Non-invasive monitoring of renal transplant recipients: Urinary excretion of soluble adhesion molecules and of the complement-split product C4d

Background: The number of inducible adhesion molecules known to be involved in cell-mediated allograft rejection is still increasing. In addition, recent data describe complement activation during acute humoral allograft rejection. The aim of this study was to assess whether specific molecules from either pathway are excreted into urine and whether they can provide useful diagnostic tools for the monitoring of renal transplant recipients. Methods: Urinary concentrations of soluble adhesion molecules (sICAM-1, sVCAM-1) and of the complement degradation product C4d were determined by standardized ELISA technique in 75 recipients of renal allografts and 29 healthy controls. Patient samples were assigned to four categories according to clinical criteria: group 1: acute steroid-sensitive rejection (ASSR, n=14), group 2: acute steroid-resistant rejection (ASRR, n=12), group 3: chronic allograft dysfunction (CAD, n=20) and group 4: stable graft function (SGF, n=29). Results: All patients with rejection episodes (groups 1-3) had significantly higher values of urinary sC4d compared with healthy controls and patients with stable graft function (p<0.05). The urinary levels of sVCAM-1 were significantly higher in group 2 (ASRR) compared with all other groups (p<0.001). Uniformly low amounts of s-VCAM-1 and complement-split product C4d were excreted by healthy controls (group 0). In contrast, urinary sICAM-1 concentration in healthy controls was almost as high as in group 2 (ASRR) whereas patients with a stable functioning graft (group 4) excreted significantly less sICAM-1 (p<0.05). Conclusion: The evaluation of sVCAM-1 and sC4d excretion in urine can provide a valuable tool with regard to the severity and type of allograft rejection. With respect to long-term allograft survival, serial measurements of these markers should have the potential to detect rejection episodes and prompt immediate treatment. Copyright (C) 2003 S. Karger AG, Basel

Reduced CD40L expression on ex vivo activated CD4+T-lymphocytes from patients with excellent renal allograft function measured with a rapid whole blood flow cytometry procedure

Background: The CD40-CD40L (CD154) costimulatory pathway plays a critical role in the pathogenesis of kidney allograft rejection. In renal transplant biopsies, CD4+ CD40L+ graft-infiltrating cells were detected during chronic rejection in contrast to acute rejection episodes. Using a rapid noninvasive FACS procedure, we were able to demonstrate CD40L upregulation in peripheral blood of patients with chronic renal allograft dysfunction. Materials and Methods: Whole blood from recipients of renal allografts was stimulated with PMA and ion-omycin and measured by flow cytometry. Patients were assigned to three groups based on transplant function. Group 1: 26 patients with excellent renal transplant function; group 2: 28 patients with impaired transplant function; group 3: 14 patients with chronic allograft dysfunction and group 4: 8 healthy controls. Results: The median percentage +/-SEM of CD4+/ CD40L+ cells stimulated ex vivo at 10 ng/ml PMA was as follows: group 1: 28.3 +/- 4.1%; group 2: 18.4 +/- 2.4%; group 3: 50.1 +/- 5.0% and group 4: 40.4 +/- 3.4%. Subdivisions of groups 2 and 3 resulted in different CD40L expression patterns. Patients with increased serum creatinine since the initial phase after transplantation ( groups 2a and 3a) revealed a higher percentage of CD4+ CD40L+ cells than patients showing a gradual increase over time ( groups 2b and 3b). Consequently, patients of group 3a exhibited a significantly reduced transplant function compared with those of group 3b. Conclusion: After PMA + ionomycin stimulation, patients with excellent kidney graft function displayed significantly reduced expression of CD40L surface molecules on CD4+ cells early after transplantation. Those with a chronic dysfunction of the renal graft showed significantly more CD4+ cells expressing CD40L compared to the other transplanted groups. These results demonstrate that the percentage of CD4+ CD40L+ cells stimulated ex vivo in peripheral blood may be a valuable marker for chronic allograft nephropathy. Copyright (C) 2004 S. Karger AG, Basel

Quantifying inter-field movements of the western corn rootworm (Diabrotica virgifera virgifera LeConte) — A Central European field study

Dispersal plays a key role in the adaptation of species. It has been suggested that even in a stable and predictable environment, it is essential for any given population to “send” a certain portion of its offspring to disperse (referred as evolutionary stable dispersal rate). Although the literature on the flight behaviour of one of the major maize pests, the western corn rootworm, is rich, relatively little is known about its inter-field movements under field conditions. In the present study, inter-field movement of adult beetles was observed in Central-Europe under quasi-isolated conditions of infested continuous and un-infested first year maize fields, and related to candidate predictor variables. Percent of immigrants (net percent of adults within a given population leaving their natal maize field and arriving in first-year maize) varied greatly across years and locations (0.4–93.3%, mean = 38.7%). Results of the study provided field evidence of the assumption that western corn rootworm performs density dependent inter-field movement. Independent from pest densities, it appeared that about 1/3 of an adult beetle population always leaves its natal maize field, which likely allows the species to find new food sources and oviposition sites. The distance between maize fields and the phenological status of maize influenced inter-field movements but at a much less extent than it could have been expected from laboratory research findings

Non-invasive monitoring of renal transplant recipients: Urinary excretion of soluble adhesion molecules and of the complement-split product C4d

Background: The number of inducible adhesion molecules known to be involved in cell-mediated allograft rejection is still increasing. In addition, recent data describe complement activation during acute humoral allograft rejection. The aim of this study was to assess whether specific molecules from either pathway are excreted into urine and whether they can provide useful diagnostic tools for the monitoring of renal transplant recipients. Methods: Urinary concentrations of soluble adhesion molecules (sICAM-1, sVCAM-1) and of the complement degradation product C4d were determined by standardized ELISA technique in 75 recipients of renal allografts and 29 healthy controls. Patient samples were assigned to four categories according to clinical criteria: group 1: acute steroid-sensitive rejection (ASSR, n=14), group 2: acute steroid-resistant rejection (ASRR, n=12), group 3: chronic allograft dysfunction (CAD, n=20) and group 4: stable graft function (SGF, n=29). Results: All patients with rejection episodes (groups 1-3) had significantly higher values of urinary sC4d compared with healthy controls and patients with stable graft function (p<0.05). The urinary levels of sVCAM-1 were significantly higher in group 2 (ASRR) compared with all other groups (p<0.001). Uniformly low amounts of s-VCAM-1 and complement-split product C4d were excreted by healthy controls (group 0). In contrast, urinary sICAM-1 concentration in healthy controls was almost as high as in group 2 (ASRR) whereas patients with a stable functioning graft (group 4) excreted significantly less sICAM-1 (p<0.05). Conclusion: The evaluation of sVCAM-1 and sC4d excretion in urine can provide a valuable tool with regard to the severity and type of allograft rejection. With respect to long-term allograft survival, serial measurements of these markers should have the potential to detect rejection episodes and prompt immediate treatment. Copyright (C) 2003 S. Karger AG, Basel

Influence of soil on the efficacy of entomopathogenic nematodes in reducing Diabrotica virgifera virgifera in maize

The use of entomopathogenic nematodes is one potential non-chemical approach to control the larvae of the invasive western corn rootworm (Diabrotica virgifera virgifera LeConte, Coleoptera: Chrysomelidae) in Europe. This study investigated the efficacy of Heterorhabditis bacteriophora Poinar (Rhabditida: Heterorhabditidae), Heterorhabditis megidis Poinar, Jackson and Klein (Rh., Heterorhabditidae) and Steinernema feltiae Filipjev (Rh., Steinernematidae) in reducing D. v. virgifera as a function of soil characteristics. A field experiment was repeated four times in southern Hungary using artificially infested maize plants potted into three different soils. Sleeve gauze cages were used to assess the number of emerging adult D. v. virgifera from the treatments and untreated controls. Results indicate that nematodes have the potential to reduce D. v. virgifera larvae in most soils; however, their efficacy can be higher in maize fields with heavy clay or silty clay soils than in sandy soils, which is in contrast to the common assumption that nematodes perform better in sandy soils than in heavy soils

Inferring introduction routes of invasive species using approximate Bayesian computation on microsatellite data

Determining the routes of introduction provides not only information about the history of an invasion process, but also information about the origin and construction of the genetic composition of the invading population. It remains difficult, however, to infer introduction routes from molecular data because of a lack of appropriate methods. We evaluate here the use of an approximate Bayesian computation (ABC) method for estimating the probabilities of introduction routes of invasive populations based on microsatellite data. We considered the crucial case of a single source population from which two invasive populations originated either serially from a single introduction event or from two independent introduction events. Using simulated datasets, we found that the method gave correct inferences and was robust to many erroneous beliefs. The method was also more efficient than traditional methods based on raw values of statistics such as assignment likelihood or pairwise F(ST). We illustrate some of the features of our ABC method, using real microsatellite datasets obtained for invasive populations of the western corn rootworm, Diabrotica virgifera virgifera. Most computations were performed with the DIYABC program (http://www1.montpellier.inra.fr/CBGP/diyabc/)

Mechanism based therapies enable personalised treatment of hypertrophic cardiomyopathy

Cardiomyopathies have unresolved genotype–phenotype relationships and lack disease-specific treatments. Here we provide a framework to identify genotype-specific pathomechanisms and therapeutic targets to accelerate the development of precision medicine. We use human cardiac electromechanical in-silico modelling and simulation which we validate with experimental hiPSC-CM data and modelling in combination with clinical biomarkers. We select hypertrophic cardiomyopathy as a challenge for this approach and study genetic variations that mutate proteins of the thick (MYH7R403Q/+) and thin filaments (TNNT2R92Q/+, TNNI3R21C/+) of the cardiac sarcomere. Using in-silico techniques we show that the destabilisation of myosin super relaxation observed in hiPSC-CMs drives disease in virtual cells and ventricles carrying the MYH7R403Q/+ variant, and that secondary effects on thin filament activation are necessary to precipitate slowed relaxation of the cell and diastolic insufficiency in the chamber. In-silico modelling shows that Mavacamten corrects the MYH7R403Q/+ phenotype in agreement with hiPSC-CM experiments. Our in-silico model predicts that the thin filament variants TNNT2R92Q/+ and TNNI3R21C/+ display altered calcium regulation as central pathomechanism, for which Mavacamten provides incomplete salvage, which we have corroborated in TNNT2R92Q/+ and TNNI3R21C/+ hiPSC-CMs. We define the ideal characteristics of a novel thin filament-targeting compound and show its efficacy in-silico. We demonstrate that hybrid human-based hiPSC-CM and in-silico studies accelerate pathomechanism discovery and classification testing, improving clinical interpretation of genetic variants, and directing rational therapeutic targeting and design

Hierarchical and stage-specific regulation of murine cardiomyocyte maturation by serum response factor

After birth, cardiomyocytes (CM) acquire numerous adaptations in order to efficiently pump blood throughout an animal’s lifespan. How this maturation process is regulated and coordinated is poorly understood. Here, we perform a CRISPR/Cas9 screen in mice and identify serum response factor (SRF) as a key regulator of CM maturation. Mosaic SRF depletion in neonatal CMs disrupts many aspects of their maturation, including sarcomere expansion, mitochondrial biogenesis, transverse-tubule formation, and cellular hypertrophy. Maintenance of maturity in adult CMs is less dependent on SRF. This stage-specific activity is associated with developmentally regulated SRF chromatin occupancy and transcriptional regulation. SRF directly activates genes that regulate sarcomere assembly and mitochondrial dynamics. Perturbation of sarcomere assembly but not mitochondrial dynamics recapitulates SRF knockout phenotypes. SRF overexpression also perturbs CM maturation. Together, these data indicate that carefully balanced SRF activity is essential to promote CM maturation through a hierarchy of cellular processes orchestrated by sarcomere assembly

Potentially inappropriate medication in older participants of the Berlin Aging Study II (BASE-II) - Sex differences and associations with morbidity and medication use

INTRODUCTION: Multimorbidity in advanced age and the need for drug treatment may lead to polypharmacy, while pharmacokinetic and pharmacodynamic changes may increase the risk of adverse drug events (ADEs). OBJECTIVE: The aim of this study was to determine the proportion of subjects using potentially inappropriate medication (PIM) in a cohort of older and predominantly healthy adults in relation to polypharmacy and morbidity. METHODS: Cross-sectional data were available from 1,382 study participants (median age 69 years, IQR 67-71, 51.3% females) of the Berlin Aging Study II (BASE-II). PIM was classified according to the EU(7)-PIM and German PRISCUS (representing a subset of the former) list. Polypharmacy was defined as the concomitant use of at least five drugs. A morbidity index (MI) largely based on the Charlson Index was applied to evaluate the morbidity burden. RESULTS: Overall, 24.1% of the participants were affected by polypharmacy. On average, men used 2 (IQR 1-4) and women 3 drugs (IQR 1-5). According to PRISCUS and EU(7)-PIM, 5.9% and 22.6% of participants received at least one PIM, while use was significantly more prevalent in females (25.5%) compared to males (19.6%) considering EU(7)-PIM (p = 0.01). In addition, morbidity in males receiving PIM according to EU(7)-PIM was higher (median MI 1, IQR 1-3) compared to males without PIM use (median MI 1, IQR 0-2, p<0.001). CONCLUSION: PIM use occurred more frequently in women than in men, while it was associated with higher morbidity in males. As expected, EU(7)-PIM identifies more subjects as PIM users than the PRISCUS list but further studies are needed to investigate the differential impact of both lists on ADEs and outcome. KEY POINTS: We found PIM use to be associated with a higher number of regular medications and with increased morbidity. Additionally, we detected a higher prevalence of PIM use in females compared to males, suggesting that women and people needing intensive drug treatment are patient groups, who are particularly affected by PIM use

Secondary contact and admixture between independently invading populations of the Western corn rootworm, diabrotica virgifera virgifera in Europe

The western corn rootworm, Diabrotica virgifera virgifera (Coleoptera: Chrysomelidae), is one of the most destructive pests of corn in North America and is currently invading Europe. The two major invasive outbreaks of rootworm in Europe have occurred, in North-West Italy and in Central and South-Eastern Europe. These two outbreaks originated from independent introductions from North America. Secondary contact probably occurred in North Italy between these two outbreaks, in 2008. We used 13 microsatellite markers to conduct a population genetics study, to demonstrate that this geographic contact resulted in a zone of admixture in the Italian region of Veneto. We show that i) genetic variation is greater in the contact zone than in the parental outbreaks; ii) several signs of admixture were detected in some Venetian samples, in a Bayesian analysis of the population structure and in an approximate Bayesian computation analysis of historical scenarios and, finally, iii) allelic frequency clines were observed at microsatellite loci. The contact between the invasive outbreaks in North-West Italy and Central and South-Eastern Europe resulted in a zone of admixture, with particular characteristics. The evolutionary implications of the existence of a zone of admixture in Northern Italy and their possible impact on the invasion success of the western corn rootworm are discussed