Posttransplant Lymphoproliferative Disorders in Neuronal Xenotransplanted Macaques.

Posttransplant lymphoproliferative disorders (PTLDs) are a heterogeneous group of lymphoid proliferations that occur in the setting of depressed T-cell function due to immunosuppressive therapy used following solid organ transplantation, hematopoietic stem cell transplantation, and also xenotransplantation. In the present study, 28 immunosuppressed parkinsonian Macaca fascicularis were intracerebrally injected with wild-type or CTLA4-Ig transgenic porcine xenografts to identify a suitable strategy to enable long-term cell survival, maturation, and differentiation. Nine of 28 (32%) immunosuppressed primates developed masses compatible with PTLD, located mainly in the gastrointestinal tract and/or nasal cavity. The masses were classified as monomorphic PTLD according to the World Health Organization classification. Immunohistochemistry and polymerase chain reaction (PCR) analyses revealed that the PTLDs were associated with macaca lymphocryptovirus as confirmed by double-labeling immunohistochemistry for CD20 and Epstein-Barr nuclear antigen 2 (EBNA-2), where the viral protein was located within the CD20+ neoplastic B cells. In sera from 3 distinct phases of the experimental life of the primates, testing by quantitative PCR revealed a progression of the viral load that paralleled the PTLD progression and no evidence of zoonotic transmission of porcine lymphotropic herpesvirus through xenoneuronal grafts. These data suggest that monitoring the variation of macaca lymphocryptovirus DNA in primates could be used as a possible early diagnostic tool for PTLD progression, allowing preemptive treatment such as immunosuppression therapy reduction.

Cost-effectiveness of noninvasive liver fibrosis tests for treatment decisions in patients with chronic hepatitis B in the UK: systematic review and economic evaluation.

We compared the cost-effectiveness of various noninvasive tests (NITs) in patients with chronic hepatitis B and elevated transaminases and/or viral load who would normally undergo liver biopsy to inform treatment decisions. We searched various databases until April 2012. We conducted a systematic review and meta-analysis to calculate the diagnostic accuracy of various NITs using a bivariate random-effects model. We constructed a probabilistic decision analytical model to estimate health care costs and outcomes quality-adjusted-life-years (QALYs) using data from the meta-analysis, literature, and national UK data. We compared the cost-effectiveness of four decision-making strategies: testing with NITs and treating patients with fibrosis stage ≥F2, testing with liver biopsy and treating patients with ≥F2, treat none (watchful waiting) and treat all irrespective of fibrosis. Treating all patients without prior fibrosis assessment had an incremental cost-effectiveness ratio (ICER) of £28,137 per additional QALY gained for HBeAg-negative patients. For HBeAg-positive patients, using Fibroscan was the most cost-effective option with an ICER of £23,345. The base case results remained robust in the majority of sensitivity analyses, but were sensitive to changes in the ≥ F2 prevalence and the benefit of treatment in patients with F0-F1. For HBeAg-negative patients, strategies excluding NITs were the most cost-effective: treating all patients regardless of fibrosis level if the high cost-effectiveness threshold of £30,000 is accepted; watchful waiting if not. For HBeAg-positive patients, using Fibroscan to identify and treat those with ≥F2 was the most cost-effective option.

Bortezomib, C1-inhibitor and plasma exchange do not prolong the survival of multi-transgenic GalT-KO pig kidney xenografts in baboons.

Galactosyl-transferase KO (GalT-KO) pigs represent a potential solution to xenograft rejection, particularly in the context of additional genetic modifications. We have performed life supporting kidney xenotransplantation into baboons utilizing GalT-KO pigs transgenic for human CD55/CD59/CD39/HT. Baboons received tacrolimus, mycophenolate mofetil, corticosteroids and recombinant human C1 inhibitor combined with cyclophosphamide or bortezomib with or without 2-3 plasma exchanges. One baboon received a control GalT-KO xenograft with the latter immunosuppression. All immunosuppressed baboons rejected the xenografts between days 9 and 15 with signs of acute humoral rejection, in contrast to untreated controls (n = 2) that lost their grafts on days 3 and 4. Immunofluorescence analyses showed deposition of IgM, C3, C5b-9 in rejected grafts, without C4d staining, indicating classical complement pathway blockade but alternate pathway activation. Moreover, rejected organs exhibited predominantly monocyte/macrophage infiltration with minimal lymphocyte representation. None of the recipients showed any signs of porcine endogenous retrovirus transmission but some showed evidence of porcine cytomegalovirus (PCMV) replication within the xenografts. Our work indicates that the addition of bortezomib and plasma exchange to the immunosuppressive regimen did not significantly prolong the survival of multi-transgenic GalT-KO renal xenografts. Non-Gal antibodies, the alternative complement pathway, innate mechanisms with monocyte activation and PCMV replication may have contributed to rejection.

Unusual serological response to hepatitis E virus in plasma donors consistent with re-infection.

Hepatitis E virus (HEV)-positive plasma donations, identified by a plasma mini-pool screening approach, were analysed using serological methods for the presence of anti-HEV IgM and IgG. Avidity testing was performed on the IgG-reactive donations. Anti-HEV IgG with high avidity was observed in two donors together with high viral loads, but with the absence of anti-HEV IgM. These data are suggestive of re-infection in a small proportion of plasma donors, which has not previously been reported.

Prevalence of hepatitis E virus in slaughter-age pigs in Scotland.

The prevalence of anti-HEV isotype-specific antibodies and viraemia were investigated in serum samples collected from slaughter-age pigs (aged 22-24 weeks) from 23 farms in Scotland. Of 176 serum samples tested, 29·0% (n = 51) were anti-HEV IgG positive, 36·9% (n = 65) anti-HEV IgA positive and 29·0% (n = 51) anti-HEV IgM positive. Overall seroprevalence (anti-HEV IgG+ and/or IgA+ and/or IgM+) was 61·4% (n = 108). HEV RNA was detected in 72/162 serum samples (44·4%). Partial sequence of ORF2 (98 nt) was obtained from eight HEV RNA-positive samples and phylogenetic analysis confirmed that they were all of genotype 3. This is the first report on the prevalence of HEV in pigs in Scotland. Given the increasing incidence of locally acquired HEV infection in the UK, evidence that HEV is a foodborne zoonosis emphasizes the need for surveillance in pigs.

Hepatitis E virus in Scottish blood donors.

BACKGROUND AND OBJECTIVES: Published prevalence figures for hepatitis E virus (HEV) reveal significant regional differences. Several studies have reported virus transmission via blood transfusion. The aim of this study was to establish HEV seroprevalence and investigate a potential HEV RNA presence in Scottish blood donors. MATERIALS AND METHODS: IgG and IgM were determined in individual serum samples. HEV RNA was investigated in plasma mini-pools corresponding to 43 560 individual donations using nested PCR. Samples amenable to reamplification with primers from a different region were considered confirmed positives, sequenced and analysed. RESULTS: A total of 73 of 1559 tested individual sera (4·7%) were IgG positive, none tested positive for IgM. Plasma mini-pool testing revealed an HEV RNA frequency of 1 in 14 520 donations. Three confirmed positives belonged, as expected to genotype 3. CONCLUSIONS: HEV IgG and RNA figures in Scottish blood donors are lower than those published for the rest of the UK, but sufficiently high to prompt further studies on potential transmission rates and effects of HEV infection, especially for immunosuppressed individuals.

New hypotheses on the function of the avian shell gland derived from microarray analysis comparing tissue from juvenile and sexually mature hens.

Activation of the shell gland region of the avian oviduct is mediated by ovarian steroids. To understand more extensively how shell glands are maintained and function, we have compared gene expression in the shell glands from juvenile and laying hens using a chicken cDNA microarray. Average expression profiles of juvenile and sexually mature shell glands were compared resulting in the identification of 266 differentially regulated genes. Reverse transcription quantitative polymerase chain reaction confirmed expression differences. The differentially expressed genes included several with known involvement in shell gland function, including ion transport and shell matrix proteins. There were also many unpredicted differentially expressed genes, and for some we propose hypotheses for their functions. These include those encoding (a) osteoprotegerin, a decoy death receptor for receptor activator of nuclear factor NFkB ligand (RANKL) and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), that in the shell gland, may prevent apoptosis and/or may have an endocrine effect by preventing RANKL's action on bone osteoclasts that mobilize stored calcium; (b) prostatic acid phosphatase (ACPP) and prostate stem cell antigen (PSCA) that could play a role in sperm physiology within the shell gland; (c) urea transporter (SLC14A2) that could provide a novel anti-microbial defence; (d) bactericidal/permeability-increasing protein-like 2 (BPIL2), and other potential anti-microbials that have not previously been documented in the chicken. These new hypotheses, if borne out experimentally, will lead to a greater understanding of shell gland function including the processes involved in eggshell formation and anti-microbial activity.

Hepatitis E: a largely underestimated, emerging threat.

Hepatitis E virus has two distinct clinical and epidemiological patterns based on the varying genotypes. Genotypes 3 and 4 cause widespread, sporadic infection in high-income countries and are emerging as the most common type of viral hepatitis in much of Europe. These infections carry significant morbidity and mortality in the growing numbers of immunosuppressed patients or in patients with established liver disease. Furthermore the growing extra-hepatic associations of the virus, including neurological and kidney injury, suggest that it may have been misnamed as a 'hepatitis' virus. This review explores current understanding of the epidemiology, virology and clinical presentations of hepatitis E infection and identifies vulnerable patient groups, who are at serious risk from infection. Guidance is offered regarding the diagnosis, treatment and prevention of this growing public health hazard.

Review article: hepatitis E-a concise review of virology, epidemiology, clinical presentation and therapy.

BACKGROUND: Hepatitis E virus (HEV) is a leading cause of acute icteric hepatitis and acute liver failure in the developing world. During the last decade, there has been increasing recognition of autochthonous (locally acquired) HEV infection in developed countries. Chronic HEV infection is now recognised, and in transplant recipients this may lead to cirrhosis and organ failure. AIM: To detail current understanding of the molecular biology of HEV, diagnostic and therapeutic strategies and propose future directions for basic science and clinical research. METHODS: PubMed was searched for English language articles using the key words "hepatitis E", "viral hepatitis", "autochthonous infection", "antiviral therapy", "liver transplantation", "acute", "chronic", "HEV", "genotype", "transmission" "food-borne", "transfusion". Additional relevant publications were identified from article reference lists. RESULTS: There has been increasing recognition of autochthonous HEV infection in Western countries, mainly associated with genotype 3. Chronic HEV infection has been recognised since 2008, and in transplant recipients this may lead to cirrhosis and organ failure. Modes of transmission include food-borne transmission, transfusion of blood products and solid organ transplantation. Ribavirin therapy is used to treat patients with chronic HEV infection, but new therapies are required as there have been reports of treatment failure with ribavirin. CONCLUSIONS: Autochthonous HEV infection is a clinical issue with increasing burden. Future work should focus on increasing awareness of HEV infection in the developed world, emphasising the need for clinicians to have a low threshold for HEV testing, particularly in immunosuppressed patients. Patients at potential risk of chronic HEV infection must also be educated and given advice regarding prevention of infection.

Hepatitis E virus in patients with decompensated chronic liver disease: a prospective UK/French study.

BACKGROUND: In developed countries, hepatitis E is a porcine zoonosis caused by hepatitis E virus (HEV) genotype 3. In developing countries, hepatitis E is mainly caused by genotype 1, and causes increased mortality in patients with pre-existing chronic liver disease (CLD). AIM: To determine the role of HEV in patients with decompensated CLD. METHODS: Prospective HEV testing of 343 patients with decompensated CLD at three UK centres and Toulouse France, with follow-up for 6 months or death. IgG seroprevalence was compared with 911 controls. RESULTS: 11/343 patients (3.2%) had acute hepatitis E infection, and three died. There were no differences in mortality (27% vs. 26%, OR 1.1, 95% CI 0.28-4.1), age (P = 0.9), bilirubin (P = 0.5), alanine aminotransferase (P = 0.06) albumin (P = 0.5) or international normalised ratio (P = 0.6) in patients with and without hepatitis E infection. Five cases were polymerase chain reaction (PCR) positive (genotype 3). Hepatitis E was more common in Toulouse (7.9%) compared to the UK cohort (1.2%, P = 0.003). HEV IgG seroprevalence was higher in Toulouse (OR 17, 95% CI 9.2-30) and Truro (OR 2.5, 95% CI 1.4-4.6) than in Glasgow, but lower in cases, compared to controls (OR 0.59, 95% CI 0.41-0.86). CONCLUSIONS: Hepatitis E occurs in a minority of patients with decompensated chronic liver disease. The mortality is no different to the mortality in patients without hepatitis E infection. The diagnosis can only be established by a combination of serology and PCR, the yield and utility of which vary by geographical location.

Economic modelling of diagnostic and treatment pathways in National Institute for Health and Care Excellence clinical guidelines: the Modelling Algorithm Pathways in Guidelines (MAPGuide) project.

BACKGROUND: National Institute for Health and Care Excellence (NICE) clinical guidelines (CGs) make recommendations across large, complex care pathways for broad groups of patients. They rely on cost-effectiveness evidence from the literature and from new analyses for selected high-priority topics. An alternative approach would be to build a model of the full care pathway and to use this as a platform to evaluate the cost-effectiveness of multiple topics across the guideline recommendations. OBJECTIVES: In this project we aimed to test the feasibility of building full guideline models for NICE guidelines and to assess if, and how, such models can be used as a basis for cost-effectiveness analysis (CEA). DATA SOURCES: A 'best evidence' approach was used to inform the model parameters. Data were drawn from the guideline documentation, advice from clinical experts and rapid literature reviews on selected topics. Where possible we relied on good-quality, recent UK systematic reviews and meta-analyses. REVIEW METHODS: Two published NICE guidelines were used as case studies: prostate cancer and atrial fibrillation (AF). Discrete event simulation (DES) was used to model the recommended care pathways and to estimate consequent costs and outcomes. For each guideline, researchers not involved in model development collated a shortlist of topics suggested for updating. The modelling teams then attempted to evaluate options related to these topics. Cost-effectiveness results were compared with opinions about the importance of the topics elicited in a survey of stakeholders. RESULTS: The modelling teams developed simulations of the guideline pathways and disease processes. Development took longer and required more analytical time than anticipated. Estimates of cost-effectiveness were produced for six of the nine prostate cancer topics considered, and for five of eight AF topics. The other topics were not evaluated owing to lack of data or time constraints. The modelled results suggested 'economic priorities' for an update that differed from priorities expressed in the stakeholder survey. LIMITATIONS: We did not conduct systematic reviews to inform the model parameters, and so the results might not reflect all current evidence. Data limitations and time constraints restricted the number of analyses that we could conduct. We were also unable to obtain feedback from guideline stakeholders about the usefulness of the models within project time scales. CONCLUSIONS: Discrete event simulation can be used to model full guideline pathways for CEA, although this requires a substantial investment of clinical and analytic time and expertise. For some topics lack of data may limit the potential for modelling. There are also uncertainties over the accessibility and adaptability of full guideline models. However, full guideline modelling offers the potential to strengthen and extend the analytical basis of NICE's CGs. Further work is needed to extend the analysis of our case study models to estimate population-level budget and health impacts. The practical usefulness of our models to guideline developers and users should also be investigated, as should the feasibility and usefulness of whole guideline modelling alongside development of a new CG. FUNDING: This project was funded by the Medical Research Council and the National Institute for Health Research through the Methodology Research Programme [grant number G0901504] and will be published in full in Health Technology Assessment; Vol. 17, No. 58. See the NIHR Journals Library website for further project information.

Xenotransplantation of galactosyl-transferase knockout, CD55, CD59, CD39, and fucosyl-transferase transgenic pig kidneys into baboons.

Galactosyl-transferase knockout (GT-KO) pigs represent the latest major progress to reduce immune reactions in xenotransplantation. However, their organs are still subject to rapid humoral rejection involving complement activation requiring the ongoing development of further genetic modifications in the pig. In a pig-to-baboon renal transplantation setting, we have used donor pigs that are not only GT-KO, but also transgenic for human CD55 (hCD55), hCD59, hCD39, and fucosyl-transferase (hHT). We studied kidney xenograft survival, physiological and immunologic parameters, xenogeneic rejection characteristics, as well as viral transmission aspects among two groups of baboons: control animals (n = 2), versus those (n = 4) treated with a cocktail of cyclophosphamide, tacrolimus, mycophenolate mofetil, steroids, and a recombinant human C1 inhibitor. Whereas control animals showed clear acute humoral rejection at around day 4, the treated animals showed moderately improved graft survival with rejection at around 2 weeks posttransplantation. Biopsies showed signs of acute vascular rejection (interstitial hemorrhage, glomerular thrombi, and acute tubular necrosis) as well as immunoglobulin (Ig)M and complement deposition in the glomerular and peritubular capillaries. The low level of preformed non-Gal-α1.3Gal IgM detected prior to transplantation increased at 6 days posttransplantation, whereas induced IgG appeared after day 6. No porcine endogenous retrovirus (PERV) transmission was detected in any transplanted baboon. Thus, surprisingly, organs from the GT-KO, hCD55, hCD59, hCD39, and hHT transgenic donors did not appear to convey significant protection against baboon anti-pig antibodies and complement activation, which obviously continue to be significant factors under a suboptimal immunosuppression regimen. The association, timing, and doses of immunosuppressive drugs remain critical. They will have to be optimized to achieve longer graft survivals.