[Transition in kidney transplantation]

Transition from pediatric to adult care in renal transplantation has emerged as a critical step in the life of a young kidney recipient. During this phase, young patients are faced with the physiological and psychological changes associated with adolescence that can lead to non-compliance and potentially graft loss. To date, there is not a unique accepted model of transition, however it has been proved that the presence of a multidisciplinary team including specialists in adolescent management and in the transition from pediatric to adult transplant care is beneficial during this at-risk phase. The goal of this team is to ensure a progressive transition of the patients according to a precise plan and time line.

Allogeneic haematopoietic stem cell transplantation for mitochondrial neurogastrointestinal encephalomyopathy.

Haematopoietic stem cell transplantation has been proposed as treatment for mitochondrial neurogastrointestinal encephalomyopathy, a rare fatal autosomal recessive disease due to TYMP mutations that result in thymidine phosphorylase deficiency. We conducted a retrospective analysis of all known patients suffering from mitochondrial neurogastrointestinal encephalomyopathy who underwent allogeneic haematopoietic stem cell transplantation between 2005 and 2011. Twenty-four patients, 11 males and 13 females, median age 25 years (range 10-41 years) treated with haematopoietic stem cell transplantation from related (n = 9) or unrelated donors (n = 15) in 15 institutions worldwide were analysed for outcome and its associated factors. Overall, 9 of 24 patients (37.5%) were alive at last follow-up with a median follow-up of these surviving patients of 1430 days. Deaths were attributed to transplant in nine (including two after a second transplant due to graft failure), and to mitochondrial neurogastrointestinal encephalomyopathy in six patients. Thymidine phosphorylase activity rose from undetectable to normal levels (median 697 nmol/h/mg protein, range 262-1285) in all survivors. Seven patients (29%) who were engrafted and living more than 2 years after transplantation, showed improvement of body mass index, gastrointestinal manifestations, and peripheral neuropathy. Univariate statistical analysis demonstrated that survival was associated with two defined pre-transplant characteristics: human leukocyte antigen match (10/10 versus <10/10) and disease characteristics (liver disease, history of gastrointestinal pseudo-obstruction or both). Allogeneic haematopoietic stem cell transplantation can restore thymidine phosphorylase enzyme function in patients with mitochondrial neurogastrointestinal encephalomyopathy and improve clinical manifestations of mitochondrial neurogastrointestinal encephalomyopathy in the long term. Allogeneic haematopoietic stem cell transplantation should be considered for selected patients with an optimal donor.

Racial Disparities in Access to and Outcomes of Kidney Transplantation in Children, Adolescents, and Young Adults: Results From the ESPN/ERA-EDTA (European Society of Pediatric Nephrology/European Renal Association-European Dialysis and Transplant Association) Registry.

BACKGROUND Racial disparities in kidney transplantation in children have been found in the United States, but have not been studied before in Europe. STUDY DESIGN Cohort study. SETTING & PARTICIPANTS Data were derived from the ESPN/ERA-EDTA Registry, an international pediatric renal registry collecting data from 36 European countries. This analysis included 1,134 young patients (aged ≤19 years) from 8 medium- to high-income countries who initiated renal replacement therapy (RRT) in 2006 to 2012. FACTOR Racial background. OUTCOMES & MEASUREMENTS Differences between racial groups in access to kidney transplantation, transplant survival, and overall survival on RRT were examined using Cox regression analysis while adjusting for age at RRT initiation, sex, and country of residence. RESULTS 868 (76.5%) patients were white; 59 (5.2%), black; 116 (10.2%), Asian; and 91 (8.0%), from other racial groups. After a median follow-up of 2.8 (range, 0.1-3.0) years, we found that black (HR, 0.49; 95% CI, 0.34-0.72) and Asian (HR, 0.54; 95% CI, 0.41-0.71) patients were less likely to receive a kidney transplant than white patients. These disparities persisted after adjustment for primary renal disease. Transplant survival rates were similar across racial groups. Asian patients had higher overall mortality risk on RRT compared with white patients (HR, 2.50; 95% CI, 1.14-5.49). Adjustment for primary kidney disease reduced the effect of Asian background, suggesting that part of the association may be explained by differences in the underlying kidney disease between racial groups. LIMITATIONS No data for socioeconomic status, blood group, and HLA profile. CONCLUSIONS We believe this is the first study examining racial differences in access to and outcomes of kidney transplantation in a large European population. We found important differences with less favorable outcomes for black and Asian patients. Further research is required to address the barriers to optimal treatment among racial minority groups.

BIOLOGICAL AND BIOCHEMICAL CHARACTERIZATION OF MURINE-TUMOR SPECIFIC TRANSPLANTATION ANTIGENS (ISOTACHOPHORESIS, IMMUNOGENICITY)

Tumor-specific transplantation antigens (TSTA) are individually distinct neoantigens expressed on the cells of chemically-induced neoplasms. TSTA are operationally defined by immunization of syngeneic mice against challenge with viable tumor cells. Immunization with cell surface or extracted TSTA induces specific resistance to transplanted tumor cells. The biological and biochemical nature of TSTA was investigated in the 3-methylcholanthrene-induced fibrosarcomas of female C3H/HeJ mice, MCA-F and MCA-D. Tumor cell suspensions were extracted by treatment with 3M KCl or 2.5% butanol solutions and the TSTA was partially purified by preparative isoelectric focusing. The isoelectric pH of TSTA purified from 3M KCl extracts was 5.8-6.0, and from butanol extracts was 6.4-6.6. Whereas immunization with 10('5) and 10('6) irradiated tumor cells induces complete rejection of tumor cell challenge over a two-fold-log dose range, immunization with ug quantities within a one-fold-log dose range of extracted TSTA induces only partial resistance to tumor challenge. Reduced immunogenicity of extracted TSTA is hypothesized to result from immunization of mice with insufficiently purified TSTA preparations. The hypothesis predicts that immunization with highly purified TSTA, free from interfering substances, induces complete rejection of tumor challenge over a broad dose range. To test the hypothesis preparative isotachophoresis (pITP) was used to purify TSTA from electrofocused TSTA fractions. Significant purification was achieved, as immunization with 15 pg to 1.5 ug (5 logs) of pITP-purified TSTA extracted from the MCA-F, or with 1 pg to 10 ng (4 logs) of TSTA from the MCA-D tumor induced specific resistance to tumor challenge. Despite 50,000 fold purification of TSTA, immunization induced partial, not complete, rejection of transplanted tumor cells. This suggests a clear dissociation of the immunogenicity and purification of extracted TSTA, indicating that the induction of partial immunity to tumor challenge is an intrinsic property of extracted TSTA.^

AUGMENTATION OF RECONSTITUTION OF GUT ASSOCIATED LYMPHOID TISSUE IN THE SYNGENEIC MURINE BONE MARROW TRANSPLANTATION MODEL

Gut was studied as a prototypical mucosal membrane in the murine BDF-1 syngeneic bone marrow transplant model. Measures of jejunal intraepithelial lymphocytes (IELs) and crypt cells were obtained by standard techniques and a method of quantifying gut lamina propria plasma cells (PCs) was developed. The degree of ablation of gut PCs and IELs after 900 rads total body irradiation with ('60)Co, and their repopulation effected by transplantation with 2.0 x 10('5) or 1.0 x 10('6) bone marrow cells demonstrated a prolonged period of profound depression in population levels of these cells which was not reflected by the extent of damage sustained to the epithelium. Differences in the depopulation and recovery of gut PCs and IELs revealed a tendency towards initial differentiation of effector cells. A positive dose response to high bone marrow cell innocula was obtained. Subsequent studies determined that gut IEL and PC repopulation was potentiated by the addition of IELs or buffy coat cells (BCs) to the bone marrow transplant. A method of isolating 1.4 - 4.0 x 10('7) viable IELs per gram of murine small bowel was devised employing intralumenal hyaluronidase digestion of the epithelial layer and centrifugation of the resulting suspension through discontinuous Percoll gradients. Irradiated mice received 2.0 x 10('5) bone marrow cells along with an equal number of IELs or BCs. The extent and duration of depression of numbers of IELs and PCs was markedly reduced by the addition of the IEL isolate to the transplantation innocula, and to a lesser degree by the addition of BCs. The augmentation of repopuation far exceeded that expected by simple lodging of cells suggesting that the additionally transplanted cells contained a subpopulation of mucosal membrane lymphoid stem cells or helper cells. Correlation analysis of PC versus IEL levels suggests a possible feedback mechanism governing the relative size of their populations. Normal ratios of IgA, IgM, and IgG bearing PCs was maintained post transplantation with all of the regimens. ^

STUDIES ON THE CLASS I GLYCOPROTEINS OF THE MURINE MAJOR HISTOCOMPATIBILITY COMPLEX (TRANSPLANTATION ANTIGEN, LYMPHOCYTE, CELL SURFACE MOLECULE)

A series of studies were undertaken to analyze and compare various aspects of murine class I glycoproteins. An initial area of investigation characterized the Qa-1 alloantigens using two-dimensional gel electrophoresis. Analysis of the products of the Qa-1('b), Qa-1('c) and Qa-1('d) alleles indicated that these were distinct molecules as determined by their lack of comigration upon comparative two-dimensional gel analysis. The importance of asparagine-linked glycosylation in the cell surface expression of class I molecules was also examined. These studies employed tunicamycin, an inhibitor of N-linked glycosylation. Tunicamycin treatment of activated T lymphocytes diminished the surface expression of Qa-1 to undetectable levels; the levels of other class I molecules exhibited little or no decrease. These results indicated that N-linked glycosylation has a differential importance in the cell surface expression of various class I molecules. The molecular weight diversity of class I molecules was also investigated. Molecular weight determination of both the fully glycosylated and unglycosylated forms of H-2 and Qa/Tla region encoded molecules established that there is a significant variation in the sizes of these forms of various class I molecules. The most significant difference ((TURN)9,000 daltons) exists between the unglycosylated forms of H-2K('b) and Qa-2, suggesting that the structural organization of these two molecules may be very different. A comparative two-dimensional gel analysis of various class I glycoproteins isolated from resting and activated T and B lymphocytes indicated that class I molecules expressed on activated T cells exhibited an isoelectrophoretic pattern that was distinct from the isoelectrophoretic pattern of class I molecules expessed on the other cell populations. This difference was attributed to a lower sialic acid content of the molecules expressed on activated T cells. Analysis of cell homogenates determined that activated T cells contained a higher level of endogenous neuraminidase activity than was detected in the other populations, suggesting that this may be the basis of the lower sialic acid content. The relationship of the Qa-4 and Qa-2 alloantigens was also examined. It was established that upon mitogen activation, the expression of Qa-4 was greatly decreased, whereas Qa-2 expression was not decreased. However, an anti-Qa-2 monoclonal antibody blocked the binding of an anti-Qa-4 monoclonal antibody to resting cells. These studies established that Qa-4 is a determinant restricted to resting cells, which is closely associated on the surface with the Qa-2 molecule. ^

Tumor immunization strategies for enhancing the graft versus tumor activity of allogeneic bone marrow transplantation

Allogeneic bone marrow transplantation (BMT) is known to induce a beneficial anti-tumor immune response called graft-versus-tumor (GVT) activity. However, GVT activity is closely associated with graft-versus-host disease (GVHD), a potentially fatal immune response against antigens on normal recipient tissues. The T-cell populations mediating these two processes are often overlapping, but studies have shown that some donor T-cells can be tumor-specific. Therefore, the goal of this study was to develop strategies for preferentially activating donor T-cells capable of mediating GVT activity but not GVHD. The three hypotheses tested were: (1) Pre-transplant immunization of BMT donors with a recipient-derived tumor cell vaccine will induce a relative increase in GVT activity as compared to GVHD. (2) Post-transplant tumor immunization of BMT recipients will enhance GVT activity without exacerbating GVHD. (3) Pre-transplant immunization of BMT donors against a tumor-specific antigen will enhance GVT activity without exacerbating GVHD. ^ To test the first two hypotheses, C3H.SW mice (MHC-matched donors) were immunized with a C57BL/6 (recipient)-derived tumor cell vaccine (leukemia or fibrosarcoma) prior to BMT, or recipients were immunized starting one month after BMT. Both donor and recipient immunization led to a significant increase in GVT activity (enhanced recipient survival and decreased tumor growth). However, donor immunization also increased fatal GVHD, which was at least partially due to activation of alloreactive T-cells recognizing the immunodominant minor histocompatibility antigen B6dom1. GVT immunity following recipient immunization was not associated with an exacerbation of GVHD or a response to B6dom1. ^ To test the third hypothesis, influenza nucleoprotein (NP) was used as a model tumor antigen. C3H.SW donors were immunized against NP prior to BMT, which led to a significant increase in GVT activity. Although recipients were not completely protected against growth of antigen loss variant tumors, there was no increase in GVHD. ^ In conclusion, (1) immunization of allogeneic BMT donors with a recipient-derived tumor cell vaccine substantially increases GVT activity but also exacerbates GVHD, (2) post-transplant tumor immunization of allogeneic BMT recipients significantly increases GVT activity and survival without exacerbating GVHD, and (3) immunization of allogeneic BMT donors against a tumor-specific antigen significantly enhances GVT activity without exacerbating GVHD. ^

Racial disparities in patient survival after liver transplantation in the United States

Background. Racial disparities in healthcare span such areas as access, outcomes after procedures, and patient satisfaction. Previous work suggested that minorities experience less healthcare and worse survival rates. In adult orthotopic liver transplantation (OLT) mixed results have been reported, with some showing African-American recipients having poor survival compared to Caucasians, and others finding no such discrepancy. ^ Purpose. This study’s purpose was to analyze the most recent United Network for Organ Sharing (UNOS) data, both before and after the implementation of the Model for End-Stage Liver Disease (MELD)/Pediatric End-Stage Liver Disease (PELD) scoring system, to determine if minority racial groups still experience poor outcomes after OLT. ^ Methods. The UNOS dataset for 1992-2001 (Era I) and 2002-2007 (Era II) was used. Patient survival rates for each Era and for adult and pediatric recipients were analyzed with adjustment. A separate multivariate analysis was performed on African-American adult patients in Era II in order to identify unique predictors for poor patient survival. ^ Results. The overall study included 66,118 OLT recipients. The majority were Caucasian (78%), followed by Hispanics (13%) and African-Americans (9%). Hispanic and African-American adults were more likely to be female, have Hepatitis C, to be in the intensive care unit (ICU) or ventilated at time of OLT, to have a MELD score ≥23, to have a lower education level, and to have public insurance when compared to Caucasian adults (all p-values < 0.05). Hispanic and African-American pediatric recipients were more likely have public insurance and less likely to receive a living donor OLT than were Caucasian pediatric OLT recipients (p <0.05). There was no difference in the likelihood of having a PELD score ≥21 among racial groups (p >0.40). African-American adults in Era I and Era II had worse patient survival rates than both Caucasians and Hispanic (pair-wise p-values <0.05). This same disparity was seen for pediatric recipients in Era I, but not in Era II. Multivariate analysis of African-American recipients revealed no unique predictors of patient death. ^ Conclusions. African-American race is still a predictor of poor outcome after adult OLT, even after adjustment for multiple clinical, demographic, and liver disease severity variables. Although African-American and Hispanic subgroups share many characteristics previously thought to increase risk of post-OLT death, only African-American patients have poor survival rates when compared to Caucasians. ^

Prevalence and risk factors for polyomavirus reactivation in solid-organ transplantation

Background. Polyomavirus reactivation is common in solid-organ transplant recipients who are given immunosuppressive medications as standard treatment of care. Previous studies have shown that polyomavirus infection can lead to allograft failure in as many as 45% of the affected patients. Hypothesis. Ubiquitous polyomaviruses when reactivated by post-transplant immunosuppressive medications may lead to impaired renal function and possibly lower survival prospects. Study Overview. Secondary analysis of data was conducted on a prospective longitudinal study of subjects who were at least 18 years of age and were recipients of liver and/or kidney transplant at Mayo Clinic Scottsdale, Arizona. Methods. DNA extractions of blinded urine and blood specimens of transplant patients collected at Mayo Clinic during routine transplant patient visits were performed at Baylor College of Medicine using Qiagen kits. Virologic assays included testing DNA samples for specific polyomavirus sequences using QPCR technology. De-identified demographic and clinical patient data were merged with laboratory data and statistical analysis was performed using Stata10. Results. 76 patients enrolled in the study were followed for 3.9 years post transplantation. The prevalence of BK virus and JC virus urinary excretion was 30% and 28%. Significant association was observed between JC virus excretion and kidney as the transplanted organ (P = 0.039, Pearson Chi-square test). The median urinary JCV viral loads were two logs higher than those of BKV. Patients that excreted both BKV and JCV appeared to have the worst renal function with a mean creatinine clearance value of 71.6 millimeters per minute. A survival disadvantage was observed for dual shedders of BKV and JCV, log-rank statistics, p = 0.09; 2/5 dual-shedders expired during the study period. Liver transplant and male sex were determined to be potential risk factors for JC virus activation in renal and liver transplant recipients. All patients tested negative for SV40 and no association was observed between polyomavirus excretion and type of immunosuppressive medication (tacrolimus, mycophenolate mofetil, cyclosporine and sirolimus). Conclusions. Polyomavirus reactivation was common after solid-organ transplantation and may be associated with impaired renal function. Male sex and JCV infection may be potential risk factors for viral reactivation; findings should be confirmed in larger studies.^

Children with acute leukemia: A comparison of outcomes and cost-effectiveness from allogeneic blood stem cell and bone marrow transplantation

The relative merits of PBSCT versus BMT for children with standard and high risk hematologic malignancies remain unclear. In a retrospective single center study, we compared allogeneic peripheral blood stem cell transplantation (PBSCT) (n=30) with bone marrow transplantation (BMT) (n=110) in children with acute leukemia. We studied recipients of HLA matched sibling stem cells, and of stem cells from alternative donors (HLA mismatched and/or unrelated) and determined whether sourcing the stem cells from PB or marrow affected engraftment, incidence of acute and chronic GvHD, and disease-free survival at 1 year. Our results show a modest reduction in time to engraftment from PB stem cells and no greater risk of GvHD, but illustrate that the severity of the underlying disease is by far the greatest determinant of 1 year survival. Patients in the BMT group had a higher treatment success rate and lower costs than the recipients of the PBSCT within the standard but not the high risk disease group, where the treatment success rate and the cumulative costs were lower in the PBSCT group compared to the BMT group. Our current incremental cost-effectiveness ratio and analysis of uncertainty suggest that allogeneic transplantation of bone marrow grafts was a more cost-effective treatment option compared to peripheral blood stem cells in patients with standard risk childhood acute leukemia disease. For high risk disease our data are less prescriptive, since the differences were more limited and the range of costs much larger. Neither option demonstrated a clear advantage from a cost-effectiveness standpoint.^

Understanding the roles of Non-homologous end joining and p53 after DNA damage

The inability to maintain genomic stability and control proliferation are hallmarks of many cancers, which become exacerbated in the presence of unrepaired DNA damage. Such genotoxic stresses trigger the p53 tumor suppressor network to activate transient cell cycle arrest allowing for DNA repair; if the damage is excessive or irreparable, apoptosis or cellular senescence is triggered. One of the major DNA repair pathway that mends DNA double strand breaks is non-homologous end joining (NHEJ). Abrogating the NHEJ pathway leads to an accumulation of DNA damage in the lymphoid system that triggers p53-mediated apoptosis; complete deletion of p53 in this system leads to aggressive lymphomagenesis. Therefore, to study the effect of p53-dependent cell cycle arrest, we utilized a hypomorphic, separation-of-function mutant, p53p/p, which completely abrogates apoptosis yet retains partial cell cycle arrest ability. We crossed DNA ligase IV deficiency, a downstream ligase crucial in mending breaks during NHEJ, into the p53p/p background (Lig4-/-p53p/p). The accumulation of DNA damage activated the p53/p21 axis to trigger cellular senescence in developing lymphoid cells, which absolutely suppressed tumorigenesis. Interestingly, these mice progressively succumb to severe diabetes. Mechanistic analysis revealed that spontaneous DNA damage accumulated in the pancreatic b-cells, a unique subset of endocrine cells solely responsible for insulin production to regulate glucose homeostasis. The genesis of adult b-cells predominantly occurs through self-replication, therefore modulating cellular proliferation is an essential component for renewal. The progressive accumulation of DNA damage, caused by Lig4-/-, activated p53/p21-dependent cellular senescence in mutant pancreatic b-cells that lead to islet involution. Insulin levels subsequently decreased, deregulating glucose homeostasis driving overt diabetes. Our Lig4-/-p53p/p model aptly depicts the dichotomous role of cellular senescence—in the lymphoid system prevents tumorigenesis yet in the endocrine system leads to the decrease of insulin-producing cells causing diabetes. To further delineate the function of NHEJ in pancreatic b-cells, we analyzed mice deficient in another component of the NHEJ pathway, Ku70. Although most notable for its role in DNA damage recognition and repair within the NHEJ pathway, Ku70 has NHEJ-independent functions in telomere maintenance, apoptosis, and transcriptional regulation/repression. To our surprise, Ku70-/-p53p/p mutant mice displayed a stark increase in b-cell proliferation, resulting in islet expansion, heightened insulin levels and hypoglycemia. Augmented b-cell proliferation was accompanied with the stabilization of the canonical Wnt pathway, responsible for this phenotype. Interestingly, the progressive onset of cellular senescence prevented islet tumorigenesis. This study highlights Ku70 as an important modulator in not only maintaining genomic stability through NHEJ-dependent functions, but also reveals a novel NHEJ-independent function through regulation of pancreatic b-cell proliferation. Taken in aggregate, these studies underscore the importance for NHEJ to maintain genomic stability in b-cells as well as introduces a novel regulator for pancreatic b-cell proliferation.

Impact of donor-recipient gender and race mismatching on patient survival in patients with end-stage liver diseases undergoing liver transplantation

Background. End-stage liver disease (ESLD) is an irreversible condition that leads to the imminent complete failure of the liver. Orthotopic liver transplantation (OLT) has been well accepted as the best curative option for patients with ESLD. Despite the progress in liver transplantation, the major limitation nowadays is the discrepancy between donor supply and organ demand. In an effort to alleviate this situation, mismatched donor and recipient gender or race livers are being used. However, the simultaneous impact of donor and recipient gender and race mismatching on patient survival after OLT remains unclear and relatively challenging to surgeons. ^ Objective. To examine the impact of donor and recipient gender and race mismatching on patient survival after OLT using the United Network for Organ Sharing (UNOS) database. ^ Methods. A total of 40,644 recipients who underwent OLT between 2002 and 2011 were included. Kaplan-Meier survival curves and the log-rank tests were used to compare the survival rates among different donor-recipient gender and race combinations. Univariate Cox regression analysis was used to assess the association of donor-recipient gender and race mismatching with patient survival after OLT. Multivariable Cox regression analysis was used to model the simultaneous impact of donor-recipient gender and race mismatching on patient survival after OLT adjusting for a list of other risk factors. Multivariable Cox regression analysis stratifying on recipient hepatitis C virus (HCV) status was also conducted to identify the variables that were differentially associated with patient survival in HCV + and HCV − recipients. ^ Results. In the univariate analysis, compared to male donors to male recipients, female donors to male recipients had a higher risk of patient mortality (HR, 1.122; 95% CI, 1.065–1.183), while in the multivariable analysis, male donors to female recipients experienced an increased mortality rates (adjusted HR, 1.114; 95% CI, 1.048–1.184). Compared to white donors to white recipients, Hispanic donors to black recipients had a higher risk of patient mortality (HR, 1.527; 95% CI, 1.293–1.804) in the univariate analysis, and similar result (adjusted HR, 1.553; 95% CI, 1.314–1.836) was noted in multivariable analysis. After the stratification on recipient HCV status in the multivariable analysis, HCV + mismatched recipients appeared to be at greater risk of mortality than HCV − mismatched recipients. Female donors to female HCV − recipients (adjusted HR, 0.843; 95% CI, 0.769–0.923), and Hispanic HCV + recipients receiving livers from black donors (adjusted HR, 0.758; 95% CI, 0.598–0.960) had a protective effect on patient survival after OLT. ^ Conclusion. Donor-recipient gender and race mismatching adversely affect patient survival after OLT, both independently and after the adjustment for other risk factors. Female recipient HCV status is an important effect modifier in the association between donor-recipient gender combination and patient survival.^