Comparison of human hepatoma HepaRG cells with human and rat hepatocytes in uptake transport assays in order to predict drug induced hepatotoxicity

Human hepatocytes are the gold standard for toxicological studies but they have several drawbacks, like scarce availability, high inter-individual variability, a short lifetime, which limits their applicability. The aim of our investigations was to determine, whether HepaRG cells could replace human hepatocytes in uptake experiments for toxicity studies. HepaRG is a hepatoma cell line with most hepatic functions, including a considerable expression of uptake transporters in contrast to other hepatic immortalized cell lines. We compared the effect of cholestatic drugs (bosentan, cyclosporinA, troglitazone,) and bromosulfophthalein on the uptake of taurocholate and estrone-3-sulfate in human and rat hepatocytes and HepaRG cells. The substrate uptake was significantly slower in HepaRG cells than in human hepatocytes, still, in the presence of drugs we observed a concentration dependent decrease in uptake. In all cell types, the culture time had a significant impact not only on the uptake process but on the inhibitory effect of drugs too. The most significant drug effect was measured at 4 h after seeding. Our report is among the first concerning interactions of the uptake transporters in the HepaRG, at the functional level. Results of the present study clearly show that concerning the inhibition of taurocholate uptake by cholestatic drugs, HepaRG cells are closer to human hepatocytes than rat hepatocytes. In conclusion, we demonstrated that HepaRG cells provide a suitable tool for hepatic uptake studies

A mezőgazdasági szervezetek gazdálkodásának vizsgálata a 2002–2009 közötti időszakban

A tanulmány a magyarországi mezőgazdasági – kettős könyvvitelt vezető – szervezetek erőforrásainak és teljesítményének vizsgálatára irányul az Agrárgazdasági Kutató Intézet (AKI) által rendelkezésünkre bocsátott adatbázis alapján, idősorok elemzése segítségével. Megállapítható, hogy a hazai mezőgazdasági szervezetekben mind a foglalkoztatottságot, az eszközállományt, a kibocsátási teljesítményt tekintve a jogi személyiségű gazdasági társaságok (rt.-k és kft.-k) a meghatározók. Ezen belül a korlátolt felelősségű társaságok dominálnak. A 2007–2009. évek átlagában a mezőgazdasági szervezetek adózás előtti eredményének közel 60%-át a kft.-k, 30%-át az rt.-k realizálták, s a maradék 10%-on osztoztak a szövetkezetek, valamint az egyéb szervezetek. Figyelemreméltó, hogy a vizsgált időszakban, kiemelkedően magas eszközkoncentráció mellett, az rt.-k jövedelmezőségi mutatóinak javulása messze meghaladta a kft.-k azonos mutatóit. 2002–2003-hoz képest a mezőgazdasági szövetkezetek társadalmi-gazdasági súlya igen jelentősen csökkent. Feltételezhető, hogy a szövetkezetek és az egyéb szervezetek egy része kft.-vé alakulva folytatta működését. Javasoljuk, hogy a Központi Statisztikai Hivatal (KSH) és az AKI a gazdálkodó szervezetek aggregált adatainak elemzésén túlmenően, törekedjen az egyes szervezeti formák adatbázisainak a közzétételére és azok elemzésére. ----------------------------------- The present study focuses on the analysis of the resources and performance of Hungarian agricultural organisations on the basis of the database provided by the Research Institute of Agricultural Economics (RIAE), by means of time series analysis. As regards employment, assets employed and volume of output in Hungarian agricultural organisations, our findings indicate that corporations with legal entity (joint stock companies and limited liability companies) are determinant. Within this category, limited liability companies dominate. In the average of the years 2007-2009, approximately 60% of income before taxes received by agricultural organisations was realised by limited liability companies, 30% by joint stock companies and the remaining 10% was shared by cooperatives and other organisations It is remarkable that in the investigated period, with outstandingly high asset concentration, the improvement of profitability indicators for joint stock companies far exceeded the same indicators for limited liability companies. Following the period of Hungary’s accession to the European Union, the socio-economic weight of agricultural organisations significantly decreased and the roles of other organisations became even more marginal than before. Presumably, some cooperatives and other organisations continued their operation as limited liability companies. Our suggestion is that in addition to the analysis of aggregated data of farming organisations, the Central Statistical Office (KSH) and RIAE should endeavour to publish and analyse the database of individual organisational forms.mezőgazdasági szervezetek, erőforrások, kibocsátások, jövedelmezőségi mutatók, agricultural organisations, resources, outputs, profitability indicators, Agribusiness, Agricultural Finance, Productivity Analysis,

UR-INE Good Hands in the Neuroscience Intensive Care Unit

Purpose: The purpose of this performance improvement project was to decrease the harm to patients related to catheter associated urinary tract infections (CAUTIs) in the Neuroscience Intensive Care Unit (NSICU). Historically, the NSICU had the highest CAUTI rates in our organization with as many as 6 CAUTIs per calendar year quarter. A core group of nurses partnered with providers, hospital infection preventionists and nursing leaders to improve practices for placing, removing, and managing urinary catheters and to implement guidelines for urine testing stewardship. Description: Significance: In 2020, the NSICU had 12 CAUTIs, the highest number of CAUTIs in the entire healthcare enterprise (n=16). The Standard Infection Ratio (SIR) reached a high of 5.12 despite a Standard Utilization Ratio (SUR) less than 1.0. Catheter associated urinary tract infection is a common healthcare associated infection. Approximately 14% of hospitalized adults will have a urinary catheter during their hospitalization and with each day the risk of CAUTI increases by 3- 7%. CAUTIs are associated with increased length of stay, increased cost and increased mortality and morbidity (NHSN, 2023). With an estimated additional cost of $14,000 to treat a CAUTI, reducing this HAI in NSICU was a top organizational priority. Strategy: The NSICU had previously implemented multiple strategies to reduce CAUTIs (male and female alternative devices, daily review for necessity, chlorhexidine (CHG) bath treatment, and utilization of the organizational nurse-led bladder management algorithm for urinary catheter removal). We included these strategies as well as compliance with CAUTI related process measures (hand hygiene, CHG bathing, daily review for necessity) in our annual education. We held skills fairs for nursing team members to demonstrate aseptic insertion technique, daily catheter care and best practices for management of the system. Each CAUTI was thoroughly investigated, and, seeing no change in our CAUTI rates, nurses were asked to redouble their efforts in catheter care and maintenance and timely removal. Due to the lack of overall improvement in CAUTI rates, we consulted with our infection preventionist (IP) who provided data on urine testing practices in the NSICU. Like many other providers, NSICU physicians routinely ordered urine cultures as part of a fever work-up. Urine cultures without the presence of indication is considered inappropriate based on CDC guidelines. Our IP noted that many of our patients did not have clinical indications for urine culture testing. It was possible that our CAUITs were the result of colonization of the catheter with the development of biofilm or catheter-associated asymptomatic bacteriuria (CDC, 2019). As this pattern of inappropriate urine testing emerged, we decided to add urine testing stewardship (UTS) to our bundle of strategies to reduce CAUTIs. Our goal was to obtain urine cultures only when indicated (i.e. recent kidney transplant recipients, neutropenia, recent genitourinary surgery, known genitourinary obstruction or stents, pregnancy, spinal cord injury with signs or symptoms of autonomic dysreflexia, organ donor or patient with classic signs of urinary tract infection without alternative explanation). Implementation: In November 2020 NSICU nurse leaders partnered with the IP and medical director to identify opportunities to implement UTS. All strategies were developed with the understanding that this was not simply a practice change but also a culture change. Team meetings (nursing and physician) were held to discuss the new approach to urine testing. Signs with testing criteria were affixed to each computer so that providers would use that information when placing orders. To ensure adherence to the change, in January 2021 all orders for urine cultures were to be reviewed by nurse leaders before being obtained. Nurse leaders were well versed in the literature supporting the reduction of unnecessary testing and frequently provided just-in-time education to nurses and providers to answer any questions and address concerns about urine testing. Evaluation and Implications for Nursing Practice and Patient Outcomes:   With the addition of urine testing stewardship, CAUTI rates dropped from a high of 5.12 in 2020 Q3 to 0.0 in 2021 Q3. Between 2020 Q1 and 2021 Q3 there were 16 CAUTIs. From May 27, 2021 to April 24, 2023 (696 days) there were no CAUTIs. As of July 2023, no patients hospitalized in the NSICU with an indwelling urinary catheter were readmitted with a UTI or urosepsis. In addition to the CAUTI in April 2023, there was a second patient with a CAUTI on July 23, 2023. In both cases, the patients did not meet testing criteria nor were nursing leaders notified of the order. Combined with previous strategies, the introduction of urine testing stewardship reduced patient harm. We eliminated the unnecessary use of antimicrobials which, in turn, increases the proliferation of resistant organisms (AHRQ, May 2023). Furthermore, providers have become more likely to approve catheter removal earlier in the course of hospitalization.   These initiatives were successful in that we reduced our CAUTI rates and changed the culture on the unit. Implementation of UTS in particular required collaboration among all team members and support from providers. Nurses are more confident in asking the “why” for other tests and procedures ordered on their patients. Nurse leader review of routine urine cultures was adopted as a best practice across the organization

Structure restraints from heteronuclear pseudocontact shifts generated by lanthanide tags at two different sites

Pseudocontact shifts (PCS) encode long-range information on 3D structures of protein backbones and side-chains. The level of structural detail that can be obtained increases with the number of different sites tagged with a paramagnetic metal ion to generate PCSs. Here we show that PCSs from two different sites can suffice to determine the structure of polypeptide chains and their location and orientation relative to the magnetic susceptibility tensor χ, provided that PCSs are available for 1H as well as heteronuclear spins. In addition, PCSs from two different sites are shown to provide detailed structural information on the conformation of methyl group-bearing amino-acid side-chains. A previously published ensemble structure of ubiquitin is shown to explain the magnetic susceptibility and alignment tensors slightly better than structures that try to explain the experimental data by a single conformation, illustrating the potential of PCSs as a tool to investigate small conformational changes.Financial support by the Australian Research Council is gratefully acknowledged

Solution conformations of a linked construct of the Zika virus NS2B-NS3 protease

The Zika virus presents a serious risk for global health. Crystal structures of different constructs of the Zika virus NS2B-NS3 protease (NS2B-NS3pro) have been determined with the aim to provide a basis for rational drug discovery. In these structures, the C-terminal β-hairpin of NS2B, NS2Bc, was observed to be either disordered (open conformation) or bound to NS3pro complementing the substrate binding site (closed conformation). Enzymatically active constructs of flaviviral NS2B-NS3 proteases commonly used for inhibitor testing contain a covalent peptide linker between NS2B and NS3pro. Using a linked construct of Zika virus NS2B-NS3pro, we studied the location of NS2Bc relative to NS3pro in solution by pseudocontact shifts generated by a paramagnetic lanthanide tag attached to NS3pro. Both closed and open conformations were observed with different inhibitors. As the NS2B co-factor is involved in substrate binding of flaviviral NS2B-NS3 proteases, the destabilization of the closed conformation in the linked construct makes it an attractive tool to search for inhibitors that interfere with the formation of the enzymatically active, closed conformation.C.N. thanks the Alexander vonHumbolt Foundation for a Feodor-Lynen fellowship. Financialsupport by the Australian Research Council is gratefully acknowl-edged. C.K. acknowledges support by the Deutsche For-schungsgemeinschaft, grant KL 1356/3

A New Concept to Secure Food Safety Standards against Fusarium Species and Aspergillus Flavus and Their Toxins in Maize

Commercial maize hybrids are exposed to different degrees of ear infection by toxigenic fungal species and toxin contamination. Their resistance to different fungi and toxin relationships are largely unknown. Without this knowledge, screening and breeding are not possible for these pathogens. Seven- to tenfold differences were found in resistance to Fusarium spp., and there was a five-fold difference in ear coverage (%) in response to A. flavus. Three hybrids of the twenty entries had lower infection severity compared with the general means for toxigenic species. Three were highly susceptible to each, and 14 hybrids reacted differently to the different fungi. Differences were also observed in the toxin content. Again, three hybrids had lower toxin content in response to all toxigenic species, one had higher values for all, and 16 had variable resistance levels. Correlations between infection severity and deoxynivalenol (DON) content were 0.95 and 0.82 (p = 0.001) for F. graminearum and F. culmorum, respectively. For fumonisin and F. verticillioides ear rot, the Pearson correlation coefficient (r) was 0.45 (p = 0.05). Two independent isolates with different aggressiveness were used, and their mean X values better described the resistance levels. This increased the reliability of the data. With the introduction of this methodological concept (testing the resistance levels separately for different fungi and with two isolates independently), highly significant resistance differences were found. The resistance to different fungal species correlated only in certain cases; thus, each should be tested separately. This is very useful in registration tests and post-registration screening and breeding. This would allow a rapid increase in food and feed safety

Prebiotic and probiotic agents enhance antibody-based immune responses to Salmonella Typhimurium infection in pigs

Salmonellosis causes significant economic losses to the pig industry and contaminated pork products are an important source of Salmonella for humans. The EU ban on the use of antibiotic growth promoters in pig production, and the emergence of antibiotic resistance has meant there is a pressing need for alternative control strategies for pathogenic bacteria such as S. Typhimurium in pigs. Here, we determined the effects of prebiotic, probiotic and synbiotic diet regimes on antibody responses to oral Salmonella challenge of pigs. The data demonstrate that the inclusion of the probiotic Lactobacillus plantarum B2984 in the diet of piglets (∼1 × 1010 cfu/animal/day) enhanced serum IgM (P<0.001), IgG (P=0.001) and IgA (P=0.039) responses to S. Typhimurium infection including cross-reacting antibodies to S. Enteritidis. Similarly, inclusion of the prebiotic lactulose at 1% (w/w) of the feed on a daily basis in the diet enhanced serum IgM (P=0.010), IgG (P=0.004) and IgA (P=0.046) responses to S. Typhimurium infection and also cross-reacting antibodies to S. Enteritidis. Inclusion of both additives in the synbiotic diet also elicited an enhanced immune response with IgM (P=0.009) and IgG (P=0.046) levels being increased, however a significant interaction of the pre and probiotics was observed when considering the immune responses to S. Typhimurium (IgM P=0.004; IgG and IgA, P<0.001 for interaction). With respect to immune responses, the effects of pre or probiotic administration were the same or reduced in the synbiotic diet compared to when used in isolation. The data support the use of Lactobacillus plantarum B2984 or lactulose as strategies to contribute to the protection of weaned piglets from zoonotic bacterial pathogens, but caution must be taken when combining dietary supplements as combinations can interact

The TATA-binding protein regulates maternal mRNA degradation and differential zygotic transcription in zebrafish

Early steps of embryo development are directed by maternal gene products and trace levels of zygotic gene activity in vertebrates. A major activation of zygotic transcription occurs together with degradation of maternal mRNAs during the midblastula transition in several vertebrate systems. How these processes are regulated in preparation for the onset of differentiation in the vertebrate embryo is mostly unknown. Here, we studied the function of TATA-binding protein (TBP) by knock down and DNA microarray analysis of gene expression in early embryo development. We show that a subset of polymerase II-transcribed genes with ontogenic stage-dependent regulation requires TBP for their zygotic activation. TBP is also required for limiting the activation of genes during development. We reveal that TBP plays an important role in the degradation of a specific subset of maternal mRNAs during late blastulation/early gastrulation, which involves targets of the miR-430 pathway. Hence, TBP acts as a specific regulator of the key processes underlying the transition from maternal to zygotic regulation of embryogenesis. These results implicate core promoter recognition as an additional level of differential gene regulation during development

Molecular pathway profiling of T lymphocyte signal transduction pathways; Th1 and Th2 genomic fingerprints are defined by TCR and CD28-mediated signaling

Contains fulltext : 108719.pdf (publisher's version ) (Open Access)BACKGROUND: T lymphocytes are orchestrators of adaptive immunity. Naive T cells may differentiate into Th1, Th2, Th17 or iTreg phenotypes, depending on environmental co-stimulatory signals. To identify genes and pathways involved in differentiation of Jurkat T cells towards Th1 and Th2 subtypes we performed comprehensive transcriptome analyses of Jurkat T cells stimulated with various stimuli and pathway inhibitors. Results from these experiments were validated in a human experimental setting using whole blood and purified CD4+ Tcells. RESULTS: Calcium-dependent activation of T cells using CD3/CD28 and PMA/CD3 stimulation induced a Th1 expression profile reflected by increased expression of T-bet, RUNX3, IL-2, and IFNgamma, whereas calcium-independent activation via PMA/CD28 induced a Th2 expression profile which included GATA3, RXRA, CCL1 and Itk. Knock down with siRNA and gene expression profiling in the presence of selective kinase inhibitors showed that proximal kinases Lck and PKCtheta are crucial signaling hubs during T helper cell activation, revealing a clear role for Lck in Th1 development and for PKCtheta in both Th1 and Th2 development. Medial signaling via MAPkinases appeared to be less important in these pathways, since specific inhibitors of these kinases displayed a minor effect on gene expression. Translation towards a primary, whole blood setting and purified human CD4+ T cells revealed that PMA/CD3 stimulation induced a more pronounced Th1 specific, Lck and PKCtheta dependent IFNgamma production, whereas PMA/CD28 induced Th2 specific IL-5 and IL-13 production, independent of Lck activation. PMA/CD3-mediated skewing towards a Th1 phenotype was also reflected in mRNA expression of the master transcription factor Tbet, whereas PMA/CD28-mediated stimulation enhanced GATA3 mRNA expression in primary human CD4+ Tcells. CONCLUSIONS: This study identifies stimulatory pathways and gene expression profiles for in vitro skewing of T helper cell activation. PMA/CD3 stimulation enhances a Th1-like response in an Lck and PKCtheta dependent fashion, whereas PMA/CD28 stimulation results in a Th2-like phenotype independent of the proximal TCR-tyrosine kinase Lck. This approach offers a robust and fast translational in vitro system for skewed T helper cell responses in Jurkat T cells, primary human CD4+ Tcells and in a more complex matrix such as human whole blood