Chronic kidney disease (CKD) results in the accumulation of metabolic waste products that are normally cleared by the kidney, known as uremia. Many of these waste products are from bacteria metabolites in the gut. Accumulation of uremic toxins in plasma and tissue, as well as the gut-plasma-tissue metabolic axis are important for understanding pathophysiological mechanisms of comorbidities in CKD. In this study, an untargeted metabolomics approach was used to determine uremic toxin accumulation in plasma, liver, heart and kidney tissue in rats with adenine-induced CKD. Rats with CKD were also given AST-120, a spherical carbon adsorbent, to assess metabolic changes in plasma and tissues with the removal of gut-derived uremic toxins. AST-120 decreased >55% of metabolites that were increased in plasma, liver and heart tissue of rats with CKD. CKD was primarily defined by 8 gut-derived uremic toxins, which were significantly increased in plasma and all tissues. These metabolites were derived from aromatic amino acids and soy protein including: indoxyl sulfate, p-cresyl sulfate, hippuric acid, phenyl sulfate, pyrocatechol sulfate, 4-ethylphenyl sulfate, p-cresol glucuronide and equol 7-glucuronide. Our results highlight the importance of diet and gut-derived metabolites in the accumulation of uremic toxins and define the gut-plasma-tissue metabolic axis in CKD.

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Patients with chronic kidney disease (CKD) require many medications. CYP2C and CYP3A drug-metabolizing enzymes play a critical role in determining the pharmacokinetics of the majority of prescribed medications. These enzymes are transcriptionally regulated by the nuclear receptors pregnane X receptor (PXR) and hepatic nuclear factor 4␣ (HNF-4␣). Expression of CYP2C and CYP3A is decreased in CKD; however, the mechanisms by which this occurs is unknown. We induced CKD in rats by 5/6 nephrectomy and used chromatin immunoprecipitation (ChIP) to determine nuclear receptor-and epigenetic alterationmediated differences in the promoter region of the CYP2C and CYP3A genes. RNA polymerase II and HNF-4␣ binding was decreased 76 and 57% in the CYP2C11 promotor and 71 and 77% in the CYP3A2 promoter, respectively (P<0.05). ChIP also revealed a 57% decrease in PXR binding to the CYP3A2 promoter in CKD rats (P<0.05). The decrease in PXR and HNF-4␣ binding was accompanied by diminished histone 4 acetylation in the CYP3A2 promoter (48%) and histone 3 acetylation in the CYP2C11 (77%) and CYP3A2 (77%) promoter loci for nuclear receptor activation (P<0.05). This study suggests that decreased nuclear receptor binding and histone acetylation may contribute to the mechanism of drug-metabolizing enzyme down-regulation and altered pharmacokinetics in CKD. Patients with chronic kidney disease (CKD) take many medications to manage their CKD and comorbidities (1). Subtherapeutic dosing and overdosing result in adverse drug events commonly associated with CKD, which translate into therapeutic ineffectiveness or druginduced toxicity. It is estimated that one medicationrelated problem occurs with every 2.7 medication exposures in patients undergoing dialysis (2). Several reports have suggested that altered pharmacokinetics in CKD at least partially mediates this increase in adverse drug events (3-5). Although it is well known that the renal clearance of drugs is altered in CKD, the effect on nonrenal drug clearance is not well understood.Hepatic drug metabolism is the major route of drug elimination and is mediated predominantly by the cytochrome P450 (CYP) superfamily of oxidizing enzymes. The CYP isoforms, CYP2C and CYP3A, metabolize the majority (43%) of clinically used drugs (6). Several studies have reported down-regulation of CYP2C and CYP3A enzymes in CKD (3,(7)(8)(9). In addition, in vitro studies in which rat primary hepatocytes were incubated with uremic serum demonstrate a decrease in expression of CYP2C11 and CYP3A2, which are rat orthologs for human CYP2C9 and CYP3A4, respectively (10). Consistent with these studies, surgical induction of CKD in rats results in a pronounced decrease of hepatic CYP2C and CYP3A function and expression (3,7,11). Although decreased CYP2C and CYP3A function and protein expression appear to be secondary to decreased mRNA expression, the molecular mechanism(s) by which this occurs in CKD is unclear.

BACKGROUND AND PURPOSEErythropoietin (EPO) is used to treat anaemia associated with chronic kidney disease (CKD). Hypoxia is associated with anaemia and is known to cause a decrease in cytochrome P450 (P450) expression. As EPO production is regulated by hypoxia, we investigated the role of EPO on P450 expression and function. EXPERIMENTAL APPROACHMale Wistar rats were subjected to a 0.7% adenine diet for 4 weeks to induce CKD. The diet continued for an additional 2 weeks while rats received EPO by i.p. injection every other day. Following euthanasia, hepatic P450 mRNA and protein expression were determined. Hepatic enzyme activity of selected P450s was determined and chromatin immunoprecipitation was used to characterize binding of nuclear receptors involved in the transcriptional regulation of CYP2C and CYP3A. KEY RESULTSEPO administration decreased hepatic mRNA and protein expression of CYP3A2 (P < 0.05), but not CYP2C11. Similarly, EPO administration decreased CYP3A2 protein expression by 81% (P < 0.001). A 32% decrease (P < 0.05) in hepatic CYP3A enzymatic activity (Vmax) was observed for the formation of 6βOH-testosterone in the EPO-treated group. Decreases in RNA pol II recruitment (P < 0.01), hepatocyte nuclear factor 4α binding (P < 0.05) and pregnane X receptor binding (P < 0.01) to the promoter region of CYP3A were also observed in EPO-treated rats. CONCLUSIONS AND IMPLICATIONSOur data show that EPO decreases the expression and function of CYP3A, but not CYP2C in rat liver. AbbreviationsAhR, aryl hydrocarbon receptor; ALT, alanine transaminase; AST, aspartate transaminase; CAR, constitutive androstane receptor; CKD, chronic kidney disease; EPO, erythropoietin; H&E, haematoxylin and eosin; HIF, hypoxia-inducible factor; HNF-4α, hepatocyte nuclear factor 4α; JAK2, Janus kinase 2; LXR, liver X receptor; P450, cytochrome P450; PXR, pregnane X receptor; RXRα, retinoid X receptor α; STAT5, signal transducer and activator of transcription 5 BJP British Journal of Pharmacology

ObjectivesThis study aims to determine the changes in expression and function of hepatic drug metabolizing enzymes (DMEs) and drug transport proteins in liver samples from patients with end‐stage renal disease (ESRD).MethodsLiver samples were collected from nine recently deceased patients with ESRD and six recently deceased controls. Relative gene expression of DMEs and transport proteins were determined by qPCR.ResultsP‐gp and CYP2E1 expression were significantly decreased (P<0.05) in ESRD livers compared to control livers (21% and 17% of control, respectively). There was a trend of decreased expression of CYP2D6, CYP2C9, OATP1B1 MRP2 and MRP3, although this failed to reach significance.ConclusionsOur results demonstrate that DMEs and transport proteins are down‐regulated in ESRD. Current studies are evaluating the enzymatic activity of DMEs in hepatic microsomes from ESRD patients using the specific probe substrates midazolam (CYP3A4), tolbutamide (CYP2C9), chlorzoxazone (CYP2E1) and dextromethorphan (CYP2D6). Our results help explain the large decrease in non‐renal drug clearance observed in patients with ESRD.This research is supported by the Lawson Health Research Institute.