Automated image analysis system for studying cardiotoxicity in human pluripotent stem cell-Derived cardiomyocytes

BackgroundCardiotoxicity, characterized by severe cardiac dysfunction, is a major problem in patients treated with different classes of anticancer drugs. Development of predictable human-based models and assays for drug screening are crucial for preventing potential drug-induced adverse effects. Current animal in vivo models and cell lines are not always adequate to represent human biology. Alternatively, human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) show great potential for disease modelling and drug-induced toxicity screenings. Fully automated high-throughput screening of drug toxicity on hiPSC-CMs by fluorescence image analysis is, however, very challenging, due to clustered cell growth patterns and strong intracellular and intercellular variation in the expression of fluorescent markers.ResultsIn this paper, we report on the development of a fully automated image analysis system for quantification of cardiotoxic phenotypes from hiPSC-CMs that are treated with various concentrations of anticancer drugs doxorubicin or crizotinib. This high-throughput system relies on single-cell segmentation by nuclear signal extraction, fuzzy C-mean clustering of cardiac α-actinin signal, and finally nuclear signal propagation. When compared to manual segmentation, it generates precision and recall scores of 0.81 and 0.93, respectively.ConclusionsOur results show that our fully automated image analysis system can reliably segment cardiomyocytes even with heterogeneous α-actinin signals.Computer Systems, Imagery and MediaAlgorithms and the Foundations of Software technolog

Prevalence of Parental Thrombophilic Defects After Fetal Death and Relation to Cause

OBJECTIVE: To estimate whether parental thrombophilic defects after fetal death, either acquired or inherited, were more prevalent than in the normal population and to estimate associations between these thrombophilic defects and different fetal death causes. METHODS: In a multicenter, prospective cohort study of 750 fetal deaths, we tested couples for antithrombin, protein C, total and free protein S, and von Willebrand factor (vWF) plasma levels. Mothers' values were compared with reference values in gestational age-matched healthy pregnant women, and fathers were compared with healthy men. Prevalence of factor V Leiden, prothrombin G20210A mutation, and lupus anticoagulant were compared with the normal population. A panel classified death cause. RESULTS: More women with fetal death had decreased antithrombin (16.8%, P <.001) and protein C (4.0%, P = .03) and increased vWF (15.5%, P <.001) plasma levels than healthy pregnant women (2.5%). However, compared with normal ranges in the nonpregnant population, we only observed more women with increased vWF (12.4%, P <.001). More fathers had decreased free protein S (6.3%, P <.001) and elevated vWF (12.1%, P <.001) than healthy men (2.5%). Prevalence of inherited thrombophilias was not higher in couples with fetal death than in the population. Neither inherited nor acquired maternal or paternal thrombophilic defects were associated with the main cause of death. Of placental causes, abruption and infarction were associated with acquired maternal defects. CONCLUSION: Except for vWF and paternal free protein S, acquired and inherited thrombophilic defects were not more prevalent after fetal death. Routine thrombophilia testing after fetal death is not advised

Allocation to highly sensitized patients based on acceptable mismatches results in low rejection rates comparable to nonsensitized patients

Contains fulltext : 208426.pdf (publisher's version ) (Open Access)Whereas regular allocation avoids unacceptable mismatches on the donor organ, allocation to highly sensitized patients within the Eurotransplant Acceptable Mismatch (AM) program is based on the patient's HLA phenotype plus acceptable antigens. These are HLA antigens to which the patient never made antibodies, as determined by extensive laboratory testing. AM patients have superior long-term graft survival compared with highly sensitized patients in regular allocation. Here, we questioned whether the AM program also results in lower rejection rates. From the PROCARE cohort, consisting of all Dutch kidney transplants in 1995-2005, we selected deceased donor single transplants with a minimum of 1 HLA mismatch and determined the cumulative 6-month rejection incidence for patients in AM or regular allocation. Additionally, we determined the effect of minimal matching criteria of 1 HLA-B plus 1 HLA-DR, or 2 HLA-DR antigens on rejection incidence. AM patients showed significantly lower rejection rates than highly immunized patients in regular allocation, comparable to nonsensitized patients, independent of other risk factors for rejection. In contrast to highly sensitized patients in regular allocation, minimal matching criteria did not affect rejection rates in AM patients. Allocation based on acceptable antigens leads to relatively low-risk transplants for highly sensitized patients with rejection rates similar to those of nonimmunized individuals

Monitoring van nationale ammoniakemissies uit de landbouw; op weg naar een verbeterde rekenmethodiek

De huidige rekenmethodiek voor het berekenen van de nationale ammoniakemissies uit landbouwkundige bronnen is aan een kritische analyse onderworpen. Aanleiding hiervoor was het grote verschil tussen de op directe wijze berekende ammoniakemissie uit deverschillende landbouwbronnen en de op indirecte wijze berekende emissie, afgeleid uit concentratiemetingen van ammoniak in lucht. De ammoniakemissies van de volgende bronnen zijn behandeld: de N-excretie van diergroepen, stal en buitenopslag van mest, mesttoediening op het land, kunstmest, beweiding en gewassen. Daarnaast is aandacht besteed aan de mestlogistiek, die van belang is voor de verdeling van geproduceerde mest over het bedrijf, binnen de regio en naar andere delen van Nederland. Aanbevelingen zijn gedaan voor de aanpassing van de huidige rekenmethodiek op de korte termijn en voor onderzoek dat na een aantal jaren moet leiden tot een structurele verbetering van de rekenmethodiek

Marine nitrogen cycling dynamics under altering redox conditions: Insights from deposition of sapropels S1 and the ambiguous S2 in the Eastern Mediterranean Sea

The eastern Mediterranean Sea (EMS) sedimentary record is periodically interspersed with organic-rich ‘sapropel’ layers. Sapropels are characteristic of basin-wide anoxic events, triggered by precession-forced insolation maxima. Relatively subdued insolation maxima, however, are not always expressed as distinct sapropel events. The EMS sedimentary record is thus useful to investigate feedbacks between marine anoxia and the nitrogen (N) cycle and offers an analogue for modern deoxygenation and past oceanic anoxic events. To this end, we investigated a ∼68 kyr sedimentary record from the EMS containing the well-established sapropel S1 (deposited in two phases: S1a [∼10.5–8.5 ka BP] and S1b [∼7.8–6.1 ka BP]) and sediments timed to the ambiguous S2 sapropel (∼53 ka BP). We used lipid biomarkers of microorganisms to reconstruct key N-cycle components: (1) anaerobic ammonium oxidation (anammox) using ladderanes and a stereoisomer of bacteriohopanetetrol (BHT-x), (2) dinitrogen gas (N2) fixation using heterocyte glycolipids, and (3) nitrification by Thaumarchaeota using crenarchaeol. Additionally, benthic foraminifera and trace metals (U, Mo, Mn) were used to reconstruct redox conditions. During S1a, abundances of Thaumarchaeota increased, likely promoted by elevated high-nutrient freshwater discharge. At this time, a combination of phosphorus supply and intensified loss of bioavailable N via water column anammox, may have reinforced anoxia by favoring diatom-diazotroph associations. During S1b, anammox is equally intense. Yet, no positive feedback on N2-fixation is observed, likely because diazotrophs were phosphorus limited. Instead, anammox may have provided negative feedback on anoxia by quenching primary production. Ladderanes suggest additional episodes of anammox between ∼69 to 39 cal ka BP, corresponding to brief periods of water column deoxygenation. Anoxia likely occurred at the sediment–water interface in S2-timed sediments (53–51 cal ka BP). During these episodes, ladderanes co-occur with the later eluting BHT-34R stereoisomer. δ13CBHT-34R indicate an anammox source, potentially synthesized by marine sedimentary anammox bacteria. No corresponding increase in diatom-diazotroph associations is observed, likely due to the oligotrophic conditions and the limited effect of sedimentary anammox on N-availability in the euphotic zone. Our results highlight various modes of operation of the N-cycle at different degrees of deoxygenation, which depend amongst others on nutrient-availability and the niche-segregation of N-loss and N2-fixating microorganisms

Paraganglioma and pheochromocytoma upon maternal transmission of SDHD mutations

The SDHD gene encodes a subunit of the mitochondrial tricarboxylic acid cycle enzyme and tumor suppressor

Evaluation of spelt germplasm for polyphenol oxidase activity and aluminium resistance

Kidney transplantation is the best treatment option for patients with end-stage renal failure. At present, approximately 800 Dutch patients are registered on the active waiting list of Eurotransplant. The waiting time in the Netherlands for a kidney from a deceased donor is on average between 3 and 4years. During this period, patients are fully dependent on dialysis, which replaces only partly the renal function, whereas the quality of life is limited. Mortality among patients on the waiting list is high. In order to increase the number of kidney donors, several initiatives have been undertaken by the Dutch Kidney Foundation including national calls for donor registration and providing information on organ donation and kidney transplantation. The aim of the national PROCARE consortium is to develop improved matching algorithms that will lead to a prolonged survival of transplanted donor kidneys and a reduced HLA immunization. The latter will positively affect the waiting time for a retransplantation. The present algorithm for allocation is among others based on matching for HLA antigens, which were originally defined by antibodies using serological typing techniques. However, several studies suggest that this algorithm needs adaptation and that other immune parameters which are currently not included may assist in improving graft survival rates. We will employ a multicenter-based evaluation on 5429 patients transplanted between 1995 and 2005 in the Netherlands. The association between key clinical endpoints and selected laboratory defined parameters will be examined, including Luminex-defined HLA antibody specificities, T and B cell epitopes recognized on the mismatched HLA antigens, non-HLA antibodies, and also polymorphisms in complement and Fc receptors functionally associated with effector functions of anti-graft antibodies. From these data, key parameters determining the success of kidney transplantation will be identified which will lead to the identification of additional parameters to be included in future matching algorithms aiming to extend survival of transplanted kidneys and to diminish HLA immunization. Computer simulation studies will reveal the number of patients having a direct benefit from improved matching, the effect on shortening of the waiting list, and the decrease in waiting time

PIRCHE-II is related to graft failure after kidney transplantation

Individual HLA mismatches may differentially impact graft survival after kidney transplantation. Therefore, there is a need for a reliable tool to define permissible HLA mismatches in kidney transplantation. We previously demonstrated that donor-derived Predicted Indirectly ReCognizable HLA Epitopes presented by recipient HLA class II (PIRCHE-II) play a role in de novo donor-specific HLA antibodies formation after kidney transplantation. In the present Dutch multi-center study, we evaluated the possible association between PIRCHE-II and kidney graft failure in 2,918 donor-recipient couples that were transplanted between 1995 and 2005. For these donors-recipients couples, PIRCHE-II numbers were related to graft survival in univariate and multivariable analyses. Adjusted for confounders, the natural logarithm of PIRCHE-II was associated with a higher risk for graft failure [hazard ratio (HR): 1.13, 95% CI: 1.04-1.23, p = 0.003]. When analyzing a subgroup of patients who had their first transplantation, the HR of graft failure for ln(PIRCHE-II) was higher compared with the overall cohort (HR: 1.22, 95% CI: 1.10-1.34, p < 0.001). PIRCHE-II demonstrated both early and late effects on graft failure in this subgroup. These data suggest that the PIRCHE-II may impact graft survival after kidney transplantation. Inclusion of PIRCHE-II in donor-selection criteria may eventually lead to an improved kidney graft survival