Immune monitoring in recipients of combined living donor kidney and hematopoietic stem/facilitating cell transplants.

Solid organ transplantation coupled with a hematopoietic stem cell transplant from the organ donor allows for the recipient to cease immunosuppressive therapy after transplant via a chimeric immune system. This beneficial effect of stem cell transplants is negatively affected by graft versus host disease (GVHD). Better understanding of the donor and recipient’s immune system is vital to mitigating graft versus host disease and induction of donor chimerism without GVHD. In this study, flow cytometry was used to characterize immune cells of the recipients’ before and up to eighteen months post transplantation. The recipients were categorized into their respective chimeric groups. Fully durable chimeric subjects did not reconstitute naïve T cells to pre-transplant levels by eighteen months post-transplant whereas the transiently chimeric subjects reconstituted. The goal was to find an immune cell biomarker for chimerism but due to the limited number of transiently chimeric subjects, the data showed no significant difference

When a Yarmulke Stands for All Jews: Navigating Shifting Signs from Synagogue to School in Luxembourg

peer reviewedIn the lives of students in Luxembourg’s Liberal Jewish complementary school, flexibility and mobility are highly valued as key characteristics of modern living. Complementary school students feel they easily meet these criteria—they are multilingual, cosmopolitan, and their approach to Jewish life is flexible, and equally importantly, they look, dress, and comport themselves “like everyone else.” These factors are understood to facilitate multiple movements and belongings in the contemporary world. The students directly contrast their ways of being with those of more observant Jews whom they refer to as “religious”; the material, embodied, and visible nature of observant Jewish life is perceived to be an impediment to participation and success in the secular sphere. However, when Jewishness appears in these students’ secular school classrooms, it is most often represented by Orthodox presenting men—often a man in a yarmulke. Further, these men and their yarmulkes are taken to represent all Jews, framed as a homogeneous group of religious adherents. For many complementary school students, these experiences can be jarring—they suddenly find themselves on the “wrong” side of the religious–secular divide and grouped together with those from whom they could not feel more distant. Drawing on ethnographic fieldwork and a material approach to religion, this article argues that the yarmulke comes to point to different levels and modes of observance and identities and enable different possible belongings in the secular public sphere as it travels across contexts that include different definitions of and attitudes toward religion and Jewishness

Defining species sensitivity and synergism potential for pesticides and pesticide mixtures through physiological traits analysis

This thesis investigates mechanisms and effects of chemical interactions on lepidopterans by analysing how toxicity is underpinned by toxicokinetic and toxicodynamic traits. Mamestra brassicae were exposed to a range of insecticides and fungicides representing several modes of actions, to identify synergistic or antagonistic effects. A novel high-throughput lepidopteran toxicity test was designed and trialled with a known toxicant, the organophosphate chlorpyrifos. This assay was suitable for assessing acute toxicity and toxicant effects on growth rates. To investigate chemical interactions, the effects of single compounds must be known. Using the novel bioassay, a range of five insecticides and four fungicides were screened for toxicity. Insecticides were toxic to larvae, except those targeting nicotinic acetylcholine receptors (nAChRs) such as imidacloprid and sulfoxaflor. Fungicide exposures caused no mortality. Of 13 binary mixture combinations tested, six showed a synergistic interaction, one an antagonistic and six no interaction (additive). The synergistic mixture of cypermethrin and prochloraz was taken forward for further testing. Sublethal and acute effects of this mixture were analysed in 2nd to 6th instar of M. brassicae larvae using the TKTD DEBTox model, highlighting synergistic effects on growth and survival. Finally, adult and larval sensitivity was compared to investigate effects of transcriptomic differences between the morphologically distinct life stages. Larvae and adults showed differential sensitivity to imidacloprid. A M. brassicae genome was sequenced and a larval and adult transcriptome generated to investigate this. Three detoxification enzyme families representing phase I, II and III metabolism were catalogued and transcriptomic differences analysed. Several of these enzymes were over 1000x upregulated in larvae, so represent potential drivers of this differential sensitivity. This work explores the mechanisms of synergistic chemical interactions via a multi-faceted approach including toxicity testing, TKTD modelling and metabolomics. This work will provide evidence to improve control of crop pests by implementing targeted pest management strategies based on life-stage sensitivities and also investigates the synergistic potential of pesticide mixtures, informing future pest management strategies and minimising risk of adverse effects to beneficial species

Developing stem-cell containing organoids as a pre-clinical model for colorectal cancer therapeutics

Colorectal cancer (CRC) is the second most common cause of cancer related deaths in the UK. Whilst identification of molecular events that contribute to the initiation and progression of CRC have facilitated the development of predictive biomarker-driven therapeutics, their success in the clinic has been restricted by the lack of anticipated responses. Furthermore, not all genetic signatures of a tumour have been linked to related drug targets, highlighting the need for novel development of compounds and better therapeutic rationales for the treatment of patient subsets. The limited success of targeted therapies, both within the clinic and drug discovery pipeline, has been attributed to a lack of effective preclinical models that are capable of capturing the complexity of deregulated signalling networks. The development of functional readouts that better represent tumour complexity, is therefore imperative to confirm the effects of hypothesis-driven therapeutics. This thesis therefore aimed to investigate whether 3D CRC organoids , which show a degree of greater complexity compared to preceding in vitro models, could be applied as suitable readouts for stratified medicine programmes and novel compounds within the drug discovery setting. To achieve this, a panel of 3D patient-derived CRC organoids cultures were generated. Suitable methodologies were established to facilitate organoids towards quantifiable, robust assay formats. This platform enabled the study of organoids within an in vitro clinical trial setting, based upon treatments administered within the ongoing FOCUS 4 stratified medicine trial, exploring organoids’ capacity to predict responses to targeted therapeutics. Organoids were differentially sensitive to therapies, irrespective of their genotypic background. It will be interesting to see whether prediction-response correlations observed in this study are typical of those seen in patients and whether functional readouts will be required to support stratified medicine approaches. Quantitative image-based analysis was also found to identify signatures of organoid responses against novel Wnt signalling inhibitors, suggesting that organoids may constitute a platform that can be used to study the effects of targeting a prospective cancer stem cell (CSC) population. Taken together, the findings in this thesis highlight the utility of patient derived organoid models as a functional model to evaluate novel therapeutic strategies, potentially generating clinically relevant hypotheses

Clinical outcomes of subcutaneous vs. transvenous implantable defibrillator therapy in a polymorbid patient cohort

BackgroundThe subcutaneous implantable cardioverter-defibrillator (S-ICD) has been designed to overcome lead-related complications and device endocarditis. Lacking the ability for pacing or resynchronization therapy its usage is limited to selected patients at risk for sudden cardiac death (SCD). ObjectiveThe aim of this single-center study was to assess clinical outcomes of S-ICD and single-chamber transvenous (TV)-ICD in an all-comers population. MethodsThe study cohort comprised a total of 119 ICD patients who underwent either S-ICD (n = 35) or TV-ICD (n = 84) implantation at the University Hospital Frankfurt from 2009 to 2017. By applying an inverse probability-weighting (IPW) analysis based on the propensity score including the Charlson Comorbidity Index (CCI) to adjust for potential extracardiac comorbidities, we aimed for head-to-head comparison on the study composite endpoint: overall survival, hospitalization, and device-associated events (including appropriate and inappropriate shocks or system-related complications). ResultsThe median age of the study population was 66.0 years, 22.7% of the patients were female. The underlying heart disease was ischemic cardiomyopathy (61.4%) with a median LVEF of 30%. Only 52.9% had received an ICD for primary prevention, most of the patients (67.3%) had advanced heart failure (NYHA class II-III) and 16.8% were in atrial fibrillation. CCI was 5 points in TV-ICD patients vs. 4 points for patients with S-ICD (p = 0.209) indicating increased morbidity. The composite endpoint occurred in 38 patients (31.9 %), revealing no significant difference between patients implanted with an S-ICD or TV-ICD (unweighted HR 1.50, 95 % confidence interval (CI) 0.78-2.90; p = 0.229, weighted HR 0.94, 95% CI, 0.61-1.50, p = 0.777). Furthermore, we observed no difference in any single clinical endpoint or device-associated outcome, neither in the unweighted cohort nor following inverse probability-weighting. ConclusionClinical outcomes of the S-ICD and TV-ICD revealed no differences in the composite endpoint including survival, freedom of hospitalization and device-associated events, even after careful adjustment for potential confounders. Moreover, the CCI was evaluated in a S-ICD cohort demonstrating higher survival rates than predicted by the CCI in young, polymorbid (S-)ICD patients

Displacement cloud point extraction procedure for preconcentration of iron(III) in water and fruit samples prior to spectrophotometric determination

ABSTRACT. For the enrichment of iron(III) prior to spectrophotometric determination, displacement cloud point extraction (D-CPE) technique was applied depending on the difference in stability constant of metal complexes. Zinc(II) as gallic acid complex was first separated into a Triton X-100 surfactant. Then, once the aqueous phase has been removed, the sample containing Fe(III) is added, and another CPE process is performed. Because Fe-GA has a higher stability than Zn-GA, Fe(III) can displace Zn(II) from the pre-extracted Zn-GA, allowing for Fe(III) separation from the complex sample matrix and its spontaneous spectrophotometric determination at 560 nm. The effects of pH, ligand, and surfactant quantities, temperature and heating time, centrifuge processes, and interferences were all studied. At the optimal conditions, the calibration graph was linear from 0.5 to 500 µg L-1 with enrichment factor of 75.0. The LOD was 0.15 µg L-1 and the RSD was 1.3% for 60 µg L-1 of Fe(III), n = 10. Accuracy was also evaluated using the standard reference substance (SRS) and flame atomic absorption spectroscopy (FAAS) approaches. This procedure was used for separation and micro-determination of Fe(III) in water and fruit tests (banana, strawberry, lemon, orange, and peaches) with recoveries ranged from 96.5 to 105%.   KEY WORDS: Displacement cloud point extraction, Iron, Spectrophotometry, Fruit samples   Bull. Chem. Soc. Ethiop. 2023, 37(1), 1-10.                                                                       DOI: https://dx.doi.org/10.4314/bcse.v37i1.1                                                        &nbsp

3D imaging of colorectal cancer organoids identifies responses to Tankyrase inhibitors

Aberrant activation of the Wnt signalling pathway is required for tumour initiation and survival in the majority of colorectal cancers. The development of inhibitors of Wnt signalling has been the focus of multiple drug discovery programs targeting colorectal cancer and other malignancies associated with aberrant pathway activation. However, progression of new clinical entities targeting the Wnt pathway has been slow. One challenge lies with the limited predictive power of 2D cancer cell lines because they fail to fully recapitulate intratumoural phenotypic heterogeneity. In particular, the relationship between 2D cancer cell biology and cancer stem cell function is poorly understood. By contrast, 3D tumour organoids provide a platform in which complex cell-cell interactions can be studied. However, complex 3D models provide a challenging platform for the quantitative analysis of drug responses of therapies that have differential effects on tumour cell subpopulations. Here, we generated tumour organoids from colorectal cancer patients and tested their responses to inhibitors of Tankyrase (TNKSi) which are known to modulate Wnt signalling. Using compounds with 3 orders of magnitude difference in cellular mechanistic potency together with image-based assays, we demonstrate that morphometric analyses can capture subtle alterations in organoid responses to Wnt inhibitors that are consistent with activity against a cancer stem cell subpopulation. Overall our study highlights the value of phenotypic readouts as a quantitative method to asses drug-induced effects in a relevant preclinical model

Improving and accelerating the differentiation and functional maturation of human stem cell-derived neurons: role of extracellular calcium and GABA

Neurons differentiated from pluripotent stem cells using established neural culture conditions often exhibit functional deficits. Recently, we have developed enhanced media which both synchronize the neurogenesis of pluripotent stem cell-derived neural progenitors and accelerate their functional maturation; together these media are termed SynaptoJuice. This pair of media are pro-synaptogenic and generate authentic, mature synaptic networks of connected forebrain neurons from a variety of induced pluripotent and embryonic stem cell lines. Such enhanced rate and extent of synchronized maturation of pluripotent stem cell-derived neural progenitor cells generates neurons which are characterized by a relatively hyperpolarized resting membrane potential, higher spontaneous and induced action potential activity, enhanced synaptic activity, more complete development of a mature inhibitory GABAA receptor phenotype and faster production of electrical network activity when compared to standard differentiation media. This entire process – from pre-patterned neural progenitor to active neuron – takes 3 weeks or less, making it an ideal platform for drug discovery and disease modelling in the fields of human neurodegenerative and neuropsychiatric disorders, such as Huntington's disease, Parkinson's disease, Alzheimer's disease and Schizophrenia

The αvβ6 integrin specific virotherapy, Ad5NULL-A20.FCU1, selectively delivers 2 potent “in-tumour” chemotherapy to Pancreatic Ductal Adenocarcinoma

Background Pancreatic ductal adenocarcinoma (PDAC) represent an unmet clinical need. Approximately 90% of PDACs express high levels of αvβ6 integrin. We have previously described Ad5NULL-A20, an adenovirus vector with ablated native means of cell entry and retargeted to αvβ6 integrin by incorporation of an A20 peptide. Methods Here, we incorporate suicide genes FCY1 and FCU1 encoding for cytosine deaminase (CDase) or a combination of CDase and UPRTase, capable of catalysing a non-toxic prodrug, 5-FC into the chemotherapeutic 5-FU and downstream metabolites, into replication-deficient Ad5 and Ad5NULL-A20. Results We show that Ad5NULL-A20 enables the transfer of suicide genes to αvβ6 integrin-positive PDAC cells which, in combination with 5-FC, results in cell death in vitro which is further mediated by a bystander effect in non-transduced cells. Intratumoural delivery of Ad5NULL-A20.FCU1 in combination with intraperitoneal delivery of 5-FC further results in tumour growth inhibition in a cell line xenograft in vivo. Using clinically-relevant 3D organoid models, we show selective transduction and therapeutic efficacy of FCU1 transgenes in combination with 5-FC. Conclusion Taken together these data provide the preclinical rationale for combined Ad5NULL-A20.FCU1 plus 5-FC as a promising targeted therapy to mediate “in-tumour chemotherapy” and merits further investigation for the treatment of PDAC patients