Der Urologe

Die Arbeitszeitbelastung von Fach- und Oberärzten in der deutschen Urologie – eine Bestandsaufnahme Zusammenfassung Hintergrund Die Arbeits- und Weiterbildungsbedingungen von urologischen Assistenzärzten in Deutschland sind bereits analysiert worden. Aber wie steht es um die urologischen Oberärzte in Deutschland? Unter welchen Bedingungen müssen diese arbeiten? Bislang existieren keine uns bekannten publizierten Daten, die diese Konditionen dieser speziellen urologischen Berufsgruppe beleuchten, daher stellen wir die Ergebnisse der vorliegenden Umfrage vor. Ziel der Arbeit Die Analyse der aktuellen Arbeitsbedingungen urologischer Fach- und Oberärzte in Deutschland möglichst umfassend zu erfragen und auszuwerten. Material und Methoden Der Arbeitskreis Angestellter Ärzte des BvDU führte eine Befragung im Rahmen des Urologischen Oberarztforums 2016 sowie online über den Email-Verteiler der Deutschen Gesellschaft für Urologie (DGU) durch. Erfragt wurden die Arbeitsbelastung, Arbeitsbedingungen und -zufriedenheit, geleistete Überstunden sowie Arbeitszeit- und Opt-out-Regelungen. Ergebnisse An der Befragung nahmen 176 Oberärzte teil, davon waren 88 % männlich und 12 % weiblich. In Deutschland gibt es 1125 Oberärzte. Das Durchschnittsalter der Befragten betrug 44,9 Jahre. Bei >80 % der Befragten lag die durchschnittliche Wochenarbeitszeit bei über 50 h, 70 % unterschrieben eine Opt-out-Regelung. Es konnte dabei ein Zusammenhang zwischen einer steigenden Jobunzufriedenheit ab einer Wochenarbeitszeit über 55 h bzw. einer durchschnittlichen Anwesenheit im Rufbereitschaftsdienst über 4 h hergestellt werden. Die Anzahl der Rufbereitschaftsdienste hatte auf die Zufriedenheit keinen Einfluss. Insgesamt gaben 43 % (70 von 162) der Befragten an, dass regelmäßig Überstunden verfallen, bei 12 % (20 von 162) verfielen alle Überstunden. Rund 30 % der Oberärzte in der deutschen Urologie sind mit ihren aktuellen Arbeitsbedingungen in der aktuellen Form unzufrieden. Schlussfolgerungen Anpassungen und Verbesserungen der Arbeitsbedingungen urologischer Oberärzte sind auf Grundlage der erhobenen Daten erforderlich, um die Attraktivität des Berufsbildes und Führungspositionen in der deutschen Urologie zu erhalten.

Konzepte zur besseren Vereinbarkeit von klinischer und wissenschaftlicher Tätigkeit – ein Thesenpapier der Arbeitsgemeinschaft „Junge Urologen" der Deutschen Gesellschaft für Urologie

Qualität statt Quantität verbessert die Weiterbildung Zusammenfassung Eine Weiterbildungsevaluation findet in Deutschland zum aktuellen Zeitpunkt zwar statt – allerdings nicht flächendeckend und regelmäßig und ohne bundesweiten Standard. Vergleiche zwischen verschiedenen Kliniken und Ausbildern können daher im Moment nicht gezogen werden. Umfragemodi, wie beispielsweise in der Schweiz seit Jahrzehnten existierend, könnten eine konstruktive Grundlage in der Erhebung des aktuellen Weiterbildungsstands urologischer Assistenzärzte sein und nachfolgend die Diskussion über Verbesserungen ermöglichen. Dazu bedarf es einer berufsgruppenübergreifenden konstruktiven Zusammenarbeit, ohne die Verantwortung lediglich den Weiterbildnern zuzuschreiben. Darüber hinaus sollten Modelle der Entlohnung guter Weiterbildung gefunden werden.

Systemwandel zu erwarten. Ein Appell an eine andere Art des Wirtschaftens aus der Sicht des Chefarztes

LUTS bei BPH-Patienten

Aktuelle Forschung pädiatrischer und adulter Keimzelltumoren

Mitteilungen der DGU

Molekulares Tumorboard Prostatakarzinom Zusammenfassung In der modernen Onkologie ist das molekulare Tumorboard die Schnittstelle zwischen Klinik und Grundlagenforschung. Durch die interdisziplinäre Zusammenarbeit von Experten verschiedener Fachdisziplinen soll die Indikation zur molekularbiologischen Tumoranalysen sinnvoll gestellt, die Untersuchungsergebnisse korrekt interpretiert und hierauf basierende personalisierte Therapieempfehlungen ausgegeben werden. Bezogen auf das metastasierte Prostatakarzinom können hiervon insbesondere Patienten mit familiärem Prostatakarzinom, jungen Manifestationsalter oder nach Ausschöpfung von zugelassenen Standardtherapien profitieren. Mit der Androgenrezeptor-Splicevariante 7 (AR-V7) steht ein prädiktiver Marker für das Ansprechen auf eine Hormontherapie mit Abirateron oder Enzalutamid zur Verfügung. Testverfahren zur Bestimmung des AR-V7-Status im Blut sind bereits kommerziell erwerblich. Mutationen in den DNA-Reparaturmechanismen stellen einen weiteren Ansatzpunkt für zielgerichtete, personalisierte Therapien dar. Defekte in der homologen Rekombination erhöhen die Empfindlichkeit von Tumorzellen gegenüber Poly(ADP-Ribose)-Polymerase-(PARP-)Inhibitoren wie Olaparib. In der TOPARP-A-Studie, eine Phase-II-Studie, lag die Ansprechrate bei Patienten mit metastasiertem Prostatakarzinom und Mutationen in DNA-Reparaturgenen bei 88 %. Mikrosatelliteninstabilität ist die Folge von Defekten in der DNA-Mismatch-Reparatur. Mit dem Immun-Checkpoint-Inhibitor Pembrolizumab steht auch für diese Patientenkollektiv eine wirksame Therapie zur Verfügung. Da weder PARP-Inhibitoren noch Pembrolizumab aktuell zur Therapie des metastasierten Prostatakarzinoms in Deutschland zugelassen sind, stärkt die Empfehlung eines molekularen Tumorboards die Erfolgsaussichten für die Genehmigung eines individuellen Heilversuches durch die Krankenkassen.

Molekulares Tumorboard – Nierenzellkarzinom Zusammenfassung Die Einführung molekularer zielgerichteter Substanzen hat die Therapie des metastasierten Nierenzellkarzinoms grundlegend verändert. Eine erste Welle der Entwicklung basiert auf dem verbesserten Verständnis der Tumorbiologie seit der Entdeckung der Bedeutung des VHL-Gens für die Progression, was den Weg für Therapieoptionen geebnet hat, die v. a. die Neoangiogenese hemmen. Relevant sind hier die Überexpression proangiogener und proliferationsfördernder Faktoren (VEGF, „vascular endothelial growth factor"; PDGF; „platelet derived growth factor") sowie eine Überaktivierung des PI3K- (Phosphatidylinositol-3-Kinase‑)Akt-Signalwegs: Zielstruktur ist hierbei das „mammalian target of rapamycin" (mTOR-)Molekül, das an der Regulation zellproliferativer Prozesse beteiligt ist. Vor allem die VEGF-, PDGF- und mTOR-Signale bzw. Signalwege sind zentrale Angriffspunkte aktueller zielgerichteter Substanzen. Eine zweite Welle ist in der Entwicklung von Therapieansätzen mit der zielgerichteten Aktivierung und Modulation des Immunsystems zu sehen, die die „Immuntherapie" wieder in den Fokus des Interesses gerückt hat. Zentrale Entwicklung ist die Anwendung von Immun-Checkpoint-Inhibitoren, mit deren Hilfe eine (Re‑)Aktivierung der zellulären Abwehr und hier v. a. von T‑Zellen erfolgt, was per se das Potenzial einer zytoreduktiven Therapie durch Abtöten der Tumorzellen in sich birgt. Auch, wenn sich die Prognose durch die rasante Entwicklung der letzten Jahre signifikant verbesserte, bleibt die Behandlung anspruchsvoll, da die meisten Patienten einen Progress erleben müssen und ein Langzeitüberleben in nur etwa 20 % der Fälle erreicht werden kann, da eben auch ein Teil der Patienten primär refraktär ist oder nicht anspricht.

Ambulante Onkologie in der Urologischen Praxis – Möglich? Machbar? Perspektiven

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Drugs

Volanesorsen: First Global Approval Abstract Volanesorsen (Waylivra®), an antisense oligonucleotide inhibitor of apolipoprotein CIII (apoCIII) mRNA, is being developed by Ionis Pharmaceuticals through its subsidiary company, Akcea Therapeutics, to treat familial chylomicronemia syndrome (FCS), hypertriglyceridemia and familial partial lipodystrophy (FPL). In May 2019, volanesorsen was approved in the EU for the treatment of adult patients with FCS based on positive results from the multinational, phase III APPROACH and COMPASS studies. Other clinical trials are ongoing to assess its utility in hypertriglyceridemia, FPL and partial lipodystrophy. This article summarizes the milestones in the development of volanesorsen leading to this first approval as an adjunct to diet in adult patients with genetically confirmed FCS and at high risk for pancreatitis, in whom response to diet and triglyceride lowering therapy has been inadequate.

Correction to: Migalastat: A Review in Fabry Disease The article Migalastat: A Review in Fabry Disease, written by Emma H. McCafferty, Lesley J. Scott, was originally published Online First without open access. After publication in volume 79, issue 5, pages 543–554 [funder] requested that the article be Open Choice to make the article an open access publication. Post-publication open access was funded by [funder]. The article is forthwith distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International License (http://creativecommons.org/licenses/by-nc/4.0/), which permits any noncommercial use, duplication, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license and indicate if changes were made.

Eltrombopag in Immune Thrombocytopenia, Aplastic Anemia, and Myelodysplastic Syndrome: From Megakaryopoiesis to Immunomodulation Abstract Eltrombopag is an orally available thrombopoietin receptor agonist indicated for the treatment of immune thrombocytopenia (ITP). Beyond the effect on megakaryopoiesis, the drug also showed a stimulating effect on the hematopoietic stem cell with consistent clinical efficacy in aplastic anemia (AA) and myelodysplastic syndromes (MDS). Eltrombopag is highly effective in ITP and less so in AA and MDS. This observation underlines the importance of residual normal hematopoiesis, which is maximal in ITP, minimal/absent in AA, and dysregulated in MDS. In ITP, the drug at 50–75 mg daily induced up to 85% responses both in clinical trials and real-life studies, with the possibility of tapering and discontinuation. In AA, eltrombopag at 150 mg daily was effective in about 40% of cases relapsed/refractory to standard immunosuppression or ineligible for bone marrow transplant. In MDS, the drug seems less effective, with responses in about a quarter of patients at various schedules. The efficacy of eltrombopag in ITP, AA, and MDS suggests the existence of common immune-pathological mechanisms in these diseases, including autoimmunity against peripheral blood cells and bone marrow precursors, as well as a possible evolution of one condition into the other. Additional mechanisms of action emerging from the clinical use of eltrombopag include modulation of T-regulatory cells, restoration of Fc-γ receptor balance in phagocytes, and an iron-mobilizing effect. In this review, we analyzed the most recent literature on eltrombopag use and efficacy in patients with ITP, AA, and MDS, exploring the basis for different dosing, combined treatments, and discontinuation in each context.

Current Treatments and New Developments in the Management of Glucocorticoid-induced Osteoporosis Abstract Glucocorticoids (GCs) are often used for improvement of quality of life, particularly in the elderly, but long-term GC use may cause harm; bone loss and fractures are among the most devastating side effects. Fracture risk is particularly high in patients with a severe underlying disease with an urgent need for treatment with high-dose GCs. Moreover, it is important to realize that these patients suffer from an augmented background fracture risk as these patients have a high presence of traditional risk factors for osteoporosis, such as high age, low body mass index (BMI), smoking and relatives with osteoporosis or hip fractures. It is thus crucial for prevention of osteoporotic fractures to use the lowest dose of GC for a short period of time to prevent fractures. Another important task is optimal treatment of the underlying disease; for instance, fracture risk is higher in patients with active rheumatoid arthritis than in patients in whom rheumatoid arthritis is in remission. Thus, fracture risk is generally highest in the early phase, when GC dosage and the disease activity of the underlying disease are high. Finally, some of the traditional risk factors can be modulated, e.g., smoking and low BMI. Life-style measures, such as adequate amounts of calcium and vitamin D and exercise therapy are also crucial. In some patients, anti-osteoporotic drugs are also indicated. In general, oral bisphosphonates (BPs) are the first choice, because of their efficacy and safety combined with the low cost of the drug. However, for those patients who do not tolerate oral BPs, alternatives ("second-line therapies") are available: BP intravenously (zoledronic acid), denosumab (Dmab), and teriparatide. Both zoledronic acid and Dmab have been proven to be superior to oral bisphosphonates like risedronate in improvement of bone mineral density. For teriparatide, vertebral fracture reduction has been shown in comparison with alendronate. Thus, to reduce the global burden of GC use and fracture risk, fracture risk management in GC users should involve at least involve life-style measures and the use of the lowest possible dose of GC. In high-risk patients, anti-osteoporotic drugs should be initiated. First choice drugs are oral BPs; however, in those with contraindications and those who do not tolerate oral BPs, second-line therapies should be started. Although this is a reasonable treatment algorithm, an unmet need is that the most pivotal (second-line) drugs are not used in daily clinical practice at the initial phase, usually characterized by high-dose GC and active underlying disease, when they are most needed. In some patients second-line drugs are started later in the disease course, with lower GC dosages and higher disease activity. As this is a paradox, we think it is a challenge for physicians and expert committees to develop an algorithm with clear indications in which specific patient groups second-line anti-osteoporotic drugs should or could be initiated as first-choice treatment.

Damoctocog Alfa Pegol: A Review in Haemophilia A Abstract Damoctocog alfa pegol (Jivi®) is approved in the USA, EU, Japan and Canada for the treatment and prophylaxis of previously treated patients aged ≥ 12 years with haemophilia A. Formulated with a 60 kDa polyethylene glycol (PEG) moiety, damoctocog alfa pegol is an intravenously (IV) administered recombinant factor VIII (rFVIII) product with a longer terminal half-life than non-PEGylated FVIII and rFVIII products. In the multinational phase II/III PROTECT VIII trial, prophylaxis with damoctocog alfa pegol reduced the likelihood of bleeding in previously treated patients aged ≥ 12 years with severe haemophilia A, with dosing schedules ranging from twice weekly to once every 7 days. Interim data from the ongoing extension phase indicated that the reduced annualized bleeding rates (ABRs) were maintained for up to 5.2 years of prophylaxis with damoctocog alfa pegol. Damoctocog alfa pegol was also effective in treating bleeding episodes and in providing haemostatic control during surgery. Damoctocog alfa pegol was generally well tolerated in adult and adolescent patients with severe haemophilia A, with most adverse events considered to be unrelated to treatment. There were no new or confirmed cases of FVIII inhibitor development and anti-PEG antibodies, observed in some patients, were of low titre and transient. Damoctocog alfa pegol extends the available treatment options in previously treated adults and adolescents with haemophilia A, offering the possibility of up to once-weekly administration for suitable patients.

Dapagliflozin: A Review in Type 2 Diabetes Abstract Dapagliflozin (Forxiga®) is a highly potent, reversible and selective sodium-glucose cotransporter-2 inhibitor indicated worldwide for the treatment of type 2 diabetes (T2D). In the EU, oral dapagliflozin once daily is approved for use as monotherapy (in patients who are intolerant of metformin) and as add-on combination therapy (with other glucose-lowering agents, including insulin) for T2D when diet and exercise alone do not provide adequate glycaemic control. In numerous well-designed clinical studies and their extensions, dapagliflozin as monotherapy and combination therapy with other antihyperglycaemic agents provided effective glycaemic control and reduced bodyweight and blood pressure (BP) across a broad spectrum of patients. Dapagliflozin reduced the rate of cardiovascular (CV) death or hospitalization for heart failure (HHF), did not adversely affect major adverse CV events (MACE) and possibly reduced progression of renal disease relative to placebo in patients with established atherosclerotic CV disease (CVD) or multiple risk factors for CVD. Dapagliflozin was generally well tolerated, with a low risk of hypoglycaemia; diabetic ketoacidosis (DKA), although rare, and genital infections were more common with dapagliflozin than placebo. Given its antihyperglycaemic, cardioprotective and possibly renoprotective properties and generally favourable tolerability profile, dapagliflozin provides an important option for the management of a broad patient population, regardless of the history of CVD.

Advances in Targeted Therapies for Triple-Negative Breast Cancer Abstract While the outcomes for patients diagnosed with hormone receptor positive (HR+) and/or human epidermal growth factor receptor 2-positive (HER2+) breast cancers have continued to improve with the development of targeted therapies, the same cannot be said yet for those affected with triple-negative breast cancer (TNBC). Currently, the mainstay of treatment for the 10–15% of patients diagnosed with TNBC remains cytotoxic chemotherapy, but it is hoped that through an enhanced characterization of TNBC biology, this disease will be molecularly delineated into subgroups with targetable oncogenic drivers. This review will focus on recent therapeutic innovations for TNBC, including poly-ADP-ribosyl polymerase (PARP) inhibitors, phosphoinositide 3-kinase (PI3K) pathway inhibitors, immune checkpoint inhibitors, and cyclin-dependent kinase (CDK) inhibitors.

Onasemnogene Abeparvovec: First Global Approval Abstract Onasemnogene abeparvovec (onasemnogene abeparvovec-xioi; formerly AVXS-101; ZOLGENSMA®) is an adeno-associated viral vector-based gene therapy designed to deliver a functional copy of the human survival motor neuron(SMN) gene to the motor neuron cells of patients with spinal muscular atrophy (SMA). It has been developed by AveXis, a Novartis company, and was approved in May 2019 in the USA for the treatment of paediatric patients aged < 2 years with SMA and bi-allelic mutations in the SMN1 gene (the primary gene encoding survival motor neuron protein). Onasemnogene abeparvovec is the first gene therapy to be approved for SMA in the USA. The recommended dose is 1.1 × 1014 vector genomes per kg of bodyweight, administered as a single intravenous infusion over 60 min. Regulatory assessments for this formulation of onasemnogene abeparvovec are underway in the EU and Japan; an intrathecal formulation is currently undergoing clinical development in the USA. This article summarizes the milestones in the development of onasemnogene abeparvovec leading to this first approval for the treatment of paediatric patients aged < 2 years with SMA and bi-allelic mutations in SMN1.

Recent Developments in LRRK2-Targeted Therapy for Parkinson's Disease Abstract Kinase activating missense mutations in leucine-rich repeat kinase 2 (LRRK2) are pathogenically linked to neurodegenerative Parkinson's disease (PD). Over the past decade, substantial effort has been devoted to the development of potent and selective small molecule inhibitors of LRRK2, as well as their preclinical testing across different Parkinson's disease models. This review outlines the genetic and biochemical evidence that pathogenic missense mutations increase LRRK2 kinase activity, which in turn provides the rationale for the development of small molecule inhibitors as potential PD therapeutics. An overview of progress in the development of LRRK2 inhibitors is provided, which in particular indicates that highly selective and potent compounds capable of clinical utility have been developed. We outline evidence from rodent- and human-induced pluripotent stem cell models that support a pathogenic role for LRRK2 kinase activity, and review the substantial experiments aimed at evaluating the safety of LRRK2 inhibitors. We address challenges still to overcome in the translational therapeutic pipeline, including biomarker development and clinical trial strategies, and finally outline the potential utility of LRRK2 inhibitors for other genetic forms of PD and ultimately sporadic PD. Collective evidence supports the ongoing clinical translation of LRRK2 inhibitors as a therapeutic intervention for PD is greatly needed.

Remogliflozin Etabonate: First Global Approval Abstract Remogliflozin, a selective sodium-glucose co-transporter subtype 2 (SGLT2) inhibitor, which is to be administered as remogliflozin etabonate (Remo™, Remozen™), the prodrug for remogliflozin, recently received its first approval as a treatment for type 2 diabetes mellitus (T2DM) in India. This article summarizes the milestones in the development of remogliflozin etabonate leading to this first approval for T2DM.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Histochemistry and Cell Biology

Immune response profile of primary tumour, sentinel and non-sentinel axillary lymph nodes related to metastasis in breast cancer: an immunohistochemical point of view Abstract Approximately 1.67 million new cases of breast cancer (BC) are diagnosed annually, and patient survival significantly decreases when the disease metastasizes. The axillary lymph nodes (ALNs) are the main doorway for BC tumoral cell escape, through which cells can disseminate to distant organs. The immune response, which principally develops in the lymph nodes, is linked to cancer progression, and its efficacy at controlling tumoral growth is compromised during the disease. Immunohistochemistry (IHC) is one of the most widely used research techniques for studying the immune response. It allows the measurement of the expression of particular markers related to the immune populations. This review focuses on the role of the immune populations in the primary tumour in the locoregional metastasis of the ALN, and the relationship of the immune response in these regions to distant metastasis. We considered only studies of immune cells using IHC techniques. In particular, lymphocytes, macrophages and dendritic cells all play important roles in BC and have been extensively studied. Although further research is needed, there is much evidence of their role in the invasion of the ALN and distant organs. Their association with tumoral growth or protection has not yet been demonstrated decisively and is very likely to be determined by a combination of factors. Moreover, even though IHC is a widely used technique in cancer diagnosis and research, there is still room for improvement, since its quantification needs to be properly standardized.

In focus in HCB

Correction to: Similarities and differences in tissue distribution of DLK1 and DLK2 during E16.5 mouse embryogenesis In the original publication of the article, some symbols in Figure 3 were not correctly aligned with the image.

Regulatory effects of PRF and titanium surfaces on cellular adhesion, spread, and cytokine expressions of gingival keratinocytes Abstract Dental implant material has an impact on adhesion and spreading of oral mucosal cells on its surface. Platelet-rich fibrin (PRF), a second-generation platelet concentrate, can enhance cell proliferation and adhesion. The aim was to examine the regulatory effects of PRF and titanium surfaces on cellular adhesion, spread, and cytokine expressions of gingival keratinocytes. Human gingival keratinocytes were cultured on titanium grade 4, titanium grade 5 (Ti5), and HA discs at 37 °C in a CO2 incubator for 6 h and 24 h, using either elutes of titanium-PRF (T-PRF) or leukocyte and platelet-rich fibrin (L-PRF), or mammalian cell culture medium as growth media. Cell numbers were determined using a Cell Titer 96 assay. Interleukin (IL)-1β, IL-1Ra, IL-8, monocyte chemoattractant protein (MCP)-1, and vascular endothelial growth factor (VEGF) expression levels were measured using the Luminex® xMAP™ technique, and cell adhesion and spread by scanning electron microscopy. Epithelial cell adhesion and spread was most prominent to Ti5 surfaces. L-PRF stimulated cell adhesion to HA surface. Both T-PRF and L-PRF activated the expressions of IL-1 β, IL-8, IL-1Ra, MCP-1, and VEGF, T-PRF being the strongest activator. Titanium surface type has a regulatory role in epithelial cell adhesion and spread, while PRF type determines the cytokine response.

Histochemistry and Cell Biology : 61 years and not tired at all

Region-specific changes in brain kisspeptin receptor expression during estrogen depletion and the estrous cycle Abstract Kisspeptin acts as a potent neuropeptide regulator of reproduction through modulation of the hypothalamic–pituitary–gonadal axis. Previous studies revealed sex differences in brain expression patterns as well as regulation of expression by estrogen. Alternatively, sex differences and estrogen regulation of the kisspeptin receptor (encoded by Kiss1r) have not been examined at cellular resolution. In the current study, we examined whether Kiss1r mRNA expression also exhibits estrogen sensitivity and sex-dependent differences using in situ hybridization. We compared Kiss1r mRNA expression between ovariectomized (OVX) rats and estradiol (E2)-replenished OVX rats to examine estrogen sensitivity, and compared expression between gonadally intact male rats and female rats in diestrus or proestrus to examine sex differences. In OVX rats, E2 replenishment significantly reduced Kiss1r expression specifically in the hypothalamic arcuate nucleus (ARC). A difference in Kiss1r expression was also observed between diestrus and proestrus rats in the hypothalamic paraventricular nucleus (PVN), but not in the ARC. Thus, estrogen appears to have region- and context-specific effects on Kiss1r expression. However, immunostaining revealed minimal colocalization of estrogen receptor alpha (ERα) in Kiss1r-expressing neuronal populations of ARC and PVN, indicating indirect or ERα-independent regulation of Kiss1r expression. Surprisingly, unlike the kisspeptin ligand, no sexual dimorphisms were observed in either the brain distribution of Kiss1r expression or in the number of Kiss1r-expressing neurons within enriched brain nuclei. The current study reveals marked differences in regulation between kisspeptin and kisspeptin receptor, and provides an essential foundation for further study of kisspeptin signaling and function in reproduction.

A temporal and spatial map of axons in developing mouse prostate Abstract Prostate autonomic and sensory axons control glandular growth, fluid secretion, and smooth muscle contraction and are remodeled during cancer and inflammation. Morphogenetic signaling pathways reawakened during disease progression may drive this axon remodeling. These pathways are linked to proliferative activities in prostate cancer and benign prostate hyperplasia. However, little is known about which developmental signaling pathways guide axon investment into prostate. The first step in defining these pathways is pinpointing when axon subtypes first appear in prostate. We accomplished this by immunohistochemically mapping three axon subtypes (noradrenergic, cholinergic, and peptidergic) during fetal, neonatal, and adult stages of mouse prostate development. We devised a method for peri-prostatic axon density quantification and tested whether innervation is uniform across the proximo–distal axis of dorsal and ventral adult mouse prostate. Many axons directly interact with or innervate neuroendocrine cells in other organs, so we examined whether sensory or autonomic axons innervate neuroendocrine cells in prostate. We first detected noradrenergic, cholinergic, and peptidergic axons in prostate at embryonic day (E) 14.5. Noradrenergic and cholinergic axon densities are uniform across the proximal–distal axis of adult mouse prostate while peptidergic axons are denser in the periurethral and proximal regions. Peptidergic and cholinergic axons are closely associated with prostate neuroendocrine cells whereas noradrenergic axons are not. These results provide a foundation for understanding mouse prostatic axon development and organization and, provide strategies for quantifying axons during progression of prostate disease.

High-throughput quantification of the effect of DMSO on the viability of lung and breast cancer cells using an easy-to-use spectrophotometric trypan blue-based assay Abstract One of the main aspects investigated in potential therapeutic compounds is their effect on cells viability and proliferative ability. Although various methods have been developed to investigate these aspects, these methods present with shortcomings in terms of either cost, availability, accuracy, precision, or throughput. This study describes a simple, economic, reproducible, and high-throughput assay to quantify cell death and proliferation. In this assay, adherent cells are fixed, stained with trypan blue, and measured for trypan blue internalization using a spectrophotometric absorbance plate reader. Corresponding cell counts to the absorbance measurements are extrapolated from a standard curve. This assay was used to measure the effect of dimethyl sulfoxide (DMSO) on the viability of breast and lung cancer cells. Decrease in cell count associated with the increase in DMSO percentage and exposure time. The assay's results closely correlated with the conventional trypan blue exclusion assay (Pearson correlation coefficient (r) > 0.99; p < 0.0001), but with higher precision. The assay developed in this study can be used for various applications such as optimization, cell treatment investigations, proliferation, and cytotoxicity studies.

Similarities and differences in tissue distribution of DLK1 and DLK2 during E16.5 mouse embryogenesis Abstract DLK1 and DLK2 are transmembrane proteins belonging to the EGF-like repeat-containing family that function as non-canonical NOTCH inhibitory ligands. DLK1 is usually downregulated after embryo development and its distribution in some adult and embryonic tissues has been described. However, the expression and role of DLK2 in embryo and adult tissues remains unclear. To better understand the relevance of both proteins during embryo development, we analyzed the expression pattern of DLK1 and DLK2 in 16.5-day-old mouse embryos (E16.5) and evaluated the possible relationship between these two proteins in embryo tissues and cell types. We found that DLK1 and DLK2 proteins exhibited a broad distribution pattern, which was detected in developing mouse organs from each of the three germ layers: ectoderm (brain, salivary glands), mesoderm (skeletal muscle, vertebral column, kidney, cartilage), and endoderm (thymus, lung, pancreas, intestine, liver). The expression pattern of DLK1 and DLK2 indicates that both proteins could play a synergistic role during cell differentiation. This study provides additional information for understanding temporal and site-specific effects of DLK1 and DLK2 during embryo morphogenesis and cell differentiation.

Quantitative pixel intensity- and color-based image analysis on minimally compressed files: implications for whole-slide imaging Abstract Current best practice in the quantitative analysis of microscopy images dictates that image files should be saved in a lossless format such as TIFF. Use of lossy files, including those processed with the JPEG algorithm, is highly discouraged due to effects of compression on pixel characteristics. However, with the growing popularity of whole-slide imaging (WSI) and its attendant large file sizes, compressed image files are becoming more prevelent. This prompted us to perform a color-based quantitative pixel analysis of minimally compressed WSI images. Sections from three tissues stained with one of three reagents representing the colors blue (hematoxylin), red (Oil-Red-O), and brown (immunoperoxidase) were scanned with a whole slide imager in triplicate at 20x, 40x, and 63x magnifications. The resulting files were in the form of a BigTIFF with a JPEG compression automatically applied during acquisition. Images were imported into analysis software, six regions of interest were applied to various morphological locations, and the areas assessed for the color of interest. Whereas the number of designated weakly or strongly positive pixels was variable across the triplicate scans for the individual regions of interest, the total number of positive pixels was consistent. These results suggest that total positivity for a specific color representing a histochemical or immunohistochemical stain can be adequately quantitated on compressed images, but degrees of positivity (e.g., weak vs. strong) may not be as reliable. However, it is important to assess individual whole-slide imagers, file compression level and algorithm, and analysis software for reproducibility.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Toxicology and Environmental Health Sciences

Level of Petroleum Hydrocarbons in Water and Sediments of Ikoli Creek Bayelsa State Nigeria Abstract Objective This study investigated the level Petroleum hydrocarbon in Ikoli creek, Bayelsa state Nigeria with focus on water and sediments. Methods Water and sediments were assessed using standard analytical procedures. Water from surface levels and sediment samples were collected from five locations in the creek. Soxhlet extraction method and liquid-liquid techniques were used in extraction of the petroleum hydrocarbons (Total Hydrocarbon Content and Total Petroleum Hydrocarbon) from the water and sediment samples collected respectively followed by column clean up. Gas Chromatography-Flame Ionization Detector (GC-FID) was use to analyzed the Target compounds and quantified by integrating the areas of both the resolved and unresolved components. Results Results of analysis of THC and TPH in water ranges between 0.010 to 0.254 mg/L and Total Petroleum Hydrocarbon (TPH) values ranges between 0.001 to 0.437 mg/L in water respectively. While in sediments the Total Hydrocarbon Content (THC) values ranges between 0.01 to 1.43 mg/kg and Total Petroleum Hydrocarbon (TPH) ranges between 0.001 to 0.44 mg/kg. The THC and TPH value of sediments increases across each sampling station in the study area and these hydrocarbons are repository in nature. Conclusions The results showed that some sampling station in Ikoli creek were slightly polluted with petroleum hydrocarbons and does not conform to acceptable international guideline, which can cause adverse effects to humans and aquatic life over time.

Ameliorating Effects of Quercetin in a Diabetic Rat Model with Social Anxiety Disorder Abstract Objective The present study aimed to investigate the effects of quercetin on diabetes-induced behavioural, metabolic and histopathological alterations in a rat model that mimics the development of social anxiety disorder in diabetic humans. Methods Diabetes was induced by an intraperitoneal injection of alloxan (150 mg/kg bw). Three weeks later, the initial animal groups (control and diabetic) were assigned to three paralleled experiments during which quercetin was orally administered at a relatively high dose (100 mg/kg bw). The behavioural assessment included the forced swim test, the elevated plus-maze, the open field and the social interaction tests. The plasma glucose, triglycerides, cholesterol, urea and creatinine levels were evaluated, whereas the histopathology of the prefrontal cerebral cortex, liver and kidneys were checked out. Results Diabetes was associated with anxiety-like, depressive-like and defective social behaviours. The metabolic parameters were significantly elevated along with a histological damage to hepatic, renal and pre-frontal cerebral tissues. Quercetin treatment in diabetic rats alleviated the behavioural disorders while improving the social interactions. A significant decrease in all metabolic parameters was found following quercetin intake, which were accompanied by a better histo-pathological profile of the studied tissues. Conclusion Our findings suggest that the pathophysiology of diabetes is tightly linked to anxiety-like, depression-like, and deteriorated social behaviours, which may originate from a severe damage to the prefrontal cerebral cortex integrity. Interestingly, a high-dose short treatment with quercetin may mitigate the diabetes-related neurobehavioural, metabolic and histological disturbances, and prevent the defective sociability in long-term diabetic subjects.

Increased Blood Lead Level Alters the Heme Biosynthesis and Haematological Parameters of Spray Painters from Western Maharashtra, India Abstract Objective To assess the present status of blood lead levels and their effect on heme biosynthesis and hematological parameters of spray painters. Methods Forty two spray painters having the age range of 25–50 years were recruited for the study from Western Maharashtra, India and compared with age matched 50 healthy male subjects. Blood lead (PbB), erythrocytes d-ALDH, Urinary d-ALA, PBG and hematological parameters were measured. Results PbB levels (p<0.001, 458%) of spray painters were significantly increased and Non-activated δ-ALAD (p<0.001, -20.21%), activated δ-ALAD (p<0.001, -28.25%) were significantly decreased and ratio of activated to non-activated δ-ALAD (p<0.001, 16%) was significantly increased in spray painters as compared to control subjects. Urinary δ-ALA (p<0.001, 160%) and PBG (p<0.001, 62.75%) levels were significantly increased in spray painters as compared to the control group. Hematological parameters of spray painters i.e. Hb (p<0.001, -21.58%) PCV (p<0.05, -4.53%) MCV (p<0.01, -5.09%) MCH (p<0.01, -6.04), MCHC (p<0.01, - 2.31) and RBC (p<0.001, - 8.06 %) were significantly decreased while total WBC count (p<0.001, 34.63%) was significantly increased as compared to the controls. Conclusion Increased PbB level in spray painters may be due to more absorption of lead thereby inhibiting heme biosynthesis enzymes and resulting in more excretion of urinary δ-ALA, PBG and alteration of hematological parameters. Therefore it is essential to monitor these spray painters and also it is necessary to create awareness for using adequate protective measures. Also necessary training and regular screening is required to minimize adverse impact of lead exposure to them.

Effect of Food Colorants and Additives on the Hematological and Histological Characteristics of Albino Rats Abstract Objective Food colorants (synthetic, inorganic and natural) represent one of the major categories of food additives. Synthetic colorants were banned due to their adverse effects on animals and human. However, synthetic dyes are still in use because they are cheap, and stable. Methods In the present in vivo study conducted on female albino rats (Rattus norvegicus), the effect of sunset yellow (SY) and sodium benzoate (NaB) combinations on the hematological and histological profile was assessed. Different combinations of SY plus NaB were dissolved in water and administered daily to experimental rat groups for 12 weeks. Group 1 (control) received only water, group 2 received 5 mg SY plus 10 mg NaB, group 3 received 5 mg SY plus 100 mg NaB, group 4 received 50 mg SY plus 100 mg NaB, group 5 received 50 mg SY plus 10 mg NaB, group 6 received 200 mg SY plus 750 mg NaB, and group 7 received 20 mg SY plus 75 mg NaB. Histopathological examinations were performed on liver and kidney of rats at the end of the experiment. Results The results revealed a decrease in RBCs count, hematocrit, WBCs, MCV and Hb levels upon the administration of SY plus NaB. The results also showed no increase in MCH, MCHC and platelet count. Liver and kidney tissues showed some lesions due to the administration of the tested compounds in comparison to the control animals. Conclusion The chemical stress caused by the SY and NaB combinations caused some degenerative changes in the liver and kidneys of rats. It could be concluded that SY and NaB combinations causes some damage in liver and kidney tissues of experimental animals. Therefore, using SY and NaB combinations should be limited.

Effects of Repeated Pulmonary Exposure to Carbon Nanotubes on Lung function Abstract Objective The development of nanotechnology has increased the risk of exposure to types of particles other than combustion-derived ones in the environment, namely, industrial nanomaterials. However, their health impacts have not been fully elucidated yet. This study examined the effects of pulmonary exposure to two types of carbon nanotube (single-walled carbon nanotubes: SWCNT, and multi-walled carbon nanotube: MWCNT) on lung physiology in mice. Methods ICR mice were divided into three experimental groups to repeatedly administer vehicle, SWCNT (50 μg/animal), or MWCNT through intratracheal route; thereafter lung function was examined. Results The values of total respiratory system resistance (R: P<0.05), elastance (E), Newtonian resistance (Rn), tissue damping (G: P<0.05), and tissue elastance (H) were higher and the value of compliance (C) was lower in the SWCNT group than in the vehicle group. On the other hand, MWCNT tended to augment the values. Conclusions Taken together, CNT themselves inhaled, can impair airway function.

Ameliorative Effect of Zinc Oxide Nanoparticles on Nicotine Induced Testicular Dysfunction; Biochemical and Histological Study Abstract Objective The aim of this study was to evaluate potential impact of zinc oxide nanoparticle on the possible testicular toxicity induced by nicotine. Methods 24 adult male albino rats were divided into four groups; negative control group, zinc oxide nanoparticle group: rats were received 5 mg/kg/day zinc oxide nanoparticles orally, nicotine treated group: rats were administered nicotine (1 mg/kg/day) intraperitoneally and nicotine with zinc oxide nanoparticles: rats were treated with nicotine and zinc oxide nanoparticles daily for 28 days. Serum testosterone level, testicular malondialdehyde, SOD and catalase activities were measured. Testes were examined by light and electron microscopes as well. Results Significant decreases of serum testosterone level, testicular SOD and catalase activity and significant increase of testicular MDA in nicotine treated group when compared with control group were detected. Zinc oxide nanoparticles administration reversed these changes. Conclusion Zinc oxide nanoparticles ameliorated nicotine induced testicular dysfunction.

Enhancement of Alcohol Metabolism by Ginseng Berry Extract and its Mixed Herbal Beverages: In vitro and in vivo Experiments Abstract Objective To investigate the enhancement of alcohol metabolism by two ginseng berry (GB) extracts and their two types of mixed herbal beverages through in vitro and in vivo experiments. Methods Two GB extract solutions and their two herbal beverages were evaluated as enhancers of alcohol metabolism in normal human embryonic liver cells (CL-48 cell line) through assays of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) activity. Cytotoxicity was also assessed in the same cell line using an MTT assay. Effects on alcohol metabolism were also observed in vivo through measurement of serum alcohol, acetaldehyde, alanine aminotransferase (ALT), and aspartate aminotransferase (AST) levels in alcohol treated rats. Blood samples were serially collected at 1, 2, 3, 4, 6 and 8 hrs after a single ethanol (EtOH) treatment. A single treatment with the test samples was administered orally 30 minutes after EtOH treatment. Results GB extract solutions effectively elevated the ADH and ALDH activity observed in vitro, while no treatment- related cytotoxic effects were found with test samples t concentrations up to 100 mg/mL. Significantly lower (p<0.01 or p<0.05) serum alcohol and acetaldehyde content was observed in samples from treated rats than in those from control rats (EtOH only) 1 or 2 hrs after EtOH treatment. In addition, noticeable decreases were observed in serum AST and ALT levels in treated samples 8 hrs after EtOH administration. HM40, an herbal mixture containing GB extract (40 mg/75 mL of ginsenoside Re), showed betters enhancement of alcohol metabolism through ADH/ALDH activation, as well as related hepatoprotective effects. Results GB extracts effectively enhanced alcohol metabolism without cytotoxicity while also providing possible hepatoprotective effects that could serve as a functional ingredient in anti-hangover alternative therapies. These extracts are expected to be more effective when made into herbal mixture beverages.

In vitro Models for Chemical Toxicity: Review of their Applications and Prospects Abstract Since it has been considered acceptable to expose animal to conditions and procedures that would not be considered acceptable for human, animals have long been used as surrogates for human beings for in vivo toxicology. This has focused mainly on gaining a better understanding of how the body's cells, organs, and systems function and are controlled, which failures could lead to pathological conditions. However, attitudes that rely on this approach are changing as the know ledge gained from animal studies is not directly related to humans, and there is a growing perception that it can also have dangerous consequences. In this re view, I collected the references about not only in vitro models for chemical toxicity, but also their applications and prospects focused on chemical hazard evaluations. In this review, I try to gather the issues and considerations that are considered necessary to develop a viable and reliable testing strategy to replace animals in toxicity testing. Most of the authors of the papers cited here share the opinion that animals will no longer be used for safety assessment, but that a more predictable human-derived in vitrosystem and in silico approach will be developed step-by-step. I collected information from peer-reviewed literature on the in vitro models for toxicological investigation of chemicals that may impact environmental or occupational diseases.

Chemical Composition and Biological Activity of Essential Oils of Centella asiatica (L.) Abstract Objective Centella asiatica (L.) is belonging the Apiaciaea family that is very popular for its culinary and medicinal use. In recent years, due to overexploitation, and ineffectual efforts to revitalize this valuable herbal source, the abundance of C. asiatica (L.) in its habitats has suffered significantly, cause to the risk of its extinction. Therefore, its in vitro production interested considerably. Methods In this study, mass production of C. asiatica (L.) was carried out through a cost effective micropropagation protocol and the chemical content of essential oils from micropropagated plants were analyzed by Gas chromatography (GC) and Gas chromatography-mass spectrometry (GC/MS) and biological properties compared with wild plants. Results In total, 45 compounds were identified and quantified in wild and micropropagated plants, representing 97.5% and 97.7% of the total oil, respectively. Also the oils were dominated by oxygenated sesquiterpenes, 70.8% and 42.6%, in the wild and micropropagated plants, respectively. Antibacterial assay showed the oil is effective regarding bacterial growth inhibition. The inhibition zones against some bacterial strains (Bacillus megaterium, Staphylococcus aureus, Klebsiella aerogenes) were even greater than those of chloramphenicol. The MIC of the oils, against the tested organisms, varied between 1 and 4μL mL−1. The lowest MIC (1 μL mL−1) was detected for the oil against Enterobacter aerugenes and Staphylococcous subrogation. Results were also indicated that the oil of micropropagated plants was more effective than the oils of wild ones. Cytotoxicity assay showed that the toxicity of the studied essential oils are not significant at concentrations of 5–15% (v/v). Conclusions It could concluded that the essential oil of C. asiatica, in micropropagated plants, have the good antibacterial activities without significant toxicity, at the applied concentration, and thus have great potentiality to be used as a resource for natural health products.

The Role of Aptamer Loaded Exosome Complexes in the Neurodegenerative Diseases Abstract Extracellular exosomes formed inside the cytoplasm of cells are small membranous vesicles that provide an important mode of cell-to-cell communication by delivering proteins, lipids and RNA to target cells. Exosomes are found associated with neurodegenerative diseases mainly Parkinson's disease (PD) and Alzheimer's diseases (AD), characterized by progressive degeneration of neurons and often linked with misfolded protein. Aptamers are single stranded DNA- and RNA-based oligonucleotides that bind target protein with high affinity and specificity and block or prevent the process of neurodegenerative diseases. In this review, we discuss the role of aptamer loaded exosomes in the neurode-generative diseases.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Der Nephrologe

Katheterinterventionen und interventionelle Shuntanlagen Zusammenfassung Hintergrund Das Leben der Dialysepatienten hängt unbestritten vom vaskulären Zugang ab. Standard ist die chirurgische Schaffung eines Shunts (arteriovenöse Fistel, AVF), was besondere Vorbereitung und Expertise erfordert. Eine besondere Herausforderung sind die Shuntreifung und die Versorgung der shuntassoziierten Probleme mit dem Ziel, den Patienten eine möglichst reibungslose Dialyse langfristig zu ermöglichen und zentrale Venenkatheter zu ersparen. Dies erfordert die Versorgung mittels endovaskulärer Techniken und chirurgischer Revisionen. Es existieren multiple Instrumente, die für die AVF-Versorgung zur Verfügung stehen. Des Weiteren wurden über die letzten Jahre Instrumentation und Techniken entwickelt, die eine endovaskuläre AVF-Anlage ermöglichen. Ziel der Arbeit Darstellung aktuell genutzter Katheterinterventionen und Beleuchtung verfügbarer Methoden der endovaskulären Shuntanlage. Material und Methoden In diesem Übersichtsartikel werden die Katheterinterventionen, Techniken, Komplikationen und Erfolge der jeweiligen Verfahren dargestellt. Ergebnisse Perkutane Angioplastien (PTA) mit verschiedenen Ballons, Stent- und Stentgraftanlage sowie endovaskuläre Thrombektomie mittels verschiedener Methoden haben das Armamentarium der Shuntversorgung über die letzten Jahre erweitert und verbessert und sind eine wertvolle Addition zu den bisherigen chirurgischen Verfahren zur Behandlung der Shuntdysfunktion. Die Möglichkeit der endovaskulären Shuntanlage eröffnet eine ganz neue Perspektive in der Kreierung des Dialysezugangs und weist bereits gute Früherfolge auf. Diskussion AVF-Anlage und -Versorgung sind ein zentrales und longitudinales Problem für Dialysepatienten. Es erfordert eine multidisziplinäre Zusammenarbeit. Kenntnis aller Verfahren und sorgfältige Selektion einer Methode für den individuellen Patienten bleiben von immenser Bedeutung.

Charakterisierung von Patienten mit atypischem hämolytisch-urämischen Syndrom (aHUS) in Deutschland

Physiologie und Pathophysiologie von FGF23 und Klotho

Von der chronischen Niereninsuffizienz zur Hämodialyse Zusammenfassung Jährlich beginnt in Deutschland eine hohe Anzahl von Patienten eine chronische Nierenersatztherapie, überwiegend in Form einer Hämodialyse(HD)-Therapie. Über diese Transitionsphase sind nur wenige gesicherte Daten verfügbar. Die ersten Monate einer HD-Therapie sind eine medizinische Hochrisikophase mit im Vergleich zu prävalenten HD-Patienten deutlich gesteigerter Mortalität. Risikofaktoren hierfür sind unter anderem Gefäßzugang via zentraler Venenkatheter und Malnutrition. Vorhandensein, längere Dauer und höhere Intensität einer nephrologischen Betreuung bei fortgeschrittener chronischer Nierenerkrankung (CKD) resultieren in einem besseren Outcome nach Transition zur HD. Studiendaten implizieren, dass ein späterer Beginn der Dialysetherapie (eGFR <10 ml/min/m2) im Vergleich zu einem früheren Beginn (eGFR <10 ml/min/m2) nicht mit einer schlechteren Prognose assoziiert ist; in die Entscheidung zur Einleitung einer Nierenersatztherapie müssen jedoch auch patientenindividuelle Aspekte einfließen. Welche weiteren Faktoren, Behandlungspraktiken und Patientencharakteristika bei fortgeschrittener CKD Einfluss auf die Prognose nach Dialysebeginn haben, ist nicht ausreichend bekannt; bisher liegen nur wenige Daten hierzu vor. Unser Ziel muss es sein, Therapiemodifikationen in der Prädialysephase zu identifizieren, die das Outcome nach Transition zur Dialysetherapie verbessern, um die mit dieser Phase verbundene exzeptionell hohe Mortalität zu reduzieren.

Genetische Nierenerkrankungen

„Übergänge" in der Nephrologie

29-jährige Patientin mit Syndrom der inadäquaten ADH-Sekretion unklarer Ursache

Der Sprung auf die Warteliste Zusammenfassung Der terminalen Niereninsuffizienz liegen ganz verschiedene Erkrankungen zugrunde, entsprechend unterscheiden sich die Dialysepatienten, die auf eine Transplantation hoffen, in weiten Bereichen. Per Gesetz ist vorgeschrieben, dass bei jedem Dialysepatienten die Eignung für eine Nierentransplantation geprüft werden muss. Aufgrund der großen bürokratischen Anforderungen des Gesetzgebers an diesen Prozess und der individuellen sehr hohen Krankheitsschwere von Dialysepatienten ist die Überprüfung der Eignung nicht leicht zu standardisieren, sehr zeitaufwändig und komplex. Zusätzlich hat ein Dialysepatient ein sehr hohes Risiko, Folgekrankheiten zu entwickeln, die eine Eignung zur Transplantation gefährden. Damit kommt der konstanten und konsequenten Überprüfung der Eignung der Patienten auch nach der Aufnahme auf die Warteliste (Wartelistenführung) eine große Bedeutung zu. Der Schlüssel für den Sprung auf die Warteliste und damit die erfolgreiche Transplantation ist eine enge und vertrauensvolle Kooperation zwischen dem Dialyse- und Transplantationszentrum. Leider führt die in Deutschland außerordentlich lange Wartezeit für ein Organ deswegen nicht nur zu einer Gefährdung des Transplantationserfolgs, sie erhöht gleichzeitig auch den Aufwand der Transplantationszentren durch eine verlängerte Wartelistenführung beträchtlich.

Einfluss des Spendenalters auf das Transplantatüberleben bei Kindern und Jugendlichen

Von der Kindersprechstunde in die Erwachsenensprechstunde Zusammenfassung Der Übergang von der Pädiatrie in die Erwachsenenmedizin stellt in der Nephrologie eine wichtige, aber auch schwierige Schnittstelle dar. Dies gilt insbesondere für Jugendliche mit chronischer Niereninsuffizienz und nach Nierentransplantation. Dieser Übersichtsartikel stellt Transitionsrichtlinien und -hürden dar. Psychosoziale und finanzielle Aspekte der Transition sowie Studien- und Berufswahl werden diskutiert. In den letzten Jahren der Betreuung in der Jugendmedizin müssen die Grundlagen für den Übergang, wie Schulungen und Förderung der Selbstständigkeit der Patienten, gelegt werden. In der Erwachsenennephrologie müssen zukünftig noch mehr Ressourcen für die Besonderheiten der transferierten Patienten geschaffen werden. Es ist insbesondere eine politische Aufgabe, die Finanzierung einer guten Transition zu ermöglichen.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Toxicology

Sphingosine 1-phosphate receptor 2/adenylyl cyclase/protein kinase A pathway is involved in taurolithocholate-induced internalization of Abcc2 in rats Abstract Taurolithocholate (TLC) is a cholestatic bile salt that induces disinsertion of the canalicular transporter Abcc2 (Mrp2, multidrug resistance-associated protein 2). This internalization is mediated by different intracellular signaling proteins such as PI3K, PKCε and MARCK but the initial receptor of TLC remains unknown. A few G protein-coupled receptors interact with bile salts in hepatocytes. Among them, sphingosine-1 phosphate receptor 2 (S1PR2) represents a potential initial receptor for TLC. The aim of this study was to evaluate the role of this receptor and its downstream effectors in the impairment of Abcc2 function induced by TLC. In vitro, S1PR2 inhibition by JTE-013 or its knockdown by small interfering RNA partially prevented the decrease in Abcc2 activity induced by TLC. Moreover, adenylyl cyclase (AC)/PKA and PI3K/Akt inhibition partially prevented TLC effect on canalicular transporter function. TLC produced PKA and Akt activation, which were blocked by JTE-013 and AC inhibitors, connecting S1PR2/AC/PKA and PI3K/Akt in a same pathway. In isolated perfused rat liver, injection of TLC triggered endocytosis of Abcc2 that was accompanied by a sustained decrease in the bile flow and the biliary excretion of the Abcc2 substrate dinitrophenyl-glutathione until the end of the perfusion period. S1PR2 or AC inhibition did not prevent the initial decay, but they accelerated the recovery of these parameters and the reinsertion of Abcc2 into the canalicular membrane. In conclusion, S1PR2 and the subsequent activation of AC, PKA, PI3K and Akt is partially responsible for the cholestatic effects of TLC through sustained internalization of Abcc2.

Role of the human N -acetyltransferase 2 genetic polymorphism in metabolism and genotoxicity of 4, 4′-methylenedianiline Abstract 4, 4′-Methylenedianiline (MDA) is used extensively as a curing agent in the production of elastomers and is classified as reasonably anticipated to be a human carcinogen based on sufficient evidence in animal experiments. Human N-acetyltransferase 1 (NAT1) and 2 (NAT2) catalyze the N-acetylation of aromatic amines and NAT2 is subjected to a common genetic polymorphism in human populations separating individuals into rapid, intermediate, and slow acetylator phenotypes. Although MDA is known to undergo N-acetylation to mono- and di-acetyl metabolites, very little is known regarding whether this metabolism is subject to the NAT2 genetic polymorphism. We investigated the N-acetylation of MDA by recombinant human NAT1, NAT2, genetic variants of NAT2, and cryoplateable human hepatocytes obtained from rapid, intermediate and slow acetylators. MDA N-acetylation was catalyzed by both recombinant human NAT1 and NAT2 exhibiting a fivefold higher affinity for human NAT2. N-acetylation of MDA was acetylator genotype dependent as evidenced via its N-acetylation by recombinant human NAT2 genetic variants or by cryoplateable human hepatocytes. MDA N-acetylation to the mono-acetyl or di-acetyl-MDA was highest in rapid, lower in intermediate, and lowest in slow acetylator human hepatocytes. MDA-induced DNA damage in the human hepatocytes was dose-dependent and also acetylator genotype dependent with highest levels of DNA damage in rapid, lower in intermediate, and lowest in slow acetylator human hepatocytes under the same MDA exposure level. In summary, the N-acetylation of MDA by recombinant human NAT2 and cryopreserved human hepatocytes support an important role for the NAT2 genetic polymorphism in modifying MDA metabolism and genotoxicity and potentially carcinogenic risk.

Highlight report: liver to bone communication

Prediction of in vivo genotoxicity of lasiocarpine and riddelliine in rat liver using a combined in vitro-physiologically based kinetic modelling-facilitated reverse dosimetry approach Abstract Pyrrolizidine alkaloids (PAs) are naturally occurring genotoxic compounds, and PA-containing plants can pose a risk to humans through contaminated food sources and herbal products. Upon metabolic activation, PAs can form DNA adducts, DNA and protein cross links, chromosomal aberrations, micronuclei, and DNA double-strand breaks. These genotoxic effects may induce gene mutations and play a role in the carcinogenesis of PAs. This study aims to predict in vivo genotoxicity for two well-studied PAs, lasiocarpine and riddelliine, in rat using in vitro genotoxicity data and physiologically based kinetic (PBK) modelling-based reverse dosimetry. The phosphorylation of histone protein H2AX was used as a quantitative surrogate endpoint for in vitro genotoxicity of lasiocarpine and riddelliine in primary rat hepatocytes and human HepaRG cells. The in vitro concentration–response curves obtained from primary rat hepatocytes were subsequently converted to in vivo dose–response curves from which points of departure (PoDs) were derived that were compared to available in vivo genotoxicity data. The results showed that the predicted PoDs for lasiocarpine and riddelliine were comparable to in vivo genotoxicity data. It is concluded that this quantitative in vitro-in silico approach provides a method to predict in vivo genotoxicity for the large number of PAs for which in vivo genotoxicity data are lacking by integrating in vitro genotoxicity assays with PBK modelling-facilitated reverse dosimetry.

Functional mismatch repair and inactive p53 drive sensitization of colorectal cancer cells to irinotecan via the IAP antagonist BV6 Abstract A common strategy to overcome acquired chemotherapy resistance is the combination of a specific anticancer drug (e.g., topoisomerase I inhibitor irinotecan) together with a putative sensitizer. The purpose of this study was to analyze the cytostatic/cytotoxic response of colorectal carcinoma (CRC) cells to irinotecan, depending on the mismatch repair (MMR) and p53 status and to examine the impact of BV6, a bivalent antagonist of inhibitors of apoptosis c-IAP1/c-IAP2, alone or combined with irinotecan. Therefore, several MSH2- or MSH6-deficient cell lines were complemented for MMR deficiency, or MSH6 was knocked out/down in MMR-proficient cells. Upon irinotecan, MMR-deficient/p53-mutated lines repaired DNA double-strand breaks by homologous recombination less efficiently than MMR-proficient/p53-mutated lines and underwent elevated caspase-9-dependent apoptosis. Opposite, BV6-mediated sensitization was achieved only in MMR-proficient/p53-mutated cells. In those cells, c-IAP1 and c-IAP2 were effectively degraded by BV6, caspase-8 was fully activated, and both canonical and non-canonical NF-κB signaling were triggered. The results were confirmed ex vivo in tumor organoids from CRC patients. Therefore, the particular MMR+/p53mt signature, often found in non-metastasizing (stage II) CRC might be used as a prognostic factor for an adjuvant therapy using low-dose irinotecan combined with a bivalent IAP antagonist.

Validation of the γH2AX biomarker for genotoxicity assessment: a review Abstract The H2AX histone protein is rapidly phosphorylated at the serine-139 position (γH2AX) in response to a broad range of DNA lesions. γH2AX induction is one of the earliest events in the DNA damage response (DDR) and plays a central role in sensing and repairing DNA damage. Since its discovery, measuring γH2AX formation using numerous methods in in vitro and in vivo experiments has been an attractive endpoint for the detection of genotoxic agents. Our review focuses on validation studies performed using this biomarker to detect the genotoxicity of model chemicals using different methods. To date, nearly two hundred genotoxic and carcinogenic model chemicals have been shown to induce in vitro γH2AX in different cell lines by numerous laboratories. Based on 27 published reports comprising 329 tested chemicals, we compared the performance of the γH2AX assay with other genotoxic endpoints (Ames assay, micronucleus, HPRT and comet) regularly used for in vitro genotoxicity assessment. Notably, the γH2AX assay performs well (91% predictivity) and efficiently differentiates aneugenic and clastogenic compounds when coupled with the pH3 biomarker. Currently, no formal guidelines have been approved for the γH2AX assay for regular genotoxicity studies, but we suggest the γH2AX biomarker could be used as a new standard genotoxicity assay and discuss its future role in genotoxicity risk assessment.

Influence of exposure dose, complex mixture, and ultraviolet radiation on skin absorption and bioactivation of polycyclic aromatic hydrocarbons ex vivo Abstract Combined exposure to complex mixtures of polycyclic aromatic hydrocarbons (PAHs) and ultraviolet radiation (UVR) is suspected to enhance PAH skin permeability and skin cancer risk depending on PAH bioactivation. The impact of PAH mixtures (exposure dose, composition, and complexity) and UVR was assessed for PAH cutaneous absorption and metabolism using realistic exposure conditions and human skin explants. PAH complex mixtures were extracted from the industrial products coal tar pitch (CTP-I) and petroleum coke (PC-I). The synthetic mixture (CTP-S) was identically reconstituted using PAH standards. The applied dose was adjusted to 1 (PC-I, CTP-I) or 10 nmol (CTP-I, CTP-S) of benzo[a]pyrene (B[a]P). Unmetabolized PAHs were recovered from the skin surface, skin and medium, and then quantified by HPLC-fluorescence detection. PAH metabolites were collected from the medium and analyzed by GC–MS/MS. B[a]P and PAH penetration was lower for the highest B[a]P dose, industrial mixtures, and CTP-I compared to PC-I. Skin irradiation increased PAH penetration only for CTP-I. PAH uptake was poorly influenced by the different experimental conditions. PAH metabolism markedly decreased in the application of mixtures, leading to unmetabolized PAH accumulation in human skin. PAH metabolism was similar between CTP-I and PC-I, but was lower for the highest dose and the industrial mixtures, suggesting a saturation of xenobiotic metabolizing enzymes, as confirmed in a time-course study. UVR strongly inhibited all PAH metabolism. Altogether, these results underline the necessity to consider the reality of human exposure (PAH complex mixtures and UVR) during in vitro experiments to properly estimate skin absorption and metabolism.

Genome-wide promoter DNA methylation profiling of hepatocellular carcinomas arising either spontaneously or due to chronic exposure to Ginkgo biloba extract (GBE) in B6C3F1/N mice Abstract Epigenetic modifications, such as DNA methylation, play an important role in carcinogenesis. In a recent NTP study, chronic exposure of B6C3F1/N mice to Ginkgo biloba extract (GBE) resulted in a high incidence of hepatocellular carcinomas (HCC). Genome-wide promoter methylation profiling on GBE-exposed HCC (2000 mg/kg group), spontaneous HCC (vehicle-control group), and age-matched vehicle control liver was performed to identify differentially methylated genes in GBE-exposed HCC and spontaneous HCC. DNA methylation alterations were correlated to the corresponding global gene expression changes. Compared to control liver, 1296 gene promoters (719 hypermethylated, 577 hypomethylated) in GBE-exposed HCC and 738 (427 hypermethylated, 311 hypomethylated) gene promoters in spontaneous HCC were significantly differentially methylated, suggesting an impact of methylation on GBE-exposed HCC. Differential methylation of promoter regions in relevant cancer genes (cMyc, Spry2, Dusp5) and their corresponding differential gene expression was validated by quantitative pyrosequencing and qRT-PCR, respectively. In conclusion, we have identified differentially methylated promoter regions of relevant cancer genes altered in GBE-exposed HCC compared to spontaneous HCC. Further study of unique sets of differentially methylated genes in chemical-exposed mouse HCC could potentially be used to differentiate treatment-related tumors from spontaneous-tumors in cancer bioassays and provide additional understanding of the underlying epigenetic mechanisms of chemical carcinogenesis.

Indomethacin-induced gut damage in a surrogate insect model, Galleria mellonella Abstract Indomethacin is a non-steroidal anti-inflammatory drug that causes gastric ulceration and increased 'leakiness' in rat models, and is used routinely as a toxicology assay to screen novel compounds for repair and restitution properties. We set out to establish conditions for indomethacin-induced gut damage in wax-moth (Galleria mellonella) larvae with a view to reducing the need for rodents in such experimentation. We administered indomethacin (0.5–7.5 µg/larva; 2–30 mg/kg) to G. mellonella via intrahaemocoelic injection and gavage (force-feeding) and monitored survival and development, blood cell (haemocyte) numbers, and changes in gut permeability. Increased levels of gut leakiness were observed within the first 4- to 24 h by tracking fluorescent microspheres in the faeces and haemolymph (blood equivalent). Additionally, we recorded varying levels of tissue damage in histological sections of the insect midgut, including epithelial sloughing and cell necrosis. Degeneration of the midgut was accompanied by significant increases in detoxification-associated activities (superoxide dismutase and glutathione-S-transferase). Herein, we present the first evidence that G. mellonella larvae force-fed indomethacin display broad symptoms of gastric damage similar to their rodent counterparts.

Comparisons of statistical models for growth curves from 90-day rat feeding studies Abstract The objective of this work was to compare several models of body weight data from 90-day rodent feeding trials. Polynomial and nonlinear functions relating time and weight were examined as were the use of Toeplitz error covariance structures and random coefficients. The models were evaluated by fitting them to five publicly available datasets from rat feeding studies. Model performance was assessed in terms of their ability to capture the complexity of the growth patterns, validity of necessary assumptions, and information criteria scores. The results demonstrated the importance of selecting a curve function that effectively reflects the mean response. Toeplitz error covariance structures resulted in superior model fit, while failing to address deviations from model assumptions. Models using the Richards function and random coefficients were generally superior to the other models evaluated and dramatically improved upon linear models with complex error structures.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com