A non-lethal malarial infection results in reduced drug metabolizing enzyme expression and drug clearance in mice.

BACKGROUND: Given the central importance of anti-malarial drugs in the treatment of malaria, there is a need to understand the effect of Plasmodium infection on the broad spectrum of drug metabolizing enzymes. Previous studies have shown reduced clearance of quinine, a treatment for Plasmodium infection, in individuals with malaria. METHODS: The hepatic expression of a large panel of drug metabolizing enzymes was studied in the livers of mice infected with the AS strain of Plasmodium chabaudi chabaudi, a nonlethal parasite in most strains of mice with several features that model human Plasmodium infections. C57BL/6J mice were infected with P. chabaudi by intraperitoneal injection of infected erythrocytes and sacrificed at different times after infection. Relative hepatic mRNA levels of various drug metabolizing enzymes, cytokines and acute phase proteins were measured by reverse transcriptase-real time PCR. Relative levels of cytochrome P450 proteins were measured by Western blotting with IR-dye labelled antibodies. Pharmacokinetics of 5 prototypic cytochrome P450 substrate drugs were measured by cassette dosing and high-resolution liquid chromatography-mass spectrometry. The results were analysed by MANOVA and post hoc univariate analysis of variance. RESULTS: The great majority of enzyme mRNAs were down-regulated, with the greatest effects occurring at the peak of parasitaemia 8 days post infection. Protein levels of cytochrome P450 enzymes in the Cyp 2b, 2c, 2d, 2e, 3a and 4a subfamilies were also down-regulated. Several distinct groups differing in their temporal patterns of regulation were identified. The cassette dosing study revealed that at the peak of parasitaemia, the clearances of caffeine, bupropion, tolbutamide and midazolam were markedly reduced by 60-70%. CONCLUSIONS: These findings in a model of uncomplicated human malaria suggest that changes in drug clearance in this condition may be of sufficient magnitude to cause significant alterations in exposure and response of anti-malarial drugs and co-medications.

Physiological Regulation of Drug Metabolism and Transport: Pregnancy, Microbiome, Inflammation, Infection, and Fasting.

This article is a report on a symposium entitled "Physiological Regulation of Drug Metabolism and Transport" sponsored by the American Society for Pharmacology and Experimental Therapeutics and held at the Experimental Biology 2017 meeting in Chicago, IL. The contributions of physiologic and pathophysiological regulation of drug-metabolizing enzymes and transporters to interindividual variability in drug metabolism are increasingly recognized but in many cases are not well understood. The presentations herein discuss the phenomenology, consequences, and mechanism of such regulation. CYP2D6 transgenic mice were used to provide insights into the mechanism of regulation of this enzyme in pregnancy, via hepatocyte nuclear factor 4α, small heterodimer partner, and retinoids. Regulation of intestinal and hepatic drug-processing enzymes by the intestinal microbiota via tryptophan and its metabolites was investigated. The potential impact of parasitic infections on human drug metabolism and clearance was assessed in mice infected with Schistosoma mansoni or Plasmodium chabaudi chabaudi AS, both of which produced widespread and profound effects on murine hepatic drug-metabolizing enzymes. Finally, the induction of Abcc drug efflux transporters by fasting was investigated. This was demonstrated to occur via a cAMP, protein kinase A/nuclear factor-E2-related factor 2/Sirtuin 1 pathway via antioxidant response elements on the Abcc genes.

EphB2 receptor tyrosine kinase promotes hepatic fibrogenesis in mice via activation of hepatic stellate cells.

Hepatic fibrosis is the result of an excessive wound-healing response subsequent to chronic liver injury. A feature of liver fibrogenesis is the secretion and deposition of extracellular matrix proteins by activated hepatic stellate cells (HSCs). Here we report that upregulation of EphB2 is a prominent feature of two mouse models of hepatic fibrosis and also observed in humans with liver cirrhosis. EphB2 is upregulated and activated in mouse HSCs following chronic carbon tetrachloride (CCl4) exposure. Moreover, we show that EphB2 deficiency attenuates liver fibrosis and inflammation and this is correlated with an overall reduction in pro-fibrotic markers, inflammatory chemokines and cytokines. In an in vitro system of HSCs activation we observed an impaired proliferation and sub-optimal differentiation into fibrogenic myofibroblasts of HSCs isolated from EphB2-/- mice compared to HSCs isolated from wild type mice. This supports the hypothesis that EphB2 promotes liver fibrosis partly via activation of HSCs. Cellular apoptosis which is generally observed during the regression of liver fibrogenesis was increased in liver specimens of CCl4-treated EphB2-/- mice compared to littermate controls. This data is suggestive of an active repair/regeneration system in the absence of EphB2. Altogether, our data validate this novel pro-fibrotic function of EphB2 receptor tyrosine kinase.

Magnetic implants in the tongue for assistive technologies: Tests of migration; oromotor function; and tissue response in miniature pigs.

OBJECTIVE: Uncertain biological consequences of titanium-magnet (Ti-mag) tongue implants constrain application of the Tongue Drive System (TDS), a brain-tongue-computer interface for individuals with severe physical impairment. Here we describe oromotor function and tongue tissue response following Ti-Mag implantation and explantation in the miniature pig, an animal model with a tongue similar in size to humans. DESIGN: A 1.8×6.2mm Ti-mag tracer was implanted into the anterior tongue in five Yucatan minipigs. X-rays were taken immediately and >six days after implantation to evaluate tracer migration. In three minipigs, the tracer was explanted >16days after implantation. Twenty-five days post-explantation, tongue tissue was harvested and processed for histological and immunohistochemical (IHC) markers of healing. In two minipigs tissue markers of healing were evaluated post-mortem following >12days implantation. Drink cycle rate (DCR) was characterized to determine the impact of procedures on oromotor function. RESULTS: Neither implantation (N=5) nor explantation (N=3) changed DCR. X-rays revealed minimal tracer migration (N=4, 0-4mm). By histology and IHC a robust capsule was present two weeks post-implantation with limited fibrosis. Explantation produced localized fibrosis and limited muscle remodeling. CONCLUSIONS: These findings suggest the safety of Ti-mag anterior tongue implants for assistive technologies in humans.

The receptor tyrosine kinase EphB2 promotes hepatic fibrosis in mice.

UNLABELLED: Beyond the well-defined role of the Eph (erythropoietin-producing hepatocellular) receptor tyrosine kinases in developmental processes, cell motility, cell trafficking/adhesion, and cancer, nothing is known about their involvement in liver pathologies. During blood-stage rodent malaria infection we have found that EphB2 transcripts and proteins were up-regulated in the liver, a result likely driven by elevated surface expression on immune cells including macrophages. This was significant for malaria pathogenesis because EphB2(-/-) mice were protected from malaria-induced liver fibrosis despite having a similar liver parasite burden compared with littermate control mice. This protection was correlated with a defect in the inflammatory potential of hepatocytes from EphB2(-/-) mice resulting in a reduction in adhesion molecules, chemokine/chemokine receptor RNA levels, and infiltration of leukocytes including macrophages/Kupffer cells, which mediate liver fibrosis during rodent malaria infections. These observations are recapitulated in the well-established carbon tetrachloride model of liver fibrosis in which EphB2(-/-) carbon tetrachloride-treated mice showed a significant reduction of liver fibrosis compared to carbon tetrachloride-treated littermate mice. Depletion of macrophages by clodronate-liposomes abrogates liver EphB2 messenger RNA and protein up-regulation and fibrosis in malaria-infected mice. CONCLUSION: During rodent malaria, EphB2 expression promotes malaria-associated liver fibrosis; to our knowledge, our data are the first to implicate the EphB family of receptor tyrosine kinases in liver fibrosis or in the pathogenesis of malaria infection.

Hepatic cytochrome P450s, phase II enzymes and nuclear receptors are downregulated in a Th2 environment during Schistosoma mansoni infection.

Inflammation and infection downregulate the activity and expression of cytochrome P450s (P450s) and other drug metabolizing enzymes (DMEs) involved in hepatic drug clearance. Schistosoma mansoni infection was reported to cause a downregulation of hepatic P450-dependent activities in mouse liver, but little is known about the specific enzymes affected or whether phase II DMEs are also affected. Here we describe the effect of murine schistosomiasis on the expression of hepatic P450s, NADPH-cytochrome P450 reductase (Cpr), phase II drug metabolizing enzymes, and nuclear receptors at 30 and 45 days postinfection (dpi). Although the hepatic expression of some of these genes was altered at 30 dpi, we observed substantial changes in the expression of the majority of P450 mRNAs and proteins measured, Cpr protein, as well as many of the UDP-glucuronosyltransferases and sulfotransferases at 45 dpi. S. mansoni infection also altered nuclear receptor expression, inducing mRNA levels at 30 dpi and depressing levels at 45 dpi. S. mansoni evoked a T helper 2 (Th2) inflammatory response at 45 dpi, as indicated by the induction of hepatic Th2 cytokine mRNAs [interleukins 4, 5, and 13], whereas the hepatic proinflammatory response was relatively weak. Thus, chronic schistosomiasis markedly and selectively alters the expression of multiple DMEs, which may be associated with Th2 cytokine release. This would represent a novel mechanism of DME regulation in disease states. These findings have important implications for drug testing in infected mice, whereas the relevance to humans with schistosomiasis needs to be determined.