ABSTRACT
GroEL, a chaperone protein responsible for peptide and denatured protein folding, undergoes substantial conformational changes driven by ATP binding and hydrolysis during folding. Utilizing these conformational changes, we demonstrated the GroEL-mediated regioselective photocyclodimerization of 2-anthracenecarboxylic acid (AC) using ATP hydrolysis as an external stimulus. We designed and prepared an optimal GroEL mutant to employ in a docking simulation that has been actively used in recent years. Based on the large difference in the motif of hydrogen bonds between AC and GroEL mutant compared with the wild-type, we predicted that GroELMEL, in which the 307â309th amino acid residues were mutated to Ala, could alter the orientation of bound AC in GroEL. The GroELMEL-mediated photocyclodimerization of AC can be used for regioselective inversion upon ATP addition to a moderate extent.
ABSTRACT
AIMS: Paraoxonase 1 (PON1) binds to high-density lipoprotein (HDL) and protects against atherosclerosis. However, the relationship between functional PON1 Q192R polymorphism, which is associated with the hydrolysis of paraoxon (POXase activity) and atherosclerotic cardiovascular disease (ASCVD), remains controversial. As the effect of PON1 Q192R polymorphism on the HDL function is unclear, we investigated the relationship between this polymorphism and the cholesterol efflux capacity (CEC), one of the biological functions of HDL, in association with the PON1 activity. METHODS: The relationship between PON1 Q192R polymorphisms and CEC was investigated retrospectively in 150 subjects without ASCVD (50 with the PON1 Q/Q genotype, 50 with the Q/R genotype, and 50 with the R/R genotype) who participated in a health screening program. The POXase and arylesterase (AREase: hydrolysis of aromatic esters) activities were used as measures of the PON1 activity. RESULTS: The AREase activity was positively correlated with CEC independent of the HDL cholesterol levels. When stratified by the PON1 Q192R genotype, the POXase activity was also positively correlated with CEC independent of HDL cholesterol. PON1 Q192R R/R genotype carriers had a lower CEC than Q/Q or Q/R genotype carriers, despite having a higher POXase activity. Moreover, in a multiple regression analysis, the PON1 Q192R genotype was associated with the degree of CEC, independent of the HDL cholesterol and POXase activity. CONCLUSIONS: The PON1 Q192R R allele is associated with reduced CEC in Japanese people without ASCVD. Further studies on the impact of this association on the severity of atherosclerosis and ASCVD development are thus called for.
ABSTRACT
It is clinically possible for patients with Alport syndrome (AS) to suffer from poststreptococcal acute glomerulonephritis (PSAGN). However, there is only one report of such a patient, and he had end-stage kidney disease. Here, we describe an 8-year-old male with X-linked AS and chronic kidney disease (CKD) stage G2. He presented with diffuse edema, gross hematuria, proteinuria, and body weight gain after streptococcal pharyngitis. Blood examination showed kidney dysfunction, hypocomplementemia, and increased anti-streptolysin-O levels. His kidney function did not improve with symptomatic treatment. Therefore, we started steroid administration on the 12th day of hospitalization. Then, his kidney function improved before he was discharged. We confirmed that his complement function had recovered at a later date. Pathological evaluation showed findings of AS and PSAGN, including cellular crescents in 3/30 glomeruli on light microscopy. In addition, electron dense deposits (EDDs) were seen in not only the visceral subepithelium but also the glomerular basement intramembrane and subendothelium, some of which were hump-like. Although AS and CKD are indicated to have a poor prognosis in PSAGN, our patient recovered after administration of steroids. Our case suggests that we can consider the administration of steroids, including pulse therapy for PSAGN, when patients have, for example, crescents on pathology, severe renal dysfunction, nephrotic proteinuria, or AS with CKD, as in our case. Kidney pathology suggested that a typical hump is not seen in patients with cooccurring AS and PSAGN. After the patient's kidney function recovered, we continued to follow him.
Subject(s)
Glomerulonephritis , Nephritis, Hereditary , Renal Insufficiency, Chronic , Streptococcal Infections , Male , Humans , Child , Glomerulonephritis/pathology , Acute Disease , ProteinuriaABSTRACT
Endonuclease V is highly conserved, both structurally and functionally, from bacteria to humans, and it cleaves the deoxyinosine-containing double-stranded DNA in Escherichia coli, whereas in Homo sapiens it catalyses the inosine-containing single-stranded RNA. Thus, deoxyinosine and inosine are unexpectedly produced by the deamination reactions of adenine in DNA and RNA, respectively. Moreover, adenosine-to-inosine (A-to-I) RNA editing is carried out by adenosine deaminase acting on dsRNA (ADARs). We focused on Arabidopsis thaliana endonuclease V (AtEndoV) activity exhibiting variations in DNA or RNA substrate specificities. Since no ADAR was observed for A-to-I editing in A. thaliana, the possibility of inosine generation by A-to-I editing can be ruled out. Purified AtEndoV protein cleaved the second and third phosphodiester bonds, 3' to inosine in single-strand RNA, at a low reaction temperature of 20-25°C, whereas the AtEndoV (Y100A) protein bearing a mutation in substrate recognition sites did not cleave these bonds. Furthermore, AtEndoV, similar to human EndoV, prefers RNA substrates over DNA substrates, and it could not cleave the inosine-containing double-stranded RNA. Thus, we propose the possibility that AtEndoV functions as an RNA substrate containing inosine induced by RNA damage, and not by A-to-I RNA editing in vivo.
Subject(s)
Arabidopsis/metabolism , Deoxyribonuclease (Pyrimidine Dimer)/metabolism , Inosine/chemistry , RNA, Plant/genetics , Arabidopsis/genetics , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Binding Sites , Deoxyribonuclease (Pyrimidine Dimer)/genetics , Gene Expression Regulation, Plant , RNA Editing , RNA, Plant/chemistry , Substrate SpecificityABSTRACT
BACKGROUND: Dermatopathy develops as a side effect in patients receiving anti-epidermal growth factor receptor antibody treatment. Topical moisturizers are used for the prevention and treatment of this dermatopathy. Active participation of patients in their own treatment is important for the appropriate application of topical preparations. We prepared a pharmaceutical instructional video for adhering to the topical application protocol. In this study, we investigated the effectiveness of this pharmaceutical instructional video on treatment adherence. METHODS: Study participants were patients with cancer receiving the anti-epidermal growth factor receptor antibody for the first time. A pharmacist instructed the patients on how to use the pharmaceutical instruction video. Daily topical preparation use following the video demonstration was assessed. The effectiveness of the pharmaceutical instruction video was evaluated by assessing the adherence of patients who did not use the pharmaceutical instruction video for the past 2 periods (26 months; controls 1 and 2). The incidence of side effects was compared between the two control groups and the group of patients who received the pharmaceutical instruction video. RESULTS: The amount of topical preparation consumed (median, g/day) by patients who received patient compliance instructions using the pharmaceutical instruction video was 9.8 g/day, as compared with control group 1 (4.5 g/day) and control group 2 (5.5 g/day) (p < 0.001). There was no difference in the incidence of side effects during the three periods. CONCLUSION: The use of visual instructional media for patient compliance by pharmacists may be effective in maintaining and improving treatment adherence.
Subject(s)
Antineoplastic Agents/adverse effects , ErbB Receptors/antagonists & inhibitors , Medication Adherence , Neoplasms/drug therapy , Patient Education as Topic , Skin Diseases/drug therapy , Aged , Female , Humans , Male , Middle Aged , Video RecordingABSTRACT
BACKGROUND: Exposure to inhalation of anticancer drugs is frequent in anticancer drug handling. Using an activated carbon mask with the ability to remove particulates and vapors of anticancer drugs may be effective. Mask fitting performance is important, because low fitting performance leads to inhalation via bypassing the mask filter (leak). This study evaluated the leak rate of multiple-shaped masks. METHODS: Activated carbon and nonactivated carbon masks of the pleated-type (like surgical mask) and cup-type were used. Four pharmacists wore the masks and a fitting tester was employed. The particle reduction rate of particles in ambient air was calculated using: particle count (outside - inside)/outside × 100 (%). Leak rate was calculated as the difference in the particle reduction rate due to the presence or the absence of a seal in the mask surroundings. RESULTS: Reduction rates of the pleated-type nonactivated carbon mask and the pleated-type activated carbon mask were 14.8% and 34.8% (mean). These values significantly increased to 85.6% and 83.3% upon sealing the mask surroundings. Particle reduction rates of the cup-type nonactivated carbon mask and activated carbon mask were 99.3% and 33.6%. When mask surroundings were sealed, these values were 99.6% and 39.2%. Leak rates of pleated-type nonactivated carbon mask, pleated-type activated carbon mask, cup-type nonactivated carbon mask, and cup-type activated carbon mask were 70.8%, 48.5%, 0.3%, and 5.6%, respectively. CONCLUSION: A difference was found in the leak rate between masks used in anticancer drug handling. Based on the low leak rate, the cup-type activated carbon mask was thought to be effective.
Subject(s)
Antineoplastic Agents/adverse effects , Carbon , Equipment Design/standards , Masks/standards , Particle Size , Equipment Design/methods , Humans , Inhalation/physiologyABSTRACT
Human antithrombin (AT) has two isoforms of which the predominant α-form is glycosylated on all four possible glycosylation sites and the lower abundant ß-isoform lacks the oligosaccharide on Asn135. The main oligosaccharide structure of human AT consists of biantennary complex-type oligosaccharides lacking a core fucose. Generally, Chinese hamster ovary (CHO) cells produce recombinant human AT (rhAT) with core-fucosylated oligosaccharides. However, rhAT lacking core-fucose oligosaccharides can be produced by POTELLIGENT® technology, which uses FUT8 knockout CHO cells in production. The rhAT has more variable glycan structures, such as tetra-antennary complex type, high-mannose type, and mannose 6-phosphate species as minor components compared to plasma-derived human AT (phAT). In addition, the site-specific glycan profile was different between two ATs. We evaluated the effect of these properties on efficacy and safety based on a comparison of rhAT made by that technology with phAT in terms of their respective oligosaccharide structures, site-specific oligosaccharide profiles, and the ratio of α- and ß-forms. Although some structural differences were found between the rhAT and phAT, we concluded that these differences have no significant effect on the efficacy and safety of rhAT.
Subject(s)
Antithrombins/chemistry , Antithrombins/metabolism , Genetic Engineering/methods , Oligosaccharides/chemistry , Plasma/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Fucosyltransferases/deficiency , Fucosyltransferases/genetics , Gene Knockout Techniques , Glycosylation , Humans , Recombinant Proteins/geneticsABSTRACT
The marine environment around Japan experienced significant changes during the Cenozoic Era. In this study, we report findings suggesting that this dynamic history left behind traces in the genome of the Japanese sand dollar species Peronella japonica and P. rubra. Although mitochondrial Cytochrome C Oxidase I sequences did not indicate fragmentation of the current local populations of P. japonica around Japan, two different types of intron sequence were found in the Alx1 locus. We inferred that past fragmentation of the populations account for the presence of two types of nuclear sequences as alleles in the Alx1 intron of P. japonica. It is likely that the split populations have intermixed in recent times; hence, we did not detect polymorphisms in the sequences reflecting the current localization of the species. In addition, we found two allelic sequences of theAlx1 intron in the sister species P. rubra. The divergence times of the two types of Alx1 intron sequences were estimated at approximately 14.9 and 4.0â¯million years ago for P. japonica and P. rubra, respectively. Our study indicates that information from the intron sequences of nuclear genes can enhance our understanding of past genetic events in organisms.
Subject(s)
Cell Nucleus/genetics , Sea Urchins/genetics , Sequence Analysis, DNA/methods , Animals , Evolution, Molecular , Introns , Japan , PhylogenyABSTRACT
Aromatase inhibitors (AIs) effectively treat hormone receptor-positive postmenopausal breast cancer, but some patients do not respond to treatment or experience recurrence. Mechanisms of AI resistance include ligand-independent activation of the estrogen receptor (ER) and signaling via other growth factor receptors; however, these do not account for all forms of resistance. Here we present an alternative mechanism of AI resistance. We ectopically expressed aromatase in MCF-7 cells expressing green fluorescent protein as an index of ER activity. Aromatase-overexpressing MCF-7 cells were cultured in estrogen-depleted medium supplemented with testosterone and the AI, letrozole, to establish letrozole-resistant (LR) cell lines. Compared with parental cells, LR cells had higher mRNA levels of steroid sulfatase (STS), which converts estrone sulfate (E1S) to estrone, and the organic anion transporter peptides (OATPs), which mediate the uptake of E1S into cells. LR cells proliferated more in E1S-supplemented medium than did parental cells, and LR proliferation was effectively inhibited by an STS inhibitor in combination with letrozole and by ER-targeting drugs. Analysis of ER-positive primary breast cancer tissues showed a significant correlation between the increases in the mRNA levels of STS and the OATPs in the LR cell lines, which supports the validity of this AI-resistant model. This is the first study to demonstrate the contribution of STS and OATPs in E1S metabolism to the proliferation of AI-resistant breast cancer cells. We suggest that E1S metabolism represents a new target in AI-resistant breast cancer treatment.
Subject(s)
Aromatase Inhibitors/pharmacology , Breast Neoplasms/pathology , Cell Proliferation/drug effects , Drug Resistance, Neoplasm/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Receptors, Estrogen/metabolism , Breast Neoplasms/drug therapy , Breast Neoplasms/metabolism , Female , Humans , Tumor Cells, CulturedABSTRACT
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with no cure. To develop effective treatments for this devastating disease, an appropriate strategy for targeting the molecule responsible for the pathogenesis of ALS is needed. We previously reported that mutant SOD1 protein causes motor neuron death through activation of ASK1, a mitogen-activated protein kinase kinase kinase. Additionally, we recently developed K811 and K812, which are selective inhibitors for ASK1. Here, we report the effect of K811 and K812 in a mouse model of ALS (SOD1(G93A) transgenic mice). Oral administration of K811 or K812 significantly extended the life span of SOD1(G93A) transgenic mice (1.06 and 1.08% improvement in survival). Moreover, ASK1 activation observed in the lumbar spinal cord of mice at the disease progression stage was markedly decreased in the K811- and K812-treated groups. In parallel, immunohistochemical analysis revealed that K811 and K812 treatment inhibited glial activation in the lumbar spinal cord of SOD1(G93A) transgenic mice. These results reinforce the importance of ASK1 as a therapeutic target for ALS treatment.
Subject(s)
Amyotrophic Lateral Sclerosis/drug therapy , Antineoplastic Agents/therapeutic use , Heterocyclic Compounds, 4 or More Rings/therapeutic use , MAP Kinase Kinase Kinase 5/antagonists & inhibitors , Administration, Oral , Animals , Antineoplastic Agents/administration & dosage , Disease Models, Animal , Heterocyclic Compounds, 4 or More Rings/administration & dosage , Male , Mice , Mice, Transgenic , Treatment OutcomeABSTRACT
Planococcus maritimus strain iso-3 was isolated from an intertidal sediment sample from the Clyde estuary in the UK. A novel red pigment glyco-carotenoic acid ester, methyl glucosyl-3,4-dehydro-apo-8'-lycopenoate has been isolated from this marine bacterium using chromatographic methods. The structure of methyl glucosyl-3,4-dehydro-apo-8'-lycopenoate was determined to be methyl 1-(beta-D-glucopyranosyloxy)-3,4-didehydro-1,2-dihydro-8'-apo-psi-caroten-8'-oate by the degradation experiment and the spectroscopic analyses. The methyl glucosyl-3,4-dehydro-apo-8'-lycopenoate showed potent antioxidative activity in the (1)O(2) suppression model.
Subject(s)
Antioxidants/isolation & purification , Antioxidants/pharmacology , Carotenoids/isolation & purification , Carotenoids/pharmacology , Geologic Sediments/microbiology , Gram-Positive Bacteria/metabolism , Antioxidants/chemistry , Carotenoids/chemistry , Gram-Positive Bacteria/chemistry , Gram-Positive Bacteria/classification , Gram-Positive Bacteria/isolation & purification , Inhibitory Concentration 50 , Phylogeny , Pigments, Biological/chemistry , Pigments, Biological/isolation & purification , Pigments, Biological/pharmacology , RNA, Bacterial/genetics , RNA, Ribosomal, 16S/genetics , Reactive Oxygen Species/antagonists & inhibitors , Spectrum Analysis , United KingdomABSTRACT
We report the structure-activity relationship of quinoline and quinazoline derivatives, which include urea, thiourea, urethane, and acylthiourea groups, as inhibitors of the platelet-derived growth factor (PDGF) receptor autophosphorylation. Our previous studies showed that the quinoline and quinazoline derivatives including urea, thiourea, and carbamate groups were highly potent compounds as the PDGF receptor autophosphorylation inhibitor, but these compounds did not exhibit receptor selectivity between the PDGF receptor and the c-kit receptor. As a result of further synthesis and biological evaluation, we have found that the quinoline and quinazoline-acylthiourea derivatives showed not only good inhibitory activity for the PDGF receptor but also receptor selectivity between the PDGF receptor and the c-kit receptor. Furthermore N-{4-[(6,7-dimethoxy-4-quinolyl)oxy]phenyl}-N'-(2-methylbenzoyl)thiourea exhibited potent oral efficacy in in vivo assay using the rat carotid balloon injury model. Therefore, the quinoline and quinazoline-acylthiourea derivatives may be expected to have potential as therapeutic agents for the treatment of restenosis.
Subject(s)
Quinazolines/chemical synthesis , Quinolines/chemical synthesis , Receptors, Platelet-Derived Growth Factor/metabolism , Thiourea/analogs & derivatives , Thiourea/chemical synthesis , Administration, Oral , Animals , Carotid Stenosis/prevention & control , Cell Line , Male , Phosphorylation , Proto-Oncogene Proteins c-kit/metabolism , Quinazolines/pharmacology , Quinolines/pharmacology , Rats , Rats, Wistar , Receptors, Platelet-Derived Growth Factor/antagonists & inhibitors , Structure-Activity Relationship , Thiourea/pharmacologyABSTRACT
The expression of CYP2C12 by GH occurs in female but not in male rat livers. Direct injection of the CYP2C12 promoter-luciferase gene into male rat livers showed that the CYP2C12 promoter was active in both male and female rats. Thus, to further examine one or more factors that regulate the gender-related expression of CYP2C12, male rats were treated with trichostatin A, a specific inhibitor of histone deacetylase capable of condensing the chromatin structure. Interestingly, the expression of CYP2C12 by GH was seen even in the livers of male rats, indicating that histone deacetylase contributes to the suppression of CYP2C12 expression in male rats. Deoxyribonuclease I hypersensitive assay using nuclei from the livers of male or female rats revealed that the chromatin structure of the CYP2C12 gene was gender specific: a hypersensitive site at a position -4.2 kb containing GH-responsive element that bound to signal transducer and activator of transcription 5 (STAT5), termed as HS (hypersensitive site) 1, was specific for female rat livers, whereas a hypersensitive site at a position -3 kb, designated as HSm (male-specific hypersensitive site), was characteristic of male rat livers. A -3425/-3275 region within HSm functioned as a negative regulatory region, when the region was inserted in front of simian virus 40 promoter. Gel shift assay demonstrated that both CCAAT/enhancer-binding protein alpha and beta bound to the -3425/-3275 region. Based on these results, we conclude that the gender-related expression of the CYP2C12 gene results from the inaccessibility of to STAT5 to the GH-responsive element by chromatin condensation seen in male rat livers, and from the presence of the male-specific HSm that acts as a silencer.