[Show abstract][Hide abstract] ABSTRACT: Adaptive CD8 T-cell immunity is the principal arm of the cellular alloimmune response, but its development requires help. This can be provided by CD4 T cells that recognize alloantigen "indirectly," as self-restricted allopeptide, but this process remains unexplained, because the target epitopes for CD4 and CD8 T-cell recognition are "unlinked" on different cells (recipient and donor antigen presenting cells (APCs), respectively). Here, we test the hypothesis that the presentation of intact and processed MHC class I alloantigen by recipient dendritic cells (DCs) (the "semidirect" pathway) allows linked help to be delivered by indirect-pathway CD4 T cells for generating destructive cytotoxic CD8 T-cell alloresponses. We show that CD8 T-cell-mediated rejection of murine heart allografts that lack hematopoietic APCs requires host secondary lymphoid tissue (SLT). SLT is necessary because within it, recipient dendritic cells can acquire MHC from graft parenchymal cells and simultaneously present it as intact protein to alloreactive CD8 T cells and as processed peptide alloantigen for recognition by indirect-pathway CD4 T cells. This enables delivery of essential help for generating cytotoxic CD8 T-cell responses that cause rapid allograft rejection. In demonstrating the functional relevance of the semidirect pathway to transplant rejection, our findings provide a solution to a long-standing conundrum as to why SLT is required for CD8 T-cell allorecognition of graft parenchymal cells and suggest a mechanism by which indirect-pathway CD4 T cells provide help for generating effector cytotoxic CD8 T-cell alloresponses at late time points after transplantation.
Proceedings of the National Academy of Sciences 09/2015; DOI:10.1073/pnas.1513533112 · 9.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The study of biliary disease has been constrained by a lack of primary human cholangiocytes. Here we present an efficient, serum-free protocol for directed differentiation of human induced pluripotent stem cells into cholangiocyte-like cells (CLCs). CLCs show functional characteristics of cholangiocytes, including bile acids transfer, alkaline phosphatase activity, γ-glutamyl-transpeptidase activity and physiological responses to secretin, somatostatin and vascular endothelial growth factor. We use CLCs to model in vitro key features of Alagille syndrome, polycystic liver disease and cystic fibrosis (CF)-associated cholangiopathy. Furthermore, we use CLCs generated from healthy individuals and patients with polycystic liver disease to reproduce the effects of the drugs verapamil and octreotide, and we show that the experimental CF drug VX809 rescues the disease phenotype of CF cholangiopathy in vitro. Our differentiation protocol will facilitate the study of biological mechanisms controlling biliary development, as well as disease modeling and drug screening.
[Show abstract][Hide abstract] ABSTRACT: The use of kidneys from controlled donation after circulatory death (DCD) donors has the potential to markedly increase kidney transplants performed. However, this potential is not being realized because of concerns that DCD kidneys are inferior to those from donation after brain-death (DBD) donors. The United Kingdom has developed a large and successful controlled DCD kidney transplant program that has allowed for a substantial increase in kidney transplant numbers. Here we describe recent trends in DCD kidney donor activity in the United Kingdom, outline aspects of the donation process, and describe donor selection and allocation of DCD kidneys. Previous UK Transplant Registry analyses have shown that while DCD kidneys are more susceptible to cold ischemic injury and have a higher incidence of delayed graft function, short- and medium-term transplant outcomes are similar in recipients of kidneys from DCD and DBD donors. We present an updated, extended UK registry analysis showing that longer-term transplant outcomes in DCD donor kidneys are also similar to those for DBD donor kidneys, and that transplant outcomes for kidneys from expanded-criteria DCD donors are no less favorable than for expanded-criteria DBD donors. Accordingly, the selection criteria for use of kidneys from DCD donors should be the same as those used for DBD donors. The UK experience suggests that wider international development of DCD kidney transplantation programs will help address the global shortage of deceased donor kidneys for transplantation.Kidney International advance online publication, 18 March 2015; doi:10.1038/ki.2015.88.
Kidney International 03/2015; 88(2). DOI:10.1038/ki.2015.88 · 8.56 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Acute umbilical hernia rupture in patients with hepatic cirrhosis and ascites is an unusual, but potentially life-threatening complication, with postoperative morbidity about 70% and mortality between 60%-80% after supportive care and 6%-20% after urgent surgical repair. Management options include primary surgical repair with or without concomitant portal venous system decompression for the control of the ascites. We present a retrospective analysis of our centre's experience over the last 6 years. Our cohort consisted of 11 consecutive patients (median age: 53 years, range: 36-63 years) with advanced hepatic cirrhosis and refractory ascites. Appropriate patient resuscitation and optimisation with intravenous fluids, prophylactic antibiotics and local measures was instituted. One failed attempt for conservative management was followed by a successful primary repair. In all cases, with one exception, a primary repair with non-absorbable Nylon, interrupted sutures, without mesh, was performed. The perioperative complication rate was 25% and the recurrence rate 8.3%. No mortality was recorded. Median length of hospital stay was 14 d (range: 4-31 d). Based on our experience, the management of ruptured umbilical hernias in patients with advanced hepatic cirrhosis and refractory ascites is feasible without the use of transjugular intrahepatic portosystemic shunt routinely in the preoperative period, provided that meticulous patient optimisation is performed.
[Show abstract][Hide abstract] ABSTRACT: Background:
We have previously shown that qualitative assessment of surface electrostatic potential of HLA class I molecules helps explain serological patterns of alloantibody binding. We have now used a novel computational approach to quantitate differences in surface electrostatic potential of HLA B-cell epitopes and applied this to explain HLA Bw4 and Bw6 antigenicity.
Protein structure models of HLA class I alleles expressing either the Bw4 or Bw6 epitope (defined by sequence motifs at positions 77 to 83) were generated using comparative structure prediction. The electrostatic potential in 3-dimensional space encompassing the Bw4/Bw6 epitope was computed by solving the Poisson-Boltzmann equation and quantitatively compared in a pairwise, all-versus-all fashion to produce distance matrices that cluster epitopes with similar electrostatics properties.
Quantitative comparison of surface electrostatic potential at the carboxyl terminal of the α1-helix of HLA class I alleles, corresponding to amino acid sequence motif 77 to 83, produced clustering of HLA molecules in 3 principal groups according to Bw4 or Bw6 epitope expression. Remarkably, quantitative differences in electrostatic potential reflected known patterns of serological reactivity better than Bw4/Bw6 amino acid sequence motifs. Quantitative assessment of epitope electrostatic potential allowed the impact of known amino acid substitutions (HLA-B*07:02 R79G, R82L, G83R) that are critical for antibody binding to be predicted.
We describe a novel approach for quantitating differences in HLA B-cell epitope electrostatic potential. Proof of principle is provided that this approach enables better assessment of HLA epitope antigenicity than amino acid sequence data alone, and it may allow prediction of HLA immunogenicity.
[Show abstract][Hide abstract] ABSTRACT: The monoclonal anti-CD20 antibody rituximab depletes B cells in the treatment of lymphoma and autoimmune disease, and contributes to alloantibody reduction in transplantation across immunological barriers. The effects of rituximab on T cells are less well described. T follicular helper cells (Tfh) provide growth and differentiation signals to germinal center (GC) B cells to support antibody production, and suppressive T follicular regulatory cells (Tfr) regulate this response. In mice, both Tfh and Tfr are absolutely dependent on B cells for their formation and on the GC for their maintenance. Here, we demonstrate that rituximab treatment results in a lack of GC B cells in human lymph nodes without affecting the Tfh or Tfr cell populations. These data demonstrate that human Tfh and Tfr do not require an on-going GC response for their maintenance. The persistence of Tfh and Tfr following rituximab treatment may permit rapid reconstitution of the pathological GC response once the B cell pool begins to recover. Strategies for maintaining remission after rituximab therapy will need to take this persistence of Tfh into account.
[Show abstract][Hide abstract] ABSTRACT: Advances in developmental biology have shown that monozygous twins may not be as phenotypically identical as once believed, and the mechanisms responsible for such differences are now becoming clearer. Whether such phenotypic differences are capable of triggering graft rejection of an organ transplanted between identical twins remains unknown but the risks seem low, and long-term transplant outcome is excellent. Available evidence to guide immunosuppressive therapy in this setting is limited but a prudent approach would include the use of steroids together with a calcineurin inhibitor after transplantation. However, once the inevitable inflammatory response associated with transplant surgery has resolved, cautious reduction and eventually withdrawal of immunosuppression should be possible.
[Show abstract][Hide abstract] ABSTRACT: Transplantation is the preferred treatment for most patients with organ failure, and equitable access to organ transplantation is therefore essential. However, large inequalities in organ donation and transplantation rates are seen between countries. Specific patient cultural groups are also disadvantaged in their access to organ transplantation within countries. These inequities are at least partly due to variations in the legal, ethical, financial, organisational, and social frameworks that enable transplants to occur. Along with the global shortage in donor organs, these variations drive transplant commercialism, organ trafficking, and transplant tourism. This chapter examines selected ‘cultural’ variations that impact on access to transplantation, and explores the issues of transplant commercialism, organ trafficking, and transplant tourism. Finally, the arguments for and against the introduction of financial incentives for organ donation are discussed.
Textbook of Organ Transplantation, 07/2014: pages 1662-1672; , ISBN: 9781118870143
[Show abstract][Hide abstract] ABSTRACT: Background: There is variation in time to listing and rates of listing for transplantation between renal units in the UK. While research has mainly focused on healthcare organization, little is known about patient perspectives of entry onto the transplant waiting list. This qualitative study aimed to explore patients' views and experiences of kidney transplant listing.
Methods: Semi-structured interviews were conducted with patients aged under 75, who were on dialysis and on the transplant waiting list, not on the waiting list, undergoing assessment for listing or who had received a transplant. Patients were recruited from a purposive sample of nine UK renal units, which included transplanting and non-transplanting units and units with high and low wait-listing patterns. Interviews were transcribed verbatim and analysed using thematic analysis.
Results: Fifty-three patients (5–7 per renal unit) were interviewed. Patients reported that they had received little information about the listing process. Some patients did not know if they were listed or had found they were not listed when they had thought they were on the list. Others expressed distress when they felt they had been excluded from potential listing based on age and/or comorbidity and felt the process was unfair. Many patients were not aware of pre-emptive transplantation and believed they had to be on dialysis before being able to be listed. There was some indication that pre-emptive transplantation was discussed more often in transplant than non-transplant units. Lastly, some patients were reluctant to consider family members as potential donors as they reported they would feel ‘guilty’ if the donor suffered subsequent negative effects.
Conclusions: Findings suggest a need to review current practice to further understand individual and organizational reasons for the renal unit variation identified in patient understanding of transplant listing. The communication of information warrants attention to ensure patients are fully informed about the listing process and opportunity for pre-emptive transplantation in a way that is meaningful and understandable to them.
[Show abstract][Hide abstract] ABSTRACT: : Renal transplantation is a successful treatment for patients with renal failure but its long-term efficacy is limited by untreatable transplant vasculopathy (TA). Endothelial damage contributes to TA and is potentially repairable by circulating endothelial progenitor cells (EPC). The frequency and function of EPC is variably influenced by end-stage renal failure (ESRF). This study aimed to characterise the late-outgrowth EPC (LO-EPC) from ESRF patients with a view to utilising autologous LO-EPC for endothelial repair following renal transplantation. LO-EPC isolated from ESRF patients and healthy volunteers were characterised phenotypically and functionally and their integrin expression profile was determined.ESRF patients generated more LO-EPC colonies than healthy controls, had higher plasma levels of IL-1rα, IL-16, IL-6, MIF, VEGF, Prolactin and PLGF. Patients' LO-EPC displayed normal endothelial cell morphology, increased secretion of PLGF, MCP-1 and IL-1β, decreased senescence and normal network formation in vitro and in vivo , but demonstrated decreased adhesion to extracellular matrix. Integrin gene profiles and protein expression were comparable in patients and healthy volunteers. In some patients, mesenchymal stem cells were co-isolated from peripheral blood and these could be differentiated into adipocytes and osteocytes in vitro . This is the first study to characterise LO-EPC from ESRF patients. Their behaviour in vitro reflects the presence of elevated trophic factors; their ability to proliferate in vitro and angiogenic function makes them candidates for prevention of transplant vasculopathy. Their impaired adhesion and the presence of MSC are areas for potential therapeutic intervention.
[Show abstract][Hide abstract] ABSTRACT: We have analyzed the relationship between donor mismatches at each HLA locus and development of HLA locus-specific antibodies in patients listed for repeat transplantation. HLA antibody screening was undertaken using single-antigen beads in 131 kidney transplant recipients returning to the transplant waiting list following first graft failure. The number of HLA mismatches and the calculated reaction frequency of antibody reactivity against 10,000 consecutive deceased organ donors were determined for each HLA locus. Two-thirds of patients awaiting repeat transplantation were sensitized (calculated reaction frequency over 15%) and half were highly sensitized (calculated reaction frequency of 85% and greater). Antibody levels peaked after re-listing for repeat transplantation, were independent of graft nephrectomy and were associated with length of time on the waiting list (odds ratio 8.4) and with maintenance on dual immunosuppression (odds ratio 0.2). Sensitization was independently associated with increasing number of donor HLA mismatches (odds ratio 1.4). All mismatched HLA loci contributed to the development of HLA locus-specific antibodies (HLA-A: odds ratio 3.2, HLA-B: odds ratio 3.4, HLA-C: odds ratio 2.5, HLA-DRB1: odds ratio 3.5, HLA-DRB3/4/5: odds ratio 3.9, and HLA-DQ: odds ratio 3.0 (all significant)). Thus, the risk of allosensitization following failure of a first renal transplant increases incrementally with the number of mismatches at all HLA loci assessed. Maintenance of re-listed patients on dual immunosuppression was associated with a reduced risk of sensitization.Kidney International advance online publication, 9 April 2014; doi:10.1038/ki.2014.106.
Kidney International 04/2014; 86(5). DOI:10.1038/ki.2014.106 · 8.56 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: It is essential to minimize the unnecessary discard of procured deceased donor kidneys, but information on discard rates and the extent to which discard can be avoided are limited. Analysis of the UK Transplant Registry revealed that the discard rate of procured deceased donor kidneys has increased from 5% in 2002-3 to 12% in 2011-12. A national offering system for hard-to-place kidneys was introduced in the UK in 2006 (the Declined Kidney Scheme), but just 13% of kidneys that were subsequently discarded until 2012 were offered through the scheme. In order to examine the appropriateness of discard, 20 consecutive discarded kidneys from 13 deceased donors were assessed to determine if surgeons agreed with the decision that they were not implantable. Donors had a median (range) age of 67 (31-80) yr. Kidneys had been offered to a median of 3 (1-12) centers before discard. Four (20%) of the discarded kidneys were thought to be usable, and nine (45%) were possibly usable. As a result of these findings, major changes to the UK deceased donor kidney offering system have been implemented, including simultaneous offering and broader entry criteria for hard-to-place kidneys. Organizational changes are necessary to improve utilization of deceased donor kidneys.