Polycyclic Aromatic Hydrocarbons Activate the Aryl Hydrocarbon Receptor and the Constitutive Androstane Receptor to Regulate Xenobiotic Metabolism in Human Liver Cells

Polycyclic aromatic hydrocarbons (PAHs) are environmental pollutants produced by incomplete combustion of organic matter. They induce their own metabolism by upregulating xenobiotic-metabolizing enzymes such as cytochrome P450 monooxygenase 1A1 (CYP1A1) by activating the aryl hydrocarbon receptor (AHR). However, previous studies showed that individual PAHs may also interact with the constitutive androstane receptor (CAR). Here, we studied ten PAHs, different in carcinogenicity classification, for their potential to activate AHR- and CAR-dependent luciferase reporter genes in human liver cells. The majority of investigated PAHs activated AHR, while non-carcinogenic PAHs tended to activate CAR. We further characterized gene expression, protein abundancies and activities of the AHR targets CYP1A1 and 1A2, and the CAR target CYP2B6 in human HepaRG hepatoma cells. Enzyme induction patterns strongly resembled the profiles obtained at the receptor level, with AHR-activating PAHs inducing CYP1A1/1A2 and CAR-activating PAHs inducing CYP2B6. In summary, this study provides evidence that beside well-known activation of AHR, some PAHs also activate CAR, followed by subsequent expression of respective target genes. Furthermore, we found that an increased PAH ring number is associated with AHR activation as well as the induction of DNA double-strand breaks, whereas smaller PAHs activated CAR but showed no DNA-damaging potential.     

Functional and Molecular Properties of DYT-SGCE Myoclonus-Dystonia Patient-Derived Striatal Medium Spiny Neurons

Myoclonus-dystonia (DYT-SGCE, formerly DYT11) is characterized by alcohol-sensitive, myoclonic-like appearance of fast dystonic movements. It is caused by mutations in the SGCE gene encoding ε-sarcoglycan leading to a dysfunction of this transmembrane protein, alterations in the cerebello-thalamic pathway and impaired striatal plasticity. To elucidate underlying pathogenic mechanisms, we investigated induced pluripotent stem cell (iPSC)-derived striatal medium spiny neurons (MSNs) from two myoclonus-dystonia patients carrying a heterozygous mutation in the SGCE gene (c.298T>G and c.304C>T with protein changes W100G and R102X) in comparison to two matched healthy control lines. Calcium imaging showed significantly elevated basal intracellular Ca²⁺ content and lower frequency of spontaneous Ca²⁺ signals in SGCE MSNs. Blocking of voltage-gated Ca²⁺ channels by verapamil was less efficient in suppressing KCl-induced Ca²⁺ peaks of SGCE MSNs. Ca²⁺ amplitudes upon glycine and acetylcholine applications were increased in SGCE MSNs, but not after GABA or glutamate applications. Expression of voltage-gated Ca²⁺ channels and most ionotropic receptor subunits was not altered. SGCE MSNs showed significantly reduced GABAergic synaptic density. Whole-cell patch-clamp recordings displayed elevated amplitudes of miniature postsynaptic currents and action potentials in SGCE MSNs. Our data contribute to a better understanding of the pathophysiology and the development of novel therapeutic strategies for myoclonus-dystonia.     

Characterization of Apo-Form Selective Inhibition of Indoleamine 2,3-Dioxygenase**

Indoleamine-2,3-dioxygenase 1 (IDO1) is a heme-containing enzyme that catalyzes the rate-limiting step in the kynurenine pathway of tryptophan (TRP) metabolism. As it is an inflammation-induced immunoregulatory enzyme, pharmacological inhibition of IDO1 activity is currently being pursued as a potential therapeutic tool for the treatment of cancer and other disease states. As such, a detailed understanding of the mechanism of action of IDO1 inhibitors with various mechanisms of inhibition is of great interest. Comparison of an apo-form-binding IDO1 inhibitor (GSK5628) to the heme-coordinating compound, epacadostat (Incyte), allows us to explore the details of the apo-binding inhibition of IDO1. Herein, we demonstrate that GSK5628 inhibits IDO1 by competing with heme for binding to a heme-free conformation of the enzyme (apo-IDO1), whereas epacadostat coordinates its binding with the iron atom of the IDO1 heme cofactor. Comparison of these two compounds in cellular systems reveals a long-lasting inhibitory effect of GSK5628, previously undescribed for other known IDO1 inhibitors. Detailed characterization of this apo-binding mechanism for IDO1 inhibition might help design superior inhibitors or could confer a unique competitive advantage over other IDO1 inhibitors vis-à-vis specificity and pharmacokinetic parameters.     

Pheromonal secretions from glands on the 5th abdominal sternite of hydropsychid and rhyacophilid caddisflies (Trichoptera)

Extracts of different body parts of adult Trichoptera were tested for electrophysiological activity. Extracts of the IVth and Vth abdominal sternites of femaleHydropsyche angustipennis, Rhyacophila nubila, andR. fasciata, containing a paired exocrine gland, elicited significant electroan-tennographic responses when tested on conspecific male antennae. The paired gland occurs also in males of all the species, and inH. angustipennis, extracts from males were more active than female extracts when tested on male antennae. Female and male extracts from all species were analyzed by gas chromatography with simultaneous flame ionization and electroantennographic detection (EAD). EAD-active peaks in female extracts, stimulating male antennae, were identified inH. angustipennis as nonan-2-one; and inR. nubila andR. fasciata as heptan-2-one, heptan-2-ol, nonan-2-one, and nonan-2-ol. EAD-active components from maleH. angustipennis stimulating male antennae were octan-2-one, nonan-2-one (major peak), (Z)-6-nonen-2-one, decan-2-one, and a methylbranched decan-2-one. Female extracts and synthetic mixtures of compounds identified from femaleH. angustipennis andR. fasciata were tested for attractivity in the field. High catches with control traps obscured the results, but a synthetic mixture of the four identified compounds was significantly attractive and not different from female extracts for attracting maleR. fasciata. InH. angustipennis, a synthetic six-component male blend, in which nonan-2-one was the major component, attracted significant numbers of male and femaleH. angustipennis. Extracts of maleR. nubila andR. fasciata contained acetophenone and hexanoic and octanoic acids but did not have any electrophysiological or behavioral activity on either male or female antennae of conspecifics. The occurrence of a female sex pheromone inRhyacophila and an aggregation pheromone inHydropsyche corresponds to earlier described differences in mating behaviors in the Rhyacophilidae and Hydropsychidae.     

A 3D analysis of the joint torques developed during driver's ingress–egress motion

Providing an easy ingress–egress (I/E) movement remains a challenge for car designers. I/E has been largely studied in kinematics, but not in dynamics. This study proposes: (1) to evaluate and describe the motor torques developed in the lower limbs and lumbar joints during I/E motions and (2) to analyse the influence of the car geometry and subject anthropometry. An experiment was performed to observe 15 subjects of three anthropometrical groups getting in and out of a car mock-up simulating three different vehicle configurations. Motor torques were extracted using an inverse dynamics analysis. Both ingress and egress motions were primarily characterised by large torques. Overall, the taller a subject and the lower the seat of the vehicle were, the larger the peak torques were. Moreover, peak torques were higher for egress than ingress. These results are discussed in regard to the current knowledge on I/E ergonomics.     

Sorption of Aniline by Montmorillonite

Sorption of aniline by montmorillonite was studied by infra-red, X-ray diffraction and differential thermal analysis methods. The amount of aniline sorbed and the type of bonding depend upon the interlayer cations: anilinium, H⁺- and Al³⁺- give anilinium aniline ions, NH₄⁴-ammonium aniline ions, alkalis and alkaline earths except Cs are bonded to aniline through water bridges and transition metal cations are coordinated to aniline partly directly and partly through water bridges. Sorption does not occur in the complete absence of water.     

Free fatty acid inhibition of the insulin induction of glucose-6-phosphate dehydrogenase in rat hepatocyte monolayers

Rat hepatocytes in monolayer culture were utilized to determine if the decrease in glucose-6-phosphate dehydrogenase (G6PD) activity resulting from the ingestion of fat can be mimicked by the addition of fatty acids to a chemically, hormonally defined medium. G6PD activity in cultured hepatocytes was induced several-fold by insulin. Dexamethasone or T3 did not amplify the insulin induction of G6PD. Glucose alone increased G6PD activity in cultured hepatocytes from fasted donors by nearly 500%. Insulin in combination with glucose induced G6PD an additional two-fold. The increase in G6PD activity caused by glucose was greater in hepatocytes isolated from 72 hr-fasted rats as compared to fed donor rats. Such a response was reminiscent of the “overshoot” phenomenon in which G6PD activity is induced well above the normal level by fasting-refeeding rats a high glucose diet. Addition of linoleate to the medium resulted in a significant suppression of insulin’s ability to induce G6PD, but linoleate had no effect on the induction of G6PD activity by glucose alone. A shift to the right in the insulin-response curve for the induction of G6PD also was detected for the induction of malic enzyme and acetyl-CoA carboxylase. Arachidonate (0.25 mM) was a significantly more effective inhibitor of the insulin action than linoleate was. Apparently rat hepatocytes in monolayer culture can be utilized as a model to investigate the molecular mechanism by which fatty acids inhibit the production of lipogenic enzymes. In part, this mechanism of fatty acid inhibition involves desensitization of hepatocytes to the lipogenic action of insulin.