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Praziquantel and Moxidectin pharmacokinetics in dogs after Helmimax administration

The purpose of the research is to study Praziquantel and Moxidectin pharmacokinetics in dogs after Helmimax administration.Materials and methods. Helmimax pharmacokinetics was studied on 8 adult male dogs of different breeds aged 2 to 5 years and weighing 15–35 kg. Helmimax was administered orally in the fasted state with a small amount of feed at a dose of 5 mg/kg for Praziquantel and 0.25 mg/kg for Moxidectin at the rate of 1 tablet per 10 kg of body weight. Blood was sampled at various periods after the administration. The collected blood underwent sample processing: formed element and protein precipitation, solid-phase extraction, and microfiltration. The active components were analyzed and detected by the HPLC-MS/MS. Active substances in the blood plasma were determined according to the developed technique which had been validated. The device was calibrated before the measurement.Results and discussion. As a result of the studies, the Praziquantel and Moxidectin pharmacokinetic parameters were calculated. The maximum concentration was 0.240 and 0.130 μg/mL, the time-to-peak concentration was 2.15 and 1.48 hours, and the elimination half-life was 8.41 and 3.61 hours for Moxidectin and Praziquantel, respectively

Фармакокинетика празиквантела и моксидектина в организме собак после применения гельмимакса

The purpose of the research is to study Praziquantel and Moxidectin pharmacokinetics in dogs after Helmimax administration.Materials and methods. Helmimax pharmacokinetics was studied on 8 adult male dogs of different breeds aged 2 to 5 years and weighing 15–35 kg. Helmimax was administered orally in the fasted state with a small amount of feed at a dose of 5 mg/kg for Praziquantel and 0.25 mg/kg for Moxidectin at the rate of 1 tablet per 10 kg of body weight. Blood was sampled at various periods after the administration. The collected blood underwent sample processing: formed element and protein precipitation, solid-phase extraction, and microfiltration. The active components were analyzed and detected by the HPLC-MS/MS. Active substances in the blood plasma were determined according to the developed technique which had been validated. The device was calibrated before the measurement.Results and discussion. As a result of the studies, the Praziquantel and Moxidectin pharmacokinetic parameters were calculated. The maximum concentration was 0.240 and 0.130 μg/mL, the time-to-peak concentration was 2.15 and 1.48 hours, and the elimination half-life was 8.41 and 3.61 hours for Moxidectin and Praziquantel, respectively.Цель исследований – изучение фармакокинетики празиквантела и моксидектина в организме собак после применения гельмимакса.Материалы и методы. Исследования фармакокинетики гельмимакса проводили на 8 взрослых собаках самцах массой тела 15–35 кг разных пород в возрасте от 2 до 5 лет. Гельмимакс вводили перорально натощак с небольшим количеством корма в дозе 5 мг/кг по празиквантелу и 0,25 мг/кг по моксидектину из расчета 1 таблетка на 10 кг массы тела. Отбор крови проводили через различные временные интервалы после введения. Отобранную кровь подвергали пробоподготовке: осаждение форменных элементов, белков, твердофазная экстракция, микрофильтрация. Анализ и детектирование действующих компонентов проводили методом ВЭЖХ МС/МС. Определение действующих веществ в плазме крови проводили по разработанной методике, которая прошла валидацию. Перед измерением осуществляли калибровку прибора.Результаты и обсуждение. В результате проведенных исследований были рассчитаны фармакокинетические показатели празиквантела и моксидектина. Максимальная концентрация составила 0,240 и 0,130 мкг/мл, время достижения максимальной концентрации 2,15 и 1,48 ч, период полувыведения 8,41 и 3,61 ч для моксидектина и празиквантела соответственно

Estrogen/Estrogen Receptor Alpha Signaling in Mouse Posterofrontal Cranial Suture Fusion

BACKGROUND: While premature suture fusion, or craniosynostosis, is a relatively common condition, the cause is often unknown. Estrogens are associated with growth plate fusion of endochondral bones. In the following study, we explore the previously unknown significance of estrogen/estrogen receptor signaling in cranial suture biology. METHODOLOGY/PRINCIPAL FINDINGS: Firstly, estrogen receptor (ER) expression was examined in physiologically fusing (posterofrontal) and patent (sagittal) mouse cranial sutures by quantitative RT-PCR. Next, the cranial suture phenotype of ER alpha and ER beta knockout (alphaERKO, betaERKO) mice was studied. Subsequently, mouse suture-derived mesenchymal cells (SMCs) were isolated; the effects of 17-beta estradiol or the estrogen antagonist Fulvestrant on gene expression, osteogenic and chondrogenic differentiation were examined in vitro. Finally, in vivo experiments were performed in which Fulvestrant was administered subcutaneously to the mouse calvaria. Results showed that increased ERalpha but not ERbeta transcript abundance temporally coincided with posterofrontal suture fusion. The alphaERKO but not betaERKO mouse exhibited delayed posterofrontal suture fusion. In vitro, addition of 17-beta estradiol enhanced both osteogenic and chondrogenic differentiation in suture-derived mesenchymal cells, effects reversible by Fulvestrant. Finally, in vivo application of Fulvestrant significantly diminished calvarial osteogenesis, inhibiting suture fusion. CONCLUSIONS/SIGNIFICANCE: Estrogen signaling through ERalpha but not ERbeta is associated with and necessary for normal mouse posterofrontal suture fusion. In vitro studies suggest that estrogens may play a role in osteoblast and/or chondrocyte differentiation within the cranial suture complex

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure fl ux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost higher eukaryotes and some protists such as Dictyostelium ) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the fi eld understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

3D replicon distributions arise from stochastic initiation and domino-like DNA replication progression.

DNA replication dynamics in cells from higher eukaryotes follows very complex but highly efficient mechanisms. However, the principles behind initiation of potential replication origins and emergence of typical patterns of nuclear replication sites remain unclear. Here, we propose a comprehensive model of DNA replication in human cells that is based on stochastic, proximity-induced replication initiation. Critical model features are: spontaneous stochastic firing of individual origins in euchromatin and facultative heterochromatin, inhibition of firing at distances below the size of chromatin loops and a domino-like effect by which replication forks induce firing of nearby origins. The model reproduces the empirical temporal and chromatin-related properties of DNA replication in human cells. We advance the one-dimensional DNA replication model to a spatial model by taking into account chromatin folding in the nucleus, and we are able to reproduce the spatial and temporal characteristics of the replication foci distribution throughout S-phase

Normal bone growth requires optimal estrogen levels: negative effects of both high and low dose estrogen on the number of growth plate chondrocytes

Endochondral bone formation at epiphyseal growth plate consists of the synchronized processes of chondrogenesis and cartilage ossification. Estrogen, the major female sex hormone, plays an important role in this process, particularly during the pubertal growth spurt. However, its effects on the growth plate are not completely understood. The aims of this study were to clarify the effects of estrogen on the kinetics of chondrocytes in the growth plates of 10- to 25-week-old female rabbits by studying the effects of ovariectomy or high-dose administration of estrogen on the balance between cell proliferation and death. Forty-eight Japanese white rabbits were divided into three groups: sham operated, ovariectomized, or ovariectomized with subsequent weekly injection of high dose estrogen from 10 weeks. The chondrocyte kinetics was investigated by histomorphometry and immunohistochemistry, using antibodies for caspase-3, a marker of apoptosis, and for proliferating cell nuclear antigen. Both ovariectomized and estrogen-injected rabbits showed a declination of the chondrocyte number although the latter animals indicated a more dramatic effect. Estrogen-injected rabbits showed a decrease in the cell proliferating ability together with an increase in chondrocytes undergoing apoptosis while ovariectomy mainly reduced the cell proliferating ability. Given the known importance of estrogen for bone growth, one would expect that ovariectomy and high-dose administration of estrogen would have opposite effects. However, the present study indicated that both low and high concentration had a similar effect: a decrease in the chondrocyte number compared with control, suggesting that estrogen has to be maintained within a narrow range for optimal bone growth