation of the serum concentration of IL1β, IL6, IL17a, IL1-RA in the programme of pre-transplant laboratory tests .

The role of FDG-PET in detecting rejection after liver transplantation

BackgroundThe activation and increased metabolic activity of T cells in acute cellular rejection could allow fluoro-2-deoxyglucose positron emission tomography to be utilized for detection of acute cellular rejection. The objective of this study was to evaluate the effectiveness of fluoro-2-deoxyglucose positron emission tomography in detecting acute cellular rejection in the clinical setting. MethodsFluoro-2-deoxyglucose positron emission tomography studies were performed on 88 orthotopic liver transplant patients at 7 and 17 days postoperatively (first positron emission tomography and second positron emission tomography, respectively). Additional studies were performed if patients had suspicion of rejection and at resolution of rejection (third positron emission tomography and fourth positron emission tomography, respectively). A circular region of interest was placed over the liver for semiquantitative evaluation of fluoro-2-deoxyglucose positron emission tomography images by means of standard uptake values. ResultsEighteen of 88 patients in our study (20.5%) had histologically proven acute cellular rejection during a 16 ± 11 day follow-up. There was no significant difference between the standard uptake values of first positron emission tomography among non-rejecters versus rejecters (2.05 ±0.46 non-rejecters versus 1.82 ± 0.40 rejecters, P = .127). Within the rejection cohort, the standard uptake values from the third positron emission tomography (rejection) were higher compared to the first positron emission tomography (baseline) (2.41 ± 0.48 third positron emission tomography versus 1.82 ± 0.41 first positron emission tomography, P < .001). ConclusionIncreased signal on fluoro-2-deoxyglucose positron emission tomography over baseline is associated with acute cellular rejection in liver transplant recipients. Additional prospective validation studies are essential to define the role of fluoro-2-deoxyglucose positron emission tomography scan as an early marker for acute cellular rejection.

Effect of Cold-Preservation on the Vessels of Vascularized Composite Allografts in Rats

Introduction We evaluated the effect of cold ischemia on the development of chronic rejection in vascularized composite allotransplantation. Methods Thirty rat hindlimbs were transplanted from LBN to WL rats and divided into two experimental groups: immediate transplantation and transplantation after 7 hours of cold preservation ischemia. The animals received daily low-dose immunosuppression with cyclosporine A for 2 months. Intimal proliferation, arterial permeability rate, leukocyte infiltration and tissue fibrosis were assessed. To elucidate the mechanism of rejection immunohistochemical and RT-PCR cytokine analysis (Roche LightCycler) were performed. Results Significant differences were found in the intimal proliferation and arterial permeability rate between the two groups (p = 0.004). The arterial permeability rate worsened in the most distal and small vessels (p = 0.047). The numbers of CD3+, CD8+, CD20+, and CD68+ were also statistically higher in the cold ischemia group (p < 0.05, all levels). A trend toward significance was observed with C4d deposition (p = 0.059). No differences were found in the RNA of cytokines. Conclusions An association between cold ischemia and chronic rejection was observed in rat hindlimb allografts in a sub-optimal immunosuppression setting. Chronic rejection intensity and distal progression were significantly related with cold ischemia. The leukocyte infiltrates were a rejection marker, but their exact implication in monitoring and their relation with cold ischemia are yet to be clarified. This work was granted by the Fundación Investigación IdiPaz, Madrid.J.B. presented this work and won the Residents Award of the Spanish Society of Plastic & Reconstructive Surgery, during the National Meeting held in Tenerife (Spain) in 2014.

Dendritic Cells as a Non-Invasive Immunological Biomarker of the Late Cellular Rejection after Kidney Transplantation

The Aim of the Study was to determine the association between peripheral blood leukocytes subpopulations and late immunological kidney transplant dysfunction. Materials and Methods This is a report of retrospective single center case-control study involving 44 patients who underwent kidney transplantation. Inclusion criteria: adult kidney transplant recipients who received first kidney transplant with a follow-up 4 and more years after KT. Exclusion criteria were infectious and oncological complications in the postoperative period, high immunological risks at the time of surgery (HLA mismatching, PRA>15%). First group (REJ) include 22 patients with chronic graft dysfunction, caused by biopsy proven late cellular rejection. Second group (STA) contained of 22 recipients, who haven’t any dysfunction in posttransplant period. Flow cytometry of peripheral blood cells was performed for establishment of immunophenotyping markers of late cellular rejection after kidney transplantation. Results The level of dendritic cells (DC) subsets significantly differed in study groups. Myeloid DCs were significantly lower in participants (REJ) with late cellular rejection than in stable recipients (STA): 0,65% (0,36-0,73) vs 1,05% (0,67-1,4), respectively (p<0,01). Absolute counts of cells were 0,039 (0,028-0,056) x109/l vs 0,063 (0,049-0,076) x109/l respectively for REJ and STA groups (p=0,003). Plasmacytoid DCs were significantly lower in participants with late cellular rejection too: 0,055% (0,04-0,085) vs 0,09% (0,05-0,12), REJ vs STA respectively (p=0,02). Absolute counts of pDC were 0,0038 (0,0021-0,0054) x109/l vs 0,005 (0,0035-0,007) x109/l in REJ and STA groups (p=0,041) Diagnostic value of each marker was assessed by receiver operating characteristic analysis. Optimal cut-off of myeloid DC, which can accurately discriminate late cellular rejection, was assessed as 0.87%, with area under ROC curve (AUC) 0.784, sensitivity 86.36%, and specificity 63.64%. For plasmacytoid DC optimal cut-off was 0.085%, with AUC 0.7, sensitivity 77.72%, and specificity 59%. Odds ratio of DC subsets frequency mitigation at time of late kidney rejection was 11.08 (p<0.01) for myeloid DC, and 4.91 (p=0.017) for plasmacytoid DC respectively. Conclusion The results of the study showed that the level of circulating myeloid and plasmacytoid dendritic cells can be effective non-invasive immunological marker of late cellular rejection after kidney transplantation.

Lymphocytes B Development After Kidney Transplantation

Introduction Alloantibodies are commonly used in kidney transplantation for donor/recipient immune matching as well as a marker of organ rejection after transplantation. In a broader context, anti-HLA antibodies development is associated with humoral response and B cells differentiation to long-lived memory cells. Recently studies on operational tolerance pointed to characteristic B cell phenotype and molecular signature. Thus, immune markers of anti-HLA antibodies development could point to early humoral immunity activation and may be beneficial for personalized risk stratification. Materials and Methods Low-risk kidney transplant recipients (n=53) were followed-up to 24 months after transplantation for alloantibodies development and organ rejection. Every three months anti-HLA antibodies, lymphocytes B phenotype, and cytokines were assessed. Memory B lymphocytes phenotype was recognized as CD27+ cells, both IgM positive and negative. In contrary naïve B cells were identified as CD27- and CD5+ B lymphocytes. Th1/Th2 and BAFF serum levels were measured with Luminex solid phase assay. Results and Discussion For the 34 recipients, alloantibodies were not present after transplantation, while for 19 others DSA antibodies were confirmed. The higher rate was observed 20 months after transplantation. Anti-HLA antibodies development was proceeded by increased in BAFF and INFgamma levels, as well as memory B lymphocytes numbers. It was found, that alloantibodies development was correlated with a number of memory B cells, Rs=0,96 (Spearman rank correlation). Conclusion Lymphocytes B phenotype monitoring after kidney transplantation is useful for alloantibodies development and may serve as an additional marker of humoral immunity activation. These could be also beneficial for individual risk stratification and tailored immunosuppression protocol development after kidney transplantation. National Centre for Research and Development, Poland (No. STRATEGMED1/233368/1/NCBR/2014). National Science Centre, Poland (No. NN402420738 and NN402 562440).

Leptin – New Marker for Rejection of Kidney Transplant?

Leptin – New Marker for Rejection of Kidney Transplant?

Polymorphisms in IMPDH2, UGT2B7, and CES2 genes influence the risk of graft rejection in kidney transplant recipients taking mycophenolate mofetil

The immunosuppressant mycophenolic acid (MPA), derived from the prodrug mycophenolate mofetil (MMF), is a drug used widely by kidney transplant recipients. This drug selectively inhibits inosine monophosphate dehydrogenase that controls the proliferation of lymphocytes, aiding in the prevention of rejection of the transplanted organ. Polymorphisms in key genes involved in MMF metabolism may alter the function of the enzymes encoded by them and contribute to interindividual variability in the response to the drug and its efficacy. The aim of this study was to investigate the association of nine polymorphic variants of eight genes involved in MMF pharmacokinetics, with rejection and adverse effects exhibited by kidney transplant recipients who use this drug. Our sample comprised 145 kidney transplant recipients undergoing post-transplant treatment whose immunosuppressive therapy consisted of MMF and corticosteroid combined or not with a calcineurin inhibitor or mTOR inhibitor. The average age of the patients was 46.9 ± 12.5 years, and they underwent transplantation 7 ± 5.71 years ago. The combination of the T/C and C/C genotypes of the polymorphism rs11706052 (IMPDH2) was associated with a 4.2-fold protection, and the combination of the genotypes A/G and G/G of the polymorphism rs7438135 (UGT2B7) showed a 2.4-fold protection, against rejection. The association of T/C and C/C genotypes in the SNP rs11706052 (IMPDH2) with the occurrence of rejection episodes considering only patients who received immunosuppressive drug MMF associated with cyclosporine or tacrolimus and corticoids, demonstrated association with a protection against rejection 15.6-fold. The T/T genotype of the polymorphism rs2241409 (CES2) was associated with a 7.2-fold increased risk of rejection. Therefore, these polymorphisms that showed a strong association with rejection episodes should be considered in future studies on new prognostic markers for rejection in patients treated with MMF.