RESUMO
The coordinated movement of organic anions (e.g., drugs, metabolites, signaling molecules, nutrients, antioxidants, gut microbiome products) between tissues and body fluids depends, in large part, on organic anion transporters (OATs) [solute carrier 22 (SLC22)], organic anion transporting polypeptides (OATPs) [solute carrier organic (SLCO)], and multidrug resistance proteins (MRPs) [ATP-binding cassette, subfamily C (ABCC)]. Depending on the range of substrates, transporters in these families can be considered multispecific, oligospecific, or (relatively) monospecific. Systems biology analyses of these transporters in the context of expression patterns reveal they are hubs in networks involved in interorgan and interorganismal communication. The remote sensing and signaling theory explains how the coordinated functions of drug transporters, drug-metabolizing enzymes, and regulatory proteins play a role in optimizing systemic and local levels of important endogenous small molecules. We focus on the role of OATs, OATPs, and MRPs in endogenous metabolism and how their substrates (e.g., bile acids, short chain fatty acids, urate, uremic toxins) mediate interorgan and interorganismal communication and help maintain and restore homeostasis in healthy and disease states.
Assuntos
Avena , Transportadores de Ânions Orgânicos , Humanos , Avena/metabolismo , Tecnologia de Sensoriamento Remoto , Proteínas de Membrana Transportadoras/metabolismo , Transportadores de Ânions Orgânicos/metabolismo , Trifosfato de AdenosinaRESUMO
Homeostasis is a prerequisite for health. When homeostasis becomes disrupted, dysfunction occurs. This is especially the case for the gut microbiota, which under normal conditions lives in symbiosis with the host. As there are as many microbial cells in and on our body as human cells, it is unlikely they would not contribute to health or disease. The gut bacterial metabolism generates numerous beneficial metabolites but also uremic toxins and their precursors, which are transported into the circulation. Barrier function in the intestine, the heart, and the kidneys regulates metabolite transport and concentration and plays a role in inter-organ and inter-organism communication via small molecules. This communication is analyzed from the perspective of the remote sensing and signaling theory, which emphasizes the role of a large network of multispecific, oligospecific, and monospecific transporters and enzymes in regulating small-molecule homeostasis. The theory provides a systems biology framework for understanding organ cross talk and microbe-host communication involving metabolites, signaling molecules, nutrients, antioxidants, and uremic toxins. This remote small-molecule communication is critical for maintenance of homeostasis along the gut-heart-kidney axis and for responding to homeostatic perturbations. Chronic kidney disease is characterized by gut dysbiosis and accumulation of toxic metabolites. This slowly impacts the body, affecting the cardiovascular system and contributing to the progression of kidney dysfunction, which in its turn influences the gut microbiota. Preserving gut homeostasis and barrier functions or restoring gut dysbiosis and dysfunction could be a minimally invasive way to improve patient outcomes and quality of life in many diseases, including cardiovascular and kidney disease.
Assuntos
Microbiota , Insuficiência Renal Crônica , Humanos , Toxinas Urêmicas , Disbiose/metabolismo , Qualidade de Vida , Rim/metabolismo , Insuficiência Renal Crônica/metabolismoRESUMO
Organic anion transporter 1 (OAT1/SLC22A6) is a drug transporter with numerous xenobiotic and endogenous substrates. The Remote Sensing and Signaling Theory suggests that drug transporters with compatible ligand preferences can play a role in "organ crosstalk," mediating overall organismal communication. Other drug transporters are well known to transport lipids, but surprisingly little is known about the role of OAT1 in lipid metabolism. To explore this subject, we constructed a genome-scale metabolic model using omics data from the Oat1 knockout mouse. The model implicated OAT1 in the regulation of many classes of lipids, including fatty acids, bile acids, and prostaglandins. Accordingly, serum metabolomics of Oat1 knockout mice revealed increased polyunsaturated fatty acids, diacylglycerols, and long-chain fatty acids and decreased ceramides and bile acids when compared with wildtype controls. Some aged knockout mice also displayed increased lipid droplets in the liver when compared with wildtype mice. Chemoinformatics and machine learning analyses of these altered lipids defined molecular properties that form the structural basis for lipid-transporter interactions, including the number of rings, positive charge/volume, and complexity of the lipids. Finally, we obtained targeted serum metabolomics data after short-term treatment of rodents with the OAT-inhibiting drug probenecid to identify potential drug-metabolite interactions. The treatment resulted in alterations in eicosanoids and fatty acids, further supporting our metabolic reconstruction predictions. Consistent with the Remote Sensing and Signaling Theory, the data support a role of OAT1 in systemic lipid metabolism.
Assuntos
Metabolismo dos Lipídeos , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Animais , Técnicas de Inativação de Genes , Genômica , Aprendizado de Máquina , Camundongos , Proteína 1 Transportadora de Ânions Orgânicos/deficiência , Proteína 1 Transportadora de Ânions Orgânicos/genéticaRESUMO
How organs sense circulating metabolites is a key question. Here, we show that the multispecific organic anion transporters of drugs, OAT1 (SLC22A6 or NKT) and OAT3 (SLC22A8), play a role in organ sensing. Metabolomics analyses of the serum of Oat1 and Oat3 knockout mice revealed changes in tryptophan derivatives involved in metabolism and signaling. Several of these metabolites are derived from the gut microbiome and are implicated as uremic toxins in chronic kidney disease. Direct interaction with the transporters was supported with cell-based transport assays. To assess the impact of the loss of OAT1 or OAT3 function on the kidney, an organ where these uptake transporters are highly expressed, knockout transcriptomic data were mapped onto a "metabolic task"-based computational model that evaluates over 150 cellular functions. Despite the changes of tryptophan metabolites in both knockouts, only in the Oat1 knockout were multiple tryptophan-related cellular functions increased. Thus, deprived of the ability to take up kynurenine, kynurenate, anthranilate, and N-formylanthranilate through OAT1, the kidney responds by activating its own tryptophan-related biosynthetic pathways. The results support the Remote Sensing and Signaling Theory, which describes how "drug" transporters help optimize levels of metabolites and signaling molecules by facilitating organ cross talk. Since OAT1 and OAT3 are inhibited by many drugs, the data implies potential for drug-metabolite interactions. Indeed, treatment of humans with probenecid, an OAT-inhibitor used to treat gout, elevated circulating tryptophan metabolites. Furthermore, given that regulatory agencies have recommended drugs be tested for OAT1 and OAT3 binding or transport, it follows that these metabolites can be used as endogenous biomarkers to determine if drug candidates interact with OAT1 and/or OAT3.
Assuntos
Rim/metabolismo , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/metabolismo , Triptofano/metabolismo , Animais , Rim/citologia , Camundongos , Estresse Oxidativo , Transporte Proteico , Transdução de SinaisRESUMO
The organic anion transporter (OAT) subfamily, which constitutes roughly half of the SLC22 (solute carrier 22) transporter family, has received a great deal of attention because of its role in handling of common drugs (antibiotics, antivirals, diuretics, nonsteroidal anti-inflammatory drugs), toxins (mercury, aristolochic acid), and nutrients (vitamins, flavonoids). Oats are expressed in many tissues, including kidney, liver, choroid plexus, olfactory mucosa, brain, retina, and placenta. Recent metabolomics and microarray data from Oat1 [Slc22a6, originally identified as NKT (novel kidney transporter)] and Oat3 (Slc22a8) knockouts, as well as systems biology studies, indicate that this pathway plays a central role in the metabolism and handling of gut microbiome metabolites as well as putative uremic toxins of kidney disease. Nuclear receptors and other transcription factors, such as Hnf4α and Hnf1α, appear to regulate the expression of certain Oats in conjunction with phase I and phase II drug metabolizing enzymes. Some Oats have a strong selectivity for particular signaling molecules, including cyclic nucleotides, conjugated sex steroids, odorants, uric acid, and prostaglandins and/or their metabolites. According to the "Remote Sensing and Signaling Hypothesis," which is elaborated in detail here, Oats may function in remote interorgan communication by regulating levels of signaling molecules and key metabolites in tissues and body fluids. Oats may also play a major role in interorganismal communication (via movement of small molecules across the intestine, placental barrier, into breast milk, and volatile odorants into the urine). The role of various Oat isoforms in systems physiology appears quite complex, and their ramifications are discussed in the context of remote sensing and signaling.
Assuntos
Regulação da Expressão Gênica/fisiologia , Transportadores de Ânions Orgânicos/genética , Transportadores de Ânions Orgânicos/metabolismo , Humanos , Transportadores de Ânions Orgânicos/química , Distribuição TecidualRESUMO
The multispecific organic anion transporters, OAT1 (SLC22A6) and OAT3 (SLC22A8), the main kidney elimination pathways for many common drugs, are often considered to have largely-redundant roles. However, whereas examination of metabolomics data from Oat-knockout mice (Oat1 and Oat3KO) revealed considerable overlap, over a hundred metabolites were increased in the plasma of one or the other of these knockout mice. Many of these relatively unique metabolites are components of distinct biochemical and signaling pathways, including those involving amino acids, lipids, bile acids, and uremic toxins. Cheminformatics, together with a "logical" statistical and machine learning-based approach, identified a number of molecular features distinguishing these unique endogenous substrates. Compared with OAT1, OAT3 tends to interact with more complex substrates possessing more rings and chiral centers. An independent "brute force" approach, analyzing all possible combinations of molecular features, supported the logical approach. Together, the results suggest the potential molecular basis by which OAT1 and OAT3 modulate distinct metabolic and signaling pathways in vivo As suggested by the Remote Sensing and Signaling Theory, the analysis provides a potential mechanism by which "multispecific" kidney proximal tubule transporters exert distinct physiological effects. Furthermore, a strong metabolite-based machine-learning classifier was able to successfully predict unique OAT1 versus OAT3 drugs; this suggests the feasibility of drug design based on knockout metabolomics of drug transporters. The approach can be applied to other SLC and ATP-binding cassette drug transporters to define their nonredundant physiological roles and for analyzing the potential impact of drug-metabolite interactions.
Assuntos
Metabolômica , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/metabolismo , Toxinas Biológicas/metabolismo , Trifosfato de Adenosina/genética , Animais , Ácidos e Sais Biliares/metabolismo , Transporte Biológico/genética , Humanos , Inativação Metabólica/genética , Túbulos Renais Proximais/metabolismo , Aprendizado de Máquina , Camundongos , Camundongos Knockout , Proteína 1 Transportadora de Ânions Orgânicos/genética , Transportadores de Ânions Orgânicos/genética , Transportadores de Ânions Orgânicos/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/genética , Transdução de SinaisRESUMO
The SLC22 transporter family consists of more than two dozen members, which are expressed in the kidney, the liver, and other tissues. Evolutionary analysis indicates that SLC22 transporters fall into at least six subfamilies: OAT (organic anion transporter), OAT-like, OAT-related, OCT (organic cation transporter), OCTN (organic cation/carnitine transporter), and OCT/OCTN-related. Some-including OAT1 [SLC22A6 or NKT (novel kidney transporter)] and OAT3 (SLC22A8), as well as OCT1 (SLC22A1) and OCT2 (SLC22A2)-are widely studied drug transporters. Nevertheless, analyses of knockout mice and other data indicate that SLC22 transporters regulate key metabolic pathways and levels of signaling molecules (e.g., gut microbiome products, bile acids, tricarboxylic acid cycle intermediates, dietary flavonoids and other nutrients, prostaglandins, vitamins, short-chain fatty acids, urate, and ergothioneine), as well as uremic toxins associated with chronic kidney disease. Certain SLC22 transporters-such as URAT1 (SLC22A12) and OCTN2 (SLC22A5)-are mutated in inherited metabolic diseases. A new systems biology view of transporters is emerging. As proposed in the remote sensing and signaling hypothesis, SLC22 transporters, together with other SLC and ABC transporters, have key roles in interorgan and interorganism small-molecule communication and, together with the neuroendocrine, growth factor-cytokine, and other homeostatic systems, regulate local and whole-body homeostasis.
Assuntos
Transporte Biológico/efeitos dos fármacos , Redes e Vias Metabólicas/efeitos dos fármacos , Transportadores de Ânions Orgânicos/metabolismo , Preparações Farmacêuticas/administração & dosagem , Preparações Farmacêuticas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Humanos , Insuficiência Renal Crônica/tratamento farmacológico , Insuficiência Renal Crônica/metabolismoRESUMO
Several SLC22 transporters in the human kidney and other tissues are thought to regulate endogenous small antioxidant molecules such as uric acid, ergothioneine, carnitine, and carnitine derivatives. These transporters include those from the organic anion transporter (OAT), OCTN/OCTN-related, and organic cation transporter (OCT) subgroups. In mammals, it has been difficult to show a clear in vivo role for these transporters during oxidative stress. Ubiquitous knockdowns of related Drosophila SLC22s-including transporters homologous to those previously identified by us in mammals such as the "Fly-Like Putative Transporters" FLIPT1 (SLC22A15) and FLIPT2 (SLC22A16)-have shown modest protection against oxidative stress. However, these fly transporters tend to be broadly expressed, and it is unclear if there is an organ in which their expression is critical. Using two tissue-selective knockdown strategies, we were able to demonstrate much greater and longer protection from oxidative stress compared to previous whole fly knockdowns as well as both parent and WT strains (CG6126: p < 0.001, CG4630: p < 0.01, CG16727: p < 0.0001 and CG6006: p < 0.01). Expression in the Malpighian tubule and likely other tissues as well (e.g., gut, fat body, nervous system) appear critical for managing oxidative stress. These four Drosophila SLC22 genes are similar to human SLC22 transporters (CG6126: SLC22A16, CG16727: SLC22A7, CG4630: SLC22A3, and CG6006: SLC22A1, SLC22A2, SLC22A3, SLC22A6, SLC22A7, SLC22A8, SLC22A11, SLC22A12 (URAT1), SLC22A13, SLC22A14)-many of which are highly expressed in the kidney. Consistent with the Remote Sensing and Signaling Theory, this indicates an important in vivo role in the oxidative stress response for multiple SLC22 transporters within the fly renal system, perhaps through interaction with SLC22 counterparts in non-renal tissues. We also note that many of the human relatives are well-known drug transporters. Our work not only indicates the importance of SLC22 transporters in the fly renal system but also sets the stage for in vivo studies by examining their role in mammalian oxidative stress and organ crosstalk.
Assuntos
Drosophila melanogaster/metabolismo , Rim/metabolismo , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Estresse Oxidativo/fisiologia , Animais , Antioxidantes/metabolismo , Transporte Biológico/fisiologia , Humanos , Transdução de Sinais/fisiologiaRESUMO
The SLC22 family of transporters is widely expressed, evolutionarily conserved, and plays a major role in regulating homeostasis by transporting small organic molecules such as metabolites, signaling molecules, and antioxidants. Analysis of transporters in fruit flies provides a simple yet orthologous platform to study the endogenous function of drug transporters in vivo. Evolutionary analysis of Drosophila melanogaster putative SLC22 orthologs reveals that, while many of the 25 SLC22 fruit fly orthologs do not fall within previously established SLC22 subclades, at least four members appear orthologous to mammalian SLC22 members (SLC22A16:CG6356, SLC22A15:CG7458, CG7442 and SLC22A18:CG3168). We functionally evaluated the role of SLC22 transporters in Drosophila melanogaster by knocking down 14 of these genes. Three putative SLC22 ortholog knockdowns-CG3168, CG6356, and CG7442/SLC22A-did not undergo eclosion and were lethal at the pupa stage, indicating the developmental importance of these genes. Additionally, knocking down four SLC22 members increased resistance to oxidative stress via paraquat testing (CG4630: p < 0.05, CG6006: p < 0.05, CG6126: p < 0.01 and CG16727: p < 0.05). Consistent with recent evidence that SLC22 is central to a Remote Sensing and Signaling Network (RSSN) involved in signaling and metabolism, these phenotypes support a key role for SLC22 in handling reactive oxygen species.
Assuntos
Proteínas de Drosophila , Proteínas de Transporte de Cátions Orgânicos , Estresse Oxidativo , Transdução de Sinais , Animais , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Proteínas de Transporte de Cátions Orgânicos/genética , Proteínas de Transporte de Cátions Orgânicos/metabolismoRESUMO
The SLC22 family of OATs, OCTs, and OCTNs is emerging as a central hub of endogenous physiology. Despite often being referred to as "drug" transporters, they facilitate the movement of metabolites and key signaling molecules. An in-depth reanalysis supports a reassignment of these proteins into eight functional subgroups, with four new subgroups arising from the previously defined OAT subclade: OATS1 (SLC22A6, SLC22A8, and SLC22A20), OATS2 (SLC22A7), OATS3 (SLC22A11, SLC22A12, and Slc22a22), and OATS4 (SLC22A9, SLC22A10, SLC22A24, and SLC22A25). We propose merging the OCTN (SLC22A4, SLC22A5, and Slc22a21) and OCT-related (SLC22A15 and SLC22A16) subclades into the OCTN/OCTN-related subgroup. Using data from GWAS, in vivo models, and in vitro assays, we developed an SLC22 transporter-metabolite network and similar subgroup networks, which suggest how multiple SLC22 transporters with mono-, oligo-, and multi-specific substrate specificity interact to regulate metabolites. Subgroup associations include: OATS1 with signaling molecules, uremic toxins, and odorants, OATS2 with cyclic nucleotides, OATS3 with uric acid, OATS4 with conjugated sex hormones, particularly etiocholanolone glucuronide, OCT with neurotransmitters, and OCTN/OCTN-related with ergothioneine and carnitine derivatives. Our data suggest that the SLC22 family can work among itself, as well as with other ADME genes, to optimize levels of numerous metabolites and signaling molecules, involved in organ crosstalk and inter-organismal communication, as proposed by the remote sensing and signaling theory.
Assuntos
Regulação da Expressão Gênica , Redes Reguladoras de Genes , Mutação , Transportadores de Ânions Orgânicos/metabolismo , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Biologia de Sistemas/métodos , Animais , Transporte Biológico , Humanos , Família Multigênica , Transportadores de Ânions Orgânicos/classificação , Transportadores de Ânions Orgânicos/genética , Proteínas de Transporte de Cátions Orgânicos/classificação , Proteínas de Transporte de Cátions Orgânicos/genética , Transdução de Sinais , Especificidade por SubstratoRESUMO
Ets-1 is a member of the Ets family of transcription factors and has critical roles in multiple biological functions. Structural kidney defects occur at an increased frequency in Jacobsen syndrome (OMIM #147791), a rare chromosomal disorder caused by deletions in distal 11q, implicating at least one causal gene in distal 11q. In this study, we define an 8.1 Mb "critical region" for kidney defects in Jacobsen syndrome, which spans ~50 genes. We demonstrate that gene-targeted deletion of Ets-1 in mice results in some of the most common congenital kidney defects occurring in Jacobsen syndrome, including: duplicated kidney, hypoplastic kidney, and dilated renal pelvis and calyces. Taken together, our results implicate Ets-1 in normal mammalian kidney development and, potentially, in the pathogenesis of some of the most common types of human structural kidney defects.
Assuntos
Síndrome da Deleção Distal 11q de Jacobsen/genética , Rim/patologia , Proteína Proto-Oncogênica c-ets-1/genética , Animais , Cromossomos Humanos Par 11 , Modelos Animais de Doenças , Deleção de Genes , Marcação de Genes , Predisposição Genética para Doença , Humanos , Síndrome da Deleção Distal 11q de Jacobsen/patologia , Rim/anormalidades , Rim/crescimento & desenvolvimento , Camundongos , Deleção de Sequência/genéticaRESUMO
The organic anion transporters OAT1 (SLC22A6) and OAT3 (SLC22A8) have similar substrate specificity for drugs, but it is far from clear whether this holds for endogenous substrates. By analysis of more than 600 metabolites in the Oat3KO (Oat3 knockout) by LC/MS, we demonstrate OAT3 involvement in the movement of gut microbiome products, key metabolites, and signaling molecules, including those flowing through the gut-liver-kidney axis. Major pathways affected included those involved in metabolism of bile acids, flavonoids, nutrients, amino acids (including tryptophan-derivatives that are uremic toxins), and lipids. OAT3 is also critical in elimination of liver-derived phase II metabolites, particularly those undergoing glucuronidation. Analysis of physicochemical features revealed nine distinct metabolite groups; at least one member of most clusters has been previously validated in transport assays. In contrast to drugs interacting with the OATs, endogenous metabolites accumulating in the Oat1KO (Oat1 knockout) versus Oat3KO have distinct differences in their physicochemical properties; they are very different in size, number of rings, hydrophobicity, and molecular complexity. Consistent with the Remote Sensing and Signaling Hypothesis, the data support the importance of the OAT transporters in inter-organ and inter-organismal remote communication via transporter-mediated movement of key metabolites and signaling molecules (e.g. gut microbiome-to-intestine-to-blood-to-liver-to-kidney-to-urine). We discuss the possibility of an intimate connection between OATs and metabolite sensing and signaling pathways (e.g. bile acids). Furthermore, the metabolomics and pathway analysis support the view that OAT1 plays a greater role in kidney proximal tubule metabolism and OAT3 appears relatively more important in systemic metabolism, modulating levels of metabolites flowing through intestine, liver, and kidney.
Assuntos
Mucosa Intestinal/metabolismo , Rim/metabolismo , Fígado/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/metabolismo , Aminoácidos/metabolismo , Animais , Ácidos e Sais Biliares/metabolismo , Dieta , Metabolismo Energético , Microbioma Gastrointestinal , Técnicas de Inativação de Genes , Intestinos/microbiologia , Ligantes , Metabolismo dos Lipídeos , Masculino , Metabolômica , Camundongos , Camundongos Endogâmicos C57BL , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/deficiência , Transportadores de Ânions Orgânicos Sódio-Independentes/genética , Especificidade por Substrato , Xenobióticos/metabolismoRESUMO
The sodium-glucose cotransporter SGLT2 inhibitor empagliflozin (plasma protein binding ~88%) may reach its target in the brush border of the early proximal tubule by glomerular filtration and tubular secretion. Here we determined whether empagliflozin is secreted by renal tubules in mice and whether genetic knockout of the basolateral organic anion transporter 3 ( Oat3-/-) affects its tubular secretion or glucosuric effect. Renal clearance studies in wild-type (WT) mice showed that tubular secretion accounted for 50-70% of empagliflozin urinary excretion. Immunostaining indicated that SGLT2 and OAT3 localization partially overlapped in proximal tubule S1 and S2 segments. Glucosuria in metabolic cage studies was reduced in Oat3-/- vs. WT mice for acute empagliflozin doses of 1, 3, and 10 mg/kg, whereas 30 mg/kg induced similar maximal glucosuria in both genotypes. Chronic application of empagliflozin (~25 mg·kg-1 ·day-1) in Oat3-/- mice was associated with lower urinary glucose-to-creatinine ratios despite maintaining slightly higher blood glucose levels than WT. On a whole kidney level, renal secretion of empagliflozin was largely unchanged in Oat3-/- mice. However, the absence of OAT3 attenuated the influence of empagliflozin on fractional glucose excretion; higher levels of plasma or filtered empagliflozin were needed to induce similar increases in fractional renal glucose excretion. We conclude that empagliflozin is excreted into the urine to similar extent by glomerular filtration and tubular secretion. The latter can occur largely independent of OAT3. However, OAT3 increases the glucosuric effect of empagliflozin, which may relate to the partial overlap of its localization with SGLT2 and thus OAT3-mediated tubular secretion of empagliflozin in the early proximal tubule.
Assuntos
Compostos Benzidrílicos/farmacologia , Glicemia/efeitos dos fármacos , Glucosídeos/farmacologia , Glicosúria/metabolismo , Túbulos Renais Proximais/efeitos dos fármacos , Transportadores de Ânions Orgânicos Sódio-Independentes/metabolismo , Eliminação Renal , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Transportador 2 de Glucose-Sódio/efeitos dos fármacos , Animais , Compostos Benzidrílicos/farmacocinética , Compostos Benzidrílicos/urina , Glicemia/metabolismo , Taxa de Filtração Glomerular , Glucosídeos/farmacocinética , Glucosídeos/urina , Glicosúria/genética , Glicosúria/prevenção & controle , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína 1 Transportadora de Ânions Orgânicos/genética , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/deficiência , Transportadores de Ânions Orgânicos Sódio-Independentes/genética , Transportador 2 de Glucose-Sódio/metabolismo , Inibidores do Transportador 2 de Sódio-Glicose/farmacocinética , Inibidores do Transportador 2 de Sódio-Glicose/urinaRESUMO
Maternal pregnancy adaptation is crucial for fetal development and long-term health. Complex interactions occur between maternal digestive and excretory systems as they interface with the developing fetus through the placenta, and transcriptomic regulation in these organs throughout pregnancy is poorly understood. Our objective is to characterize transcriptomic changes across gestation in maternal organs and placenta. Gene expression was quantified in the kidney, liver, and small intestine harvested from nonpregnant and pregnant FVB mice at four time points and placenta at three time points (N = 5/time point) using Affymetrix Mouse Gene 1.0 ST arrays. In maternal organs, we identified 476 genes in the liver, 207 genes in the kidney, and 27 genes in the small intestine that were differentially expressed across gestation (False Discovery Rate [FDR] adjusted q < 0.05). The placenta had a total of 1576 differentially expressed genes between the placenta at either/gd15 or gd19 compared to gd10. We identified a number of pathways enriched for genes differentially expressed across gestation, including 5 pathways in the placenta, 9 pathways in the kidney, and 28 pathways in the liver, including the citrate cycle, retinol metabolism, bile acid synthesis, and steroid bile synthesis, which play functional roles in fetal development and pregnancy maintenance. Characterization of normal longitudinal changes that occur in pregnancy provides context to understand how perturbations in these biochemical pathways and perturbations in nutrient signaling may impact pregnancy.
Assuntos
Perfilação da Expressão Gênica , Placenta/metabolismo , Transcriptoma/fisiologia , Animais , Feminino , Regulação da Expressão Gênica , Intestino Delgado/metabolismo , Rim/metabolismo , Fígado/metabolismo , Camundongos , Análise Multivariada , GravidezRESUMO
PURPOSE OF REVIEW: Uric acid homeostasis in the body is mediated by a number of SLC and ABC transporters in the kidney and intestine, including several multispecific 'drug' transporters (e.g., OAT1, OAT3, and ABCG2). Optimization of uric acid levels can be viewed as a 'systems biology' problem. Here, we consider uric acid transporters from a systems physiology perspective using the framework of the 'Remote Sensing and Signaling Hypothesis.' This hypothesis explains how SLC and ABC 'drug' and other transporters mediate interorgan and interorganismal communication (e.g., gut microbiome and host) via small molecules (e.g., metabolites, antioxidants signaling molecules) through transporters expressed in tissues lining body fluid compartments (e.g., blood, urine, cerebrospinal fluid). RECENT FINDINGS: The list of uric acid transporters includes: SLC2A9, ABCG2, URAT1 (SLC22A12), OAT1 (SLC22A6), OAT3 (SLC22A8), OAT4 (SLC22A11), OAT10 (SLC22A13), NPT1 (SLC17A1), NPT4 (SLC17A3), MRP2 (ABCC2), MRP4 (ABCC4). Normally, SLC2A9, - along with URAT1, OAT1 and OAT3, - appear to be the main transporters regulating renal urate handling, while ABCG2 appears to regulate intestinal transport. In chronic kidney disease (CKD), intestinal ABCG2 becomes much more important, suggesting remote organ communication between the injured kidney and the intestine. SUMMARY: The remote sensing and signaling hypothesis provides a useful systems-level framework for understanding the complex interplay of uric acid transporters expressed in different tissues involved in optimizing uric acid levels under normal and diseased (e.g., CKD, gut microflora dysbiosis) conditions.
Assuntos
Transporte Biológico , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Ácido Úrico/metabolismo , Animais , Humanos , Mucosa Intestinal/metabolismo , Bombas de Íon/genética , Bombas de Íon/metabolismo , Rim/metabolismo , Proteína 2 Associada à Farmacorresistência Múltipla , Transdução de Sinais , Biologia de SistemasRESUMO
There has been a recent interest in the broader physiological importance of multispecific "drug" transporters of the SLC and ABC transporter families. Here, a novel multi-tiered systems biology approach was used to predict metabolites and signaling molecules potentially affected by the in vivo deletion of organic anion transporter 1 (Oat1, Slc22a6, originally NKT), a major kidney-expressed drug transporter. Validation of some predictions in wet-lab assays, together with re-evaluation of existing transport and knock-out metabolomics data, generated an experimentally validated, confidence ranked set of OAT1-interacting endogenous compounds enabling construction of an "OAT1-centered metabolic interaction network." Pathway and enrichment analysis indicated an important role for OAT1 in metabolism involving: the TCA cycle, tryptophan and other amino acids, fatty acids, prostaglandins, cyclic nucleotides, odorants, polyamines, and vitamins. The partly validated reconstructed network is also consistent with a major role for OAT1 in modulating metabolic and signaling pathways involving uric acid, gut microbiome products, and so-called uremic toxins accumulating in chronic kidney disease. Together, the findings are compatible with the hypothesized role of drug transporters in remote inter-organ and inter-organismal communication: The Remote Sensing and Signaling Hypothesis (Nigam, S. K. (2015) Nat. Rev. Drug Disc. 14, 29). The fact that OAT1 can affect many systemic biological pathways suggests that drug-metabolite interactions need to be considered beyond simple competition for the drug transporter itself and may explain aspects of drug-induced metabolic syndrome. Our approach should provide novel mechanistic insights into the role of OAT1 and other drug transporters implicated in metabolic diseases like gout, diabetes, and chronic kidney disease.
Assuntos
Metaboloma/fisiologia , Modelos Biológicos , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Animais , CamundongosRESUMO
Statistical analysis was performed on physicochemical descriptors of â¼250 drugs known to interact with one or more SLC22 "drug" transporters (i.e., SLC22A6 or OAT1, SLC22A8 or OAT3, SLC22A1 or OCT1, and SLC22A2 or OCT2), followed by application of machine-learning methods and wet laboratory testing of novel predictions. In addition to molecular charge, organic anion transporters (OATs) were found to prefer interacting with planar structures, whereas organic cation transporters (OCTs) interact with more three-dimensional structures (i.e., greater SP3 character). Moreover, compared with OAT1 ligands, OAT3 ligands possess more acyclic tetravalent bonds and have a more zwitterionic/cationic character. In contrast, OCT1 and OCT2 ligands were not clearly distinquishable form one another by the methods employed. Multiple pharmacophore models were generated on the basis of the drugs and, consistent with the machine-learning analyses, one unique pharmacophore created from ligands of OAT3 possessed cationic properties similar to OCT ligands; this was confirmed by quantitative atomic property field analysis. Virtual screening with this pharmacophore, followed by transport assays, identified several cationic drugs that selectively interact with OAT3 but not OAT1. Although the present analysis may be somewhat limited by the need to rely largely on inhibition data for modeling, wet laboratory/in vitro transport studies, as well as analysis of drug/metabolite handling in Oat and Oct knockout animals, support the general validity of the approach-which can also be applied to other SLC and ATP binding cassette drug transporters. This may make it possible to predict the molecular properties of a drug or metabolite necessary for interaction with the transporter(s), thereby enabling better prediction of drug-drug interactions and drug-metabolite interactions. Furthermore, understanding the overlapping specificities of OATs and OCTs in the context of dynamic transporter tissue expression patterns should help predict net flux in a particular tissue of anionic, cationic, and zwitterionic molecules in normal and pathophysiological states.
Assuntos
Biologia Computacional/métodos , Aprendizado de Máquina , Proteínas de Membrana Transportadoras/metabolismo , Preparações Farmacêuticas/metabolismo , Concentração de Íons de Hidrogênio , Proteínas de Membrana Transportadoras/química , Modelos Moleculares , Proteína 1 Transportadora de Ânions Orgânicos/química , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/química , Transportadores de Ânions Orgânicos Sódio-Independentes/metabolismo , Transportador 1 de Cátions Orgânicos/química , Transportador 1 de Cátions Orgânicos/metabolismo , Preparações Farmacêuticas/química , Ligação Proteica , Conformação Proteica , Especificidade por SubstratoRESUMO
The hepatocyte nuclear factors, Hnf1a and Hnf4a, in addition to playing key roles in determining hepatocyte fate, have been implicated as candidate lineage-determining transcription factors in the kidney proximal tubule (PT) [Martovetsky et. al., (2012) Mol Pharmacol 84:808], implying an additional level of regulation that is potentially important in developmental and/or tissue-engineering contexts. Mouse embryonic fibroblasts (MEFs) transduced with Hnf1a and Hnf4a form tight junctions and express multiple PT drug transporters (e.g., Slc22a6/Oat1, Slc47a1/Mate1, Slc22a12/Urat1, Abcg2/Bcrp, Abcc2/Mrp2, Abcc4/Mrp4), nutrient transporters (e.g., Slc34a1/NaPi-2, Slco1a6), and tight junction proteins (occludin, claudin 6, ZO-1/Tjp1, ZO-2/Tjp2). In contrast, the coexpression (with Hnf1a and Hnf4a) of GATA binding protein 4 (Gata4), as well as the forkhead box transcription factors, Foxa2 and Foxa3, in MEFs not only downregulates PT markers but also leads to upregulation of several hepatocyte markers, including albumin, apolipoprotein, and transferrin. A similar result was obtained with primary mouse PT cells. Thus, the presence of Gata4 and Foxa2/Foxa3 appears to alter the effect of Hnf1a and Hnf4a by an as-yet unidentified mechanism, leading toward the generation of more hepatocyte-like cells as opposed to cells exhibiting PT characteristics. The different roles of Hnf4a in the kidney and liver was further supported by reanalysis of ChIP-seq data, which revealed Hnf4a colocalization in the kidney near PT-enriched genes compared with those genes enriched in the liver. These findings provide valuable insight, not only into the developmental, and perhaps organotypic, regulation of drug transporters, drug-metabolizing enzymes, and tight junctions, but also for regenerative medicine strategies aimed at restoring the function of the liver and/or kidney (acute kidney injury, AKI; chronic kidney disease, CKD).
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Diferenciação Celular , Linhagem da Célula , Hepatócitos/metabolismo , Túbulos Renais Proximais/metabolismo , Fígado/metabolismo , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Proteínas de Junções Íntimas/metabolismo , Fatores de Transcrição/metabolismo , Transportadores de Cassetes de Ligação de ATP/genética , Animais , Biomarcadores/metabolismo , Fibroblastos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Células HEK293 , Humanos , Túbulos Renais Proximais/citologia , Fígado/citologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Proteína 2 Associada à Farmacorresistência Múltipla , Proteínas de Transporte de Cátions Orgânicos/genética , Ratos , Proteínas de Junções Íntimas/genética , Fatores de Transcrição/genética , TransfecçãoRESUMO
Bicuspid aortic valves calcify at a significantly higher rate than normal aortic valves, a process that involves increased inflammation. Because we have previously found that bicuspid aortic valve experience greater stretch, we investigated the potential connection between stretch and inflammation in human aortic valve interstitial cells (AVICs). Microarray, quantitative PCR (qPCR), and protein assays performed on AVICs exposed to cyclic stretch showed that stretch was sufficient to increase expression of interleukin and metalloproteinase family members by more than 1.5-fold. Conditioned medium from stretched AVICs was sufficient to activate leukocytes. microRNA sequencing and qPCR experiments demonstrated that miR-148a-3p was repressed in both stretched AVICs (43% repression) and, as a clinical correlate, human bicuspid aortic valves (63% reduction). miR-148a-3p was found to be a novel repressor of IKBKB based on data from qPCR, luciferase, and Western blot experiments. Furthermore, increasing miR-148a-3p levels in AVICs was sufficient to decrease NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) signaling and NF-κB target gene expression. Our data demonstrate that stretch-mediated activation of inflammatory pathways is at least partly the result of stretch-repression of miR-148a-3p and a consequent failure to repress IKBKB. To our knowledge, we are the first to report that cyclic stretch of human AVICs activates inflammatory genes in a tissue-autonomous manner via a microRNA that regulates a central inflammatory pathway.
Assuntos
Valva Aórtica/anormalidades , Biomarcadores/metabolismo , Doenças das Valvas Cardíacas/metabolismo , Quinase I-kappa B/metabolismo , Inflamação/genética , MicroRNAs/genética , NF-kappa B/metabolismo , Valva Aórtica/imunologia , Valva Aórtica/metabolismo , Doença da Válvula Aórtica Bicúspide , Western Blotting , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Perfilação da Expressão Gênica , Doenças das Valvas Cardíacas/imunologia , Humanos , Quinase I-kappa B/genética , Inflamação/imunologia , Inflamação/patologia , Monócitos/citologia , Monócitos/metabolismo , NF-kappa B/genética , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Estresse FisiológicoRESUMO
The multispecific organic anion drug transporters OAT6 (SLC22A20) and OAT1 (SLC22A6) are expressed in nasal epithelial cells and both can bind odorants. Sequence analysis of OAT6 revealed an evolutionarily conserved 79-amino acid (AA) fragment present not only in OAT6 but also in other SLC22 transporters, such as the organic anion transporter (OAT), organic carnitine transporter (OCTN), and organic cation transporter (OCT) subfamilies. A similar fragment is also conserved in some odorant receptors (ORs) in both humans and rodents. This fragment is located in regions believed to be important for ligand/substrate preference and recognition in both classes of proteins, raising the possibility that it may be part of a potential common ligand/substrate recognition site in certain ORs and SLC22 transporters. In silico screening of an odorant database containing known OR ligands with a pharmacophore hypothesis (generated from a set of odorants known to bind OAT6 and/or OAT1), followed by in vitro uptake assays in transfected cells, identified OR ligands capable of inhibiting OAT6- and/or OAT1-mediated transport, albeit with different affinities. The conservation of the AA fragments between these two different classes of proteins, together with their coexpression in olfactory as well as other tissues, suggests the possibility that ORs and SLC22 transporters function in concert, and raises the question as to whether these transporters function in remote sensing and signaling and/or as transceptors.