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International Journal of Biological... 2020Hyperuricemia (HUA) is a metabolic disease characterized by elevated serum uric acid (SUA). Empagliflozin, a kind of sodium-glucose cotransporter 2 inhibitors, has...
Hyperuricemia (HUA) is a metabolic disease characterized by elevated serum uric acid (SUA). Empagliflozin, a kind of sodium-glucose cotransporter 2 inhibitors, has recently emerged as a new antidiabetic agent by facilitating glucose excretion in urine. Moreover, there was evidence of SUA reduction following treatment with empagliflozin in addition to glycaemic control, while the molecular mechanisms remain unknown. To investigate the potential mechanisms, the model of type 2 diabetes (T2DM) with HUA was established by combination of peritoneal injection of potassium oxonate and intragastric administration of hypoxanthine in KK-Ay mice. A series of method such as RT-PCR, western blot, immunochemistry, immunofluorescence were conducted to explore the mechanism. Our results showed that empagliflozin significantly ameliorated the levels of SUA and blood glucose in T2DM mice with HUA. Furthermore, in both kidney and ileum, empagliflozin obviously promoted protein expression of uric acid (UA) transporter ABCG2, p-AMPK, p-AKT and p-CREB. The same trend was observed in human tubular epithelial (HK-2) cells. Additionally, through application of an AMPK inhibitor (Compound C), it was further confirmed empagliflozin exerted its anti-hyperuricemic effects in an AMPK dependent manner. Meanwhile, with the help of ChIP assay and luciferase reporter gene assay, we found that CREB further activated ABCG2 via binding to the promoter of ABCG2 to induce transcription. Taken together, our study demonstrated that empagliflozin treatment played an essential role in attenuating HUA by upregulation of ABCG2 via AMPK/AKT/CREB signaling pathway.
Topics: AMP-Activated Protein Kinases; ATP Binding Cassette Transporter, Subfamily G, Member 2; Animals; Benzhydryl Compounds; Blotting, Western; Cell Line; Chromatin Immunoprecipitation; Cyclic AMP Response Element-Binding Protein; Diabetes Mellitus, Type 2; Glucosides; HEK293 Cells; Humans; Hyperuricemia; Immunohistochemistry; Male; Mice; Mice, Inbred C57BL; Proto-Oncogene Proteins c-akt; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction
PubMed: 32015688
DOI: 10.7150/ijbs.33007 -
Nanotoxicology May 2019Ligands that accelerate nanoceria dissolution may greatly affect its fate and effects. This project assessed the carboxylic acid contribution to nanoceria dissolution in...
Ligands that accelerate nanoceria dissolution may greatly affect its fate and effects. This project assessed the carboxylic acid contribution to nanoceria dissolution in aqueous, acidic environments. Nanoceria has commercial and potential therapeutic and energy storage applications. It biotransforms . Citric acid stabilizes nanoceria during synthesis and in aqueous dispersions. In this study, citrate-stabilized nanoceria dispersions (∼4 nm average primary particle size) were loaded into dialysis cassettes whose membranes passed cerium salts but not nanoceria particles. The cassettes were immersed in iso-osmotic baths containing carboxylic acids at pH 4.5 and 37 °C, or other select agents. Cerium atom material balances were conducted for the cassette and bath by sampling of each chamber and cerium quantitation by ICP-MS. Samples were collected from the cassette for high-resolution transmission electron microscopy observation of nanoceria size. In carboxylic acid solutions, nanoceria dissolution increased bath cerium concentration to >96% of the cerium introduced as nanoceria into the cassette and decreased nanoceria primary particle size in the cassette. In solutions of citric, malic, and lactic acids and the ammonium ion ∼15 nm, ceria agglomerates persisted. In solutions of other carboxylic acids, some select nanoceria agglomerates grew to ∼1 micron. In carboxylic acid solutions, dissolution half-lives were 800-4000 h; in water and horseradish peroxidase they were ≥55,000 h. Extending these findings to and environmental systems, one expects acidic environments containing carboxylic acids to degrade nanoceria by dissolution; two examples would be phagolysosomes and in the plant rhizosphere.
Topics: Carboxylic Acids; Cerium; Hydrogen-Ion Concentration; Ligands; Microscopy, Electron, Transmission; Nanoparticles; Oxidation-Reduction; Particle Size; Solubility; Surface Properties
PubMed: 30729879
DOI: 10.1080/17435390.2018.1553251 -
Frontiers in Immunology 2021Congenital Zika virus (ZIKV) infection can induce fetal brain abnormalities. Here, we investigated whether maternal ZIKV infection affects placental physiology and...
Congenital Zika virus (ZIKV) infection can induce fetal brain abnormalities. Here, we investigated whether maternal ZIKV infection affects placental physiology and metabolic transport potential and impacts the fetal outcome, regardless of viral presence in the fetus at term. Low (10 PFU-ZIKV; low ZIKV) and high (5x10 PFU-ZIKV; high ZIKV) virus titers were injected into immunocompetent (ICompetent C57BL/6) and immunocompromised (ICompromised A129) mice at gestational day (GD) 12.5 for tissue collection at GD18.5 (term). High ZIKV elicited fetal death rates of 66% and 100%, whereas low ZIKV induced fetal death rates of 0% and 60% in C57BL/6 and A129 dams, respectively. All surviving fetuses exhibited intrauterine growth restriction (IUGR) and decreased placental efficiency. High-ZIKV infection in C57BL/6 and A129 mice resulted in virus detection in maternal spleens and placenta, but only A129 fetuses presented virus RNA in the brain. Nevertheless, pregnancies in both strains produced fetuses with decreased head sizes (p<0.05). Low-ZIKV-A129 dams had higher IL-6 and CXCL1 levels (p<0.05), and their placentas showed increased CCL-2 and CXCL-1 contents (p<0.05). In contrast, low-ZIKV-C57BL/6 dams had an elevated CCL2 serum level and increased type I and II IFN expression in the placenta. Notably, less abundant microvilli and mitochondrial degeneration were evidenced in the placental labyrinth zone (Lz) of ICompromised and high-ZIKV-ICompetent mice but not in low-ZIKV-C57BL/6 mice. In addition, decreased placental expression of the drug transporters P-glycoprotein (P-gp) and breast cancer resistance protein (Bcrp) and the lipid transporter Abca1 was detected in all ZIKV-infected groups, but Bcrp and Abca1 were only reduced in ICompromised and high-ZIKV ICompetent mice. Our data indicate that gestational ZIKV infection triggers specific proinflammatory responses and affects placental turnover and transporter expression in a manner dependent on virus concentration and maternal immune status. Placental damage may impair proper fetal-maternal exchange function and fetal growth/survival, likely contributing to congenital Zika syndrome.
Topics: ATP-Binding Cassette Transporters; Animals; Apoptosis; Biomarkers; Female; Gene Expression; Host-Pathogen Interactions; Immunity; Immunocompromised Host; Immunohistochemistry; Male; Mice; Placenta; Pregnancy; Pregnancy Complications, Infectious; Zika Virus; Zika Virus Infection
PubMed: 34093581
DOI: 10.3389/fimmu.2021.680246 -
Frontiers in Bioscience (Landmark... Jun 2009The liver plays a key role in the metabolic conversion and elimination of endo- and xenobiotics. Hepatobiliary transport of many of these compounds is mediated by... (Review)
Review
The liver plays a key role in the metabolic conversion and elimination of endo- and xenobiotics. Hepatobiliary transport of many of these compounds is mediated by several ATP-binding cassette (ABC) transporters expressed at the canalicular membrane of the hepatocyte. Impaired function of these ABC transporters leads to impaired bile formation or cholestasis and mutations in these genes are associated with a variety of hereditary cholestatic syndromes. At the transcriptional level, these ABC transporters and the metabolizing enzymes involved in processing of their substrates are coordinately regulated by members of the nuclear receptor (NR) family of ligand-modulated transcription factors. In this review we will focus on ABC transporters involved in hepatobiliary excretion and how they are associated with hepatic physiology and disease states. We will also examine how NRs, acting as intracellular sensors for lipophilic molecules, regulate these ABC transporters and maintain metabolic homeostasis.
Topics: ATP-Binding Cassette Transporters; Animals; Bile Acids and Salts; Biliary Tract; Cholestasis, Intrahepatic; Humans; Liver; Liver Diseases; Models, Biological; Receptors, Cytoplasmic and Nuclear
PubMed: 19482594
DOI: 10.2741/3576 -
The Journal of Investigative Dermatology Feb 2016SLURP1, a member of the lymphocyte antigen 6 protein family, is secreted by suprabasal keratinocytes. Mutations in SLURP1 cause a palmoplantar keratoderma (PPK) known as... (Comparative Study)
Comparative Study
SLURP1, a member of the lymphocyte antigen 6 protein family, is secreted by suprabasal keratinocytes. Mutations in SLURP1 cause a palmoplantar keratoderma (PPK) known as mal de Meleda. SLURP2, another secreted lymphocyte antigen 6 protein, is encoded by a gene located ?20 kb downstream from SLURP1. SLURP2 is produced by suprabasal keratinocytes. To investigate the importance of SLURP2, we first examined Slurp2 knockout mice in which exon 2-3 sequences had been replaced with lacZ and neo cassettes. Slurp2(-/-) mice exhibited hyperkeratosis on the volar surface of the paws (i.e., palmoplantar keratoderma), increased keratinocyte proliferation, and an accumulation of lipid droplets in the stratum corneum. They also exhibited reduced body weight and hind limb clasping. These phenotypes are similar to those of Slurp1(-/-) mice. To solidify a link between Slurp2 deficiency and palmoplantar keratoderma and to be confident that the disease phenotypes in Slurp2(-/-) mice were not secondary to the effects of the lacZ and neo cassettes on Slurp1 expression, we created a new line of Slurp2 knockout mice (Slurp2X(-/-)) in which Slurp2 was inactivated with a simple nonsense mutation. Slurp2X(-/-) mice exhibited the same disease phenotypes. Thus, Slurp2 deficiency and Slurp1 deficiencies cause the same disease phenotypes.
Topics: Adaptor Proteins, Signal Transducing; Animals; Antigens, Ly; Cells, Cultured; Codon, Nonsense; Disease Models, Animal; GPI-Linked Proteins; Gene Expression Regulation; Immunohistochemistry; Keratinocytes; Keratoderma, Palmoplantar; Mice; Mice, Knockout; Phenotype; Random Allocation; Real-Time Polymerase Chain Reaction; Urokinase-Type Plasminogen Activator
PubMed: 26967477
DOI: 10.1016/j.jid.2015.11.003 -
Journal of Neurochemistry Sep 2018The blood-brain barrier (BBB) maintains brain homeostasis by tightly regulating the exchange of molecules with systemic circulation. It consists primarily of...
The blood-brain barrier (BBB) maintains brain homeostasis by tightly regulating the exchange of molecules with systemic circulation. It consists primarily of microvascular endothelial cells surrounded by astrocytic endfeet, pericytes, and microglia. Understanding the make-up of transporters in rat BBB is essential to the translation of pharmacological and toxicological observations into humans. In this study, experimental workflows are presented in which the optimization of (a) isolation of rat brain microvessels (b) enrichment of endothelial cells, and (c) extraction and digestion of proteins were evaluated, followed by identification and quantification of BBB proteins. Optimization of microvessel isolation was indicated by 15-fold enrichment of endothelial cell marker Glut1 mRNA, whereas markers for other cell types were not enriched. Filter-aided sample preparation was shown to be superior to in-solution sample preparation (10251 peptides vs. 7533 peptides). Label-free proteomics was used to identify nearly 2000 proteins and quantify 1276 proteins in isolated microvessels. A combination of targeted and global proteomics was adopted to measure protein abundance of 6 ATP-binding cassette and 27 solute carrier transporters. Data analysis using proprietary Progenesis and open access MaxQuant software showed overall agreement; however, Abcb9 and Slc22a8 were quantified only by MaxQuant, whereas Abcc9 and Abcd3 were quantified only by Progenesis. Agreement between targeted and untargeted quantification was demonstrated for Abcb1 (19.7 ± 1.4 vs. 17.8 ± 2.3) and Abcc4 (2.2 ± 0.7 vs. 2.1 ± 0.4), respectively. Rigorous quantification of BBB proteins, as reported in this study, should assist with translational modeling efforts involving brain disposition of xenobiotics.
Topics: Animals; Biological Transport; Blood-Brain Barrier; Brain; Chromatography, Liquid; Glucose Transporter Type 1; In Vitro Techniques; Male; Mass Spectrometry; Microvessels; RNA, Messenger; Rats; Rats, Sprague-Dawley
PubMed: 29675872
DOI: 10.1111/jnc.14446 -
Xenobiotica; the Fate of Foreign... 20161. The umbilical cord is a direct conduit to the fetus hence transporters could have roles in partitioning substances between the maternal-placental-fetal units. Here we...
1. The umbilical cord is a direct conduit to the fetus hence transporters could have roles in partitioning substances between the maternal-placental-fetal units. Here we determined the expression and localization of the ATP-Binding Cassette (ABC) transporters BCRP (ABCG2), P-gp (ABCB1) and MRP1 (ABCC1) in human umbilical cords. 2. The mRNA for BCRP and MRP1 was detected in 25/25 samples, but P-gp was detected in only 5/25. ABC transporter mRNA expression relative to 18S was 25.6 ± 0.3, 26.5 ± 0.6 and 22.2 ± 0.2 cycles for BCRP, MRP1 and P-gp respectively. 3. Using a subset of 10 umbilical cords, BCRP protein was present in all samples (immunoblot) with positive correlation between mRNA and proteins (p = 0.07, r = 0.62) and between immunoblotting and immunohistochemistry (IHC) (p = 0.03, r = 0.67). P-gp protein was observed in 4/10 samples by both immunoblot and IHC, with no correlation between mRNA and protein (p = 0.45, r = 0.55) or immunoblotting and IHC (p = 0.2, r = 0.72), likely due to small sample size. MRP1 protein was not observed. 4. Localization of BCRP and P-gp proteins was to Wharton's jelly with no specific staining in arterial or venous endothelia. 5. Understanding ABC transporter expression in the umbilical cord may be useful for determining fetal exposures to xenobiotics if functional properties can be defined.
Topics: ATP Binding Cassette Transporter, Subfamily B, Member 1; ATP Binding Cassette Transporter, Subfamily G, Member 2; Cohort Studies; Demography; Female; Gene Expression Regulation; Humans; Immunohistochemistry; Multidrug Resistance-Associated Proteins; Neoplasm Proteins; Pregnancy; Umbilical Cord
PubMed: 26407213
DOI: 10.3109/00498254.2015.1091118 -
The Journal of Biological Chemistry Feb 2020The RNA helicase bad response to refrigeration 2 homolog (BRR2) is required for the activation of the spliceosome before the first catalytic step of RNA splicing. BRR2...
The RNA helicase bad response to refrigeration 2 homolog (BRR2) is required for the activation of the spliceosome before the first catalytic step of RNA splicing. BRR2 represents a distinct subgroup of Ski2-like nucleic acid helicases whose members comprise tandem helicase cassettes. Only the N-terminal cassette of BRR2 is an active ATPase and can unwind substrate RNAs. The C-terminal cassette represents a pseudoenzyme that can stimulate RNA-related activities of the N-terminal cassette. However, the molecular mechanisms by which the C-terminal cassette modulates the activities of the N-terminal unit remain elusive. Here, we show that N- and C-terminal cassettes adopt vastly different relative orientations in a crystal structure of BRR2 in complex with an activating domain of the spliceosomal Prp8 protein at 2.4 Å resolution compared with the crystal structure of BRR2 alone. Likewise, inspection of BRR2 structures within spliceosomal complexes revealed that the cassettes occupy different relative positions and engage in different intercassette contacts during different splicing stages. Engineered disulfide bridges that locked the cassettes in two different relative orientations had opposite effects on the RNA-unwinding activity of the N-terminal cassette, with one configuration enhancing and the other configuration inhibiting RNA unwinding compared with the unconstrained protein. Moreover, we found that differences in relative positioning of the cassettes strongly influence RNA-stimulated ATP hydrolysis by the N-terminal cassette. Our results indicate that the inactive C-terminal cassette of BRR2 can both positively and negatively affect the activity of the N-terminal helicase unit from a distance.
Topics: Adenosine Triphosphatases; Catalysis; Crystallography, X-Ray; Humans; Protein Conformation; RNA Splicing; RNA-Binding Proteins; Ribonucleoproteins, Small Nuclear; Spliceosomes; Substrate Specificity
PubMed: 31914407
DOI: 10.1074/jbc.RA119.010964 -
International Journal of Molecular... Sep 2017One of the main chemotherapeutic drugs used on a routine basis in patients with metastatic colorectal cancer ((m)CRC) is the topoisomerase-1 inhibitor, irinotecan.... (Review)
Review
BACKGROUND
One of the main chemotherapeutic drugs used on a routine basis in patients with metastatic colorectal cancer ((m)CRC) is the topoisomerase-1 inhibitor, irinotecan. However, its usefulness is limited by the pre-existing or inevitable development of resistance. The ATP-binding cassette (ABC) transporter ABCG2/breast cancer resistance protein (BRCP) through its function in xenobiotic clearance might play an important role in irinotecan resistance. With a goal to evaluate the clinical significance of ABCG2 measurements, we here review the current literature on ABCG2 in relation to irinotecan treatment in CRC patients.
RESULTS
Few studies have evaluated the association between gene or protein expression and prognosis in CRC patients. Discordant results were reported. The discrepancies might be explained by the use of different criteria for interpretation of results in the immunohistochemistry studies. Only one large study evaluated the ABCG2 protein expression and efficacy of irinotecan in mCRC (CAIRO study, = 566). This study failed to demonstrate any correlation between ABCG2 protein expression in the primary tumor and response to irinotecan-based treatment. We recently raised questions on how to evaluate ABCG2 immunoreactivity patterns, and the results in the CAIRO study might be influenced by using a different scoring protocol than the one proposed by us. In contrast, our recent exploratory study of mRNA expression in 580 patients with stage III primary CRC (subgroup from the randomized PETACC-3 study) indicated that high tumor tissue mRNA expression might be predictive for lack of efficacy of irinotecan.
CONCLUSION
The biological role of ABCG2 in predicting clinical irinotecan sensitivity/resistance in CRC is uncertain. In particular, the significance of ABCG2 cellular localization needs to be established. Data concerning mRNA expression and prediction of adjuvant irinotecan efficacy are still sparse and need to be confirmed.
Topics: ATP Binding Cassette Transporter, Subfamily G, Member 2; Animals; Camptothecin; Colorectal Neoplasms; Humans; Immunohistochemistry; Irinotecan
PubMed: 28880238
DOI: 10.3390/ijms18091926 -
Canadian Journal of Gastroenterology &... 2019ATP-binding cassette (ABC) transporters are the members of the efflux pumps that are responsible for the removal of cytotoxic substances by active transport. ABCB11, the...
Expression Analysis of ATP-Binding Cassette Transporters ABCB11 and ABCB4 in Primary Sclerosing Cholangitis and Variety of Pediatric and Adult Cholestatic and Noncholestatic Liver Diseases.
ATP-binding cassette (ABC) transporters are the members of the efflux pumps that are responsible for the removal of cytotoxic substances by active transport. ABCB11, the bile salt efflux pump of hepatocytes, coordinates cellular excretion of numerous conjugated bile salts into the bile canaliculi, whereas ABCB4 acts as an ATP-dependent floppase translocating phosphatidylcholine from the inner to the outer leaflet of the bile canalicular membrane. Loss of functional ABCB11 and ABCB4 proteins causes early-onset refractory cholestasis or cholangiopathy. In this study, we investigated the expression and localization pattern of ABCB11 and ABCB4 using immunohistochemistry and RNA profiling in liver samples from patients with different types and stages of chronic cholestatic liver disease, with emphasis on primary sclerosing cholangitis (PSC), compared to a variety of cholestatic and noncholestatic hepatopathies. Therefore, ABCB11 and ABCB4 expressions were investigated on formalin-fixed and paraffin-embedded (FFPE) material in a patient cohort of total 43 patients with or without cholestatic liver diseases, on protein level using immunohistochemistry and on RNA level using nanoString technology. Intriguingly, our results demonstrated increased expression of ABCB11 and ABCB4 on protein as well as RNA level in PSC, and the expression pattern correlated with disease progression. We concluded from our study that patients with PSC demonstrate altered expression levels and pattern of ABCB11 and ABCB4 which correlated with disease progression; thereby, ABCB11 and ABCB4 analysis may be a useful tool for assessment of disease stages in PSC.
Topics: ATP Binding Cassette Transporter, Subfamily B; ATP Binding Cassette Transporter, Subfamily B, Member 11; Adult; Bile Acids and Salts; Bile Canaliculi; Biological Transport; Case-Control Studies; Child; Cholangitis, Sclerosing; Cholestasis; Disease Progression; Female; Hepatocytes; Humans; Immunohistochemistry; Liver; Liver Diseases; Male; RNA
PubMed: 31886153
DOI: 10.1155/2019/1085717