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While it is recognized that non-alcoholic fatty liver disease (NAFLD) is associated with cardiovascular disease (CVD), how NAFLD affects the development and progression of CVD remains unclear and debatable. Hence, we aimed to determine the role of steatotic hepatocyte-derived small extracellular vesicles (sEVs) in foam cell formation and atherosclerosis progression. sEVs from steatotic hepatocytes were isolated and characterized. MicroRNA (miRNA) deep sequencing was utilized to identify functional miRNA in sEVs. Lastly, we conducted a cross-sectional study on patients with NAFLD to validate these findings. Treatment of sEVs from steatotic hepatocytes promoted macrophage-derived foam cell formation and atherosclerosis progression via inhibition of ABCA1-mediated cholesterol efflux. Macrophage-specific deletion of Abca1 in ApoE -/- mice abolished the role of steatotic hepatocyte-derived sEVs in atherosclerosis progression. In addition, hepatocyte-specific deletion of Rab27a , which is the key GTPase regulating sEV release, significantly ameliorated high-fat, high-cholesterol diet-induced atherosclerosis progression in ApoE -/- mice. The miRNA deep sequencing results showed that miR-30a-3p was enriched in sEVs from steatotic hepatocytes. miR-30a-3p directly targeted the 3' untranslated region of ABCA1 to inhibit ABCA1 expression and cholesterol efflux. Treatment with antagomiR-30a-3p significantly attenuated atherosclerosis progression in high-fat, high-cholesterol diet-fed ApoE -/- mice. Moreover, serum sEVs from patients with NAFLD and sEV-miR-30a-3p expression were associated with decreased cholesterol efflux levels in foam cells. Steatotic hepatocyte-derived sEVs promote foam cell formation and facilitate atherogenesis via the miR-30a-3p/ABCA1 axis. Reducing sEV secretion by steatotic hepatocytes or targeting miR-30a-3p may be potential therapeutic approaches to slow the progression of NAFLD-driven atherosclerosis. The presence of hepatic steatosis is strongly correlated with the risk of cardiovascular disease and cardiovascular events, yet the molecular mechanisms linking steatosis to progression of atherosclerosis are unclear. Herein, we identified small extracellular vesicles from steatotic hepatocytes as a trigger that accelerated the progression of atherosclerosis. Steatotic hepatocyte-derived small extracellular vesicles promoted foam cell formation via the miR-30a-3p/ABCA1 axis. Our findings not only provide mechanistic insight into non-alcoholic fatty liver disease-driven atherosclerosis but also provide potential therapeutic targets for patients with atherosclerosis. [Display omitted] • Extracellular vesicles from steatotic hepatocytes promote foam cell formation in atherosclerosis. • Steatotic hepatocyte-derived extracellular vesicles inhibit ABCA1-mediated cholesterol efflux via miR-30a-3p. • Inhibition of extracellular vesicle release from hepatocytes ameliorates atherosclerosis progression in ApoE -/- mice. • Treatment of antagomiR-30a-3p attenuates high-fat, high-cholesterol diet-induced atherosclerosis progression in ApoE -/- mice. [ABSTRACT FROM AUTHOR]

• miR-495-3p inhibits leukemic cell growth and is downregulated in BCR-ABL1 -expressing cell lines. • miR-495-3p modulates responses to tyrosine kinase inhibitor treatment, including those in UT-7 cells expressing T315I. • Overexpression of miR-495-3p leads to a decrease ofin MDR1 expression and drug efflux activity. • Bioinformatics analyses revealed that miR-495-3p targets genes that are upregulated in blast crisis. Chronic myeloid leukemia (CML) is a clonal hematopoietic malignancy driven by the BCR-ABL1 fusion oncoprotein. The development of tyrosine kinase inhibitors (TKIs) has deeply increased long-term survival of CML patients. Nonetheless, one patient out of four will switch TKI off owing either to drug intolerance or resistance partly due to amplification or mutations of BCR-ABL1 oncogene and alteration in ATP-binding cassette (ABC) transporters. Increasing evidence suggests the involvement of the microRNA miR-495-3p in cancer-associated chemoresistance through multidrug resistance 1 (MDR1) gene, which encodes an ATP-dependent efflux pump. Our study aimed at investigating the potential role of miR-495-3p in CML TKI chemo-sensitivity and determining the underlying molecular circuitry involved. We first observed that miR-495-3p expression was lower in BCR-ABL1 -expressing cellular models in vitro. Notably, loss-of-function experiments showed increased proliferation associated with a decreased number of nondividing cells (G0/G1) and resistance to Imatinib. Conversely, our data showed that miR-495-3p overexpression hindered leukemic cell growth and TKI resistance in Imatinib-resistant T315I-mutant cells, as well as drug efflux activity through MDR1 regulation. Further investigating the role of miR-495-3p in CML patients, we found that predicted miR-495-3p targets were upregulated in patients in blast crisis that were involved in protein phosphorylation and associated with the worst prognosis. Taken together, our results demonstrate that downregulation of miR-495-3p expression is important in the malignant phenotype of CML and TKI resistance mechanisms and could be a useful biomarker and a potential therapeutic target to eradicate CML. [Display omitted] [ABSTRACT FROM AUTHOR]

• A higher prevalence of MDR-NTHi strains is observed in northern Taiwan. • New transposons conferring AMP, TET, and CHL resistance are identified. • Efflux pumps contribute to NTHi resistance to cephems, carbapenems, and macrolides. Nontypeable Haemophilus influenzae (NTHi), once considered a harmless commensal, has emerged as a significant concern due to the increased prevalence of multidrug-resistant (MDR) strains and their association with invasive infections. This study aimed to explore the epidemiology and molecular resistance mechanisms of 51 NTHi isolates collected from patients with invasive infections in northern Taiwan between 2011 and 2020. This investigation revealed substantial genetic diversity, encompassing 29 distinct sequence types and 18 clonal complexes. Notably, 68.6% of the isolates exhibited ampicillin resistance, with 28 categorised as MDR and four isolates were even resistant to up to six antibiotic classes. Among the MDR isolates, 18 pulsotypes were identified, indicating diverse genetic lineages. Elucidation of their resistance mechanisms revealed 18 β-lactamase-producing amoxicillin–clavulanate-resistant (BLPACR) isolates, 12 β-lactamase-producing ampicillin-resistant (BLPAR) isolates, and 5 β-lactamase-nonproducing ampicillin-resistant (BLNAR) isolates. PBP3 analysis revealed 22 unique substitutions in BLPACR and BLNAR, potentially contributing to cephem resistance. Notably, novel transposons, Tn 7736 -Tn 7739 , which contain critical resistance genes, were discovered. Three strains harboured Tn 7739 , containing seven resistance genes [ aph(3′)-Ia, bla TEM-1 , catA, sul2, strA, strB , and tet (B)], while four other strains carried Tn 7736 , Tn 7737 , and Tn 7738 , each containing three resistance genes [ bla TEM-1 , catA , and tet (B)]. The emergence of these novel transposons underscores the alarming threat posed by highly resistant NTHi strains. Our findings indicated that robust surveillance and comprehensive genomic studies are needed to address this growing public health challenge. [Display omitted] [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is a common cause of skin infections, food poisoning and severe life-threatening infections. Methicillin-Resistant Staphylococcus aureus (MRSA) is known to cause chronic nosocomial infections by virtue of its multidrug resistance and biofilm formation mechanisms. The antimicrobial resistance owned by S. aureus is primarily due to efflux pumps and formation of microbial biofilms. These drug resistant, sessile and densely packed microbial communities possess various mechanisms including quorum sensing and drug efflux. Quorum sensing is a cooperative physiological process which is used by bacterial cells for social interaction and signal transduction in biofilm formation whereas efflux of drugs is derived by efflux pumps. Apart from their significant role in multidrug resistance, efflux pumps also contribute to transporting cell signalling molecules and due to their occurrence; we face the frightening possibility that we will enter the pre-antibiotic era soon. Compounds that modulate efflux pumps are also known as efflux pump inhibitors (EPI's) that act in a synergistic manner and potentiate the antibiotics efficacy which has been considered as a promising approach to encounter bacterial resistance. EPIs inhibit the mechanism of drug efflux s as well as transport of quorum sensing signalling molecules which are the supreme contributors of miscellaneous virulence factors. This review presents an accomplishments of the recent investigations allied to efflux pump inhibitors against S. aureus and also focus on related correspondence between quorum sensing system and efflux pump inhibitors in terms of S. aureus and MRSA biofilms that may open a new avenue for controlling MRSA infections. [ABSTRACT FROM AUTHOR]

• ABCA1 and ABCG1 promote cholesterol efflux to suppress foam cell generation. • lncRNAs regulate cholesterol efflux to influence atherosclerosis development. • ABCA1 and ABCG1 are regulated by different lncRNAs. • Some lncRNAs play dual roles in ABCA1 expression and atherosclerosis. • Some lncRNAs form a feedback loop with target gene. Coronary heart disease (CHD) with atherosclerosis is the leading cause of death worldwide. ABCA1 and ABCG1 promote cholesterol efflux to suppress foam cell generation and reduce atherosclerosis development. Long noncoding RNAs (lncRNAs) are emerging as a unique group of RNA transcripts that longer than 200 nucleotides and have no protein-coding potential. Many studies have found that lncRNAs regulate cholesterol efflux to influence atherosclerosis development. ABCA1 is regulated by different lncRNAs, including MeXis, GAS5, TUG1, MEG3, MALAT1, Lnc-HC, RP5-833A20.1, LOXL1-AS1, CHROME, DAPK1-IT1, SIRT1 AS lncRNA, DYNLRB2-2, DANCR, LeXis, LOC286367, and LncOR13C9. ABCG1 is also regulated by different lncRNAs, including TUG1, GAS5, RP5-833A20.1, DYNLRB2-2, ENST00000602558.1, and AC096664.3. Thus, various lncRNAs are associated with the roles of ABCA1 and ABCG1 on cholesterol efflux in atherosclerosis regulation. However, some lncRNAs play dual roles in ABCA1 expression and atherosclerosis, and the functions of some lncRNAs in atherosclerosis have not been investigated in vivo. In this article, we review the roles of lncRNAs in atherosclerosis and focus on new insights into lncRNAs associated with the roles of ABCA1 and ABCG1 on cholesterol efflux and the potential of these lncRNAs as novel therapeutic targets in atherosclerosis. [ABSTRACT FROM AUTHOR]

O -linked-N-acetylglucosaminylation (O -GlcNAcylation), a dynamic post-translational modification (PTM), holds profound implications in controlling various cellular processes such as cell signaling, metabolism, and epigenetic regulation that influence cancer progression and therapeutic resistance. From the therapeutic perspective, O -GlcNAc modulates drug efflux, targeting and metabolism. By integrating signals from glucose, lipid, amino acid, and nucleotide metabolic pathways, O -GlcNAc acts as a nutrient sensor and transmits signals to exerts its function on genome stability, epithelial-mesenchymal transition (EMT), cell stemness, cell apoptosis, autophagy, cell cycle. O -GlcNAc also attends to tumor microenvironment (TME) and the immune response. At present, several strategies aiming at targeting O -GlcNAcylation are under mostly preclinical evaluation, where the newly developed O -GlcNAcylation inhibitors markedly enhance therapeutic efficacy. Here we systematically outline the mechanisms through which O -GlcNAcylation influences therapy resistance and deliberate on the prospects and challenges associated with targeting O -GlcNAcylation in future cancer treatments. • O -GlcNAc regulates multiple key biological processes, contributing to cancer's therapeutic resistance. • O -GlcNAcylation targets essential elements within the tumor microenvironment and immune response. • Plenty of current strategies target O -GlcNAcylation in cancer. • O -GlcNAcylation shows great potential for clinical use, with challenges needing future research. [ABSTRACT FROM AUTHOR]

• EPIs bring a major decrease in resistance to antibiotics. • Broad spectrum EPIs development would be an important tactic for MDR infections. • EPIs can be used to enhance antibiotics efficacy in the of biofilm treatment. • Few EPIs from natural source show less toxicity but same efficiency as synthetic one. Bacterial multidrug resistance (MDR) has become a major public health threat. Along with enzymatic modifications, overexpression of MDR efflux pumps is a significant antibiotic resistance mechanism. Efflux pumps are the main mediators of resistance mechanisms to many classes of drug and biofilm structure. Identification of natural substrates and inhibitors is an emerging research discipline. Efflux pump inhibition enables drug accumulation inside bacterial cells, allowing more effective delivery to the intended targets, potentially enhancing the antibacterial action. In this review, we highlight the role of efflux pumps and natural molecules as their inhibitors, both in Gram-positive and Gram-negative bacteria. We also discuss the structure of the AcrAB-TolC efflux pump in Enterobacteriaceae , its mechanisms of action and antibiotic synergetic effects with natural molecules. Antibiotic resistance has become a global concern for healthcare workers and physicians. Efflux pumps are one of the major mechanisms of resistance. Hence, we describe examples of natural efflux pump inhibitors used to combat antibiotic resistance. [ABSTRACT FROM AUTHOR]

Polyethylene glycols (PEGs) in general use are polydisperse molecules with molecular weight (MW) distributed around an average value applied in their designation e.g. , PEG 4000. Previous research has shown that PEGs can act as P-glycoprotein (P-gp) inhibitors with the potential to affect the absorption and efflux of concomitantly administered drugs. However, questions related to the mechanism of cellular uptake of PEGs and the exact role played by P-gp has not been addressed. In this study, we examined the mechanism of uptake of PEGs by MDCK-mock cells, in particular, the effect of MW and interaction with P-gp by MDCK-hMDR1 and A549 cells. The results show that: (a) the uptake of PEGs by MDCK-hMDR1 cells is enhanced by P-gp inhibitors; (b) PEGs stimulate P-gp ATPase activity but to a much lesser extent than verapamil; and (c) uptake of PEGs of low MW (<2000 Da) occurs by passive diffusion whereas uptake of PEGs of high MW (>5000 Da) occurs by a combination of passive diffusion and caveolae-mediated endocytosis. These findings suggest that PEGs can engage in P-gp-based drug interactions which we believe should be taken into account when using PEGs as excipients and in PEGylated drugs and drug delivery systems. PEGs are taken up by cells and act as P-gp substrates. Low molecular weight (MW) PEGs cross cell membranes by passive diffusion, whereas those high MW PEGs enter cells by a combination of passive diffusion and caveolae-mediated endocytosis. Image 1 [ABSTRACT FROM AUTHOR]

• Glymphatic transport is impaired in the cerebral small vessel disease animal model. • Enlarged perivascular spaces and impaired glymphatic transport coexist. • Enlarged perivascular spaces may be a manifestation of impaired glymphatic system. To observe glymphatic transport and evaluate enlarged perivascular spaces (PVSs) in spontaneously hypertensive rats (SHRs). SHRs were used as an animal model of cerebral small vessel disease, and Wistar Kyoto (WKY) rats were used as the control group. Histopathology was used to evaluate the enlargement of PVSs. A fluorescent tracer was infused into the cisterna magna of rats, and the proportion of the brain parenchyma area exposed to the fluorescent tracer was later quantified to evaluate the influx and efflux function of the glymphatic system. The global and polarized expression of aquaporin protein 4 (AQP4) was analyzed by immunofluorescence. Compared with WKY rats, SHRs exhibited obviously enlarged PVSs and significantly decreased influx and efflux function of the glymphatic system. The results showed a significant decrease in AQP4 polarity in SHRs, but a difference in global AQP4 expression was not observed between SHRs and WKY rats. Impaired glymphatic transport may be involved in the pathogenesis of arteriolosclerotic cerebral small vessel disease, and enlarged PVSs may be a manifestation of the impaired glymphatic system. [ABSTRACT FROM AUTHOR]

The upsurge of multiple drug resistance (MDR) bacteria substantially diminishes the effectiveness of antibiotic arsenal and therefore intensifies the rate of therapeutic failure. The major factor in MDR is efflux pump-mediated resistance. A unique pump can make bacteria withstand a wide range of structurally diverse compounds. Therefore, their inhibition is a promising route to eliminate resistance phenomenon in bacteria. Phytochemicals are excellent alternatives as resistance-modifying agents. They can directly kill bacteria or interact with the crucial events of pathogenicity, thereby decreasing the ability of bacteria to develop resistance. Numerous botanicals display noteworthy efflux pumps inhibitory activities. Edible plants are of growing interest. Likewise, some plant families would be excellent sources of efflux pump inhibitors (EPIs) including Apocynaceae, Berberidaceae, Convolvulaceae, Cucurbitaceae, Fabaceae, Lamiaceae, and Zingiberaceae. Easily applicable methods for screening plant-derived EPIs include checkerboard synergy test, berberine uptake assay and ethidium bromide test. In silico high-throughput virtual detection can be evaluated as a criterion of excluding compounds with efflux substrate-like characteristics, thereby improving the selection process and extending the identification of EPIs. To ascertain the efflux activity inhibition, real-time PCR and quantitative mass spectrometry can be applied. This review emphasizes on efflux pumps and their roles in transmitting bacterial resistance and an update plant-derived EPIs and strategies for identification. Image 1 • Active efflux as the main resistance strategy in bacteria. • Phytochemicals as promising alternatives against efflux pumps-mediated MDR. • Herbals-based efflux pump inhibitors screening, the methods. [ABSTRACT FROM AUTHOR]

Drones or unmanned aerial vehicles are autonomous or remotely controlled multipurpose aerial vehicles driven by aerodynamic forces and capable of carrying a payload. Whereas initially used exclusively for military purposes, the use of drones has gradually spread into other areas. Given their great flexibility and favourable costs, the use of drones has also been piloted in various healthcare settings. We briefly summarize current knowledge regarding the use of drones in healthcare, focusing on infectious diseases and/or microbiology when applicable. Information was sought through PubMed and extracted from peer-reviewed literature published between January 2010 and August 2019 and from reliable online news sources. The search terms 'drones', 'unmanned aerial vehicles', 'microbiology' and 'medicine' were used. Peer-reviewed literature on the use of drones in healthcare has steadily increased in recent years. Drones have been successfully evaluated in various pilot programmes and are already implemented in some settings for transporting samples and delivering blood, vaccines, medicines, organs, life-saving medical supplies and equipment. In addition, a promising proof-of-concept 'lab-on-a-drone' was recently presented, as well as several pilot studies showing the benefits of drone use in surveillance and epidemiology of infectious diseases. The potential for drone use in clinical microbiology, infectious diseases and epidemiology is vast. Drones may help to increase access to healthcare for individuals that might otherwise not benefit from appropriate care due to remoteness and lack of infrastructure or funds. However, factors such as national airspace legislation and legal medical issues, differences in topography and climates, cost-effectiveness, and community attitudes and acceptance in different cultures and societies currently impede the widespread use of drones. Significant cost savings compared with ground transportation, speed and convenience of delivery, and the booming drone sector will probably drive drone implementation in various areas of medicine in the next 5 years. [ABSTRACT FROM AUTHOR]

The tumor microenvironment (TME) is an attractive target to develop novel strategies for hormone-dependent cancers. Several molecules in the TME can favor tumor development and progression, including lipoproteins. Lipoproteins are taken up by cancer cells, providing them with cholesterol and fatty acids. Cholesterol regulates cell signaling and it is converted into a series of bioactive metabolites, including hormones. The conflicting results of epidemiological and interventional studies suggest that the local availability of lipoproteins in the TME is more relevant for cancer biology than their circulating levels. Thus, reducing lipoprotein uptake and stimulating cell cholesterol efflux to high-density lipoproteins (HDLs) can represent a novel adjuvant strategy for cancer management. HDL-like particles can also act as drug delivery systems for tumor targeting. The TME can favor tumor progression and TME signature can predict disease outcome. Sources of lipids, proinflammatory, and pro-oxidant molecules are present in the TME, including lipoproteins. Cholesterol affects cell proliferation as a component of lipid rafts regulating cell signaling, and as building block for the local synthesis of hormones and oxysterols. VLDL and LDL in the TME can provide cancer cells with lipids and exert proinflammatory and pro-oxidant activities. HDL can promote the removal of cholesterol and its metabolites from cancer cells and blunt inflammation and oxidative stress. Boosting the HDL system by the infusion of reconstituted HDL could represent a promising approach to affect cancer cell viability. [ABSTRACT FROM AUTHOR]

• Polymyxin B combinations were tested against carbapenemase-producing Klebsiella pneumoniae. • Synergy was frequently observed with fosfomycin, minocycline, and rifampicin. • Wildtype mgrB was associated with synergy of polymyxin B and aztreonam. • Absence of arr-2 / arr-3 was associated with synergy of polymyxin B and rifampicin. • Bacterial genetic determinants of combination effects warrant further investigation. Combination therapy can enhance the activity of available antibiotics against multidrug-resistant Gram-negative bacteria. This study assessed the effects of polymyxin B combinations against carbapenemase-producing Klebsiella pneumoniae (K. pneumoniae). Twenty clinical K. pneumoniae strains producing NDM-1 (n = 8), OXA-48-like (n = 10), or both NDM-1 and OXA-48-like (n = 2) carbapenemases were used. Whole-genome sequencing was applied to detect resistance genes (e.g. encoding antibiotic-degrading enzymes) and sequence alterations influencing permeability or efflux. The activity of polymyxin B in combination with aztreonam, fosfomycin, meropenem, minocycline, or rifampicin was investigated in 24-hour time-lapse microscopy experiments. Endpoint samples were spotted on plates with and without polymyxin B at 4 x MIC to assess resistance development. Finally, associations between synergy and bacterial genetic traits were explored. Synergistic and bactericidal effects were observed with polymyxin B in combination with all other antibiotics: aztreonam (11 of 20 strains), fosfomycin (16 of 20), meropenem (10 of 20), minocycline (18 of 20), and rifampicin (15 of 20). Synergy was found with polymyxin B in combination with fosfomycin, minocycline, or rifampicin against all nine polymyxin-resistant strains. Wildtype mgrB was associated with polymyxin B and aztreonam synergy (P = 0.0499). An absence of arr-2 and arr-3 was associated with synergy of polymyxin B and rifampicin (P = 0.0260). Emergence of populations with reduced polymyxin B susceptibility was most frequently observed with aztreonam and meropenem. Combinations of polymyxin B and minocycline or rifampicin were most active against the tested NDM-1 and OXA-48-like-producing K. pneumoniae. Biologically plausible genotype-phenotype associations were found. Such information might accelerate the search for promising combinations and guide individualised treatment. [ABSTRACT FROM AUTHOR]

The long-term persistence of antibiotic resistance in the environment, especially in drinking water, is a public health concern. Expression of an efflux pump, an important mechanism of resistance to antibiotics, usually confers a fitness cost in bacteria. In this study, we aimed to determine why antibiotic resistance conferred by overexpression of an efflux pump persisted in low-nutrient environments (TOC < 10 mg/L) such as drinking and source water in which antibiotic selective pressure might be very low or even absent. Competition experiments between wild-type Pseudomonas aeruginosa and ciprofloxacin-resistant mutants revealed that the fitness cost of ciprofloxacin resistance significantly decreased (p < 0.05) under low-nutrient (0.5 mg/L total organic carbon (TOC)) relative to high-nutrient (500 mg/L TOC) conditions. Mechanisms underlying this fitness cost were analyzed. The mexD gene expression in resistant bacteria (cip_3 strain) was significantly lower (p < 0.05) in low-nutrient conditions, with 10 mg/L TOC ((8.01 ± 0.82)-fold), than in high-nutrient conditions, with 500 mg/L TOC ((48.89 ± 4.16)-fold). Moreover, rpoS gene expression in resistant bacteria ((1.36 ± 0.13)-fold) was significantly lower (p < 0.05) than that in the wild-type strain ((2.78 ± 0.29)-fold) under low-nutrient conditions (10 mg/L TOC), suggesting a growth advantage. Furthermore, the difference in metabolic activity between the two competing strains was significantly smaller (p < 0.05) in low-nutrient conditions (5 and 0.5 mg/L TOC). These results suggest that nutrient levels are a key factor in determining the persistence of antibiotic resistance conferred by efflux pumps in the natural environment with trace amounts or no antibiotics. Unlabelled Image [ABSTRACT FROM AUTHOR]

Long non-coding RNAs (lncRNAs) have proven to be involved in the progression of atherosclerosis and dyslipidemia. In addition, vascular smooth muscle cells (VSMCs) phenotype switching, including VSMCs-derived foam cells formation, plays a key role in the pathogenesis of atherosclerosis. LncRNA ENST00000602558.1, one of the differentially expressed lncRNAs between coronary artery disease (CAD) patients and healthy controls identified by our previous study, was located to TG and HDL susceptibility loci, but its role and underlying mechanism in the pathogenesis of atherosclerosis remain unclear. The present study aims to explore the role and underlying mechanism of ENST00000602558.1 in the regulation of cholesterol efflux from VSMCs. ABCG1 mRNA and protein expression in VSMCs was detected using qRT-PCR and Western blot, respectively. ABCG1-mediated cholesterol efflux to HDL from VSMCs was measured by means of NBD-cholesterol fluorescence intensity. The binding of ENST00000602558.1 to p65 and p65 to ABCG1 promoter region was detected by RNA immunoprecipitation (RIP) assay and chromatin immunoprecipitation (ChIP) assay, respectively. Overexpression of ENST00000602558.1 downregulated ABCG1 mRNA and protein expression, while knockdown of ENST00000602558.1 upregulated ABCG1 mRNA and protein expression. Consistently, ENST00000602558.1 overexpression decreased ABCG1-mediated cholesterol efflux to HDL from VSMCs by 30.38% (p < 0.001), and knockdown of ENST00000602558.1 increased ABCG1-mediated cholesterol efflux to HDL from VSMCs by 30.41% (p = 0.001). In addition to cholesterol efflux, overexpression of ENST00000602558.1 increased lipid accumulation and TC/TG levels, while knockdown of ENST00000602558.1 decreased lipid accumulation and TC/TG levels in VSMCs. Furthermore, we confirmed that ENST00000602558.1 regulated ABCG1 expression and ABCG1-mediated cholesterol efflux from VSMCs through binding to p65. In conclusion, ENST00000602558.1 played an important role in mediating cholesterol efflux to HDL from VSMCs by regulating ABCG1 expression through binding to p65. Image 1 • ENST00000602558.1 specifically regulated ABCG1 expression in VSMCs. • ENST00000602558.1 regulated ABCG1-mediated cholesterol efflux to HDL from VSMCs, but not ABCA1-mediated cholesterol efflux to apoA1. • ENST00000602558.1 regulated ABCG1 expression and ABCG1-mediated cholesterol efflux from VSMCs through binding to p65. [ABSTRACT FROM AUTHOR]

Highlights • Responses of soil respiration to rainfall pulse were analyzed in an oak forest. • Drought affects the response of soil respiration to rainfall event. • Larger soil respiration response to rainfall is found under severe drought. • Drought rather than precipitation size affects rainfall induced total C loss. Abstract As a consequence of climate change, a higher frequency of spring drought periods followed by episodic precipitation events is predicted for Central China. Little is known about the effect of drought intensity on the response of soil CO 2 efflux to discrete precipitation events in forest ecosystems. A field experiment was conducted to assess the responses of soil CO 2 efflux to spring drought and precipitation pulse in an oak forest located at climatic transitional zone in Central China. Soil respiration (SR) and heterotrophic respiration (HR) under 30-day drought showed higher responses to precipitation pulse than those under control and 20- day drought treatments. Regardless of drought intensity, precipitation events of 5, 10, and 20 mm levels significantly increased SR by 86.0, 103.4, and 175.2%, and HR by 108.4, 129.5, and 161.5% when compared with non-water addition plots, respectively. Severe drought treatment (30-day drought) amplified the contribution of HR to SR in response to precipitation pulse (84.5% vs 78.3% for the control). Drought intensity rather than precipitation amount showed substantial influence on total carbon loss through SR and HR. These findings highlight the importance of incorporating both drought intensity and precipitation events and their impacts on soil carbon cycling into future predictions of forest ecosystems under climate change. [ABSTRACT FROM AUTHOR]

Highlights • Girdling significantly affected stem CO 2 efflux by altered substrate availability. • Effects of girdling on stem CO 2 efflux differed in degree and timing with season. • Apparent Q 10 of the stem CO 2 efflux increased above the girdle. Abstract The effect of carbohydrate supply on stem CO 2 efflux and its temperature sensitivity is crucial in predicting forest carbon cycling under climate change and forest management but has been rarely reported. In this study, we altered carbohydrate supply via stem girdling and investigated the effect of nonstructural carbohydrate concentration on stem CO 2 efflux and its temperature sensitivity in Chinese fir and sweetgum trees in spring and winter. The stem CO 2 efflux of Chinese fir increased significantly above the girdle in spring and winter, but decreased below the girdle in winter. The stem CO 2 efflux of sweetgum increased above the girdle and decreased below the girdle in spring but not in winter. Seasonal variations in the response of stem CO 2 efflux to girdling were associated with the changes in stem soluble sugar and starch concentration, especially in sweetgum tree. Girdling altered Q 10 of the stem CO 2 efflux, and this effect was dependent on location, season and species. Compared with the Q 10 of the stem CO 2 efflux in non-girdled trees, that of CO 2 efflux in Chinese fir increased by 45% above the girdle but decreased by 16% below the girdle in winter. These values also exhibited 6% and 11% responses in spring. By contrast, the Q 10 of the stem CO 2 efflux in sweetgum increased by 95% above the girdle and decreased by 45% below the girdle in spring. However, these values were nearly constant in winter. We concluded that the effect of carbohydrate supply on stem CO 2 efflux and its temperature sensitivity depends on season and species. Therefore, the vital role of carbohydrate supply that regulates stem CO 2 efflux and its temperature sensitivity should be considered for the understanding and modeling of forest carbon cycling response to forest management, especially under global climate changes. [ABSTRACT FROM AUTHOR]

Abstract Polyethylene glycol (PEG)-decorated doxorubicin (Dox)/carboxymethyl chitosan (CMC)/gold nanoparticles (AuNPs) have been developed for cancer therapy. CMC was used as a reducing and stabilizing agent for the fabrication of AuNPs and Dox was loaded onto AuNPs as a chemotherapeutic agent. Dox-loaded CMC-stabilized AuNPs (Dox/CMC-AuNPs) with a mean diameter of 104.0 nm, zeta potential of −48.32 mV, and drug loading efficiency of 60.14% were prepared. PEG was attached to CMC-AuNPs for enhancing systemic drug exposure and prolonging the circulation in blood stream. Compared with Dox/CMC-AuNPs, Dox-loaded PEGylated CMC-AuNPs (Dox/CMC-AuNPs-PEG) showed a reduced hydrodynamic size (71.2 nm), less negative zeta potential (−12.83 mV), and an enhanced Dox loading efficiency (73.14%). Dox/CMC-AuNPs and Dox/CMC-AuNPs-PEG exhibited sustained and pH-dependent drug release profiles and exhibited antiproliferation effects against the A549 cells. In a bi-directional transport study of Caco-2 cell monolayers, AuNPs reduced the efflux ratio, which indicated that the P-glycoprotein-mediated multidrug resistance (MDR) was overcome. Dox/CMC-AuNPs-PEG resulted in reduced drug clearance (CL) and improved half-life (t 1/2), compared with Dox/CMC-AuNPs, in rats after intravenous administration. These results suggest that Dox/CMC-AuNPs-PEG could be a promising nanotherapeutic approach to overcome MDR in cancer and prolong their circulation in the blood stream. [ABSTRACT FROM AUTHOR]

Abstract Tuberculosis is a major cause of mortality and morbidity in developing countries, and the emergency of multidrug and extensive drug resistance cases is an utmost issue on the control of the disease. Despite the efforts on the development of new antibiotics, eventually there will be strains resistant to them as well. Efflux plays an important role in the evolution of resistance in Mycobacterium tuberculosis. Tap is an important efflux pump associated with tuberculosis resistant to isoniazid, rifampicine and ofloxacin and with multidrug resistance. The development of efflux inhibitors for Tap could raise the effectiveness of second line drugs and reduce the duration of the current treatment. Therefore the objective of this study is to build a reliable molecular model of Tap efflux pump and test the possible competitive inhibition between efflux inhibitors and antibiotics in the optimized structure. We built twenty five Tap models with molecular modelling to elect the best according to the results of the validation analysis. The elected model went through to a 100 ns molecular dynamics simulation in a lipid bilayer, and the resulting optimized structure was used in docking studies to test if the used efflux inhibitors may act via competitive inhibition on antibiotics. The validation results pointed the model built by Phyre2 as the closest to a possible native Tap structure, and therefore it was the elected model. RSMD analysis revealed the model is stable, where the predicted binding site stabilized between 15 and 20 ns, maintaining the RMSD at around 0.35 Å throughout the molecular dynamics simulation in a lipid bilayer. Therefore this model is reliable and can also be used for further studies. The docking studies showed a possibility of competitive inhibition by NUNL02 on ofloxacin and bedaquiline, and by verapamil on ofloxacin and rifampicin. This presents the possibility that NUNL02 and verapamil are possible inhibitors of Tap efflux and highlights the importance of including efflux inhibitors as adjuvants to the tuberculosis therapy, as it indicates a possible extrusion of ofloxacin, rifampicin and bedaquilin by Tap. Graphical abstract Image 1 Highlights • Tap-Phyre2 was stable in a molecular dynamics simulation in a lipid bilayer. • Tap-Phyre2 is reliable to be used for further structure-based studies. • Tap may extrude rifampicin, ofloxacin and bedaquiline. • Verapamil may inhibit Tap's extrusion of ofloxacin and rifampicin via competition. • NUNL02 may inhibit Tap's efflux of ofloxacin and bedaquiline via competition. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract Quizartinib, a second-generation FLT3 inhibitor, is in clinical development for the treatment of acute myeloid leukemia. We studied its pharmacokinetic interactions with the multidrug efflux transporters ABCB1 and ABCG2 and the multidrug metabolizing enzyme CYP3A, using in vitro transport assays and knockout and transgenic mouse models. Quizartinib was transported by human ABCB1 in vitro , and by mouse (m)Abcb1 and mAbcg2 in vivo. Upon oral administration, the brain accumulation of quizartinib was 6-fold decreased by mAbcb1 and 2-fold by mAbcg2 (together: 12-fold). Unexpectedly, the absence of mAbcb1 resulted in a ∼2-fold lower plasma exposure in Abcb1a/1b −/− and Abcb1a/1b;Abcg2 −/− mice, suggesting that loss of mAbcb1 causes compensatory alterations in alternative quizartinib elimination or uptake systems. mAbcb1 and mAbcg2 themselves did not appear to restrict quizartinib oral availability. Oral and intravenous pharmacokinetics of quizartinib were not substantially altered between wild-type, Cyp3a knockout and CYP3A4-humanized mice. All three strains showed relatively high (33–51%) oral bioavailability. If this also applies in humans, this would suggest a limited risk of CYP3A-related inter-individual variation in exposure for this drug. Our results provide a possible rationale for using pharmacological ABCB1/ABCG2 inhibitors together with quizartinib when treating malignant lesions situated in part or in whole behind the blood-brain barrier. [ABSTRACT FROM AUTHOR]

Highlights • A screening method based on gene overexpression was developed. • The efflux and influx transporters for MCFA were identified. • Efficient efflux pump system was constructed for MCFA. Abstract Medium-chain fatty acids (C 6 -C 10 , MCFAs) represent valuable molecules for food and pharmaceutical industries. However, MCFAs in Escherichia coli induce stresses and toxicity, reducing final yields. These phenotypes could be alleviated if MCFA export from cells was accelerated, yet information on proteins responsible for exporting MCFAs remains limited. Here, a panel of 17 genes or operons was screened. Overexpressing resistance nodulation cell division family transporter acrE , mdtE and mdtC increased MCFA titer by 46.4%, 65.2% and 33.8%, whereas overexpressing transcriptional activators soxS , marA and multidrug efflux pumps cmr , fadL decreased MCFA titer by 42.5%, 27.8%, 37.6% and 29.5%, respectively. Furthermore, the effect of multiple transporter engineering was investigated. Overexpressing acrE , mdtE and mdtC together with deleting cmr increased MCFA production by more than two-fold. This study demonstrated the efficient efflux pump specifically for MCFAs and this multiple transporter engineering strategy can be employed to synthesize other bio-products toxic to microorganisms. [ABSTRACT FROM AUTHOR]

Multidrug resistance is highly conserved in mammalian, fungal, and bacterial cells, is characterized by resistance to several unrelated xenobiotics, and poses significant challenges to managing infections and many cancers. Eukaryotes use a highly conserved set of drug efflux transporters that confer pleiotropic drug resistance (PDR). To interrogate the regulation of this critical process, here we developed a small molecule-responsive biosensor that couples transcriptional induction of PDR genes to growth rate in the yeast Saccharomyces cerevisiae. Using diverse PDR inducers and the homozygous diploid deletion collection, we applied this biosensor system to genome-wide screens for potential PDR regulators. In addition to recapitulating the activity of previously known factors, these screens identified a series of genes involved in a variety of cellular processes with significant but previously uncharacterized roles in the modulation of yeast PDR. Genes identified as down-regulators of the PDR included those encoding theMADfamily of proteins involved in the mitotic spindle assembly checkpoint (SAC) complex. Of note, we demonstrated that genetic disruptions of the mitotic spindle assembly checkpoint elevate expression of PDRmediating efflux pumps in response to exposure to a variety of compounds that themselves have no known influence on the cell cycle. These results not only establish our biosensor system as a viable tool for investigating PDR in a high-throughput fashion, but also uncover critical control mechanisms governing the PDR response and a previously uncharacterized link between PDR and cell cycle regulation in yeast. [ABSTRACT FROM AUTHOR]

Highlights • RFMS practice significantly increased CO 2 efflux and maize yield simultaneously. • Soil respiration was much higher under the ridge soils than the furrow soils. • Increasing soil respiration under RFMS practice did not induce a decline in SOC. • Carbon emission efficiency was significantly higher in RFMS than in CK practice. Abstract Ridge–furrow mulching system (RFMS) is widely used in arid and semi–arid areas, but its effect on soil respiration (R s) and its components, including heterotrophic (R h) and autotrophic respiration (R a) are still poorly understood. In this study, CO 2 flux from the soil of furrows and ridges was measured across different RFMS practices (i.e., three different ridge/furrow ratios) and conventional flat planting (CK). A trenching method was used to estimate the contribution of R h to R s. Compared with CK, RFMS significantly increased soil temperature of the ridge, promoted soil moisture of the furrow, and enhanced microbial diversity at the early crop growth stage, resulting in increased R s and its components. The ridge soils exhibited much higher R s (3.43 μmol m–2 s–1) than the furrow soils (2.98 μmol m–2 s–1) under all three RFMS practices. The contribution ratios of R h to R s across the different practices ranged from 50.4% to 59.6%. Soil temperature rather than soil moisture explained the seasonal variation of R s and its components for both CK and RFMS. Nonetheless, high R s and R h values in RFMS did not induce a decline of soil organic carbon during the two–year experimental period. Improved root growth in RFMS practices may provide more exudates to the soil, thus offsetting soil carbon decomposition. Compared with CK, RFMS with ridge/furrow ratios of 40:70 cm, 55:55 cm, and 70:40 cm, significantly increased soil CO 2 emissions by 10.6%, 19.6%, and 20.4%, respectively, while increasing maize yield by 26.1%, 36.4%, and 50.3%, respectively. Carbon emission efficiency (CEE) was significantly higher in RFMS than in CK in both years. This study suggests that, due to its high CEE, RFMS with a ridge/furrow ratio of 70:40 cm could be a highly promising strategy for sustaining crop productivity while minimizing environmental impacts. [ABSTRACT FROM AUTHOR]

Highlights • Cyclodextrin inclusion complex of nintedanib prepared using different cyclodextrins. • Sulfobutyl ether β-CD demonstrated higher inclusion and better stability. • Inclusion complex improved transport across clinically relevant EpiIntestinal tissue model. • Inclusion complex also improved bio-activity of nintedanib against lung fibroblast cells. • Shows promise to use via oral or pulmonary route in novel dosage forms. Abstract Cyclodextrin complex of nintedanib was prepared aiming for increased bio-activity and improved transport across intestinal membrane with reduced p-glycoprotein (p-gp) efflux. Based on preliminary phase solubility studies and molecular modeling, sulfobutyl ether derivative of β-cyclodextrin (SBE-β-CD, Captisol®) was selected to prepare inclusion complex. Complexation was confirmed using FTIR, 1H NMR, DSC, and XRD. Bioactivity of the formed complex was tested using lung fibroblast cells, WI-38 for anti-proliferative activity and effect on collagen deposition and cells migration. In-vitro permeability studies were performed using epiIntestinal tissue model to assess the effect of complexation on transport and p-gp efflux. Results of the study demonstrated that cyclodextrin complexation increased stability of nintedanib in PBS (pH 7.4) and simulated intestinal fluid (SIF). Further, bioactivity of nintedanib also improved. Interestingly, complexation has increased transport of nintedanib across intestinal membrane and reduced efflux ratio, suggesting the role of cyclodextrin complexation in modulating p-gp efflux. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Synthesis of amino acid conjugates of 3-(1-chloro-3,4-dihydronaphthalen-2-yl) acrylic acid. • Screening for NorA inhibitors against Staphylococcus aureus. • The potent inhibitors displayed MEC of 1.56 μg/mL against NorA overexpressing strain SA-1199B. • Mechanism for NorA specificity elucidated by ethidium bromide efflux. Abstract Inhibitors for NorA efflux pump of Staphylococcus aureus have attracted the attention of many researchers towards the discovery and development of novel efflux pump inhibitors (EPIs). In an attempt to find specific potent inhibitors of NorA efflux pump of S. aureus , a total of 15 amino acid conjugates of 3-(1-chloro-3,4-dihydronaphthalen-2-yl)acrylic acid (4 – 18) were synthesized using a simple convenient synthetic approach and bioevaluated against NorA efflux pump. Two compounds 7 and 8 (each having MEC of 1.56 µg/mL) were found to restore the activity of ciprofloxacin through reduction of the MIC elucidated by comparing the ethidium bromide efflux in dose dependent manner in addition to ethidium bromide efflux inhibition and accumulation study using NorA overexpressing strain SA-1199B. Most potent compounds among these were able to restore the antibacterial activity of ciprofloxacin completely against SA-1199B. Structure activity relationship (SAR) studies and docking study of potent compounds 7 and 8 could elucidate the structural requirements necessary for interaction with the NorA efflux pumps. On the whole, compounds 7 and 8 have ability to reverse the NorA efflux mediated resistance and could be further optimized for development of potent efflux pump inhibitors. [ABSTRACT FROM AUTHOR]

Increasing atmospheric CO 2 concentration and nitrogen deposition are predicted to increase fresh carbon (C) inputs in soils because of higher net primary productivity. However, relationship between soil C cycling and increased inputs of fresh C in soils is not clear. Moreover, most of the studies on soil organic carbon (SOC) focus on upper 20 cm although >50% SOC is present below this depth. We designed this study to unravel the response of SOC to 0% (control), 0.3%, 0.6%, 1.2% and 2.4% addition of maize litter (C4) to a surface (0–15 cm) and deep layers (55–75 cm) of a C3 soil. The isotopic difference between the maize litter and soil allowed distinguishing soil-derived (native) CO 2 -C from litter-derived CO 2 -C and calculating the priming effect (PE). Total respiration was measured over 78 d incubation whereas the PE was determined 39 d and 78 d after incubation. Our results showed that all the litter addition levels induced PE -stimulation of mineralization of the extant SOC in response to availability of fresh organic matter (FOM)- in surface and deep soil layers. Moreover, the PE was significantly higher in the deep than surface soil layer in response to all litter additions, except 2.4% treatment during the 2nd harvest. This result suggests that the deep SOC would mineralize at higher rates under similar supply of FOM to surface and deep soils in this agroecosystem. We also showed a significantly positive linear relationship between PE and litter addition rates for both soils implying continuous loss of SOC under increasing organic matter inputs. In conclusion, our study reveals that the deep soil C would be more vulnerable to priming effect in response to litter addition than the surface soil. [ABSTRACT FROM AUTHOR]

Highlights • Greater transfer of thermal energy into subsoil than in temperate or boreal forests. • Warming increased surface soil CO 2 efflux by 40% in young Chinese fir plantation. • Warming reduced moisture, increased soluble N and fine root turnover in deep soils. • Increased surface soil CO 2 efflux was due mainly to CO 2 production in deep soils. Abstract How warming affects the magnitude of CO 2 fluxes within the soil profile remains an important question, with implications for modeling the response of ecosystem carbon balance to changing climate. Information on belowground responses to warming is especially limited for the tropics and subtropics because the majority of manipulative studies have been conducted in temperate and boreal regions. We examined how artificial warming affected CO 2 gas production and exchange across soil profiles in a replicated mesocosms experiment relying on heavily weathered subtropical soils and planted with Chinese fir (Cunninghamia lanceolata). Half of 2 × 2 m mesocosms (5 replications) was heated with cables buried at a 10 cm depth, which increased temperature in the whole soil profile by 4.5, 3.6 and 2.5 °C at 15, 30 and 60 cm soil depths, respectively. Using a combination of chamber-based and concentration gradient method (CGM) approaches, we found that warming increased soil CO 2 efflux across the whole profile by 40%. Changes were unevenly distributed across soil depth: mean CO 2 production rate decreased from 0.74 to 0.67 μmol CO 2 m−2 s−1 in topsoils (0–15 cm depth) whereas it increased from 0.26 to 0.73 μmol CO 2 m−2 s−1 in subsoils (15–60 cm depth). Warming reduced moisture more strongly in subsurface than surface soils and increased subsoil soluble N concentrations as well as fine root turnover, in line with previous temperate and boreal warming studies. This consistency indicates that overall responses of subtropical forests to warming may be similar to forests in higher latitudes. [ABSTRACT FROM AUTHOR]

Abstract The changing landscape of fuel consumption related, in part, to increased engine efficiency and the inexpensive supply of natural gas, has led to the closure of multiple refineries. As the operational lifetime of many refineries exceeds 100 years, historical releases of oil and refined products is common. To evaluate remediation and rehabilitation options, there is a need to understand the rate and distribution of natural hydrocarbon mass losses across these large properties. Here, surficial CO 2 flux measurements were used to evaluate naturally occurring hydrocarbon mass losses at a large-scale former refinery that has been closed since 1982. Natural source zone depletion (NSZD) rates over a five-year period (2012–2016) were derived from surficial CO 2 efflux measurements on a high-resolution grid (N > 80). Results demonstrate substantial variations of mass loss rates across the site. Average site-wide mass loss rates ranged from 1.1–5.4 g TPH m−2 d−1 as C 10 H 22 with a multi-year average of 4.0 g TPH m−2 d−1 as decane (C 10 H 22), consistent with observations at other sites. Statistical analysis demonstrated that the same average mass loss rates would have been obtained with fewer measurement locations (N = 20–30). Comparing NSZD rates to site metadata show CO 2 fluxes to be a reasonably good proxy for zones of subsurface hydrocarbon contamination – particularly with respect to vadose zone impacts. It is hypothesized that the observed decline of NSZD rates over the study period is related to rise of groundwater levels, leading to increased submergence of the smear zone. Overall, mass loss rates calculated from CO 2 fluxes show NSZD can result in substantial contaminant removal, which may rival that obtained from engineered remediation, under some conditions. Highlights • CO 2 effluxes were measured at a large-scale former refinery site. • Sampling density required for calculating representative NSZD rates was evaluated. • CO 2 effluxes were correlated with subsurface source zone characteristics, indicating they can be used for contaminant delineation. • Site-wide NSZD rates calculated using background correction ranged from 1.1 to 5.4 g TPH m−2 day−1. • Site-wide NSZD mass losses were comparable to those achieved by engineered systems in localized areas. [ABSTRACT FROM AUTHOR]

Abstract Aims Hydrogen sulfide (H 2 S), an important gasotransmitter, is involved in a variety of cellular functions and pathophysiologic processes. Drug resistance due to alterations in drug trafficking and metabolism severely limits the effectiveness of cancer therapy. This study examined the role of H 2 S in drug resistance in liver cancer cells. Materials and methods Human primary hepatocellular carcinoma cell line (HepG2) and doxorubicin (Dox)-resistant cells were used in this study. Cell survival was analyzed by MTT, Annexin V-FITC/propidium iodide staining and clonogenic assay. Western blotting was used for analysis of protein expression, and immunoprecipitation was used to determine interactions of LXR/RXR. Key findings The expression of H 2 S-generating enzyme cystathionine gamma-lyase (CSE) was inhibited by doxorubicin treatment in HepG2 cells, and H 2 S sensitized Dox-inhibited cell survival and colony formation. In addition, H 2 S promoted cellular retention of Dox by suppressing the expressions of ABCA1 and ABCG8. H 2 S significantly blocked Dox-induced heterodimer formation between LXRα and RXRβ and attenuated the binding of LXRα/RXRβ to the promoters of ABCA1 and ABCG8 genes. RXRβ but not LXRα was S -sulfhydrated by H 2 S, and blockage of RXRβ S -sulfhydration abolished the inhibitory role of H 2 S on LXRα/RXRβ heterodimer formation. CSE expression was reduced in Dox-resistant cells in comparison with their parental cells, while H 2 S could reverse drug resistance in Dox-resistant cells. Significance Our study provides a novel solution for reversing drug resistance in cancer cells by targeting H 2 S signalling. Graphical abstract Unlabelled Image [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • A novel series of covalent Btk inhibitors was discovered. • Hit molecules showed high ER. • Optimization of ER led to the discovery of the lead molecule. Abstract Bruton's tyrosine kinase (Btk) is a member of the Tec kinase family that is expressed in cells of hematopoietic lineage (e.g. B cells, macrophages, monocytes, and mast cells). Small molecule covalent irreversible Btk inhibitors targeting Cys481 within the ATP-binding pocket have been applied in the treatment of B-cell malignancies. Starting from a fragment, we discovered a novel series of potent covalent irreversible Btk inhibitors that bear N -linked groups occupying the solvent accessible pocket (SAP) of the active site of the Btk kinase domain. The hit molecules, however, displayed high P-gp mediated efflux ratio (ER) and poor A-B permeability in Caco-2 assay. By decreasing tPSA, installing steric hindrance and adjusting clogP, one top molecule 9 was discovered, which showed a 99% decrease in efflux ratio and a 90-fold increase in A-B permeability compared to hit molecule 1. [ABSTRACT FROM AUTHOR]

Abstract This study aimed to investigate the cellular disposition of liquiritigenin via the sulfonation pathway and the role of efflux transporters in liquiritigenin sulfate excretion. The sulfonation disposition of liquiritigenin was investigated using SULT1A3 overexpressed HEK293 cells (HEK-SULT1A3 cells). Liquiritigenin generated one mono-sulfate metabolite (7-O-sulfate) in HEK-SULT1A3 cell lysate. And the sulfonation followed the Michaelis-Menten kinetic (V max = 0.84 nmol/min/mg and K m = 7.12 μM). Expectedly, recombinant SULT1A3 (hSULT1A3) showed a highly similar kinetic profile with cell lysate. Furthermore, 7-O-sulfate was rapidly generated and excreted in HEK-SULT1A3 cells. Ko143 (a BCRP-selective inhibitor) at 20 μM significantly decreased the excretion rate of liquiritigenin sulfate (>42.5%, p < 0.001). Moreover, the pan-MRPs inhibitor MK-571 at 20 μM essentially abolished the liquiritigenin sulfate effluxion, resulting in the marked reduction of excretion rate (>97.4%, p < 0.001). Furthermore, knockdown of BCRP led to moderate reduction in sulfate excretion (15.9%–16.9%, p < 0.05). Silencing of MRP4 caused significant decreased in sulfate excretion (20.2%–32.5%, p < 0.01). In conclusion, one sulfate metabolite was generated from liquiritigenin in HEK-SULT1A3 cells. BCRP and MRP4 should be the key factors for the cellular excretion of liquiritigenin sulfate. Graphical abstract Unlabelled Image [ABSTRACT FROM AUTHOR]

Abstract Background and aims Uremia patients have impaired high-density lipoprotein (HDL) function and a high risk of coronary artery disease (CAD). Increased lipoprotein electronegativity can compromise lipoprotein function, but the effect of increased HDL electronegativity on HDL function and its association with CAD in uremia patient are not clear. We aimed to assess HDL electronegativity and various properties of HDL in uremia patients and investigate whether electronegative HDL is a risk factor for CAD in these individuals. Methods HDL from 60 uremia patients and 43 healthy controls was separated into 5 subfractions (H1 H5) with increasing electronegativity by using anion-exchange chromatography. Lipoprotein content was analyzed by gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight-mass spectrometry. HDL anti-oxidant, anti-apoptosis and cholesterol efflux activities were examined by fluorescence-based assays. Results The percentage of H5 HDL (H5%) was significantly higher in uremia patients than in controls (p < 0.001). The concentration of apolipoprotein (Apo) AI was lower and apolipoprotein modifications were more prevalent in uremia HDL subfractions than in control HDL subfractions. Carbamylation of ApoAI and ApoCIII was increased in more electronegative HDL subfractions from uremia patients. Anti-oxidant activity, anti-apoptotic activity, and cholesterol efflux capability were reduced in HDL subfractions from uremia patients when compared with control HDL subfractions. Multiple logistic regression analysis showed that H5% was associated with CAD risk in uremia patients. Conclusions In HDL of uremia patients, increased electronegativity is accompanied by compositional changes and impaired function. Our findings indicate that increased H5% is associated with increased CAD risk in uremia patients. Graphical abstract Image 1 Highlights • Antioxidant lipoproteins were less abundant in uremia H5 than in control H5. • Carbamylation of ApoA1/ApoCIII contributes to increased electronegativity of HDL. • Antioxidant, antiapoptotic, and cholesterol efflux of HDL were reduced in uremia H5. • H5% could be a prediction marker for the development of CAD in uremia patients. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the use, cost, postoperative urinary tract infection (UTI) rates, and complications of dextrose instillation during cystoscopy.Methods: The medical records of patients who underwent cystoscopy during pelvic reconstructive surgery between June 2016 and June 2017 were reviewed. Patients were divided into two groups: patients who had one ampule of dextrose 50% (D50) directly instilled and patients who did not have D50 instilled during cystoscopy. Preoperative demographics, UTI rates, and postoperative complications were compared. Pharmaceutical cost and availability were reported by the pharmacy at our institution.Results: Out of 63 patients identified, dextrose instillation was used in 20 patients and no dextrose was used in 43 patients. Each ampule of D50 cost $2.18 and there were no problems with supply shortage. As D50 was directly instilled into the bladder, there was immediate visualization of ureteral efflux at the time of surgery. Three patients (15%) in the dextrose group and 10 patients (23%) in the nondextrose group developed postoperative UTIs. There was no statistically significant difference in postoperative UTI rates between the two groups (p = 0.43) and there were no differences in postoperative complications.Conclusion: Dextrose is a safe, cost-effective, readily available agent that provides instantaneous visualization of ureteral efflux without an increased risk of postoperative UTI. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract The expression and activity of human placental transporters during pregnancy could be altered by several factors including pathological changes associated with preeclampsia. The aims of this study were to identify the placental efflux transporters involved in the bio-disposition of pravastatin, determine the protein expression of these transporters and their encoding genes as well as the activity of pravastatin uptake in placentas obtained from patients with preeclampsia. ATP-dependent uptake of [3H]-pravastatin by trophoblast tissue apical and basal membrane vesicles exhibited sigmoidal kinetics. The curved shapes of Eadie-Hofstee plots indicate that more than one placental transporter are involved in the uptake of pravastatin. ATP-dependent uptake of [3H]-pravastatin into vesicles expressing MRP1-5, BCRP, and P-gp, as well as the results of inhibition studies suggest that BCRP and MRP1 are the major placental efflux transporters responsible for the in vitro uptake of pravastatin. Compared to placentas from healthy pregnancies, preeclamptic placentas had increased number of syncytial knots with increased expression of BCRP in their apical membrane and increased expression of MRP1 in the cytoplasm of the syncytiotrophoblast and in cytoplasm of syncytial knots. There was a concomitant increase in ABCC1 but not in ABCG2 gene expressions in preeclamptic placentas. ATP-dependent uptake of [3H]-pravastatin by vesicles prepared from apical membranes of preeclamptic placentas was similar to the uptake by vesicles prepared from placentas obtained after uncomplicated pregnancies (13.9 ± 6.5 vs 14.1 ± 5.8 pmol·mg protein−1 min−1). The transporter-specific changes in the expression of BCRP and MRP1 in preeclamptic placentas did not affect the efflux activity of transporters localized on the apical membrane of the syncytiotrophoblast. [ABSTRACT FROM AUTHOR]

This study was carried out to test the essential oil from C. ambrosioides leaves and its main constituent, α-Terpinene, in an antibacterial activity assay. As well, it was evaluated ability reduce resistance to norfloxacin and ethidium bromide was compared the Staphylococcus aureus 1199B whith 1199 wild type strain. The MIC of the C. ambrosioides essential oil and α-Terpinene were determined by microdilution method. The MIC of the essential oil and α-Terpinene presented a value ≥ 1024 μg/mL. However, when associated with antibacterials, the essential oil from C. ambrosioides leaves significantly reduced the MIC of antibiotics and ethidium bromide, characterizing an efflux pump inhibition. The C. ambrosioides essential oil, despite having no direct antibacterial activity against the S. aureus 1199B strain, showed a potentiating action when associated with antibacterial agents, this being attributed to an inhibition of efflux pumps. [ABSTRACT FROM AUTHOR]

The efflux velocity is the basis for the prediction of turbine wake. A novel energy coefficient is defined to propose a new theoretical equation to predict the efflux velocity of tidal current turbine in this paper. Several CFD cases with different tip speed ratio and solidity is conducted using the DES-SA model. In order to overcome the limitations of the axial momentum theory, the effects of tip speed ratio and solidity on the efflux velocity are studied and the energy coefficients with different tip speed ratio and solidity are determined using the proposed equation based on the CFD results. Several semi-empirical efflux velocity equations are finally proposed by fitting the equation of the energy coefficient with tip speed ratio. The application of these equations in the prediction of wake flow and the power calculation of tidal turbine are also introduced in this paper. [ABSTRACT FROM AUTHOR]

Soil alkalization severely restricts agricultural production and economic development worldwide, this problem is far more serious in Songnen Plain, the largest commodity grain base of China. However, little research has been done concerning the mechanisms of plant responses to alkaline stress to date. In this study, we isolated an alkali inducible gene GsBOR2 from Glycine soja on the basis of RNA seq data. GsBOR2 sh high protein sequence similarity with the known boron transporters in other species. The expression of GsBOR2 was highly up-regulated by 50 mM NaHCO 3 treatment and displayed tissue specificity. We then generated the transgenic Arabidopsis overexpressing GsBOR2 and found that the transgenic lines exhibited enhanced alkaline tolerance compared to wild type plants, as illustrated by longer roots and greater shoot biomass. Moreover, GsBOR2 overexpression was also capable of increasing plant resistance to KHCO 3 treatment but not to high-pH stress. Functional complementation of Scbor1 mutant yeasts suggested that GsBOR2 could likely mediate the efflux of boron from cells. Taken together, the alkali responsive gene GsBOR2 is a positive regulator of plant bicarbonate tolerance. [ABSTRACT FROM AUTHOR]

Abstract The putative mechanism by which bacterial RND-type multidrug efflux pumps recognize and transport their substrates is a complex and fascinating enigma of structural biology. How a single protein can recognize a huge number of unrelated compounds and transport them through one or just a few mechanisms is an amazing feature not yet completely unveiled. The appearance of cooperativity further complicates the understanding of structure-dynamics-activity relationships in these complex machineries. Experimental techniques may have limited access to the molecular determinants and to the energetics of key processes regulating the activity of these pumps. Computer simulations are a complementary approach that can help unveil these features and inspire new experiments. Here we review recent computational studies that addressed the various molecular processes regulating the activity of RND efflux pumps. [ABSTRACT FROM AUTHOR]

Resistance of bacteria to antibiotics is a public health concern worldwide due to the increasing failure of standard antibiotic therapies. Antimicrobial photodynamic inactivation (aPDI) is a promising non-antibiotic alternative for treating localized bacterial infections that uses non-toxic photosensitizers and harmless visible light to produce reactive oxygen species and kill microbes. Phenothiazinium photosensitizers like methylene blue (MB) and toluidine blue O are hydrophobic cations that are naturally expelled from bacterial cells by multidrug efflux pumps, which reduces their effectiveness. We recently reported the discovery of a NorA efflux pump inhibitor-methylene blue (EPI-MB) hybrid compound INF55-(Ac)en–MB that shows enhanced photodynamic inactivation of the Gram-positive bacterium methicillin-resistant Staphylococcus aureus (MRSA) relative to MB, both in vitro and in vivo . Here, we report the surprising observation that INF55-(Ac)en–MB and two related hybrids bearing the NorA efflux pump inhibitors INF55 and INF271 also show enhanced aPDI activity in vitro (relative to MB) against the Gram-negative bacteria Escherichia coli and Acinetobacter baumannii , despite neither species expressing the NorA pump. Two of the hybrids showed superior effects to MB in murine aPDI infection models. The findings motivate wider exploration of aPDI with EPI-MB hybrids against Gram-negative pathogens and more detailed studies into the molecular mechanisms underpinning their activity. [ABSTRACT FROM AUTHOR]

Soil CO 2 efflux and its carbon isotopic composition are undoubtedly important for estimating ecosystem carbon budgets and for partitioning respiration sources at various spatial and temporal scales. Under natural conditions, non-steady processes will strongly influence the diffusive fluxes of 13 CO 2 and 12 CO 2 between soil and the atmosphere, which results in variations of effluxed soil δ 13 CO 2 and will lead to bias in respiration source partitioning. In this study, we present a set of quantitative relationships between soil CO 2 efflux and its δ 13 C by solving the diffusion equation. The results showed that the effluxed δ 13 CO 2 converged toward the respiratory δ 13 CO 2 with an increasing efflux rate but that the values greatly differed at low efflux rates. Both our own experiments and data from the literature verified this convergence pattern of the effluxed δ 13 CO 2 , which implies that most of the variations in the δ 13 C of soil effluxed CO 2 may derive from diffusive fractionation rather than from biological causes. Our results explain the isotopic flux patterns of CO 2 under natural environmental variations and are vitally important for isotope-based modeling of ecosystem carbon exchange under changing climatic regimes. [ABSTRACT FROM AUTHOR]

Background and aims Tolerance to soil acidity was studied in two species of Ericaceae that grow in mine-contaminated soils (S Portugal, SW Spain) to find out if there are interspecific variations in H + tolerance which might be related to their particular location. Methods Tolerance to H + toxicity was tested in nutrient solutions using seeds collected in SW Spain. Plant growth and nutrient contents in leaves, stems and roots were determined. Viability tests and proton exchange were studied in roots exposed, short-term, to acidic conditions. Membrane ATPase activity and the cell-wall pectic polysaccharide domain rhamnogalacturonan-II (RG-II) were analysed to find out interspecific differences. Results Variation in survival, growth and mineral composition was found between species. The H + -tolerant species ( Erica andevalensis ) showed greater concentration of nutrients than E. australis . Very low pH (pH 2) produced a significant loss of root nutrients (K, P, Mg) in the sensitive species. Root ATPase activity was slightly higher in the tolerant species with a correspondingly greater H + efflux capacity. In both species, the great majority of the RG-II domains were in their boron-bridged dimeric form. However, shifting to a medium of pH 2 caused some of the boron bridges to break in the sensitive species. Conclusions Variation in elements linked to the cell wall-membrane complex and the stability of their components (RG-II, H + -ATPases) are crucial for acid stress tolerance. Thus, by maintaining root cell structure, active proton efflux avoided toxic H + build-up in the cytoplasm and supported greater nutrient acquisition in H + -tolerant species. [ABSTRACT FROM AUTHOR]

A multi-step procedure has been described which afforded satisfactory yields of N,N′ -disubstituted cinnamamides derived from N -Boc-protected amino acids (Boc-Gly, Boc-Val, Boc-Phe). The key step of this synthesis was a regioselective RedAl reduction of an amide function in presence of a carbamate group. Next, these cinnamamides were evaluated in co-admnistration with ciprofloxacin as efflux pump inhibitors against two S. aureus strains, NorA overexpressing SA1199B and wild type SA1199. In parallel, their intrinsic toxicity was appreciated on human lung fibroblast MRC5 cells. Therefore, the cinnamamide combining both carbamate and indol-3-yl groups, was found to be the most active and one of the less toxic EPI and constituted a promising hit. [ABSTRACT FROM AUTHOR]

Background Efflux pumps of the Resistance-Nodulation-cell Division superfamily confer multi-drug resistance to Gram-negative bacteria. The most-studied polyspecific transporter belonging to this class is the inner-membrane trimeric antiporter AcrB of Escherichia coli . In previous studies, a functional rotation mechanism was proposed for its functioning, according to which the three monomers undergo concerted conformational changes facilitating the extrusion of substrates. However, the molecular determinants and the energetics of this mechanism still remain unknown, so its feasibility must be proven mechanistically. Methods A computational protocol able to mimic the functional rotation mechanism in AcrB was developed. By using multi-bias molecular dynamics simulations we characterized the translocation of the substrate doxorubicin driven by conformational changes of the protein. In addition, we estimated for the first time the free energy profile associated to this process. Results We provided a molecular view of the process in agreement with experimental data. Moreover, we showed that the conformational changes occurring in AcrB enable the formation of a layer of structured waters on the internal surface of the transport channel. This water layer, in turn, allows for a fairly constant hydration of the substrate, facilitating its diffusion over a smooth free energy profile. Conclusions Our findings reveal a new molecular mechanism of polyspecific transport whereby water contributes by screening potentially strong substrate-protein interactions. General significance We provided a mechanistic understanding of a fundamental process related to multi-drug transport. Our results can help rationalizing the behavior of other polyspecific transporters and designing compounds avoiding extrusion or inhibitors of efflux pumps. [ABSTRACT FROM AUTHOR]

Estrone, estradiol and estriol are endogenous human estrogens that are rapidly conjugated with glucuronic acid in both intestinal and hepatic epithelial cells. The resulting glucuronides, estrone-3-glucuronide (E 1 -G), estradiol-3- and 17-glucuronides (E 2 -3G and E 2 -17G), as well as estriol-3- and 16-glucuronides (E 3 -3G and E 3 -16G) are found in human plasma and urine. Unlike E 2 -17G, the efflux transport of other estrogen glucuronides by human transporters has not yet been investigated comprehensively. We have studied the transport of E 1 -G, E 2 -3G, E 3 -3G, E 3 -16G and estrone-3-sulfate (E 1 -S), another important estrogen conjugate, using the vesicular transport assay with recombinant human MRP2, MRP3, MRP4, MDR1 and BCRP that were expressed in insect cells. The transport screening assays revealed that whereas E 1 -S was a good and specific substrate for BCRP, the less transporter-specific conjugates, E 1 -G and E 2 -3G, were still transported by BCRP at 10-fold higher rates than E 1 -S. BCRP also transported E 3 -16G at higher rates than the studied MRPs, while it transported E 3 -3G at lower rates than MRP3. MRP2 exhibited lower or equal transport rates of E 1 -G, E 2 -3G, E 3 -3G and E 3 -16G in comparison to MRP3 and BCRP in the screening assays, mainly due to its high K m values, between 180 and 790 μM. MRP3 transported all the tested glucuronides at rather similar rates, at K m values below 20 μM, but lower V max values than other transporters. In the case of E 3 -3G, MRP3 was the most active transporter in the screening assay. MRP4 transported only E 3 -16G at considerable rates, while none of the tested estrogen conjugates was transported by MDR1 at higher rates than control vesicles. These new results, in combination with previously reported in vivo human data, stimulate our understanding on the substrate specificity and role of efflux transporters in disposition of estrogen glucuronides in humans. [ABSTRACT FROM AUTHOR]

Drug-induced liver injury (DILI) is a common reason for drug withdrawal from the market. An important cause of DILI is drug-induced cholestasis. One of the major players involved in drug-induced cholestasis is the bile salt efflux pump (BSEP; ABCB11 ). Inhibition of BSEP by drugs potentially leads to cholestasis due to increased (toxic) intrahepatic concentrations of bile acids with subsequent cell injury. In order to investigate the possibilities for in silico prediction of cholestatic effects of drugs, we developed a mechanistic biokinetic model for human liver bile acid handling populated with human in vitro data. For this purpose we considered nine groups of bile acids in the human bile acid pool, i.e. chenodeoxycholic acid, deoxycholic acid, the remaining unconjugated bile acids and the glycine and taurine conjugates of each of the three groups. Michaelis-Menten kinetics of the human uptake transporter Na + -taurocholate cotransporting polypeptide (NTCP; SLC10A1 ) and BSEP were measured using NTCP-transduced HEK293 cells and membrane vesicles from BSEP-overexpressing HEK293 cells. For in vitro-in vivo scaling, transporter abundance was determined by LC-MS/MS in these HEK293 cells and vesicles as well as in human liver tissue. Other relevant human kinetic parameters were collected from literature, such as portal bile acid levels and composition, bile acid synthesis and amidation rate. Additional empirical scaling was applied by increasing the excretion rate with a factor 2.4 to reach near physiological steady-state intracellular bile acid concentrations (80 μM) after exposure to portal vein bile acid levels. Simulations showed that intracellular bile acid concentrations increase 1.7 fold in the presence of the BSEP inhibitors and cholestatic drugs cyclosporin A or glibenclamide, at intrahepatic concentrations of 6.6 and 20 μM, respectively. This simplified model provides a tool for a first indication whether drugs at therapeutic concentrations might cause cholestasis by inhibiting BSEP. [ABSTRACT FROM AUTHOR]

CHF6001 is a new and potent PDE4 inhibitor for the treatment of human lung diseases, designed for topical administration by inhalation. In preclinical assessment CHF6001 appeared safe and devoid of emetic effect, which is typical side effect of PDE4 inhibitors in humans. CHF6001 absorption, distribution and excretion were evaluated in rats by PO and IV administration of [14C]CHF6001; additionally the role of transporters was investigated by using transfected cells expressing either human transporters or MDR1 and BCRP KO mice. [14C]CHF6001 intravenously administered as bolus distributed in all the tissues (with very low levels in brain and fetus) and it was mainly eliminated in bile. Following oral administration [14C]CHF6001 about half of the dose was absorbed through the gut. In vitro, CHF6001 was a substrate of human membrane transporters MDR1 and BCRP. In wild and BCRP KO mice CHF6001 was not detectable in brain, whereas it was measurable in Mdr1a/b KO mice. Therefore, in animal species Mdr1a/b plays a significant role in CHF6001 disposition, limiting its distribution into brain and contributing to the safety profile observed in preclinical evaluation. This behavior was confirmed by the results of the first human studies, where CHF6001 was safe and with no emetic effect at all the evaluated doses. [ABSTRACT FROM AUTHOR]

Low phosphorus (P) availability elicits efflux of organic substances viz. carboxylic acids, phenolics, proteins, amino acids, sugars and other low molecular weight compounds in many leguminous crops including soybean ( Glycine max (L.) Merr.). The potential for root exudation varies widely among soybean genotypes, as synthesis and secretion of root exudates place additional burden on the carbon demand of the plant. Hence, efficient photosynthetic machinery may attribute to the differential root exudation potential of soybean genotypes in response to low soil P availability. An attempt was made to understand the varietal differences in photo-biochemical processes of soybean genotypes identified previously with contrasting root exudation potential under low P (Vengavasi and Pandey, 2016). Genotypes EC-232019 (P-efficient) and EC-113396 (P-inefficient) were grown in soil with low (2 mg P kg −1 soil) and sufficient (25 mg P kg −1 soil) P levels under natural environment and observations were recorded at anthesis. The genotype EC-232019 exhibited higher maximal carboxylation rate (V cmax ), maximal photosynthesis (A max ), apparent quantum efficiency (Φ), mesophyll conductance (g m ), triose phosphate utilization rate (TPU), photochemical quenching (qP) and electron transport rate (ETR), along with higher chlorophyll a , total chlorophyll and total carotenoid concentration as compared to the P-inefficient EC-113396. Low P-induced reduction in maximal electron transport rate (J max ) and Φ was higher in EC-113396 rather than EC-232019, suggesting superior photo-biochemical efficiency in the latter. The observed variation in P uptake and growth responses might be attributed in part to the improved photo-biochemical processes exhibited by the P-efficient genotype EC-232019. [ABSTRACT FROM AUTHOR]

The fruits of Euodia rutaecarpa (Euodiae Fructus, EF), the widely used traditional Chinese medicine, have various central nervous system effects. Alkaloids following as evodiamine (EDM), rutaecarpine (RCP) and dehydroevodiamine (DEDM) are the major substances in EF. The MDCK-pHaMDR cell monolayer model was utilized as a blood-brain barrier (BBB) surrogate model to study their BBB permeability. The transport samples were analyzed by high performance liquid chromatography and the apparent permeability coefficients ( P app ) were calculated. EDM and RCP showed high permeability through BBB by passive diffusion, while DEDM showed moderate permeability with efflux mechanism related to P -glycoprotein ( P -gp). EDM and RCP could also reduce the efflux of DEDM probably by inhibiting P -gp. The neuroprotective effects of the three alkaloids were then studied on the PC12 cell line injured by 1-methyl-4-phenylpyridinium ion (MPP + ) or hydrogen peroxide (H 2 O 2 ). EDM could significantly reduce MPP + or H 2 O 2 -induced cell injury dose-dependently. RCP could increase the cell viability in MPP + treated group while DEDM showed a protective effect against H 2 O 2 injury. This study predicted the permeability of EDM, RCP and DEDM through BBB and discovered the neuroprotective substance basis of EF as a potential encephalopathy drug. [ABSTRACT FROM AUTHOR]

A novel series of 4-substituted 2-naphthamide derivatives were designed, synthesized and evaluated for their biological activity. In particular, the ability of the compounds to potentiate the action of antibiotics, to inhibit Nile Red efflux and to target AcrB specifically was investigated. The results indicated that most of the 4-substituted 2-naphthamide derivatives were able to synergize with the antibiotics tested, and inhibit Nile Red efflux by AcrB in the resistant phenotype. Subsequent exclusion of compounds with off target effects such as outer- or inner membrane permeabilization identified compounds 7c , 7g, 12c , 12i and 13g as efflux pump inhibitors (EPIs). Particularly, compounds 7c , 7g and 12i were found to be the most potent EPIs, which synergized with the two substrates tested at lower concentrations than that of parent A3, demonstrating an improvement in potency as compared to A3. Additionally, when the outer membrane of E. coli was permeabilized, compound 12c displayed a huge increase in efficacy and was able to synergize with erythromycin at a concentration that was 16 times lower than that of the parent A3. Hence we were able to design and synthesize compounds that displayed significant increase in efficacy as EPIs against AcrB. [ABSTRACT FROM AUTHOR]