Moderation of alcohol consumption as a recommendation in European hypertension management guidelines: a survey on awareness, screening and implementation among European physicians

Objectives: Moderation of alcohol consumption is included as a class I, level of evidence A recommendation in the current European guidelines for the management of hypertension. We investigated its awareness and self-reported implementation among European physicians across different specialties and workplaces. Design and setting: A cross-sectional survey study conducted in two annual German meetings (German Society of Cardiology and the German Society of Internal Medicine) and two annual European meetings (European Society of Hypertension and European Society Cardiology) in 2015. Participants: 1064 physicians attending the European meetings were interviewed including 52.1% cardiologists, 29.2% internists and 8.8% general practitioners. Main outcome measures: Physician screening of alcohol consumption, awareness and self-implementation of the recommendation of the current European guidelines about moderation of alcohol consumption for the management of hypertension. Results: Overall, 81.9% of physicians reported to generally quantify alcohol consumption in patients with hypertension. However, only 28.6% and 14.5% of participants reported screening alcohol consumption in their patients with newly detected or treatment-resistant hypertension. Physicians recommended a maximum alcohol intake of 13.1 +/- 11.7 g/day for women (95% CI 12.3 to 13.8) and 19.9 +/- 15.6 g/day for men (95% CI 18.8 to 20.9). In case of moderate to high alcohol consumption, 10.3% would manage only hypertension without addressing alcohol consumption, while 3.7% of the physicians would do so in case of alcohol dependence (p<0.001). Conclusions: The average amount of alcohol intake per day recommended by European physicians in this survey was in agreement with the guidelines. The low number of physicians that screen for alcohol consumption in patients with newly detected and with treatment-resistant hypertension indicates an important deficit in the management of hypertension

Herb-Induced Liver Injury in the Berlin Case-Control Surveillance Study

Herb-induced liver injury (HILI) has recently attracted attention due to increasing reports of hepatotoxicity associated with use of phytotherapeutics. Here, we present data on HILI from the Berlin Case-Control Surveillance Study. The study was initiated in 2000 to investigate the serious toxicity of drugs including herbal medicines. Potential cases of liver injury were ascertained in more than 180 Departments of all 51 Berlin hospitals from October 2002 to December 2011. Drug or herb intake was assessed through a standardized face- to-face interview. Drug or herbal aetiology was assessed based on the updated Council for International Organizations of Medical Sciences scale. In ten of all 198 cases of hepatotoxicity included in the study, herbal aetiology was assessed as probable (once ayurvedic herb) or possible (Valeriana five times, Mentha piperita once, Pelargonium sidoides once, Hypericum perforatum once, Eucalyptus globulus once). Mean age was 56.4 ± 9.7 years, and the predominant pattern of liver injury was hepatocellular. No cases of acute liver failure or death were observed. This case series corroborates known risks for ayurvedic herbs, supports the suspected association between Valeriana use and liver injury, and indicates a hepatotoxic potential for herbs such as Pelargonium sidoides, Hypericum perforatum or Mentha piperita that were rarely associated with liver injury before. However, given that possible causality does not prove clinical significance, further studies in this field are needed

Effect of a common UMOD variant on kidney function, blood pressure, cognitive and physical function in a community-based cohort of older adults

In genome-wide association studies, genetic variants in the UMOD gene associate with kidney function, blood pressure (BP), and hypertension. Elevated BP is linked to kidney function and impaired cognitive as well as physical performance in later life. We investigated the association between UMOD rs4293393-A > G and kidney function, BP, cognitive and physical function in the Berlin Aging Study II (BASE-II). Data of 1556 older BASE-II participants (mean age 68.2 +/- 3.7 years) were analyzed. BP was determined by standardized automated measurements, estimated glomerular filtration rate (eGFR) by CKD Epidemiology Collaboration creatinine equation. Cognitive function was assessed by Mini-Mental State Examination and Digit Symbol Substitution Test, while physical function by Handgrip Strength and Timed Up and Go-Test. Association analyses were performed by covariance and logistic regression models adjusting for sex. G-allele carriers at UMOD rs4293393 exhibited significantly higher eGFR values compared to non-carriers (AA, 76.4 ml/min/1.73 m(2), CI: 75.7-77.2 vs. AG, 78.4 ml/min/1.73 m(2), CI: 77.3-79.5 vs. GG, 78.5 ml/min/1.73 m(2), CI: 75.4-81.7; P = 0.010), and a lower risk of eGFR < 60 mL/min/1.73 m(2) (AG, OR: 0.63, CI: 0.41-0.97, P = 0.033). However, UMOD rs4293393 genotypes were not associated with BP, diagnosis of hypertension or cognitive and physical function parameters. Our data corroborate previous findings on the association of UMOD rs4293393-G with better kidney function in older adults. However, no association between UMOD and BP or physical and cognitive parameters in these community-dwelling older adults was detected

Anticontractile Effect of Perivascular Adipose Tissue and Leptin are Reduced in Hypertension

Leptin causes vasodilatation both by endothelium-dependent and -independent mechanisms. Leptin is synthesized by perivascular adipose tissue (PVAT). The hypothesis of this study is that a decrease of leptin production in PVAT of spontaneously hypertensive rats (SHR) might contribute to a diminished paracrine anticontractile effect of the hormone. We have determined in aorta from Wistar-Kyoto (WKY) and SHR (i) leptin mRNA and protein levels in PVAT, (ii) the effect of leptin and PVAT on contractile responses, and (iii) leptin-induced relaxation and nitric oxide (NO) production. Leptin mRNA and protein expression were significantly lower in PVAT from SHR. Concentration-response curves to angiotensin II were significantly blunted in presence of PVAT as well as by exogenous leptin (10−9 M) only in WKY. This anticontractile effect was endothelium-dependent. Vasodilatation induced by leptin was smaller in SHR than in WKY, and was also endothelium-dependent. Moreover, release of endothelial NO in response to acute leptin was higher in WKY compared to SHR, but completely abolished in the absence of endothelium. In conclusion, the reduced anticontractile effect of PVAT in SHR might be attributed to a reduced PVAT-derived leptin and to an abrogated effect of leptin on endothelial NO release probably due to an impaired activation of endothelial NO synthase

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

Key Learning Outcomes for Clinical Pharmacology and Therapeutics Education in Europe: A Modified Delphi Study.

Harmonizing clinical pharmacology and therapeutics (CPT) education in Europe is necessary to ensure that the prescribing competency of future doctors is of a uniform high standard. As there are currently no uniform requirements, our aim was to achieve consensus on key learning outcomes for undergraduate CPT education in Europe. We used a modified Delphi method consisting of three questionnaire rounds and a panel meeting. A total of 129 experts from 27 European countries were asked to rate 307 learning outcomes. In all, 92 experts (71%) completed all three questionnaire rounds, and 33 experts (26%) attended the meeting. 232 learning outcomes from the original list, 15 newly suggested and 5 rephrased outcomes were included. These 252 learning outcomes should be included in undergraduate CPT curricula to ensure that European graduates are able to prescribe safely and effectively. We provide a blueprint of a European core curriculum describing when and how the learning outcomes might be acquired

Genetics and pharmacogenetics of renal diseases – implications for a personalized therapy

Ein gemeinsames Ziel pharmakogenetischer und pharmakogenomischer Forschung besteht darin, die Effekte der individuellen genetischen Ausstattung auf die Sicherheit und Effektivität einer etablierten Arzneimitteltherapie zu analysieren [1,3,7]. Im Hinblick auf die klinische Anwendung soll dadurch für den zu behandelnden Patienten ein optimaler Therapieeffekt bei einem Minimum an unerwünschten Arzneimittelreaktionen erzielt werden [1]. Im klassischen Sinn beschäftigt sich die Pharmakogenetik dabei mit Genvarianten in Proteinen, die für pharmakokinetische und –dynamische Prozesse bedeutsam sind [3]. Die genomweite Betrachtungsweise ermöglicht neben der Berücksichtigung der Summeneffekte genetischer Variabilität [1] die Identifizierung von Varianten, die mit der Prädisposition gegenüber Erkrankungen assoziiert sind und damit als Wegweiser für neue therapeutische Zielstrukturen in Frage kommen [12]. Die vorliegende Habilitationsschrift fasst fünf Arbeiten zusammen, in denen diese Aspekte bei renalen und kardiovaskulären Erkrankungen am Beispiel von zwei Kandidatengenen untersucht wurden. Die ersten drei Arbeiten stellen dabei klassische pharmakogenetische Analysen zur Genetik der Expression des Phase I-Enzyms CYP3A5 und ihrer Bedeutung bei Patienten nach Nierentransplantation dar. Da die systemische und lokale Exposition gegenüber dem CYP3A5-Substrat Ciclosporin und seinen Metaboliten die Toxizität des Calcineurin-Inhibitors beeinflusst [44,63], wurde zunächst (2.1.1) untersucht, ob der CYP3A5-Genotyp in Analogie zur hepatischen Expression auch die renale CYP3A5-Enzymexpression differentiell beeinflusst [81]. Die Arbeit bestätigte die in kleineren Analysen bereits vorbeschriebene renale Genotyp-Phänotyp-Korrelation für CYP3A5 in gesundem Nierengewebe [35,65]. Der Genotyp steuert dabei die Höhe der CYP3A5-Proteinexpression in den Zellen des proximalen Tubulus. Diese Ergebnisse konnten wir nachfolgend (2.1.2) auf die pathophysiologische Situation bei akuter Nierentransplantat-Rejektion erweitern [82]. Das CYP3A5*1-Allel des Spenders determinierte auch in Rejektionsbiopsaten sowohl eine höhere mRNA-Expression als auch eine stärkere Proteinexpression im proximalen Tubulus. Als neuen Befund detektierten wir zudem eine Assoziation des Spender-Genotyps und der CYP3A5-Expressionsspiegel im Nierentransplantat mit dem Erfolg einer Glukokortikoid-Therapie. Im Rahmen der dritten Arbeit (2.1.3) zeigte sich ein positiver Effekt des CYP3A5*1-Allels des Empfängers auf das Patientenüberleben nach Nierentransplantation [83]. Dies erschien plausibel, da der CYP3A5-Genotyp sowohl das Ciclosporin-Metabolitenprofil lokal in der Niere [58] als auch im systemischen Kreislauf beeinflusst [62]. Zwei sehr umfangreiche mittlerweile publizierte Studien zum Einfluss verschiedener genetischer Varianten auf das Nierentransplantatüberleben und die Mortalität der Empfänger [100,101] konnten unsere Befunde für den CYP3A5-Genotyp jedoch nicht bestätigen. Auf der Grundlage von Assoziationsstudien gelten Promotorvarianten im Uromodulin-Gen (UMOD) als vielversprechende Kandidaten für verschiedene renale Erkrankungen [67] und die arterielle Hypertonie [104]. Die vierte (2.2.1) und fünfte (2.2.2) hier vorgestellte Arbeit zur Bedeutung der UMOD-Variante rs12917707 für die Nierenfunktion [84,85] stellen daher beispielhaft den zweiten Aspekt genetischer Untersuchungen – die Analyse eines krankheitsmodifizierenden Einflusses – dar. In der Arbeit 2.2.1 bei Patienten nach Nierentransplantation [84] hatte das UMOD-Allel rs12917707-T der Spenderniere keinen Einfluss auf die Höhe der Serumkreatinin-Konzentration nach Transplantation. Grund hierfür könnte das geringe mittlere Alter (< 45 Jahre) in unserem Patientenkollektiv sein, da sich UMOD-Effekte auf Parameter der Nierenfunktion altersabhängig deutlicher darstellen [77,80]. Ein signifikanter positiver Effekt des protektiven UMOD-Allels des Spenders auf das Transplantatüberleben zeigte sich erst in der adjustierten multivariaten Analyse nach Ausschluss der Todesfälle mit funktionsfähigem Transplantat. In der letzten hier eingeschlossenen Publikation (2.2.2) konnten wir den in populationsbasierten Studien detektierten günstigen Einfluss des rs12917707-T Allels auf die glomeruläre Filtrationsrate auch bei Patienten mit arterieller Hypertonie und manifester kardiovaskulärer Erkrankung nachweisen [85]. Als neuer Befund zeigte sich zudem eine Assoziation mit dem linksatrialen Durchmesser. Dies deutet auf ein Wechselspiel kardialer und renaler Funktionen hin, das durch UMOD moduliert werden könnte. Die hier zusammengeführten Arbeiten verweisen am Beispiel von CYP3A5 und UMOD auf das Potential genetischer Untersuchungen für eine individualisierte Therapie. Der Erfolg dieser Ansätze ist jedoch unter anderem abhängig von der Anzahl mitbeteiligter Varianten, deren Allelfrequenzen und jeweiligen Effektgrößen [1]. Darüber hinaus stellen Varianten in Einzelgenen lediglich eine Variable unter vielen Faktoren mit Einfluss auf eine definierte Arzneimitteltherapie dar. Auch deshalb sollten unsere neuartigen Befunde in weiteren Studien verifiziert werden.A common objective of both pharmacogenetic and pharmacogenomic research is to analyze the effects of an individual's genetic composition on safety and efficacy of an established drug therapy [1,3,7]. With regard to the clinical application, this approach is meant to achieve an optimal drug response while reducing adverse drug reactions for the individual patient [1]. In its conventional meaning, pharmacogenetics deals with single genetic variations for proteins involved in pharmacokinetic and pharmacodynamic processes [3]. The genome-wide approach enables both consideration of sum effects of genetic variability [1] and identification of variants associated with disease susceptibility, the latter serving as markers for possible new therapeutic targets [12]. The professorial dissertation presented here combines five studies in which these aspects were investigated in renal and cardiovascular diseases using the example of two candidate genes. The first three publications represent pharmacogenetic analyses on the genotype-dependent expression of the phase I-metabolizing enzyme CYP3A5 and its relevance for patients after kidney transplantation. As systemic and local exposure to the CYP3A5-substrate cyclosporine and its metabolites affects the toxicity of this calcineurin inhibitor [44,63], we initially (2.1.1) investigated whether the CYP3A5-genotype differentially affects the renal CYP3A5 enzyme expression comparable to the hepatic expression [81]. Our work corroborated a renal genotype-phenotype-correlation for CYP3A5 in normal human kidney samples as had been described in smaller analyses [35,65]. The genotype determines the level of CYP3A5 protein expression in epithelial cells of the proximal tubule. Afterwards (2.1.2), we were able to extend these results to the pathophysiological situation of acute kidney transplant rejection [82]. In renal biopsies during acute rejection, the donor CYP3A5*1-allele likewise determines higher intragraft mRNA levels and a more intense protein expression in proximal tubules. As a novel finding we detected an association of responsiveness to steroid treatment during acute rejection with the donor CYP3A5*1-allele and CYP3A5 expression levels within the renal allograft. The third work (2.1.3) revealed a protective effect of the recipient CYP3A5*1-allele on patient survival after kidney transplantation [83]. This seemed plausible as the CYP3A5-genotype affects the profile of cyclosporine metabolites both locally in the kidney [58] and systemic circulation [62]. By now, however, two large studies on the effect of several genetic variants on kidney graft survival and recipient survival have been published [100,101] which did not confirm our results for the CYP3A5-genotype. Based on genome- wide association studies, variants in the promoter region of the uromodulin gene (UMOD) are promising candidates for different renal diseases [67] as well as arterial hypertension [104]. Thus, the fourth (2.2.1) and fifth (2.2.2) work on the impact of the UMOD-variant rs12917707 on renal function presented here [84,85] exemplify the second aspect of genetic research by analyzing the potential disease-modifying effect of genetic variants. In patients after kidney transplantation (2.2.1), the donor UMOD-allele rs12917707-T had no effect on serum creatinine concentrations after transplantation [84]. This may be attributable to the young age of our patients (mean < 45 years) as UMOD- effects on parameters of renal function are more pronounced with increasing age [77,80]. A significant positive influence of the protective donor UMOD- allele on graft survival was only observed in multivariate adjusted analysis after exclusion of deaths with a functioning graft. In the last study (2.2.2), we were able to extend the documented role of the rs12917707-T allele on glomerular filtration rate to patients with arterial hypertension and clinically apparent cardiovascular disease [85]. The work moreover revealed a novel association between rs12917707 and left atrial diameter. This suggests a cardiorenal interplay which may be modulated by UMOD. The studies presented here point out the potential impact of genetic research for an individualized therapy as exemplified for CYP3A5 and UMOD. However, the success of these approaches depends – amongst others – on the number of contributing genetic variants, their allele frequencies, and respective effect sizes [1]. Furthermore, variants in single genes are only one among many factors influencing a given drug therapy. Hence, our novel findings should be verified in further studies accounting for these aspects

Cloning of the promoter of human endothelin-converting enzyme (ECE)-1c and identification of regulatory promoter elements

Titelblatt, Inhaltsverzeichnis,Danksagung und Lebenslauf Abkürzungsverzeichnis Einleitung Fragestellung Material und Methoden Ergebnisse Diskussion Zusammenfassung Literatur VeröffentlichungenIn der vorliegenden Arbeit konnte das c-spezifische Exon des humanen Gens für ECE-1 genomisch lokalisiert, die Startpunkte der Transkription von ECE-1c mRNA lokalisiert und der ECE-1c Promotor kloniert sowie funktionell charakterisiert werden. Das Exon ECE-1c liegt 55 kbp 5´ des b-spezifischen Exons und ist damit das am weitesten 5´ lokalisierte Exon des humanen ECE-1 Gens. Der ECE-1c- spezifische Promotor enthält keine TATA-Box und kein Inr-Element, eine zusammen mit dem GC-Reichtum des Promotors typische Konstellation für einen housekeeping-Promotor. Durch das Fehlen eindeutiger Kernpromotorelemente verläuft die Transkriptionsinitiation bei TATA(Inr(-Promotoren häufig unpräzise. Auch im ECE-1c Promotor existieren multiple Transkriptionsstartpunkte, die mittels RPA an den Positionen (110, (140 und (350 bp relativ zum Translationsstartkodon in Exon 1c detektiert werden konnten, wobei die Subklonierung und anschließende Sequenzierung der RACE- Produkte eng benachbarte Startpunkte bei (101 und (106 bp bestätigte. Als mögliche cis-Elemente, die anstelle der TATA-Box und eines Inr die basale Transkription der ECE-1c mRNA initiieren und damit den Transkriptionsstart festlegen können, finden sich in dem Sequenzabschnitt zwischen (142 und (240 Konsensussequenzen für E2F ((154), Sp1 ((180) sowie zwei CAAT-Boxen ((196 und (220). Die Transfektionsexperimente in der vorliegenden Arbeit zeigten für diesen Promotorbereich den stärksten Anstieg in der RLA als Ausdruck der Promotoraktivität. Beide CAAT-Boxen im ECE-1c Promotor sind in passendem Abstand zu den ermittelten Transkriptionsstartpunkten bei etwa (110 und (140 lokalisiert. Die Konsensussequenz für E2F bei (154 bp vom Translationsstartkodon in Exon 1c liegt direkt 5´ einer der detektierten Transkriptionsstartpunkte bei etwa (140 bp. Die durch Punktmutation dieses cis-Elementes nachgewiesene starke Reduktion der Promotoraktivität spricht für die zentrale Bedeutung von E2F bei der basalen Transkriptionsregulation von ECE-1c. Im ECE-1c Promotor liegen die Konsensussequenzen für die Bindung von Sp1 40 bp bzw. 70 bp 5´ der Startpunkte bei (140 bzw. (110. GATA und ETS kommen als regulatorische Transkriptionsfaktoren des ECE-1c Promotors in Betracht. In der distal gelegenen Promotorregion von ECE-1c finden sich zwei CA-Mikrosatelliten an den Positionen (507 bis (542 und (797 bis (836 sowie ein CG-Mikrosatellit zwischen (543 und (564, deren funktionell wirksame Polymorphismen später gezeigt werden konnten. Bedenkt man die Schlüsselrolle von ECE-1 im Rahmen der Endothelinbiosynthese, könnte diesem Promotorpolymorphismus eine pathophysiologische Bedeutung bei der Ausprägung kardiovaskulärer Erkrankungen zukommen. Der ECE-1c Promotor zeigte im Vergleich zu den ECE-1a, -1b und -1d Promotoren eine deutlich stärkere Reporteraktivität, die sowohl zelltyp-, als auch speziesunabhängig nachgewiesen werden konnte. Die vorliegende Arbeit liefert somit erste Hinweise auf die transkriptionelle Regulation der am stärksten exprimierten ECE-1 Isoform, was zu einem besseren Verständnis der Regulation des Endothelinsystems während der Entwicklung und unter pathophysiologischen Bedingungen beitragen kann.In this work the c-isoform specific exon of the human ECE-1 gene was localized in the genome and the start points of ECE-1c mRNA transcription were determined, followed by subcloning and functional characterization of the ECE- 1c promoter. The c-specific exon is located 55 kbp upstream of the b-specific exon and represents the leading exon of the human ECE-1 gene. The c-specific promoter lacks a TATA-box and an Inr, a constellation - in conjunction with its GC-abundance - which is typical for housekeeping promoters. The lack of classical core promoter elements in TATA(Inr(-promoters often leads to imprecise initiation of transcription. RNase protection assay revealed multiple transcriptional start points for the ECE-1c promoter at -110, -140 and -350 bp relative to the translational start codon in exon 1c. Subcloning and sequencing of the 5´-RACE products confirmed two adjacent transcriptional start points at -101 and -106. Putative cis-elements which could initiate the basal transcription of ECE-1c mRNA are E2F (-154), Sp1 (-180) and two CAAT- boxes (-196 and -220). Transfection experiments showed the strongest increase in promoter activity in association with this promoter region. Both CAAT-boxes are located in appropriate distance to the transcriptional start points at -110 and -140, respectively. The consensus element E2F at -154 bp upstream of the translational start codon in exon 1c is located immediately 5´ of the transcriptional start point at -140 bp. Site-directed mutagenesis of this E2F consensus dramatically decreased ECE-1c promoter activity which strongly supports the importance of this element for initiation of transcription. There are also two consensus sites for Sp1 binding in the ECE-1c promoter, located 40 bp and 70 bp upstream of the transcriptional start points at -140 and -110, respectively. Additionaly, GATA and ETS factors are potentially involved in promoter regulation. Two CA-microsatellites (-507 to -542 and -797 to -836) and one CG-microsatellite (-543 to -564) are located in the more distal promoter region. Their functional polymorphisms were demonstrated in subsequent studies. The ECE-1c promoter showed distinctly stronger activity compared to the alternative ECE-1 promoters, irrespective of cell-type or species tested. In conclusion, this work provides first insights into the transcriptional regulation of the major ECE-1 isoform which may lead to a better understanding of the regulation of the endothelin system in developmental and pathophysiological conditions