Regulation of Proteome Maintenance Gene Expression by Activators of Peroxisome Proliferator-Activated Receptor α

The nuclear receptor peroxisome proliferator-activated receptor α (PPARα) is activated by a large number of xenobiotic and hypolipidemic compounds called peroxisome proliferator chemicals (PPCs). One agonist of PPARα (WY-14,643) regulates responses in the mouse liver to chemical stress in part by altering expression of genes involved in proteome maintenance (PM) including protein chaperones in the heat shock protein (Hsp) family and proteasomal genes (Psm) involved in proteolysis. We hypothesized that other PPARα activators including diverse hypolipidemic and xenobiotic compounds also regulate PM genes in the rat and mouse liver. We examined the expression of PM genes in rat and mouse liver after exposure to 7 different PPCs (WY-14,643, clofibrate, fenofibrate, valproic acid, di-(2-ethylhexyl) phthalate, perfluorooctanoic acid, and perfluorooctane sulfonate) using Affymetrix microarrays. In rats and mice, 174 or 380 PM genes, respectively, were regulated by at least one PPC. The transcriptional changes were, for the most part, dependent on PPARα, as most changes were not observed in similarly treated PPARα-null mice and the changes were not consistently observed in rats treated with activators of the nuclear receptors CAR or PXR. In rats and mice, PM gene expression exhibited differences compared to typical direct targets of PPARα (e.g., Cyp4a family members). PM gene expression was usually delayed and in some cases, it was transient. Dose-response characterization of protein expression showed that Hsp86 and Hsp110 proteins were induced only at higher doses. These studies demonstrate that PPARα, activated by diverse PPC, regulates the expression of a large number of genes involved in protein folding and degradation and support an expanded role for PPARα in the regulation of genes that protect the proteome

Hepatic Xenobiotic Metabolizing Enzyme and Transporter Gene Expression through the Life Stages of the Mouse

Differences in responses to environmental chemicals and drugs between life stages are likely due in part to differences in the expression of xenobiotic metabolizing enzymes and transporters (XMETs). No comprehensive analysis of the mRNA expression of XMETs has been carried out through life stages in any species.Using full-genome arrays, the mRNA expression of all XMETs and their regulatory proteins was examined during fetal (gestation day (GD) 19), neonatal (postnatal day (PND) 7), prepubescent (PND32), middle age (12 months), and old age (18 and 24 months) in the C57BL/6J (C57) mouse liver and compared to adults. Fetal and neonatal life stages exhibited dramatic differences in XMET mRNA expression compared to the relatively minor effects of old age. The total number of XMET probe sets that differed from adults was 636, 500, 84, 5, 43, and 102 for GD19, PND7, PND32, 12 months, 18 months and 24 months, respectively. At all life stages except PND32, under-expressed genes outnumbered over-expressed genes. The altered XMETs included those in all of the major metabolic and transport phases including introduction of reactive or polar groups (Phase I), conjugation (Phase II) and excretion (Phase III). In the fetus and neonate, parallel increases in expression were noted in the dioxin receptor, Nrf2 components and their regulated genes while nuclear receptors and regulated genes were generally down-regulated. Suppression of male-specific XMETs was observed at early (GD19, PND7) and to a lesser extent, later life stages (18 and 24 months). A number of female-specific XMETs exhibited a spike in expression centered at PND7.The analysis revealed dramatic differences in the expression of the XMETs, especially in the fetus and neonate that are partially dependent on gender-dependent factors. XMET expression can be used to predict life stage-specific responses to environmental chemicals and drugs

Analysis of the heat shock response in mouse liver reveals transcriptional dependence on the nuclear receptor peroxisome proliferator-activated receptor α (PPARα)

<p>Abstract</p> <p>Background</p> <p>The nuclear receptor peroxisome proliferator-activated receptor alpha (PPARα) regulates responses to chemical or physical stress in part by altering expression of genes involved in proteome maintenance. Many of these genes are also transcriptionally regulated by heat shock (HS) through activation by HS factor-1 (HSF1). We hypothesized that there are interactions on a genetic level between PPARα and the HS response mediated by HSF1.</p> <p>Results</p> <p>Wild-type and PPARα-null mice were exposed to HS, the PPARα agonist WY-14,643 (WY), or both; gene and protein expression was examined in the livers of the mice 4 or 24 hrs after HS. Gene expression profiling identified a number of <it>Hsp </it>family members that were altered similarly in both mouse strains. However, most of the targets of HS did not overlap between strains. A subset of genes was shown by microarray and RT-PCR to be regulated by HS in a PPARα-dependent manner. HS also down-regulated a large set of mitochondrial genes specifically in PPARα-null mice that are known targets of PPARγ co-activator-1 (PGC-1) family members. Pretreatment of PPARα-null mice with WY increased expression of PGC-1β and target genes and prevented the down-regulation of the mitochondrial genes by HS. A comparison of HS genes regulated in our dataset with those identified in wild-type and HSF1-null mouse embryonic fibroblasts indicated that although many HS genes are regulated independently of both PPARα and HSF1, a number require both factors for HS responsiveness.</p> <p>Conclusions</p> <p>These findings demonstrate that the PPARα genotype has a dramatic effect on the transcriptional targets of HS and support an expanded role for PPARα in the regulation of proteome maintenance genes after exposure to diverse forms of environmental stress including HS.</p

Transcriptional ontogeny of the developing liver

<p>Abstract</p> <p>Background</p> <p>During embryogenesis the liver is derived from endodermal cells lining the digestive tract. These endodermal progenitor cells contribute to forming the parenchyma of a number of organs including the liver and pancreas. Early in organogenesis the fetal liver is populated by hematopoietic stem cells, the source for a number of blood cells including nucleated erythrocytes. A comprehensive analysis of the transcriptional changes that occur during the early stages of development to adulthood in the liver was carried out.</p> <p>Results</p> <p>We characterized gene expression changes in the developing mouse liver at gestational days (GD) 11.5, 12.5, 13.5, 14.5, 16.5, and 19 and in the neonate (postnatal day (PND) 7 and 32) compared to that in the adult liver (PND67) using full-genome microarrays. The fetal liver, and to a lesser extent the neonatal liver, exhibited dramatic differences in gene expression compared to adults. Canonical pathway analysis of the fetal liver signature demonstrated increases in functions important in cell replication and DNA fidelity whereas most metabolic pathways of intermediary metabolism were under expressed. Comparison of the dataset to a number of previously published microarray datasets revealed 1) a striking similarity between the fetal liver and that of the pancreas in both mice and humans, 2) a nucleated erythrocyte signature in the fetus and 3) under expression of most xenobiotic metabolism genes throughout development, with the exception of a number of transporters associated with either hematopoietic cells or cell proliferation in hepatocytes.</p> <p>Conclusions</p> <p>Overall, these findings reveal the complexity of gene expression changes during liver development and maturation, and provide a foundation to predict responses to chemical and drug exposure as a function of early life-stages.</p

Inhaled ozone (O3)-induces changes in serum metabolomic and liver transcriptomic profiles in rats

Air pollution has been linked to increased incidence of diabetes. Recently, we showed that ozone (O3) induces glucose intolerance, and increases serum leptin and epinephrine in Brown Norway rats. In this study, we hypothesized that O3 exposure will cause systemic changes in metabolic homeostasis and that serum metabolomic and liver transcriptomic profiling will provide mechanistic insights. In the first experiment, male Wistar Kyoto (WKY) rats were exposed to filtered air (FA) or O3 at 0.25, 0.50, or 1.0 ppm, 6 h/day for two days to establish concentration-related effects on glucose tolerance and lung injury. In a second experiment, rats were exposed to FA or 1.0 ppm O3, 6 h/day for either one or two consecutive days, and systemic metabolic responses were determined immediately after or 18 h post-exposure. O3 increased serum glucose and leptin on day 1. Glucose intolerance persisted through two days of exposure but reversed 18 h-post second exposure. O3 increased circulating metabolites of glycolysis, long-chain free fatty acids, branched-chain amino acids and cholesterol, while 1,5-anhydroglucitol, bile acids and metabolites of TCA cycle were decreased, indicating impaired glycemic control, proteolysis and lipolysis. Liver gene expression increased for markers of glycolysis, TCA cycle and gluconeogenesis, and decreased for markers of steroid and fat biosynthesis. Genes involved in apoptosis and mitochondrial function were also impacted by O3. In conclusion, short-term O3 exposure induces global metabolic derangement involving glucose, lipid, and amino acid metabolism, typical of a stress–response. It remains to be examined if these alterations contribute to insulin resistance upon chronic exposure

Characterization of Peroxisome Proliferator–Activated Receptor α—Independent Effects of PPARα Activators in the Rodent Liver: Di-(2-ethylhexyl) phthalate also Activates the Constitutive-Activated Receptor

Peroxisome proliferator chemicals (PPC) are thought to mediate their effects in rodents on hepatocyte growth and liver cancer through the nuclear receptor peroxisome proliferator–activated receptor (PPAR) α. Recent studies indicate that the plasticizer di-(2-ethylhexyl) phthalate (DEHP) increased the incidence of liver tumors in PPARα-null mice. We hypothesized that some PPC, including DEHP, induce transcriptional changes independent of PPARα but dependent on other nuclear receptors, including the constitutive-activated receptor (CAR) that mediates phenobarbital (PB) effects on hepatocyte growth and liver tumor induction. To determine the potential role of CAR in mediating effects of PPC, a meta-analysis was performed on transcript profiles from published studies in which rats and mice were exposed to PPC and compared the profiles to those produced by exposure to PB. Valproic acid, clofibrate, and DEHP in rat liver and DEHP in mouse liver induced genes, including Cyp2b family members that are known to be regulated by CAR. Examination of transcript changes by Affymetrix ST 1.0 arrays and reverse transcription-PCR in the livers of DEHP-treated wild-type, PPARα-null, and CAR-null mice demonstrated that (1) most (∼94%) of the transcriptional changes induced by DEHP were PPARα-dependent, (2) many PPARα-independent genes overlapped with those regulated by PB, (3) induction of genes Cyp2b10, Cyp3a11, and metallothionine-1 by DEHP was CAR dependent but PPARα-independent, and (4) induction of a number of genes (Cyp8b1, Gstm4, and Gstm7) was independent of both CAR and PPARα. Our results indicate that exposure to PPARα activators including DEHP leads to activation of multiple nuclear receptors in the rodent liver

The Physicochemistry of Capped Nanosilver Predicts Its Biological Activity in Rat Brain Endothelial Cells (RBEC4)

The “capping” or coating of nanosilver (nanoAg) extends its potency by limiting its oxidation and aggregation and stabilizing its size and shape. The ability of such coated nanoAg to alter the permeability and activate oxidative stress pathways in rat brain endothelial cells (RBEC4) was examined in the present study. The aggregate size and zeta potential of nanoAg with different sizes (10 and 75 nm) and coatings (PVP and citrate) were measured in cell culture media. Results indicated that both PVP-coated nanoAg were less electronegative than their citrate-coated counterparts over all exposure times, but only the PVP-coated 10 nm particles retained their initial electronegativity over all exposure times. In addition, only the PVP-coated particles retained their initial sizes throughout the 3 h measurement. PVP-coated 10 nm nanoAg selectively altered the permeability of RBEC4 monolayers within a 15 min exposure, although high resolution microscopy indicated that all coated nanoAg distributed throughout the cell’s cytoplasm within the 3 h exposure. Reporter genes for AP-1 and NRF2/ARE, transfected into RBEC4, were selectively stimulated by the PVP-coated 10 nm nanoAg. Global gene arrays indicated that only PVP-coated nanoAg significantly altered gene expressions in the RBEC4, and those altered by 10 nm PVP-coated nanoAg were qualitatively similar but quantitatively much higher than those of its 75 nm counterpart. IPA and DAVID analyses indicated that the altered pathways affected by both PVP-coated nanoAg were primarily associated with a NRF2-mediated oxidative stress response, endocytosis, and bioenergetics. Together, these data suggest that the physicochemical features of surface coating aggregate size and surface charge contribute to capped nanoAg’s permeability and oxidative stress responses in RBEC4

The Physicochemistry of Capped Nanosilver Predicts Its Biological Activity in Rat Brain Endothelial Cells (RBEC4)

The “capping” or coating of nanosilver (nanoAg) extends its potency by limiting its oxidation and aggregation and stabilizing its size and shape. The ability of such coated nanoAg to alter the permeability and activate oxidative stress pathways in rat brain endothelial cells (RBEC4) was examined in the present study. The aggregate size and zeta potential of nanoAg with different sizes (10 and 75 nm) and coatings (PVP and citrate) were measured in cell culture media. Results indicated that both PVP-coated nanoAg were less electronegative than their citrate-coated counterparts over all exposure times, but only the PVP-coated 10 nm particles retained their initial electronegativity over all exposure times. In addition, only the PVP-coated particles retained their initial sizes throughout the 3 h measurement. PVP-coated 10 nm nanoAg selectively altered the permeability of RBEC4 monolayers within a 15 min exposure, although high resolution microscopy indicated that all coated nanoAg distributed throughout the cell’s cytoplasm within the 3 h exposure. Reporter genes for AP-1 and NRF2/ARE, transfected into RBEC4, were selectively stimulated by the PVP-coated 10 nm nanoAg. Global gene arrays indicated that only PVP-coated nanoAg significantly altered gene expressions in the RBEC4, and those altered by 10 nm PVP-coated nanoAg were qualitatively similar but quantitatively much higher than those of its 75 nm counterpart. IPA and DAVID analyses indicated that the altered pathways affected by both PVP-coated nanoAg were primarily associated with a NRF2-mediated oxidative stress response, endocytosis, and bioenergetics. Together, these data suggest that the physicochemical features of surface coating aggregate size and surface charge contribute to capped nanoAg’s permeability and oxidative stress responses in RBEC4