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Dendritic cells from the mesenteric lymph nodes (MLN) contain dense esterase-positive inclusions that may originate in effete intestinal epithelial cells and reach MLN without degradation. The MLN esterases have the electrophoretic mobilities of both intestinal and mononuclear cells. Cryptosporidium parvum (CP)-infected mice have CP Ag-positive cells in MLN and also increased numbers of dense esterase-positive cells, but the CP Ag-positive cells do not stain for esterase. To characterize the handling of epithelial cell products by dendritic cells, we analyzed mRNAs in the MLN of control and CP-infected recombination-activating gene−/−DO11.10 mice by oligoarrays. mRNAs for 115 proteins were increased in MLN after CP infection, of which the principal increases in trypsin and chymotrypsin approximated to 250-fold. Colipase, reg-1, C-reactive protein-ductin, and amyloid were also up-regulated >10-fold and all returned to baseline by 28 days after infection. mRNAs for the same proteins were detected in intestinal epithelial cells of infected mice by oligoarrays and RT-PCR after infection. mRNA for CP β-tubulin was detectable in intestinal epithelial cells between 5 and 18 days after infection but was not detected in the MLN throughout the observation period. It appears that host response to CP infection includes expression of mRNA for some pancreatic enzymes by intestinal epithelial cells and their subsequent transport to the MLN. The esterase and trypsin, and mRNAs for chymotrypsin, colipase, and others that may derive from uninfected epithelial cells, appear to be transported to the MLN intact, while mRNA for CP β-tubulin that is derived from infected cells is degraded.

Although CD4 T cells are required for recovery from cryptosporidial infection, mice with severe combined immunodeficiency (SCID) remain infected for long periods without ill effect. In contrast, mice whose ability to use interferon(IFN)‐g is impaired, by neutralization or gene knockout, experience heavy cryptosporidial infection that may lead to death. To determine whether the innate immunity of SCID mice to Cryptosporidium parvum (CP) requires IFNg, doubly immunodeficient C57BL/6 SCID‐IFN-g knockout mice were bred. These mice experienced heavy CP infections of the gut; a significantly greater number became moribund or died, compared with mice carrying the SCID mutation alone or carrying disrupted IFNg genes alone. Mice with gene disruptions of inducible nitric oxide synthetase or Fas/Fas ligand recovered normally from CP infection. The results indicate that mice unable to produce specific immune responses because of the SCID mutation require IFN-g to avoid death after infection with CP. Cryptosporidium parvum (CP) is an apicomplexan parasite that infects epithelial cells in the intestine and the biliary tree. CP has attracted attention recently because it is difficult to eradicate from water supplies and can cause large outbreaks of infection. CP causes only transient diarrhea in healthy individuals, but it is responsible for severe enteritis and cholangitis in immunodeficient individuals, including individuals infected with AIDS. Experience in humans indicates that CD4 cells and an intact CD40‐CD154 pathway [1] are required for recovery from CP infections. These conclusions are confirmed in mice, where cytokine neutralization studies have identified interleukin (IL)‐12 and interferon (IFN)‐g as important or essential cytokines [2‐4]. Roles of these potentially separate effector mechanisms have been explored, in part, by infecting gene-knockout mice with CP. B6 mice with disrupted IFN-g genes have been

Healthy, varicella-zoster virus (VZV)-seropositive subjects, aged 55-89 years (mean age 66 years), received either 4000 PFU of live, attenuated VZV vaccine (n=85) or an equal volume of this vaccine that was heat-inactivated (n=82). Both vaccines significantly boosted VZV antibody (enzyme immunoassay) and gamma-interferon production by peripheral blood mononuclear cells stimulated by VZV antigen. These responses returned to baseline by 12 months. Circulating mononuclear cells that proliferated in response to VZV antigen were significantly more numerous (responder cell frequency assay) after either vaccine, and persisted with a half-life of 17. 5-21.3 months. There were no differences in immune response to either vaccine in this older age cohort.

The authors describe a study to plan and implement an information system for nurses. The objectives were to (1) determine the clinical information needs of nurses; (2) adapt an existing clinical information system (CLINT) to address their expressed needs; and (3) evaluate nurses’ use of and satisfaction with the enhanced system. Thirty-nine nurses on a medical teaching unit in a tertiary hospital in Canada participated in the project. A needs assessment influenced the design of the nursing interface to CLINT and the development of educational and participatory strategies to promote its use. Data were collected before, after, and throughout the implementation period. Qualitative and quantitative methods, including focus groups, online questionnaires, and automated usage data collection, were used to describe nurses’ use of and satisfaction with the system. The results suggested that peer mentorship, organizational support, and collaboration were the most effective strategies for promoting system use. The hospital information system (IHIS), Netscape, drug information and basic texts were the most frequently used databases. Nurses were satisfied with the system and reported progress in changing clinical practice. CLINT helped them to keep up with educational and professional development. In conclusions, nurses are willing to use information systems that are relevant to their needs and user friendly. There is, however, a paucity of resources available for evidence-based clinical decision making.

CD154 is necessary for mice to clear aCryptosporidium parvuminfection, but whether this ligand has to be expressed on T cells with specificity forC. parvumhas not been determined. We infected DO11.10 (ovalbumin specific) T-cell receptor transgenic mice that had been bred to a RAG−/−background withC. parvumand found that the infection was cleared within 6 weeks, while RAG−/−controls were unable to clearC. parvuminfection. Recovery was accompanied by an increase in the number of splenic T cells with the CD44highphenotype that characterizes memory cells. To determine whether aC. parvum-infected environment sufficed to activate transgenic T cells, we reconstitutedC. parvum-infected BALB/c SCID mice with DO11.10 RAG−/−splenocytes. Fecal excretion ofC. parvumantigen ceased in the 12 weeks following the adoptive transfer, unless the mice were also injected with tolerizing doses of ovalbumin. DO11.10 T cells were found in the submucosa ofC. parvum-infected, but not uninfected, BALB/c SCID hosts within 48 h of injection. The transferred DO11.10 T cells divided and acquired a CD44highmemory phenotype inC. parvum-infected, but not uninfected, recipients. DO11.10 splenocytes from CD154 knockout donors failed to clear aC. parvuminfection, confirming a requirement for CD154 in recovery. In vitro, the DO11.10 cells did not proliferate in response toC. parvumantigen, and a tBlast GenBank search revealed no matches between the ovalbumin peptide andC. parvumDNA sequences.C. parvum-infected SCID mice given RAG−/−CD8+T cells with aListeria-specific transgene did not recover fromC. parvuminfection. Our data suggest that antigen-nonspecific CD4+T-cell effector mechanisms in combination with the innate arm of the immune system are sufficient for the eradication ofC. parvuminfection.

The Bonnet monkey model of respiratory syncytial virus (RSV) infection may be a useful nonhuman primate model for studying RSV disease in humans because Bonnet monkeys can predictably be infected to obtain an orderly sequence of morphologic, cytologic, virologic, serologic, and inflammatory changes related to time of infection. Young feral Bonnet monkeys, Macaca radiata, were infected endotracheally with 106plaque-forming units (pfu) of the Long strain of RSV. RSV was recovered from the animals' lungs at necropsy on days 3, 5, and 7 with the highest viral titer obtained on day 3 (1.1 and 5.2 X 103pfu/g of tissue in the upper and lower lobes, respectively). RSV antigen and F protein mRNA were detected 3–5 days after infection in alveolar macrophages and in the epithelium of bronchi, terminal bronchioles, and alveoli. Histologic analysis of RSV-infected lungs at necropsy revealed progressive bronchiolar mucosal and submucosal inflammation, periarterial mononuclear interstitial inflammation, and focal alveolitis, with a maximal response at 7 days after infection. Cell counts in bronchoalveolar lavage (BAL) increased with time with neutrophils and macrophages predominating on day 3 (6.47 and 5.85 x 105/mm3, respectively) and lymphocytes predominating on day 9 (4.18 X 105/mm3). Serum-neutralizing antibody appeared on day 5 and IgG antibody to RSV was detected on day 9. This sequence of morphologic, cytologic, virologic, serologic, and inflammatory change following RSV infection creates a useful model in the study of experimentally induced RSV disease with a potential for testing future vaccine-induced alterations in RSV disease response.

Estimates of responder cell frequency (RCF) based on limiting dilution analyses are laborious, and alternative means to quantitate cell-mediated immunity to immunogens are desirable. It was shown that levels of interleukin (IL)-2 in the supernatant of varicella-zoster virus-stimulated blood lymphocytes from immune adults peaked at 48 h of culture and correlated partially with estimates of RCF (r = .74, P = .003). Levels of gamma-interferon, IL-4, and IL-10 increased through the first 4 days of culture, and gamma-interferon levels showed some correlation with peak IL-2 levels (r = .48, P = .03). Nevertheless, correlations between levels of these cytokines and RCF did not reach statistically significant levels.

Severe combined immunodeficiency disease (SCID) consists of a group of heterogeneous genetic disorders. The most severe phenotype, T-B- SCID, is inherited as an autosomal recessive trait and is characterized by a profound deficiency of both T cell and B cell immunity. There is a uniquely high frequency of T-B- SCID among Athabascan-speaking Native Americans (A-SCID). To localize the A-SCID gene, we conducted a genomewide search, using linkage analysis of approximately 300 microsatellite markers in 14 affected Athabascan-speaking Native American families. We obtained conclusive evidence for linkage of the A-SCID locus to markers on chromosome 10p. The maximum pairwise LOD scores 4.53 and 4.60 were obtained from two adjacent markers, D10S191 and D10S1653, respectively, at a recombination fraction of straight theta=.00. Recombination events placed the gene in an interval of approximately 6.5 cM flanked by D10S1664 and D10S674. Multipoint analysis positioned the gene for the A-SCID phenotype between D10S191 and D10S1653, with a peak LOD score of 5.10 at D10S191. Strong linkage disequilibrium was found in five linked markers spanning approximately 6.5 cM in the candidate region, suggesting a founder effect with an ancestral mutation that occurred sometime before 1300 A.D.

Background We examined whether socioeconomic status and rurality influenced outcomes after coronary surgery. Methods We identified 14,150 patients undergoing isolated coronary surgery. Socioeconomic and rurality data was obtained from the Australian Bureau of Statistics and linked to patients’ postcodes. Outcomes were compared between categories of socioeconomic disadvantage (highest versus lowest quintiles, n= 3150 vs. 2469) and rurality (major cities vs. remote, n=9598 vs. 839). Results Patients from socioeconomically-disadvantaged areas experienced a greater burden of cardiovascular risk factors including diabetes, obesity and current smoking. Thirty-day mortality (disadvantaged 1.6% vs. advantaged 1.6%, p>0.99) was similar between groups as was late survival (7 years: 83±0.9% vs. 84±1.0%, p=0.79). Those from major cities were less likely to undergo urgent surgery. There was similar 30-day mortality (major cities: 1.6% vs. remote: 1.5%, p=0.89). Patients from major cities experienced improved survival at seven years (84±0.5% vs. 79±2.0%, p=0.010). Propensity-analysis did not show socioeconomic status or rurality to be associated with late outcomes. Conclusion Patients presenting for coronary artery surgery from different socioeconomic and geographic backgrounds exhibit differences in their clinical profile. Patients from more rural and remote areas appear to experience poorer long-term survival, though this may be partially driven by the population's clinical profile.

Salmonella enterica serovars Derby and Mbandaka are isolated from different groups of livestock species in the UK. S. Derby is predominantly isolated from pigs and turkeys and S. Mbandaka is predominantly isolated from cattle and chickens. Alignment of the genome sequences of two isolates of each serovar led to the discovery of a new putative Salmonella pathogenicity island, SPI-23, in the chromosome sequence of S. Derby isolates. SPI-23 is 37 kb in length and contains 42 ORFs, ten of which are putative type III effector proteins. In this study we use porcine jejunum derived cell line IPEC-J2 and in vitro organ culture of porcine jejunum and colon, to characterise the association and invasion rates of S. Derby and S. Mbandaka, and tissue tropism of S. Derby respectively. We show that S. Derby invades and associates to an IPEC-J2 monolayer in significantly greater numbers than S. Mbandaka, and that S. Derby preferentially attaches to porcine jejunum over colon explants. We also show that nine genes across SPI-23 are up-regulated to a greater degree in the jejunum compared to the colon explants. Furthermore, we constructed a mutant of the highly up-regulated, pilV-like gene, potR, and find that it produces an excess of surface pili compared to the parent strain which form a strong agglutinating phenotype interfering with association and invasion of IPEC-J2 monolayers. We suggest that potR may play a role in tissue tropism.

We conducted a meta-analysis of Parkinson's disease genome-wide association studies using a common set of 7,893,274 variants across 13,708 cases and 95,282 controls. Twenty-six loci were identified as having genome-wide significant association; these and 6 additional previously reported loci were then tested in an independent set of 5,353 cases and 5,551 controls. Of the 32 tested SNPs, 24 replicated, including 6 newly identified loci. Conditional analyses within loci showed that four loci, including GBA, GAK-DGKQ, SNCA and the HLA region, contain a secondary independent risk variant. In total, we identified and replicated 28 independent risk variants for Parkinson's disease across 24 loci. Although the effect of each individual locus was small, risk profile analysis showed substantial cumulative risk in a comparison of the highest and lowest quintiles of genetic risk (odds ratio (OR) = 3.31, 95% confidence interval (CI) = 2.55-4.30; P = 2 × 10-16). We also show six risk loci associated with proximal gene expression or DNA methylation. © 2014 Nature America, Inc. All rights reserved.

Selective T cell deficiency is a rare immune deficiency characterized by the absence of CD8 + T lymphocytes and depressed/absent T cell function. This syndrome has been associated with mutations in the gene for ZAP-70, a tyrosine kinase that has profound effects on signaling via the T cell receptor. In this paper we describe a patient with selective T cell deficiency and certain phenotypic features that are unique among the small number of patients described. The patient had virtually absent T cell function, hypogammaglobulinemia, and no response to vaccination. The T lymphocytes failed to respond to mitogenic stimuli, even in the presence of exogenous interleukin 2. Similar to other patients with this disorder, the T cells were capable of proliferating when stimulated by pharmacologic agents such as phorbol ester and ionomycin. While peripheral blood T cells had limited capability to increase cytosolic Ca 2+ levels in response to mitogenic stimulation, thymocytes responded to a large panel of antibodies and mitogens. This report broadens the spectrum of clinical presentations associated with selective T cell deficiency and, for the first time, compares the responses of both peripheral T cells and thymocytes. The data support the concept that the defect in signal transduction resulting from the absence of ZAP-70 is primarily manifested following export of T lymphocytes from the thymus and that selection of CD8-positive T cells is dependent on the presence of ZAP-70.

Several mutations involving the fibroblast growth factor receptor (FGFR) gene family have been identified in association with phenotypically distinct forms of craniosynostosis. One such point mutation, resulting in the substitution of proline by arginine in a critical region of the linker region between the first and second immunoglobulin-like domains, is associated with highly specific phenotypic consequences in that mutation at this point in FGFR1 results in Pfeiffer syndrome and analogous mutation in FGFR2 results in Apert syndrome. We now show that a much more variable clinical presentation accompanies analogous mutation in the FGFR3 gene. Specifically, mental retardation, apparently unrelated to the management of the craniosynostosis, appears to be a variable clinical consequence of this FGFR3 mutation.

We report a case of thrombotic thrombocytopenic purpura in a woman post mitral valve repair who presented with unexplained thrombocytopenia, intermittent fever, acute renal failure and severe digital ischaemia. The diagnosis of thrombotic thrombocytopenic purpura was confirmed after exclusion of many of the aetiological factors of postoperative digital ischaemia, a positive haemolytic screen and blood film examination which showed schistocytes and fragmented red cells. Plasma ADAMTS-13 activity measured by enzyme-linked immunosorbent assay was reduced. Treatment of thrombotic thrombocytopenic purpura with exchange plasmapheresis and methylprednisolone was of paramount importance and the patient was discharged home on day 30 with complete recovery of haematological, neurological and renal function. In order to increase the awareness of this rare multisystem process, we report our experience in managing this life-threatening condition. Our discussion covers the diagnostic challenges, theories on aetiology, pathogenesis and treatment of this condition in the context of cardiac surgery.

The use of the radial artery as a second arterial graft during coronary surgery has grown in popularity due to high patency and low harvest site complication rates. We sought to assess whether higher risk patients derive prognostic benefit.From 2001 to 2009, 11,388 patients underwent isolated primary multivessel coronary surgery. We identified a higher risk subgroup (n = 2581) according to emergent status, coronary instability, low ejection fraction and/or aortic counterpulsation. Among these, 1832 (71%) received at least one radial artery graft in addition to a left internal thoracic artery (LITA). The remaining 749 (29%) received LITA and veins only.Patients not receiving a radial artery were more likely to be elderly, female, have poor left ventricular function or be of emergent status. These patients experienced higher unadjusted 30-day mortality (radial: 2% vs vein: 8%, P0.0001) with lower unadjusted 7-year survival (80 ± 1.3 vs 67 ± 2.4%, P0.0001). Subsequently, 515 patients in the radial group were propensity-matched to 515 receiving LITA + veins (mean logistic EuroSCORE, radial: 11.6 ± 9.7% vs vein: 11.6 ± 10.3%, P = 0.99). At 30 days, there were comparable rates of mortality (radial: 4% vs vein: 3%, P0.99), stroke (1 vs 1%, P0.99), myocardial infarction (1 vs 2%, P = 0.79), and any morbidity/mortality (34 vs 35%, P = 0.95). At 7 years, survival rates between the radial and vein groups were similar (radial: 75 ± 2.6% vs vein: 74 ± 2.9%, P = 0.65).Patients with the greatest coronary instability, urgency of surgery or impairment of ventricular function are not disadvantaged in early outcomes or mid-term survival by the use of only a single arterial graft.

The live attenuated varicella vaccine offers some hope that the frequency or severity of herpes zoster might be reduced. Universal immunization with this vaccine should result in less latent varicella-zoster virus in dorsal root and cranial nerve ganglia than that which occurs following varicella. Moreover, the vaccine virus is not well adapted for growth in human cells at normal body temperature. Thus, reduced virus for reactivation, and less robust replication, may lessen the problem of herpes zoster in vaccinees. For those individuals who have already had varicella, the risk of herpes zoster is closely related to the loss of varicella-zoster virus cell-mediated immune responses, which decline with aging (or immune suppression). In aging individuals these immune responses can be enhanced by booster immunization with the varicella vaccine, suggesting that a vaccine to prevent herpes zoster is feasible.

Subjectsor = 55 years of age were immunized with attenuated varicella zoster virus (VZV) vaccine (live) or with the same vaccine, which had been heated to 56 degrees C for 7 days (killed). The ability of subjects' blood lymphocytes to lyse target cells infected with VZV was determined before and 3 months after immunization using autologous Epstein-Barr virus (EBV) lymphoblasts as targets for human leukocyte antigen (HLA) class I restricted cytotoxicity and human fibroblasts as targets for unrestricted (natural killer [NK]) cytotoxicity. The live vaccine recipients showed an increase in their class I-restricted lysis of targets compared with the recipients of the killed vaccine. The two populations showed equivalent increase in their NK-dependent lysis of fibroblast targets. The results support the view that both the live and killed vaccines stimulate cytotoxicity by VZV-specific lymphocytes but that the live vaccine stimulates relatively more class I-restricted killing.

Background\ud Despite the frequent isolation of Salmonella enterica sub. enterica serovars Derby and Mbandaka from livestock in the UK and USA little is known about the biological processes maintaining their prevalence. Statistics for Salmonella isolations from livestock production in the UK show that S. Derby is most commonly associated with pigs and turkeys and S. Mbandaka with cattle and chickens. Here we compare the first sequenced genomes of S. Derby and S. Mbandaka as a basis for further analysis of the potential host adaptations that contribute to their distinct host species distributions. \ud \ud Results\ud Comparative functional genomics using the RAST annotation system showed that predominantly mechanisms that relate to metabolite utilisation, in vivo and ex vivo persistence and pathogenesis distinguish S. Derby from S. Mbandaka. Alignment of the genome nucleotide sequences of S. Derby D1 and D2 and S. Mbandaka M1 and M2 with Salmonella pathogenicity islands (SPI) identified unique complements of genes associated with host adaptation. We also describe a new genomic island with a putative role in pathogenesis, SPI-23. SPI-23 is present in several S. enterica serovars, including S. Agona, S. Dublin and S. Gallinarum, it is absent in its entirety from S. Mbandaka. \ud \ud Conclusions\ud We discovered a new 37 Kb genomic island, SPI-23, in the chromosome sequence of S. Derby, encoding 42 ORFS, ten of which are putative TTSS effector proteins. We infer from full-genome synonymous SNP analysis that these two serovars diverged, between 182kya and 625kya coinciding with the divergence of domestic pigs. The differences between the genomes of these serovars suggest they have been exposed to different stresses including, phage, transposons and prolonged externalisation. The two serovars possess distinct complements of metabolic genes; many of which cluster into pathways for catabolism of carbon sources.

Transgenic and knockout mice usefully model the mechanisms that result in the clearance of Cryptosporidium parvum from the gut. CD4+ cells, cells expressing MHC class II, and CD154/CD40 interactions are essential. Unexpectedly, AND RAG-/- and DO11.10 RAG-/- mice with single specificities of T cells successfully clear Cryptosporidium infection. Clearance is accompanied by activation of CD4+ cells in the MLN. The ability of T cells bearing receptors for apparently irrelevant and non-cross reactive antigens to activate and to clear infection is surprising. The requirement for class II MHC expression for Cryptosporidium clearance raises the alternative possibilities that (a) class II MHC is required to present a peptide that is loaded as a consequence of infection or (b) that the cytokine environment engendered by a Cryptosporidium infection allows affinity for self MHC to activate naive T cells. In order to test the hypothesis that peptide loading is necessary, we used A betaE alpha-/-Ii-/- mice that express a hybrid IA-IE MHC molecule. They also carry a transgene that makes an E alpha peptide while disruption of their invariant chain blocks the loading of a foreign peptide on to their MHC class II molecules. After oral gavage, the course of infection was followed by ELISA. CD4+ cells in the MLN of these mice were activated to express CD69 and the infection was cleared. We conclude that the loading of a Cryptosporidium or other infection-dependent peptide onto the MHC class II molecules of APCs is not necessary for clearance of Cryptosporidium. Instead the TcR affinity for self-MHC must suffice for T cell activation in the cytokine environment resulting from infection.

Objectives A single or dual-dose strategy for myocardial protection is attractive in long operations, in avoiding the need to interrupt the procedure to re-administer cardioplegia. We hypothesized that a single administration of Bretschneider histidine-tryptophan-ketoglutarate (HTK) crystalloid solution (Custodiol) offers myocardial protection comparable with repeated tepid blood cardioplegia. Methods We reviewed a prospectively compiled single-centre database containing all adult cardiac procedures performed from January 2005 to January 2011. Preoperative demographic and investigative data, operative variables and postoperative (30-day) mortality and morbidity were compared between the Custodiol and blood cardioplegia groups. The study primary endpoints were 30-day mortality, return to the operating theatre, myocardial infarction, stroke, postoperative requirement for an intra-aortic balloon pump, new renal failure, prolonged ventilation and re-admission to hospital within 30 days. Propensity score matching was performed to correct for any bias that may have been associated with the usage of Custodiol. Results A total of 1900 cardiac surgical procedures were identified of which 126 (7%) utilized Custodiol and 1774 (93%) used blood cardioplegia as the primary cardioplegic agent. After propensity-score matching, we were able to match 71 Custodiol cases one-to-one to those receiving blood cardioplegia. There were no statistically significant differences noted for any of the endpoints studied after propensity-score matching. In particular, the proportion of mortality (blood cardioplegia: 1 vs Custodiol 4%, P = 0.63) any mortality/morbidity (blood cardioplegia: 35 vs Custodiol: 39% P = 0.46) was similar between the groups. Conclusions The use of Custodiol is convenient, simple and at least as safe as tepid blood cardioplegia for myocardial protection in complex cardiac operations. A randomized prospective comparison of myocardial protection strategies is warranted.

SUMMARY Human blood lymphocytes were stimulated with varicella zoster virus (VZV) antigen in limiting dilution cultures and the amounts of interferon-gamma (IFN-γ) and IL-4 measured in the supernatants. The results indicate that up to 85% of proliferating cells of young adults produce IFN-γ and up to 10% make IL-4. At limiting dilution, few if any wells were positive for both IFN-γ and IL-4. The amount of IFN-γ per well increased in the presence of antibody to IL-4, but anti-IFN-γ did not increase IL-4 production. The frequency of wells containing IFN-γ was lower in subjects < 19 or > 55 years of age, and the amounts of IFN-γ in positive wells was significantly lower in cultures of the older subjects' lymphocytes. The frequency of IL-4-making cells did not fall significantly with age. The data suggest that the age-related decline in the frequency of blood T cells which responds to VZV affects mainly the cells with a Th1-like cytokine phenotype.

Current criteria for the histological diagnosis of Creutzfeldt-Jakob disease (CJD) include features such as spongiform change, neuronal loss and reactive gliosis which are shared to a varying extent with other neuro-degenerative disorders. Reliable visualization of prion protein (PrP) has substantial potential value in diagnostic practice and as a research tool, since accumulation of the disease-associated isoform of this protein is apparently specific for spongiform encephalopathies. A number of antisera against PrP have previously been employed in conjunction with a range of pre-treatments designed to optimize the specificity of immunostaining; such varied usage makes the comparison and interpretation of results difficult. This study was undertaken to identify optimal combinations of each of three PrP antisera and five pre-treatments designed to specifically demonstrate disease-specific PrP in a series of seven CJD cases, six cases of Alzheimer-type dementia and six non-demented control cases. Specific staining of amyloid plaques, spongiform neuropil, neurons and, occasionally, astrocytes was achieved in CJD cases. Alzheimer and control cases were unstained. Use of formic acid with guanidine thiocyanate, and hydrolytic autoclaving with IB3 and SP30 antisera proved most effective and can be recommended for future immunocytochemical studies. PrP immunocytochemistry revealed a greater extent of subcortical neural involvement than routine histological techniques in CJD; the relationship between classical neuropathology in CJD and PrP accumulation as revealed by immunocytochemistry is not clear cut and requires further investigation. These findings may help to broaden our understanding of human spongiform encephalopathies, and have implications for diagnostic practices in neuropathology.

A 10-day-old infant with stridor was found to have herpes simplex virus type 2 infection of the larynx. The infant's poor clinical response to both acyclovir and foscarnet prompted extensive clinical and virologic evaluations, which revealed acyclovir-resistant herpes simplex virus.

Limiting dilution cultures have been used to estimate the frequency of T cells which respond to antigen stimulation in vitro, but nothing is known of the reproducibility of this assay when applied to human blood. We developed a simplified form of limiting dilution culture in which blood mononuclear cells were diluted in round bottom 96-well plates and cultured for 10 days. The assay was used to estimate the frequency of blood mononuclear cells proliferating in response to varicella-zoster virus antigen. 250 subjects aged 25-87 years were studied: 95 of these subjects had annual measurements repeated over a 4 year period. The coefficients of variation for intra-assay replicates was 5%, and for inter-assay comparisons was 17.6% and 26% for tests at a 1 or 3 month interval respectively. For subjects studied at 1 year intervals the coefficient of variation was 42% with the total variability equally distributed between the variation between subjects and the variation within subjects. These data provide the first quantitative data to validate limiting dilution cultures for the long term in vitro measurement of human T cell responses.

Guinea pigs were immunized with glycoprotein gD-2t in SAF-m or saline, then challenged with herpes simplex virus, type 2 (HSV-2). Animals given gD-2t in SAF-m had higher anti-gD-2t antibodies, fewer and less severe vaginal lesions, and decreased ganglionic latency compared to animals given gD-2t in saline. Leucocytes from animals vaccinated with gD-2t in SAF-m had greater proliferative responses to gD-2t in vitro than cells from control animals. MHC II-restricted, gD-2t-specific cytotoxic T cells were induced in guinea pigs vaccinated with gD-2t in SAF-m. Thus, immunization of guinea pigs with gD-2t in SAF-m markedly reduced the incidence and severity of primary HSV-2 by eliciting both humoral and cell-mediated responses.

The Radial Artery Patency and Clinical Outcomes Study (RAPCO) was devised and implemented in Melbourne in order to establish the appropriate place of the radial artery in the hierarchy of conduits available to the modern coronary bypass surgeon. Designed as a biological comparison with minimisation of other confounding variables, it compares this free arterial graft with the right internal thoracic artery and saphenous vein, with all conduits used in an identical fashion in two parallel cohorts of different age ranges. Enrolment was completed in 2004 and 10-year follow-up is in progress, with mean duration of about seven years at present. The midterm clinical and angiographic results to date are reviewed here, but definitive conclusions will not be possible until full completion angiographic data is available. The trial data provides a number of potential substudies of conduits, risk factors for failure and the natural history of treated coronary disease.

Aortic allograft fibrocalcification and valvular insufficiency have been observed in patients less than 3 years of age at initial replacement of the left ventricular outflow tract. From June 1985 through May 1992, 47 children have undergone aortic root replacement with cryopreserved aortic valve allografts. Thirty-three children were 3 years of age or older and 14 were less than 3 years of age at operation. In the older patient group, there were three (9%) hospital deaths and one child underwent cardiac transplantation 30 hours after aortic root replacement because of left ventricular failure. Clinical follow-up of the 29 surviving older children is from 4 months to 6.6 years (mean 3.0 years). One patient was lost to follow-up. Two children (7%) have required reoperation, but primary allograft degeneration was not observed. In the younger patient group, there were three (21%) hospital deaths. Follow-up ranged from 2.5 months to 4.7 years (mean 2.3 years). Among 11 operative survivors, one late death resulted from a pulmonary embolus. Seven of 10 (70%) remaining allograft recipients had progressive allograft calcification or insufficiency. Six of them have required reoperation to explant the allograft, and one child is currently receiving cyclosporine therapy with the original valve allograft. The cause of allograft failure is possibly immunologic. The prevalence of early aortic valve allograft degeneration has prompted the consideration of nonviable allografts or xenografts, pulmonary autografts, or minimal immunosuppression as alternatives when left ventricular outflow tract reconstruction is necessary in children less than 3 years of age.

The distribution of ubiquitin was studied by immunocytochemistry in eight cases of human spongiform encephalopathy and compared with the findings in seven age- and sex-matched cases of Alzheimer's disease and six non-demented control cases. The results were also compared with the immunocytochemical distribution of prion protein and the lysosomal aspartic protease cathepsin D. In the human spongiform encephalopathies, ubiquitin immunoreactivity was found in a punctate distribution at the periphery of prion protein amyloid plaques and in a finely granular pattern in the neuropil around and within areas of spongiform change. Cortical nerve cells contained scanty ubiquitinated dot-like inclusions, and occasional microglia around the areas of spongiform change also gave a positive staining reaction for ubiquitin, as did multiple irregular thread-like structures in the neuropil and white matter. The ubiquitin-containing structures at the plaque periphery in human spongiform encephalopathies resemble the neuritic processes at the periphery of the senile plaque in Alzheimer's disease. The granular positivity for ubiquitin associated with areas of spongiform change closely resembles the pattern of immunostaining seen with the antibodies to the prion protein and cathepsin D, consistent with the reported accumulation of ubiquitinated proteins and prion protein in lysosomes in the murine scrapie model. Further studies are required to investigate the role of lysosomes in this group of disorders, and to study the localization of other cell stress proteins and prion protein in spongiform encephalopathies.

HSV is a successful human pathogen that causes severe infections in immunocompromised adults and newborn humans. The possibility that HSV might evade host immune responses by interfering with accessory cell function was investigated in vitro using newborns' monocytes adhered to plastic. These cells lost their ability to present staphylococcal enterotoxin B to resting T cells in a stimulatory form after overnight culture with HSV. The interference with T cell proliferation required live virus and was abolished by heat- or UV inactivation. The T cell proliferative response was restored by the addition of IL-2. Furthermore, HSV-precultured monocytes had a reduced production of IL-1 alpha and TNF-beta after phorbol-ionomycin stimulation. Wild-type HSV and a HSV mutant lacking the primary virion host shutoff protein both interfered with IL-1 synthesis and presentation of staphylococcal enterotoxin B for T cell proliferation. These results suggest that HSV can interfere with the provision by human monocytes of costimulator factors that are essential for T cell stimulation. This effect of HSV may be due to secondary shutoff mechanisms that decrease host protein synthesis or secretion after HSV infection.

Objective: Mitral valve surgery may be regarded as less favourable for training, due to greater mortality risk, technical complexity, and difficultyforthesupervisortoobserve.We examinedthisperceptionbyreviewingamulticentreexperience.Methods:We analyseda multicentre database over a 7-year period containing 2216 isolated and combined mitral procedures. Of these, 2048 were performed by consultants and 168 by trainees (92% vs 8%) of varying seniority. Preoperative characteristics, early postoperative outcomes and 6-year survival were compared between groups. Propensity-score matching was performed to correct for group differences. Results: Trainees were less likely to operate on patients, who had previously undergone coronary surgery (consultant 4.3% vs trainee 1.2%, p = 0.043) and those with moderate to severe mitral regurgitation (86% vs 81%, p = 0.012). There were no other statistically significant differences in preoperative variables, such as urgency, endocarditis and left-ventricular dysfunction. There were similar rates of mitral valve repair (48% vs 51%, p = 0.48). Trainees were more likely to operate on rheumatic valve pathology (20% vs 28%, p = 0.012). Intra-operatively, trainees had longer aortic cross-clamp times (119 52 vs 136 50 min, p = 0.0001). At 30 days, mortality was comparable (4.5% vs 3.6%, p = 0.56) with a trend towards higher any mortality/morbidity in consultant procedures (33% vs 26%, p = 0.059). At 6 years, survival was similar (79 1.4% vs 78 4.0%, p = 0.73). After derivation of 142 propensity-score-matched patient pairs, trainees cases still experienced longer cross-clamp times (121 58 vs 137 52 min, p = 0.023), but there was similar 30-day mortality (4.2% vs 3.5%, p > 0.99) and any mortality/morbidity (28% vs 24%, p = 0.52). Six-year survival between matched pairs was also similar (74 7.2% vs 80 4.4%, p = 0.64). Trainee status did not predict early or late adverse events after multivariate Cox regression with and without propensity-score adjustment. Conclusions: Trainee outcomes are not inferior even when corrected for risk. This suggests that excellent operative training and supervision can be achieved in mitral valve surgery. # 2011 European Association for Cardio-Thoracic Surgery. Published by Elsevier B.V. All rights reserved.

Cytotoxic, proliferative, and serum IgG antibody responses to varicella-zoster virus (VZV) were measured monthly in 29 young adults for 3 months following primary immunization with the Oka strain attenuated VZV vaccine. No subjects had lymphocytes stimulated to VZV-specific cytotoxicity at study entry although lymphocytes from 2 subjects proliferated when stimulated by VZV antigen. The percentages of subjects with positive major histocompatibility complex class II-restricted cytotoxic responses at 1, 2, and 3 months after immunization were 54%, 69%, and 66%, respectively. Correlations between cytotoxic, proliferative, and antibody responses were highest in the first month after immunization; lower but still statistically significant after the second month; and not apparent by the third month. The data suggest that antibody and cell-mediated immune responses to VZV immunization develop in parallel after immunization, but these effector mechanisms are independently regulated by 3 months after immunization.

The Oka strain live attenuated varicella-zoster virus (VZV) vaccine was administered subcutaneously to 202 VZV-immune individuals who were 55 to greater than 87 years old. The dose administered varied from 1100 to 12,000 pfu. One cohort received 3000 pfu with a 3000 pfu booster 3 months later. The vaccine was well tolerated. VZV-specific immunologic responses were evaluated over a 24-month period. The mean anti-VZV antibody level was significantly increased for 12 months after vaccination. Interferon-gamma production in vitro by peripheral blood mononuclear cells (PBMC) of vaccinees was also increased for 6 months after vaccination. Most significantly, VZV-specific proliferating T cells in PBMC of vaccinees were increased in frequency from 1 in 68,000 to 1 in 40,000. This vaccine-enhanced frequency of VZV-responding T cells is similar to the frequency observed in 35- to 40-year-old adults. Dose and age of the vaccinees did not significantly influence the magnitude of the mean cell-mediated immune response. The data indicate that VZV immunity in the elderly can be boosted by active immunization. If the increased incidence of herpes zoster that accompanies aging results from the natural waning of immunity, active immunization may prevent or attenuate zoster in the elderly.