Novel Metabolites Are Associated With Thrombosis After Cardiac Surgery in Children With Congenital Heart Disease.

BACKGROUND: Thrombosis is a major complication after cardiac surgery in children with congenital heart disease. The mechanisms underlying thrombosis development remain poorly understood. We aimed to identify novel circulating metabolites before cardiac surgery that are associated with thrombosis after surgery in children with congenital heart disease. METHODS: In this prospective cohort study, all blood samples were drawn right before surgical incision and after the induction of anesthesia, and plasma was separated immediately under 4 °C. Untargeted metabolomic data were measured by Metabolon in plasma from children (age range, 0 days-18 years) with congenital heart disease undergoing cardiac surgery. The primary outcome was thrombosis within 30 days of surgery or before discharge. Associations of individual metabolites with thrombosis were assessed with logistic regression with false discovery rate correction for multiple comparison and adjustment for clinical characteristics; elastic net regression was used to select a prediction model. RESULTS: Out of 1115 metabolites measured in samples from 203 children, 776 met the quality control criteria. In total, 25 children (12.3%) developed thrombosis. Among the 776 metabolites, 175 were significantly associated with thrombosis (false discovery rate Q<0.05). The top 3 metabolites showing the strongest associations with thrombosis were eicosapentaenoate, stearidonate, and andro steroid monosulfate C19H28O6S (false discovery rate, 0.01 for all). Pathway analysis showed that the pathways of nicotinate and nicotinamide metabolism and glycerophospholipid metabolism were enriched (false discovery rate, 0.003 for both) and had significant impact on the development of thrombosis. In elastic net regression analysis, the area under the receiver operating-characteristic curve of a prediction model for thrombosis was 0.969 in the training sample (70% of the total sample) and 0.833 in the testing sample (the remaining 30%). CONCLUSIONS: We have identified promising novel metabolites and metabolic pathways associated with thrombosis. Future studies are warranted to confirm these findings and examine the mechanistic pathways to thrombosis.

The Effect of Infectious Complications During Ventricular Assist Device Use on Outcomes of Pediatric Heart Transplantation.

To describe the impact of infectious adverse events (IAEs) during ventricular assist device (VAD) support on graft loss, infection, and rejection after pediatric heart transplant (HT). Pedimacs data were linked to Pediatric Heart Transplant Society (PHTS) data for patients receiving a VAD followed by HT between September 2012 and December 2016. Linked patients were categorized into IAE on VAD (group A) and no IAE on VAD (group B). Infectious adverse event locations included nondevice, device (external or internal), and sepsis. Post-HT outcomes for analysis were graft loss, infection, and rejection. Time-dependent analysis included Kaplan-Meier and multiphase parametric hazard function analysis. We linked 207 patients (age 9.4 ± 6.3 years). Post-HT follow-up was 19.4 patient-months (<8 days-4.1 years). Group A included 42 patients (20%) with 62 IAEs. Group B included 165 patients without an IAE. Group A patients were younger (7.4 ± 6.1 vs. 9.5 ± 6.3 years; p = 0.03), waited longer for HT (5.3 ± 4.1 vs. 2.9 ± 2.5 months; p = 0.0005), and were hospitalized longer post-HT (42 ± 59 vs. 23 ± 22 days; p = 0.05). VAD-related IAEs were rare (N = 11). Groups A and B had similar freedom from first post-HT infection, rejection, and graft loss (all p > 0.1). However, patients with VAD-related IAE were somewhat more likely to experience rejection (p = 0.03) and graft loss (p = 0.01). Children with an IAE on VAD who survive to HT are younger, wait longer for HT, and remain hospitalized longer than those without an IAE on VAD. Overall, IAE on VAD did not impact post-HT outcomes, but VAD-related IAE may be associated with graft loss and rejection.

Resource utilization in children with paracorporeal continuous-flow ventricular assist devices.

BACKGROUND: Paracorporeal continuous-flow ventricular assist devices (PCF VAD) are increasingly used in pediatrics, yet PCF VAD resource utilization has not been reported to date. METHODS: Pediatric Interagency Registry for Mechanically Assisted Circulatory Support (PediMACS), a national registry of VADs in children, and Pediatric Health Information System (PHIS), an administrative database of children's hospitals, were merged to assess VAD implants from 19 centers between 2012 and 2016. Resource utilization, including hospital and intensive care unit length of stay (LOS), and costs are analyzed for PCF VAD, durable VAD (DVAD), and combined PCF-DVAD support. RESULTS: Of 177 children (20% PCF VAD, 14% PCF-DVAD, 66% DVAD), those with PCF VAD or PCF-DVAD are younger (median age 4 [IQR 0-10] years and 3 [IQR 0-9] years, respectively) and more often have congenital heart disease (44%; 28%, respectively) compared to DVAD (11 [IQR 3-17] years; 14% CHD); p < 0.01 for both. Median post-VAD LOS is prolonged ranging from 43 (IQR 15-82) days in PCF VAD to 72 (IQR 55-107) days in PCF-DVAD, with significant hospitalization costs (PCF VAD $450,000 [IQR $210,000-$780,000]; PCF-DVAD $770,000 [IQR $510,000-$1,000,000]). After adjusting for patient-level factors, greater post-VAD hospital costs are associated with LOS, ECMO pre-VAD, greater chronic complex conditions, and major adverse events (p < 0.05 for all). VAD strategy and underlying cardiac disease are not associated with LOS or overall costs, although PCF VAD is associated with higher daily-level costs driven by increased pharmacy, laboratory, imaging, and clinical services costs. CONCLUSION: Pediatric PCF VAD resource utilization is staggeringly high with costs primarily driven by pre-implantation patient illness, hospital LOS, and clinical care costs.

Less Is More in Post Pediatric Heart Transplant Care.

BACKGROUND: Historically, steroids and endomyocardial biopsies have, respectively, been part of standard immunosuppression for preventing cardiac transplant rejection and monitoring for rejection. However, these treatments come with numerous adverse effects. Some transplant programs have questioned whether the risks and costs outweigh the benefits or whether they may interfere with patient outcomes. METHODS: Pediatric cardiac transplantations over 15 years (n = 49) were examined in a single-center retrospective study. Two groups of patients were formed: group 1 received induction steroids and underwent routine protocol biopsy (n = 18), and group 2 neither received steroids nor underwent routine biopsy (n = 13). RESULTS: The 1-year survival rate was similar between the two approaches: group 1 survival was 94% and group 2 survival was 92%. However, differences between the two groups were observed for comorbidities. Group 1 had 11 patients that exhibited rejection, and group 2 had only 1 patient (p = 0.003). Group 2 had fewer cases of posttransplant hypertension (p = 0.001) and insulin dependence (p = 0.02). CONCLUSIONS: This study suggests a less-invasive posttransplant approach that avoids biopsies and steroids was safely implemented in this single center. Both groups had similar survival. However, group 2 had statistically significant less posttransplant rejection, hypertension, and diabetes. Overall, this study shows no increased risk associated with steroid and biopsy avoidance in posttransplant patients, but with some clear benefits.

Resource Utilization in Pediatric Patients Supported With Ventricular Assist Devices in the United States: A Multicenter Study From the Pediatric Interagency Registry for Mechanically Assisted Circulatory Support and the Pediatric Health Information System.

BACKGROUND: Few data exist on resource utilization with pediatric ventricular assist devices (VADs). We tested the hypothesis that device type and adverse events are associated with increased resource utilization in pediatric patients supported with VADs. METHODS AND RESULTS: The Pediatric Interagency Registry for Mechanically Assisted Circulatory Support, a national registry of VADs in patients <19 years old, and the Pediatric Health Information System, an administrative database, were merged. Univariate analysis was performed assessing the association of all factors with the total cost and length of stay first. Significant variables (P<0.05) were subjected to multivariable analysis. The study included 142 patients from 19 centers with VAD implants from October 2012 to June 2016. The median age was 9 years (interquartile range [IQR] 2-15), 84 (59%) supported with a continuous-flow VAD. Overall median hospital costs were $750 000 (IQR $539 000 to $1 100 000) with a median hospital length of stay of 81 days (IQR 54-128). On multivariable analysis, device type and postoperative complications were not associated with resource utilization. Factors associated with increased costs included patient age, lower-volume VAD center, being intubated, being on extracorporeal membrane oxygenation, number of complex chronic medical conditions, and length of stay. Among continuous-flow VAD patients, discharge to home before transplant versus remaining hospitalized was associated with lower hospital costs (median $600 000 [IQR $400 000 to $820 000] versus median $680 000 [IQR $500 000 to $970 000], P=0.03). CONCLUSION: VADs in pediatric patients are associated with high resource utilization. Increased resource utilization was associated with lower-volume VAD centers, disease severity at VAD implantation, and the presence of complex chronic medical conditions. Further study is needed to develop cost-effective strategies in this complex population.

Development of a Pediatric Cardiac Mechanical Support Program.

The development of a pediatric cardiac support program is a complex, multidisciplinary project. This study describes the University of Iowa Congenital Heart Program's experience from its inception to the present. In, we examine those specific factors that have led to substantial improvements in the program, additionally identifying where further gains can be made. We retrospectively reviewed all pediatric patients who received mechanical cardiac support at the University of Iowa from the inception of the program in 1991. In total, 29 patients received mechanical support between December 1991 and December 2015 and are included in the study. Twelve patients received continuous flow devices and 17 patients received pulsatile flow devices. Median age at implant was 12.8 years (range 0.1-18.2 years). Median weight at implant was 40.5 kg (3.2-123.4 kg). Factors examined included: operating room (OR) time, intensive care unit and hospital length of stay, intubation days, blood product usage, pre- and post-operative bilirubin, creatinine, natriuretic peptide B (NPPB), and device implanted. Categorical and continuous variables were compared using Chi-squared and Wilcoxon rank-sum tests, respectively. Of the 29 patients who received mechanical support, 17 (58.6%) were discharged home, 11 (37.9%) died during their hospitalization, and 1 (3.5%) remains hospitalized. Median length of ventricular assist device support was 59.5 days (range 1-653 days). Between December 1991 and December 2011, in-hospital mortality was 64.3%. Following this period, significant changes were made to patient management with in-hospital mortality decreasing to 13.3% between February 2013 and December 2015. Comparison between deceased and living patients revealed several significant factors including: median number of packed red blood cells transfused, 8 versus 4 units (P = 0.048), median OR time, 396 versus 299 min (P = 0.003), and device implanted. During the early stages of the mechanical support program, higher than expected mortality rates prompted changes in the management of pediatric cardiac patients, specifically, the development of a dedicated management team. These changes significantly improved outcomes and we suggest can be used as a model for similar cardiac support programs, especially in smaller volume programs.

Fontan Patient Survival After Pediatric Heart Transplantation Has Improved in the Current Era.

BACKGROUND: Historically, patients with a prior Fontan procedure for complex congenital heart disease (CHD) have been considered at higher risk for death after heart transplant (HT) compared with other HT transplant candidates. With the overall trend of improved survival of pediatric HT recipients, it is unclear of Fontan patient post-HT survival has also improved in the current era. METHODS: Data from the Pediatric Heart Transplant Study database for Fontan patients who underwent HT was compared between the early era (1993 to 2006, n = 150) and late era (2007 to 2014, n = 252). Post-HT survival and pre-HT characteristics were compared among eras and also with non-Fontan CHD patients. RESULTS: At time of HT, Fontan patients in the late era were more likely to require inotropic support, have protein-losing enteropathy, have failure to thrive, and be further from time of Fontan, although less likely to be on ventilator support. Only ventilator support and earlier year of HT were significant risk factors for death in the multivariate analysis. Post-HT Fontan patient survival significantly improved from the early to late era (p = 0.02), particularly in the early phase, with 1-year survival of 77% in the early era and 89% in the late era. Late era non-Fontan CHD patient 1-year post-HT survival was similar to Fontan patients at 92%. CONCLUSIONS: Survival of Fontan patients after HT has significantly improved in the current era. Currently, expected post-HT survival for Fontan patients is on par with other CHD patients. Fontan patients should not be excluded from consideration for HT solely on a history of Fontan.

Use of HeartWare Ventricular Assist System for systemic ventricular support of a pediatric patient after Mustard procedure.

BACKGROUND: The HeartWare Ventricular Assist System is indicated to provide mechanical circulatory support of patients with intractable heart failure as a bridge to cardiac transplantation. We describe the use of this device to support the systemic right ventricle (RV) of a pediatric patient with New York Heart Association class IIIC congestive heart failure who had undergone Mustard procedure for D-transposition of the great vessels as an infant. CASE REPORT: A HeartWare ventricular assist device was implanted in the left chest of a 16-year-old female patient (body surface area 1.43 m(2)) who presented with edema and later deteriorated, developing acute kidney injury, dysrhythmia, and pulmonary edema. RESULTS: The patient's edema and acute kidney injury resolved after device placement. She was discharged home and successfully underwent device removal with heart transplant five months later. CONCLUSION: The HeartWare device may be used for extended support as a systemic RV in a pediatric patient. It is feasible to consider using the device in this patient population.

Early predictors of survival to and after heart transplantation in children with dilated cardiomyopathy.

BACKGROUND: The importance of clinical presentation and pretransplantation course on outcome in children with dilated cardiomyopathy listed for heart transplantation is not well defined. METHODS AND RESULTS: The impact of age, duration of illness, sex, race, ventricular geometry, and diagnosis of myocarditis on outcome in 261 children with dilated cardiomyopathy enrolled in the Pediatric Cardiomyopathy Registry and Pediatric Heart Transplant Study was studied. End points included listing as United Network for Organ Sharing status 1, death while waiting, and death after transplantation. The median age at the time of diagnosis was 3.4 years, and the mean time from diagnosis to listing was 0.62±1.3 years. Risk factors associated with death while waiting were ventilator use and older age at listing in patients not mechanically ventilated (P=0.0006 and P=0.03, respectively). Shorter duration of illness (P=0.04) was associated with listing as United Network for Organ Sharing status 1. Death after transplantation was associated with myocarditis at presentation (P=0.009), nonwhite race (P<0.0001), and a lower left ventricular end-diastolic dimension z score at presentation (P=0.04). In the myocarditis group, 17% (4 of 23) died of acute rejection after transplantation. CONCLUSIONS: Mechanical ventilator use and older age at listing predicted death while waiting, whereas nonwhite race, smaller left ventricular dimension, and myocarditis were associated with death after transplantation. Although 97% of children with clinically or biopsy-diagnosed myocarditis at presentation survived to transplantation, they had significantly higher posttransplantation mortality compared with children without myocarditis, raising the possibility that preexisting viral infection or inflammation adversely affects graft survival.

Comparison of risk factors and outcomes for pediatric patients listed for heart transplantation after bidirectional Glenn and after Fontan: an analysis from the Pediatric Heart Transplant Study.

BACKGROUND: Patients listed for transplant after the bidirectional Glenn (BDG) may have better outcomes than patients listed after Fontan. This study examined and compared outcomes after listing for BDG and Fontan patients. METHODS: All patients listed for transplant after the BDG in the Pediatric Heart Transplant Study between January 1993 and December 2008 were evaluated. Comparisons were made with Fontan patients and with a matched cohort of congenital heart disease patients. Competing outcomes analysis and actuarial survival were evaluated for the study populations, including an examination of various risk factors. RESULTS: Competing outcomes analysis for BDG and Fontan patients after listing were similar. There was no difference in actuarial survival after listing or transplant among the 3 cohorts. Mechanical ventilation, United Network of Organ Sharing status, and age were risk factors for death after listing in BDG and Fontan patients, but ventilation at the time of transplant was significant only for the Fontan patients. Mortality was increased in Fontan patients listed < 6 months after surgery compared with patients listed > 6 months after surgery, but no difference was observed in BDG patients. There was a trend toward improved survival after listing for both populations across 3 eras of the study, but this did not reach statistical significance. CONCLUSION: Outcomes after listing for BDG and Fontan patients are similar. Mechanical ventilation at the time of transplant remains a significant risk factor for death in the Fontan population, as does listing for transplant soon after the Fontan, suggesting that some patients may benefit from transplant instead of Fontan completion.

Outcomes of Pneumocystis jiroveci pneumonia infections in pediatric heart transplant recipients.

PJP is known to cause significant morbidity and rarely death in immunosuppressed patients. The prevalence and outcomes of PJP in pediatric solid-organ transplant patients are not well established. This study utilizes data from the PHTS to establish the prevalence and outcome of PJP in pediatric heart transplant recipients. We conducted a retrospective cohort study using data from the PHTS, including data from 24 institutions between January 1, 1993, and December 31, 2004. Infections that occur in PHTS subjects are recorded in a standardized data collection form. The prevalence and outcomes of PJP in pediatric heart transplant recipients were determined. There were a total of 18 patients (1%) with PJP out of the 1854 pediatric heart transplant recipients in the PHTS database. A majority of PJP occurred two months to two yr post-transplant, and patients with PJP had a significantly decreased mortality compared with other fungal infections. PJP is an infrequent complication experienced by pediatric heart transplant recipients. Patients that have experienced PJP have an increased survival compared to patients with other fungal infections, and most PJP occurred within two yr of transplant.

Allosensitization and outcomes in pediatric heart transplantation.

BACKGROUND: Allosensitization among children being considered for heart transplantation remains a great challenge. Controversy exists as to the best approach for those with elevated panel-reactive antibody (PRA) titers. We sought to define the association between elevated PRA and outcomes using data from the multi-institutional Pediatric Heart Transplant Study Group. METHODS: Between January 1993 and December 2008, 3,016 patients (>1 month of age) were listed for heart transplantation. PRA data at listing were available for 2,500 (83%) patients, and 2,237 underwent transplantation with PRA data being available for 1,904 (85%). Because various PRA assays were employed (e.g., cell-based and solid phase) we entered the highest value regardless of methodology. RESULTS: Among the factors associated with high PRA at transplant were Status 1 at listing, previous sternotomy and prior Norwood procedure. An elevated PRA at listing was associated with higher risk of death while waiting. Of subjects with PRA ≥ 50% only 57% were transplanted by 1 year on the waitlist, as compared with 76% of those with PRA <10%. Waitlist mortality for the highly allosensitized subjects (≥ PRA 50%) was 19% by 12 months. Survival at 1 year after transplantation was significantly lower in those with PRA ≥ 50% versus those with PRA <10% (73% vs 90%, respectively, p < 0.0001). Those with elevated PRA who had a negative prospective crossmatch had no difference in survival compared with those without allosensitization. There was no significant association between PRA levels and time to first rejection or development of coronary allograft vasculopathy. CONCLUSIONS: Significant allosensitization is associated with more than a 2-fold increased risk of death within the first transplant year. Although prospective crossmatching abrogates the risk of post-transplant mortality, it may contribute to higher pre-transplant attrition due to longer waitlist times. There is a critical need for strategies to minimize the impact of allosensitization and antibody-mediated rejection immediately after transplantation.

Outcomes after listing for primary transplantation for infants with unoperated-on non-hypoplastic left heart syndrome congenital heart disease: a multi-institutional study.

BACKGROUND: Although heart transplantation has been used as the primary therapy for congenital heart lesions in infants other than hypoplastic left heart syndrome (HLHS), the outcomes in this group of patients have not been determined. METHODS: We used the Pediatric Heart Transplant Study database (1993 to 2006) to compare outcomes of 388 infants aged < 6 months listed for HLHS, 161 with other congenital heart diseases (non-HLHS), and 145 with cardiomyopathy in early (1993 to 1999) and recent (2000 to 2006) eras. RESULTS: The cardiomyopathy group was significantly (p < 0.001) different from the HLHS and non-HLHS groups at listing: more girls, older age, and a greater need for high-dose inotropes, mechanical ventilation, and/or mechanical circulatory support. Survival after listing was similar among the groups in the early era. Although outcomes after listing in HLHS and cardiomyopathy patients improved in the recent era, outcomes in non-HLHS patients did not. Survival at 1 and 5 years after listing was significantly worse (p < 0.001) for non-HLHS patients (51%, 48%) vs HLHS (71%, 61%), with age- and sex-adjusted hazard ratio (HR) of 1.79 (95% confidence interval, 1.15-2.77, p = 0.009) and CM (80%, 74%; HR, 2.72; 95% confidence interval, 1.59-4.67, p < 0.001) in the recent era. Post-transplant survival in both eras was not significantly different among the groups. CONCLUSION: Use of heart transplantation as primary therapy for non-HLHS infants has not improved over time and currently is associated with significantly poorer results vs HLHS and cardiomyopathy due to a higher risk for death before transplant.

Orthotopic heart transplantation in a child with hereditary spherocytosis.

Hereditary spherocytosis (HS) is a genetic, frequently familial hemolytic blood disease that presents with varying degrees of hemolytic anemia, splenomegaly, and jaundice.The disease arises as a result of defects in any of a number of proteins responsible for maintaining the shape and flexibility of the red blood cell, resulting in an osmotically fragile and characteristically spherical red blood cell. Theoretically, cardiopulmonary bypass can exacerbate hemolysis and subsequent renal dysfunction.There are few reports of open heart surgery for patients with HS and none for orthotopic heart transplantation.We report a 6-year-old boy with HS who underwent orthotopic heart transplantation.

Histone deacetylase inhibitor uses p21(Cip1) to maintain anergy in CD4+ T cells.

T cell anergy defined as antigen-specific proliferative unresponsiveness was induced in CD4+ T cells exposed to antigen (Ag) in the presence of the histone deacetylase (HDAC) inhibitors n-butyrate, trichostatin A or scriptaid. However, the ability of HDAC inhibitors to induce anergy in Th1 cells was not due to general histone hyperacetylation. Instead, the anergy induced by HDAC inhibitors was associated with upregulation of p21(Cip1), a secondary effect of histone acetylation. Induction of p21(Cip1) in the absence of histone hyperacetylation by exposure to okadaic acid also resulted in T cell anergy. In addition, Ag-specific p21(Cip1)-deficient CD4+ T cells were much less susceptible to anergy induction by n-butyrate. Thus, p21(Cip1) appears to mediate the proliferative unresponsiveness found in CD4+ T cell anergized by exposure to Ag in the presence of HDAC inhibitors.

Takotsubo-like cardiomyopathy in a 17-year-old male with a pheochromocytoma.

We report a case of a 17-year-old adolescent male with Takotsubo-like cardiomyopathy in the setting of pheochromocytoma who presented with hematemesis, hypertension, and pallor. Takotsubo-like cardiomyopathy is rarely reported in the pediatric population, and this is the first report in the pediatric literature of Takotsubo-like cardiomyopathy associated with both pheochromocytoma and an elongated course of the left anterior descending coronary artery.

Cardiac transplant outcomes in pediatric patients with pre-formed anti-human leukocyte antigen antibodies and/or positive retrospective crossmatch.

BACKGROUND: Children undergoing heart transplantation who have preformed anti-human leucocyte antigen (HLA) panel reactive antibodies (PRA) or positive retrospective crossmatch (XM) may be at increased risk for rejection and graft failure. We assessed outcomes of transplant recipients with either positive PRA before transplant or positive retrospective XM. METHODS: A review of 148 heart transplant patients between 1990 and 2006 was undertaken, identifying transplants in patients with pre-transplant PRA > 1% and/or a positive XM. Demographic information and detailed post-transplant outcomes including episodes of rejection, infection, and graft failure were recorded. RESULTS: There were 11 PRA positive (PRA+) transplants, 135 PRA negative (PRA-) transplants, and no PRA data on 2. There were 14 XM+ transplants, 115 XM- transplants, and no XM data on 19. Kaplan-Meier graft survival was better in XM- than XM+ patients (p < 0.015), but not different between PRA+ and PRA- Groups. Timing of first rejection and number of rejection episodes were not different between XM+ and XM- Groups or between PRA+ and PRA- Groups. Infections were not different between PRA or XM Groups. Four patients were PRA+/XM- (all PRA, 1%-10%), 7 were PRA-/XM+, and 7 were PRA+/XM+ (6 of 7 PRA >10%). CONCLUSIONS: Pediatric heart transplant patients whose retrospective XM is positive are at significantly increased risk for graft failure. Elevated pre-transplant PRA may not predict increased risk of graft failure, although markedly positive PRA (>10%) predicts a positive retrospective XM. Improved treatment for pediatric transplant patients with a positive retrospective XM is needed.

Histone deacetylase inhibitors induce antigen specific anergy in lymphocytes: a comparative study.

Induction of immune tolerance to transplanted tissue continues to be a challenge for organ transplantation. In the present study, six widely used histone deacetylase inhibitors (HDAI), sodium butyrate (n-butyrate), Trichostatin A, Oxamflatin, Scriptaid, HDAC I and HDAC III, were examined for ability to induce antigen-specific immune anergy in cloned and naïve murine CD4(+) T cells. When first compared for their ability to inhibit histone deacetylation Trichostatin A was found to be 10 times more potent than HDAC III, Oxamflatin and Scriptaid and 10(4) times more potent than n-butyrate. When we compared ability to inhibit CD4(+) T cell proliferation in response to IL-2 stimulation, Trichostatin A was the most potent with 100% inhibition using 100 nM Trichostatin A, while 1 muM of HDAC III, Oxamflatin and Scriptaid and 1 mM of n-butyrate were required for this effect. When the tolerogenic activity of Trichostatin A, Scriptaid and n-butyrate were compared using cloned Th1 cells specific for keyhole limpet hemocyanin (KLH), all three HDAI were effective, but Trichostatin A was again the most potent. Finally, Trichostatin A (0.05 mM) was shown to induce anergy in OT-II ovalbumin-specific naïve CD4(+) T-cells. We concluded that Trichostatin A was the most potent HDAI with regard to inhibition of histone deacetylation and the ability to induce antigen-specific anergy in both cloned and naïve CD4(+) T cells. These results will guide future studies examining HDAIs for ability to induce clinical tolerance in organ transplantation.

Kinetic studies on the interactions of heparin and complement proteins using surface plasmon resonance.

Heparin is a naturally occurring polysaccharide known to interact with complement proteins and regulate multiple steps in the complement cascade. Quantitative information, in the form of affinity constants for heparin-complement interactions, is not generally available and there are no reports of a comprehensive analysis using the same interaction method. Such information should improve our understanding of how exogenously administered pharmaceutical heparin and the related endogenous polysaccharide, heparan sulfate, regulate complement activation. The current study provides the first comprehensively analysis of the binding of various complement proteins to heparin using surface plasmon resonance (SPR). Complement proteins C1, C2, C3, C4, C5, C6, C7, C8, C9, C1INH, factor I, factor H, factor B and factor P all bind heparin but exhibit different binding kinetics and dissociation constants (Kd) ranging from 2 to 320 nM. By taking into account these Kd values and the serum concentrations of these complement proteins, the percentage of each binding to exogenously administered heparin was calculated and found to range from 2% to 41%. This study provides essential information required for the rational design of new therapeutic agents capable of regulating the complement activation.