Mutations in LOXHD1 gene can cause auditory neuropathy spectrum disorder.

Objectives: The aim of this paper was to study the auditory phenotype of three related children with sensorineural hearing loss (2 sisters and their cousin) following genetic analysis revealing mutations in LOXHD1. Methods: Genetic testing was conducted on three related children. They were assessed with a standard clinical test battery including distortion otoacoustic emissions, auditory brainstem responses and audiometry. Results: We identified heterozygous variants in LOXHD1 in a family of Irish/German and Italian/Irish ancestry with autosomal recessive auditory neuropathy spectrum disorder (ANSD). Mutations in LOXHD1 (MIM #613072) have been linked to an autosomal recessive nonsyndromic hearing loss (DFNB77), mapped to the locus 18q12-q21. All three subjects had evidence of some, albeit few, functioning cochlear hair cells as revealed by the presence of a cochlear microphonic and/or partial otoacoustic emissions early in life. Conclusion: To our knowledge, this is the first association between LOXHD1 mutations and ANSD in two patients who have been successfully managed with cochlear implants.

Minor differences in perceived sheepmeat eating quality scores of Australian, Chinese and American consumers.

Understanding consumer sensory perceptions of sheepmeat is essential for consumer satisfaction post-purchase. Meat Standards Australia (MSA) sensory protocols have been effectively utilised in beef for international consumers however, to date sheepmeat testing is largely limited to Australian consumers. This study measured the sensory responses (liking of odour, tenderness, juiciness, liking of flavour, and overall liking) of 2160 untrained American, Australian and Chinese consumers to grilled longissimus lumborum (LL) and semimembranosus (SM) muscles from 164 lambs and 168 yearlings. Across countries there was no difference in juiciness or overall liking sensory scores. American consumers scored tenderness, flavour and odour slightly higher than Australian consumers, and Chinese consumer scores were lowest. Consistently for all countries, sensory scores were greatest in the LL muscle, in lambs compared to yearlings particularly for the LL, and Merino sired and female lambs. These results indicate that cultural background has minimal impact on sensory perceptions of sheepmeat, and provides valuable information for future eating quality prediction models.

The Prevalence of Pseudomonas Aeruginosa Infection Over a Ten-Year Period in Children with Cystic Fibrosis

Background Pseudomonas aeruginosa (PA) infection is associated with an increased morbidity and adverse prognosis in children with Cystic Fibrosis(CF). The aim of the study was to evaluate the prevalence and characteristics of PA over a ten year period at a single paediatric tertiary referral centre in Ireland. Methods This was a retrospective cross-sectional study. Patient's case notes, microbiology laboratory results and CF Registry of Ireland(CFRI) data were used to collect the data. Results The overall chronic PA infection prevalence was 28.1%(45/160) in 2004 and 21.3%(35/164) in 2014. In 2004, 54/160(33.8%) patients were never infected with PA, 27/160(16.9%) were free for 12 months and 34/160(21.3%) were intermittently infected. In 2014; 80/164(49%) patients, 38/164(23.2%) and 11/164(6.7%) were never infected, free for 12 months and intermittently infected respectively. Conclusion There has been a decline in the overall prevalence of PA infection and a change in the pattern of prevalence over the last decade at our Centre.

Factors affecting lamb eating quality and the potential for their integration into an MSA sheepmeat grading model.

Major efforts in the sheep industry to control eating quality have resulted in reduced product variability. Yet inconsistent eating quality for consumers remains, due to a degree of inaccurate representation of cut quality. Eating quality defined through a complex interplay of different factors can be predicted for individual cuts, and Meat Standards Australia (MSA) grading schemes have been developed to achieve these defined quality outcomes. This review outlines the justifications to refine the current sheepmeat MSA pathways system to transition into a cuts-based prediction model and details some of the factors affecting sheepmeat eating quality as key factors under consideration into the new model. The development of the new sheepmeat MSA prediction model will allow for more efficient carcass sorting to underpin a value based payment system throughout the supply chain. However it requires the inclusion of individual carcass yield and eating quality measurements (i.e. IMF). Furthermore, the adoption challenges internationally of an MSA like model are discussed.

Post-zygotic genomic changes in glutamate and dopamine pathway genes may explain discordance of monozygotic twins for schizophrenia.

BACKGROUND: Monozygotic twins are valuable in assessing the genetic vs environmental contribution to diseases. In the era of complete genome sequences, they allow identification of mutational mechanisms and specific genes and pathways that offer predisposition to the development of complex diseases including schizophrenia. METHODS: We sequenced the complete genomes of two pairs of monozygotic twins discordant for schizophrenia (MZD), including one representing a family tetrad. The family specific complete sequences have allowed identification of post zygotic mutations between MZD genomes. It allows identification of affected genes including relevant network and pathways that may account for the diseased state in pair specific patient. RESULTS: We found multiple twin specific sequence differences between co-twins that included small nucleotides [single nucleotide variants (SNV), small indels and block substitutions], copy number variations (CNVs) and structural variations. The genes affected by these changes belonged to a number of canonical pathways, the most prominent ones are implicated in schizophrenia and related disorders. Although these changes were found in both twins, they were more frequent in the affected twin in both pairs. Two specific pathway defects, glutamate receptor signaling and dopamine feedback in cAMP signaling pathways, were uniquely affected in the two patients representing two unrelated families. CONCLUSIONS: We have identified genome-wide post zygotic mutations in two MZD pairs affected with schizophrenia. It has allowed us to use the threshold model and propose the most likely cause of this disease in the two patients studied. The results support the proposition that each schizophrenia patient may be unique and heterogeneous somatic de novo events may contribute to schizophrenia threshold and discordance of the disease in monozygotic twins.

Long-term prognosis for 1-year relapse-free survivors of CD34+ cell-selected allogeneic hematopoietic stem cell transplantation: a landmark analysis.

CD34+ cell selection significantly improves GvHD-free survival in allogeneic hematopoietic stem cell transplantation (allo-HSCT). However, specific information regarding long-term prognosis and risk factors for late mortality after CD34+ cell-selected allo-HSCT is lacking. We conducted a single-center landmark analysis in 276 patients alive without relapse 1 year after CD34+ cell-selected allo-HSCT for AML (n=164), ALL (n=33) or myelodysplastic syndrome (n=79). At 5 years' follow-up after the 1-year landmark (range 0.03-13 years), estimated relapse-free survival (RFS) was 73% and overall survival (OS) 76%. The 5-year cumulative incidence of relapse and non-relapse mortality (NRM) were 11% and 16%, respectively. In multivariate analysis, Hematopoietic Cell Transplantation Comorbidity Index score⩾3 correlated with marginally worse RFS (hazard ratio (HR) 1.78, 95% confidence interval (CI) 0.97-3.28, P=0.06) and significantly worse OS (HR 2.53, 95% CI 1.26-5.08, P=0.004). Despite only 24% of patients with acute GvHD within 1 year, this also significantly correlated with worse RFS and OS, with increasing grades of acute GvHD associating with increasingly poorer survival on multivariate analysis (P<0.0001). Of 63 deaths after the landmark, GvHD accounted for 27% of deaths and was the most common cause of late mortality, followed by relapse and infection. Although prognosis is excellent for patients alive without relapse 1 year after CD34+ cell-selected allo-HSCT, risks of late relapse and NRM persist, particularly due to GvHD.

Probing the causes of thermal hysteresis using tunable Nagg micelles with linear and brush-like thermoresponsive coronas.

Self-assembled thermoresponsive polymers in aqueous solution have great potential as smart, switchable materials for use in biomedical applications. In recent years, attention has turned to the reversibility of these polymers' thermal transitions, which has led to debate over what factors influence discrepancies in the transition temperature when heating the system compared to the temperature obtained when cooling the system, known as the thermal hysteresis. Herein, we synthesize micelles with tunable aggregation numbers (Nagg) whose cores contain poly(n-butyl acrylate-co-N,N-dimethylacrylamide) (p(nBA-co-DMA)) and four different thermoresponsive corona blocks, namely poly(N-isopropylacrylamide) (pNIPAM), poly(N,N-diethylacrylamide) (pDEAm), poly(diethylene glycol monomethyl ether methacrylate) (pDEGMA) and poly(oligo(ethylene glycol) monomethyl ether methacrylate) (pOEGMA). By studying their thermoresponsive behavior, we elucidate the effects of changing numerous important characteristics both in the thermoresponsive chain chemistry and architecture, and in the structure of their self-assemblies. Our findings demonstrate large deviations in the reversibility between the self-assemblies and the corresponding thermoresponsive homopolymers; specifically we find that micelles whose corona consist of polymers with a brush-like architecture (pDEGMA and pOEGMA) exhibit irreversible phase transitions at a critical chain density. These results lead to a deeper understanding of stimuli-responsive self-assemblies and demonstrate the potential of tunable Nagg micelles for uncovering structure-property relationships in responsive polymer systems.

Fluorescent polymeric nanovehicles for neural stem cell modulation.

Nanomaterials are emerging as strong candidates for applications in drug delivery and offer an alternative platform to modulate the differentiation and activity of neural stem cells. Herein we report the synthesis and characterization of two different classes of polymeric nanoparticles: N-isopropylacrylamide-based thermoresponsive nanogels RM1 and P(TEGA)-b-P(d,lLA)2 nano-micelles RM2. We covalently linked the nanoparticles with fluorescent tags and demonstrate their ability to be internalized and tracked in neural stem cells from the postnatal subventricular zone, without affecting their proliferation, multipotency and differentiation characteristics up to 150 µg ml-1. The difference in chemical structure of RM1 and RM2 does not appear to impact toxicity however it influences the loading capacity. Nanogels RM1 loaded with retinoic acid improve solubility of the drug which is released at 37 °C, resulting in an increase in the number of neurons, comparable to what can be obtained with a solution of the free drug solubilised with a small percentage of DMSO.

Soluble and membrane-bound interleukin (IL)-15 Rα/IL-15 complexes mediate proliferation of high-avidity central memory CD8+ T cells for adoptive immunotherapy of cancer and infections.

The lack of persistence of infused T cells is a principal limitation of adoptive immunotherapy in man. Interleukin (IL)-15 can sustain memory T cell expansion when presented in complex with IL-15Rα (15Rα/15). We developed a novel in-vitro system for generation of stable 15Rα/15 complexes. Immunologically quantifiable amounts of IL-15 were obtained when both IL-15Rα and IL-15 genes were co-transduced in NIH 3T3 fibroblast-based artificial antigen-presenting cells expressing human leucocyte antigen (HLA) A:0201, ß2 microglobulin, CD80, CD58 and CD54 [A2-artificial antigen presenting cell (AAPC)] and a murine pro-B cell line (Baf-3) (A2-AAPC15Rα/15 and Baf-315Rα/15 ). Transduction of cells with IL-15 alone resulted in only transient expression of IL-15, with minimal amounts of immunologically detectable IL-15. In comparison, cells transduced with IL-15Rα alone (A2-AAPCRα ) demonstrated stable expression of IL-15Rα; however, when loaded with soluble IL-15 (sIL-15), these cells sequestered 15Rα/15 intracellularly and also demonstrated minimal amounts of IL-15. Human T cells stimulated in vitro against a viral antigen (CMVpp65) in the presence of 15Rα/15 generated superior yields of high-avidity CMVpp65 epitope-specific T cells [cytomegalovirus-cytotoxic T lymphocytes (CMV-CTLs)] responding to ≤ 10- 13 M peptide concentrations, and lysing targets cells at lower effector : target ratios (1 : 10 and 1 : 100), where sIL-15, sIL-2 or sIL-7 CMV-CTLs demonstrated minimal or no activity. Both soluble and surface presented 15Rα/15, but not sIL-15, sustained in-vitro expansion of CD62L+ and CCR7+ central memory phenotype CMV-CTLs (TCM ). 15Rα/15 complexes represent a potent adjuvant for augmenting the efficacy of adoptive immunotherapy. Such cell-bound or soluble 15Rα/15 complexes could be developed for use in combination immunotherapy approaches.

Virus-specific T-cell banks for 'off the shelf' adoptive therapy of refractory infections.

Adoptive immunotherapy with transplant donor-derived virus-specific T cells has emerged as a potentially curative approach for the treatment of drug-refractory EBV+lymphomas as well as CMV and adenovirus infections complicating allogeneic hematopoietic cell transplants. Adoptive transfer of HLA partially matched virus-specific T cells from healthy third party donors has also shown promise in the treatment of these conditions, with disease response rates of 50-76% and strikingly low incidences of toxicity or GVHD recorded in initial trials. In this review, we examine the reported experience with transplant donor and third party donor-derived virus-specific T cells, identifying characteristics of the viral pathogen, the T cells administered and the diseased host that contribute to treatment response or failure. We also describe the characteristics of virus-specific T-cell lines in our center's bank and the frequency with which in vitro culture promotes expansion of immunodominant T cells specific for epitopes that are presented by a limited array of prevalent HLA alleles, which facilitates their broad applicability for treatment.

Late effects in patients with Fanconi anemia following allogeneic hematopoietic stem cell transplantation from alternative donors.

Hematopoietic stem cell transplantation (HSCT) is curative for hematological manifestations of Fanconi anemia (FA). We performed a retrospective analysis of 22 patients with FA and aplastic anemia, myelodysplastic syndrome or acute myelogenous leukemia who underwent a HSCT at Memorial Sloan Kettering Cancer Center and survived at least 1 year post HSCT. Patients underwent either a TBI- (N=18) or busulfan- (N=4) based cytoreduction followed by T-cell-depleted transplants from alternative donors. Twenty patients were alive at time of the study with a 5- and 10-year overall survival of 100 and 84% and no evidence of chronic GvHD. Among the 18 patients receiving a TBI-based regimen, 11 (61%) had persistent hemochromatosis, 4 (22%) developed hypothyroidism, 7 (39%) had insulin resistance and 5 (27%) developed hypertriglyceridemia after transplant. Eleven of 16 evaluable patients (68%), receiving TBI, developed gonadal dysfunction. Two patients who received a TBI-based regimen died of squamous cell carcinoma. One patient developed hemochromatosis, hypothyroidism and gonadal dysfunction after busulfan-based cytoreduction. TBI appears to be a risk factor for malignant and endocrine late effects in the FA host. Multidisciplinary follow-up of patients with FA (including cancer screening) is essential for early detection and management of late complications, and improving long-term outcomes.

Integration of DNA sequence and DNA methylation changes in monozygotic twin pairs discordant for schizophrenia.

Schizophrenia is a complex mental disorder with high heritability (80%), extensive genetic heterogeneity, environmental contributions and only 50% concordance in discordant monozygotic (MZ) twins. Discordant MZ twins provide an exceptional opportunity to assess patient specific genome-wide genetic and epigenetic changes that may account for the disease phenotype. A combined analysis of genetic and epigenetic changes on the same twin pairs is expected to provide a more effective approach for two reasons. First, it is now possible to generate relatively reliable complete genome sequences as well as promoter methylation states on an individual level and second, the unaffected twin that originated from the same zygote provides a near perfect genetic match for contrast and comparison. This report deals with the combined analysis of DNA sequence data and methylation data on two pairs of discordant MZ twins that have been clinically followed for over 20 years. Results on Family 1 show that 58 genes differ in DNA sequence as well as promoter methylation in a schizophrenia-affected twin as compared to her healthy co-twin. The corresponding number for family 2 was 13. The two lists are over represented by neuronal genes and include a number of known schizophrenia candidate genes and drug targets. The results argue that changes in multiple genes via co-localized genetic and epigenetic alteration contribute to a liability threshold that is necessary for development of schizophrenia. This novel hypothesis, although logical, remains to be validated.

T-cell depleted allogeneic hematopoietic cell transplants as a platform for adoptive therapy with leukemia selective or virus-specific T-cells.

Allogeneic hematopoietic cell transplants adequately depleted of T-cells can reduce or prevent acute and chronic GVHD in both HLA-matched and haplotype-disparate hosts, without post-transplant prophylaxis with immunosuppressive drugs. Recent trials indicate that high doses of CD34+ progenitors from G-CSF mobilized peripheral blood leukocytes isolated and T-cell depleted by immunoadsorption to paramagnetic beads, when administered after myeloablative conditioning with TBI and chemotherapy or chemotherapy alone can secure consistent engraftment and abrogate GVHD in patients with acute leukemia without incurring an increased risk of a recurrent leukemia. Early clinical trials also indicate that high doses of in vitro generated leukemia-reactive donor T-cells can be adoptively transferred and can induce remissions of leukemia relapse without GVHD. Similarly, virus-specific T-cells generated from the transplant donor or an HLA partially matched third party, have induced remissions of Rituxan-refractory EBV lymphomas and can clear CMV disease or viremia persisting despite antiviral therapy in a high proportion of cases. Analyses of treatment responses and failures illustrate both the advantages and limitations of donor or banked, third party-derived T-cells, but underscore the potential of adoptive T-cell therapy in the absence of ongoing immunosuppression.

Affinity maturation of T-cell receptor-like antibodies for Wilms tumor 1 peptide greatly enhances therapeutic potential.

WT1126 (RMFPNAPYL) is a human leukocyte antigen-A2 (HLA-A2)-restricted peptide derived from Wilms tumor protein 1 (WT1), which is widely expressed in a broad spectrum of leukemias, lymphomas and solid tumors. A novel T-cell-receptor (TCR)-like single-chain variable fragment (scFv) antibody specific for the T-cell epitope consisting of the WT1/HLA-A2 complex was isolated from a human scFv phage library. This scFv was affinity-matured by mutagenesis combined with yeast display and structurally analyzed using a homology model. This monovalent scFv showed a 100-fold affinity improvement (dissociation constant (KD)=3 nm) and exquisite specificity towards its targeted epitope or HLA-A2(+)/WT1(+) tumor cells. Bivalent scFv-huIgG1-Fc fusion protein demonstrated an even higher avidity (KD=2 pm) binding to the T-cell epitope and to tumor targets and was capable of mediating antibody-dependent cell-mediated cytotoxicity or tumor lysis by chimeric antigen receptor-expressing human T- or NK-92-MI-transfected cells. This antibody demonstrated specific and potent cytotoxicity in vivo towards WT1-positive leukemia xenograft that was HLA-A2 restricted. In summary, T-cell epitopes can provide novel targets for antibody-based therapeutics. By combining phage and yeast displays and scFv-Fc fusion platforms, a strategy for developing high-affinity TCR-like antibodies could be rapidly explored for potential clinical development.

Comparison of outcomes at two institutions of patients with ALL receiving ex vivo T-cell-depleted or unmodified allografts.

We compared outcomes of adult patients receiving T-cell-depleted (TCD) hematopoietic SCT (HCT) without additional GVHD prophylaxis at Memorial Sloan Kettering Cancer Center (MSKCC, N=52), with those of patients receiving conventional grafts at MD Anderson Cancer Center (MDACC, N=115) for ALL in CR1 or CR2. Patients received myeloablative conditioning. Thirty-nine patients received anti-thymocyte globulin at MSKCC and 29 at MDACC. Cumulative incidence of grades 2-4 acute (P=0.001, 17.3% vs 42.6% at 100 days) and chronic GVHD (P=0.006, 13.5% vs 33.4% at 3 years) were significantly lower in the TCD group. The non-relapse mortality at day 100, 1 and 3 years was 15.4, 25.0 and 35.9% in the TCD group and 9.6, 23.6 and 28.6% in the unmodified group (P=0.368). There was no difference in relapse (P=0.107, 21.3% vs 35.5% at 3 years), OS (P=0.854, 42.6% vs 43.0% at 3 years) or RFS (P=0.653, 42.8% vs 35.9% at 3 years). In an adjusted model, age >50, cytogenetics and CR status were associated with inferior RFS (hazard ratio (HR)=2.16, P=0.003, HR=1.77, P=0.022, HR=2.47, P<0.001), whereas graft type was NS (HR=0.90, P=0.635). OS and RFS rates are similar in patients undergoing TCD or conventional HCT, but TCD effectively reduces the rate of GVHD.

Artificial Antigen Presenting Cells: An Off the Shelf Approach for Generation of Desirable T-Cell Populations for Broad Application of Adoptive Immunotherapy.

Adoptive transfer of antigen specific T-cells can lead to eradication of cancer and viral infections. The broad application of this approach has further been hampered by the limited availability of adequate numbers of T-cells for treatment in a timely manner. This has led to efforts for the development of efficient methods to generate large numbers of T-cells with specificity for tumor or viral antigens that can be harnessed for use in cancer therapy. Recent studies have demonstrated that during encounter with tumor antigen, the signals delivered to T-cells by professional antigen-presenting cells can affect T-cell programming and their subsequent therapeutic efficacy. This has stimulated efforts to develop artificial antigen-presenting cells that allow optimal control over the signals provided to T-cells. In this review, we will discuss the cellular artificial antigen-presenting cell systems and their use in T-cell adoptive immunotherapy for cancer and infections.

Impacts on rural livelihoods in Cambodia following adoption of best practice health and husbandry interventions by smallholder cattle farmers.

To better understand how smallholder farmers whom own the majority of Cambodian cattle can contribute to efforts to address food security needs in the Mekong region, a five-year research project investigating methods to improve cattle health and husbandry practices was conducted. Cattle production in Cambodia is constrained by transboundary animal diseases (TADs) including foot-and-mouth disease (FMD) and haemorrhagic septicaemia (HS) plus poor nutrition, reproduction and marketing knowledge. The project worked in six villages in Kandal, Takeo and Kampong Cham province during 2007-12. Farmers from three 'high intervention' (HI) villages incrementally received a participatory extension programme that included FMD and HS vaccination, forage development and husbandry training. Evaluation of project impacts on livelihoods was facilitated by comparison with three 'low intervention' (LI) villages where farmers received vaccinations only. Results of knowledge, attitude and practice (KAP) and socio-economic surveys conducted in 2012 of 120 participating farmers identified that farmer knowledge in the HI project sites exceeded LI sites on the topics of biosecurity, internal parasites, nutrition and reproduction. HI farmers adopted biosecurity practices including a willingness to vaccinate for FMD and HS at their own cost, separate sick from healthy cattle, grow and feed forages and displayed awareness of the benefits of building fattening pens. HI farmers that grew forages observed time savings exceeding two hours per day each for men, women and children, enabling expansion of farm enterprises, secondary employment and children's schooling. Logistic regression analysis revealed that farmers in the HI group significantly increased annual household income (P < 0.001), with 53% reporting an increase of 100% or more. We conclude that improving smallholder KAP of cattle health and production can lead to improved livelihoods. This strategy should be of interest to policymakers, donors, researchers and extension workers interested in addressing TAD control, food insecurity and rural poverty in Southeast Asia.

Graft-versus-host disease after double-unit cord blood transplantation has unique features and an association with engrafting unit-to-recipient HLA match.

Manifestations of and risk factors for graft-versus-host disease (GVHD) after double-unit cord blood transplantation (DCBT) are not firmly established. We evaluated 115 DCBT recipients (median age, 37 years) who underwent transplantation for hematologic malignancies with myeloablative or nonmyeloablative conditioning and calcineurin inhibitor/mycophenolate mofetil immunosuppression. Incidence of day 180 grades II to IV and III to IV acute GVHD (aGVHD) were 53% (95% confidence interval, 44 to 62) and 23% (95% confidence interval, 15 to 31), respectively, with a median onset of 40 days (range, 14 to 169). Eighty percent of patients with grades II to IV aGVHD had gut involvement, and 79% and 85% had day 28 treatment responses to systemic corticosteroids or budesonide, respectively. Of 89 engrafted patients cancer-free at day 100, 54% subsequently had active GVHD, with 79% of those affected having persistent or recurrent aGVHD or overlap syndrome. Late GVHD in the form of classic chronic GVHD was uncommon. Notably, grades III to IV aGVHD incidence was lower if the engrafting unit human leukocyte antigen (HLA)-A, -B, -DRB1 allele match was >4/6 to the recipient (hazard ratio, 0.385; P = .031), whereas engrafting unit infused nucleated cell dose and unit-to-unit HLA match were not significant. GVHD after DCBT was common in our study, predominantly affected the gut, and had a high therapy response, and late GVHD frequently had acute features. Our findings support the consideration of HLA- A,-B,-DRB1 allele donor-recipient (but not unit-unit) HLA match in unit selection, a practice change in the field. Moreover, new prophylaxis strategies that target the gastrointestinal tract are needed.