Populationspharmakokinetik von Indomethacin bei Kindern mit Salzverlusttubulopathien

Da es wenige fundierte populationspharmakokinetische Studien zu Indomethacin insgesamt und nur eine bei Kindern gibt, sollte in der hier vorliegenden Arbeit die Populationspharmakokinetik von Indomethacin bei Kindern mit Salzverlusttubulopathien näher untersucht werden. Aufgrund der großen interindividuellen Unterschiede in der Pharmakokinetik des Indomethacins bieten sich populationspharmakokinetische Methoden besonders an. Zur populationspharmakokinetischen Auswertung der vorliegenden Patientendaten wurde das Software-Programm NONMEM verwendet. Als Strukturmodell war ein 1-Kompartiment-Modell mit Absorption erster Ordnung zur Beschreibung der Pharmakokinetik des Indomethacins am geeignetsten. Dabei wurde die Absorptionskonstante ka mit 1,5 h-1 festgesetzt. Dieser Wert wurde aus der Literatur (Oberbauer et al. 1993)entnommen. Als pharmakokinetische Parameter wurden die Clearance (CL) und das Verteilungsvolumen (V) von Indomethacin bestimmt.Zur Beschreibung sowohl der CL als auch des V wurde die Covariable Körperoberfläche(BSA) in das Modell integriert. Daraus ergab sich eine deutlich bessere Modellanpassung,was anhand der Objektivierungsfunktion und Diagnostikgraphiken zur Güte der Anpassung gezeigt wurde. Der Konzentrationszeitverlauf des Indomethacins konnte deutlich exakter durch das Modell vorhergesagt werden als ohne Integration dieser zusätzlichen Covariable. Die Covariable BSA erscheint besonders geeignet, da sie die Körpergröße (HT) und das Körpergewicht (WT) miteinander vereint, für die beide ein Einfluss auf die Pharmakokinetik des Indomethacins nachgewiesen werden konnte. Für andere demographische Faktoren der Patienten, wie Geschlecht, Gesamtproteinkonzentration im Serum oder Nierenfunktion konnte kein Einfluss auf die pharmakokinetischen Parameter nachgewiesen werden. Die Integration des Körpergewichts (WT) nach dem von Holford propagierten Modell führte zu keiner besseren Beschreibung der Pharmakokinetik des Indomethacins gegenüber dem BSA-Modell. Auch die Integration einer IOV in das BSA-Modell führte zu keiner Verbesserung despharmakokinetischen Modells. Als statistisches Modell zur Beschreibung interindividuellen Variabilität (η) in den Modellparametern wurde das so genannte exponentielle Fehlermodell verwendet und für die intraindividuelle Variabilität (ε) das proportionale Fehlermodell.Aus dem so entwickelten pharmakokinetischen Modell ließen sich folgende Werte für die Clearance und das Verteilungsvolumen, sowie der HWZ des Indomethacins berechnen: CL= 2,67 (l/m²*h) V = 23,6 (l/m²) t1/2 = 6,0 Stunden Nach Berechnung des Coefficient of variation (CV) kann die Clearance durch das pharmakokinetische Modell mit einer Präzision von 58 % vorhergesagt werden. Für das Verteilungsvolumen ergibt sich eine Präzision von 64 %. Es bleibt also weiterhin noch eine interindividuelle Variabilität bestehen, welche durch das Modell nicht näher erklärt werden kann. Der CV für die intraindividuelle Variabilität beträgt 69 %. Das bedeutet,dass die Indomethacinkonzentrationen mit einer Präzision von 31% durch das pharmakokinetische Modell vorhergesagt werden können. Dennoch sind die Ergebnisse mit denen in vorhergehenden Studien ermittelten vergleichbar,was nach Durchsicht der Literatur zur Pharmakokinetik des Indomethacins gezeigt werden konnte. Es ist sinnvoll, sie in die Überlegungen zur richtigen Dosisfindung im klinischen Alltag heranzuziehen. Zusammenfassend lässt sich festhalten, dass populationspharmakokinetische Modellesehr geeignet sind, um pharmakokinetische Studien durchzuführen, da sie bei geringerem Aufwand und deutlich patientenschonenderem Umgang ähnlich valide Ergebnisse,wie klassische pharmakokinetische Studien liefern. Gerade auch im Hinblick auf die neue Arzneimittelverordnung eignet sich die Populationspharmakokinetik den neuen Anforderungen gerecht zu werden und somit zur Arzneimittelsicherheit im kindlichen Kollektiv beizutragen

A SAM analogue-utilizing ribozyme for site-specific RNA alkylation in living cells

Post-transcriptional RNA modification methods are in high demand for site-specific RNA labelling and analysis of RNA functions. In vitro-selected ribozymes are attractive tools for RNA research and have the potential to overcome some of the limitations of chemoenzymatic approaches with repurposed methyltransferases. Here we report an alkyltransferase ribozyme that uses a synthetic, stabilized S-adenosylmethionine (SAM) analogue and catalyses the transfer of a propargyl group to a specific adenosine in the target RNA. Almost quantitative conversion was achieved within 1 h under a wide range of reaction conditions in vitro, including physiological magnesium ion concentrations. A genetically encoded version of the SAM analogue-utilizing ribozyme (SAMURI) was expressed in HEK293T cells, and intracellular propargylation of the target adenosine was confirmed by specific fluorescent labelling. SAMURI is a general tool for the site-specific installation of the smallest tag for azide-alkyne click chemistry, which can be further functionalized with fluorophores, affinity tags or other functional probes

Therapy preferences of patients with lung and colon cancer: A discrete choice experiment

Objectives: There is increasing interest in studies that examine patient preferences to measure health-related outcomes. Understanding patients’ preferences can improve the treatment process and is particularly relevant for oncology. In this study, we aimed to identify the subgroup-specific treatment preferences of German patients with lung cancer (LC) or colorectal cancer (CRC). Methods: Six discrete choice experiment (DCE) attributes were established on the basis of a systematic literature review and qualitative interviews. The DCE analyses comprised generalized linear mixed-effects model and latent class mixed logit model. Results: The study cohort comprised 310 patients (194 with LC, 108 with CRC, 8 with both types of cancer) with a median age of 63 (SD =10.66) years. The generalized linear mixed-effects model showed a significant (P<0.05) degree of association for all of the tested attributes. “Strongly increased life expectancy” was the attribute given the greatest weight by all patient groups. Using latent class mixed logit model analysis, we identified three classes of patients. Patients who were better informed tended to prefer a more balanced relationship between length and health-related quality of life (HRQoL) than those who were less informed. Class 2 (LC patients with low HRQoL who had undergone surgery) gave a very strong weighting to increased length of life. We deduced from Class 3 patients that those with a relatively good life expectancy (CRC compared with LC) gave a greater weight to moderate effects on HRQoL than to a longer life. Conclusion: Overall survival was the most important attribute of therapy for patients with LC or CRC. Differences in treatment preferences between subgroups should be considered in regard to treatment and development of guidelines. Patients’ preferences were not affected by sex or age, but were affected by the cancer type, HRQoL, surgery status, and the main source of information on the disease

TGFβ-signaling and FOXG1-expression are a hallmark of astrocyte lineage diversity in the murine ventral and dorsal forebrain

Heterogeneous astrocyte populations are defined by diversity in cellular environment, progenitor identity or function. Yet, little is known about the extent of the heterogeneity and how this diversity is acquired during development. To investigate the impact of TGF (transforming growth factor) β-signaling on astrocyte development in the telencephalon we deleted the TGFBR2 (transforming growth factor beta receptor 2) in early neural progenitor cells in mice using a FOXG1 (forkhead box G1)-driven CRE-recombinase. We used quantitative proteomics to characterize TGFBR2-deficient cells derived from the mouse telencephalon and identified differential protein expression of the astrocyte proteins GFAP (glial fibrillary acidic protein) and MFGE8 (milk fat globule-EGF factor 8). Biochemical and histological investigations revealed distinct populations of astrocytes in the dorsal and ventral telencephalon marked by GFAP or MFGE8 protein expression. The two subtypes differed in their response to TGFβ-signaling. Impaired TGFβ-signaling affected numbers of GFAP astrocytes in the ventral telencephalon. In contrast, TGFβ reduced MFGE8-expression in astrocytes deriving from both regions. Additionally, lineage tracing revealed that both GFAP and MFGE8 astrocyte subtypes derived partly from FOXG1-expressing neural precursor cells.Fil: Weise, Stefan Christopher. Universität Freiburg Im Breisgau; AlemaniaFil: Villarreal, Alejandro. Universität Freiburg Im Breisgau; Alemania. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Biología Celular y Neurociencia "Prof. Eduardo de Robertis". Universidad de Buenos Aires. Facultad de Medicina. Instituto de Biología Celular y Neurociencia; ArgentinaFil: Heidrich, Stefanie. Universität Freiburg Im Breisgau; AlemaniaFil: Dehghanian, Fariba. University Of Isfahan; Irán. Universität Freiburg Im Breisgau; AlemaniaFil: Schachtrup, Christian. Universität Freiburg Im Breisgau; AlemaniaFil: Nestel, Sigrun. Universität Freiburg Im Breisgau; AlemaniaFil: Schwarz, Jennifer. Universität Freiburg Im Breisgau; AlemaniaFil: Thedieck, Kathrin. Universität Oldenburg; Argentina. University of Groningen; Países BajosFil: Vogel, Tanja. Universität Freiburg Im Breisgau; Alemani

Diagnosis of congenital heart malformations -possibilities for the employment of telepathology

Goal: In a study of 10 autopsy cases with congenital cardiac malformations we investigated whether obtaining a second opinion by means of telepathology could satisfy quality standards for the diagnosis of cardiac malformations and what the advantages and disadvantages of such a procedure might be. Material: The investigatory samples were 10 formalinfixed hearts with complex malformations from 9 fetuses and one newborn on which autopsies had been performed at the Pathological Institute of the Charité Hospital. The requests for a second opinion, which included text and image data, were sent in the form of Microsoft PowerPoint presentations to 5 experts in 4 countries. Per case the number of images that were sent was between 3 and 7. The size of the files was between 439 and 942 kb. The time required for preparation of the cases for sending them to the specialists was between 1 and 2 hours: this encompassed the time for putting the notation on the images, compressing them, creating a file that included both the images and the clinical data and then sending the case file. Results: All 10 cardiac malformations were correctly identified. In 8 of the 10 cases at least one expert had questions. After these questions had been answered and further images had been sent final correct diagnoses were made in all cases. All experts said that the quality of the images was very good. Use of a standardized findings questionnaire, which also included the marking of anatomic structures and of pathological findings in the images, proved useful. Standardized findings forms facilitate orientation during interpretation of the cases and should be used generally to avoid misunderstandings in telepathological communication. Conclusions: In general it is possible to obtain an effective and reliable diagnosis of congenital heart malformations by means of telepathology. It is far quicker to get a second opinion by this means than by conventional means

Deletion of the Na-K-2Cl cotransporter NKCC1 results in a more severe epileptic phenotype in the intrahippocampal kainate mouse model of temporal lobe epilepsy

Increased neuronal expression of the Na-K-2Cl cotransporter NKCC1 has been implicated in the generation of seizures and epilepsy. However, conclusions from studies on the NKCC1-specific inhibitor, bumetanide, are equivocal, which is a consequence of the multiple potential cellular targets and poor brain penetration of this drug. Here, we used Nkcc1 knockout (KO) and wildtype (WT) littermate control mice to study the ictogenic and epileptogenic effects of intrahippocampal injection of kainate. Kainate (0.23 ?g in 50 nl) induced limbic status epilepticus (SE) in both KO and WT mice with similar incidence, latency to SE onset, and SE duration, but the number of intermittent generalized convulsive seizures during SE was significantly higher in Nkcc1 KO mice, indicating increased SE severity. Following SE, spontaneous recurrent seizures (SRS) were recorded by continuous (24/7) video/EEG monitoring at 0-1, 4-5, and 12-13 weeks after kainate, using depth electrodes in the ipsilateral hippocampus. Latency to onset of electrographic SRS and the incidence of electrographic SRS were similar in WT and KO mice. However, the frequency of electrographic seizures was lower whereas the frequency of electroclinical seizures was higher in Nkcc1 KO mice, indicating a facilitated progression from electrographic to electroclinical seizures during chronic epilepsy, and a more severe epileptic phenotype, in the absence of NKCC1. The present findings suggest that NKCC1 is dispensable for the induction, progression and manifestation of epilepsy, and they do not support the widely held notion that inhibition of NKCC1 in the brain is a useful strategy for preventing or modifying epilepsy.Peer reviewe

Melanocortin-4 receptor gene: case-control study and transmission disequilibrium test confirm that functionally relevant mutations are compatible with a major gene effect for extreme obesity

We initially performed a mutation screen of the coding region of the MC4R in 808 extremely obese children and adolescents and 327 underweight or normal-weight controls allowing for a case-control study. A total of 16 different missense, nonsense, and frameshift mutations were found in the obese study group; five of these have not been observed previously. In vitro assays revealed that nine [the haplotype (Y35X; D37V) was counted as one mutation] of the 16 mutations led to impaired cAMP responses, compared with wild-type receptor constructs. In contrast, only one novel missense mutation was detected in the controls, which did not alter receptor function. The association test based on functionally relevant mutations was positive (P = 0.006, Fisher's exact test, one-sided). We proceeded by screening a total of 1040 parents of 520 of the aforementioned obese young index patients to perform transmission disequilibrium tests. The 11 parental carriers of functionally relevant mutations transmitted the mutation in 81.8% (P = 0.033; exact one-sided McNemar test). These results support the hypothesis that these MC4R mutations represent major gene effects for obesity

TGFβ-Signaling and FOXG1-Expression Are a Hallmark of Astrocyte Lineage Diversity in the Murine Ventral and Dorsal Forebrain

Heterogeneous astrocyte populations are defined by diversity in cellular environment, progenitor identity or function. Yet, little is known about the extent of the heterogeneity and how this diversity is acquired during development. To investigate the impact of TGF (transforming growth factor) β-signaling on astrocyte development in the telencephalon we deleted the TGFBR2 (transforming growth factor beta receptor 2) in early neural progenitor cells in mice using a FOXG1 (forkhead box G1)-driven CRE-recombinase. We used quantitative proteomics to characterize TGFBR2-deficient cells derived from the mouse telencephalon and identified differential protein expression of the astrocyte proteins GFAP (glial fibrillary acidic protein) and MFGE8 (milk fat globule-EGF factor 8). Biochemical and histological investigations revealed distinct populations of astrocytes in the dorsal and ventral telencephalon marked by GFAP or MFGE8 protein expression. The two subtypes differed in their response to TGFβ-signaling. Impaired TGFβ-signaling affected numbers of GFAP astrocytes in the ventral telencephalon. In contrast, TGFβ reduced MFGE8-expression in astrocytes deriving from both regions. Additionally, lineage tracing revealed that both GFAP and MFGE8 astrocyte subtypes derived partly from FOXG1-expressing neural precursor cells