Kathleen M. Bottum

Aryl Hydrocarbon Receptor Deficiency Enhances Insulin Sensitivity and Reduces PPAR-α Pathway Activity in Mice

Background: Numerous man-made pollutants activate the aryl hydrocarbon receptor (AhR) and are risk factors for type 2 diabetes. AhR signaling also affects molecular clock genes to influence glucose metabolism. Objective: We investigated mechanisms by which AhR activation affects glucose metabolism. Methods: Glucose tolerance, insulin resistance, and expression of peroxisome proliferator–activated receptor-α (PPAR-α) and genes affecting glucose metabolism or fatty acid oxidation and clock gene rhythms were investigated in wild-type (WT) and AhR-deficient [knockout (KO)] mice. AhR agonists and small interfering RNA (siRNA) were used to examine the effect of AhR on PPAR-α expression and glycolysis in the liver cell line Hepa-1c1c7 (c7) and its c12 and c4 derivatives. Brain, muscle ARNT-like protein 1 (Bmal1) siRNA and Ahr or Bmal1 expression plasmids were used to analyze the effect of BMAL1 on PPAR-α expression in c7 cells. Results: KO mice displayed enhanced insulin sensitivity and improved glucose tolerance, accompanied by decreased PPAR-α and key gluconeogenic and fatty acid oxidation enzymes. AhR agonists increased PPAR-α expression in c7 cells. Both Ahr and Bmal1 siRNA reduced PPAR-α and metabolism genes. Moreover, rhythms of BMAL1 and blood glucose were altered in KO mice. Conclusions: These results indicate a link between AhR signaling, circadian rhythms, and glucose metabolism. Furthermore, hepatic activation of the PPAR-α pathway provides a mechanism underlying AhR-mediated insulin resistance.

ARYL HYDROCARBON RECEPTOR DEFICIENCY PROTECTS MICE FROM DIET-INDUCED ADIPOSITY AND METABOLIC DISORDERS THROUGH INCREASED ENERGY EXPENDITURE

Background/Objectives:Epidemics of obesity and diabetes are escalating. High-calorie/high-fat food is a major cause for these global health issues, but molecular mechanisms underlying high-fat, diet-induced obesity are still not well understood. The aryl hydrocarbon receptor (AhR), a transcription factor that acts as a xenobiotic sensor, mediates environmental toxicant-induced obesity, insulin resistance and development of diabetes. AhR also influences lipid metabolism and diet-induced obesity. The effects of AhR deficiency on diet-induced obesity, hepatic steatosis and insulin resistance were examined.Methods:Male wild-type (WT), AhR null (AhR−/−) and AhR heterozygote (AhR+/−) mice were fed a normal chow diet (NCD, 10% kcal from fat) or a high-fat diet (HFD, 60% kcal from fat) for up to 14 weeks. Adiposity, adipose and liver morphology, insulin signaling, metabolic parameters and gene profiles were assessed.Results:AhR deficiency protected against HFD-induced obesity, hepatic steatosis, insulin resistance and inflammation. Moreover, AhR deficiency preserved insulin signaling in major metabolic tissues. These protective effects result from a higher energy expenditure in AhR-deficient mice compared with WT. Levels of transcript for both the thermogenic gene, uncoupling protein 1 (Ucp1), in brown adipose tissue and mitochondrial β-oxidation genes in muscle were significantly higher in AhR−/− and AhR+/− mice compared with WT.Conclusions:This work documents a physiologically relevant function for AhR in regulation of body weight, hepatic fat deposition, insulin sensitivity and energy expenditure under HFD exposure, suggesting that AhR signaling may be developed as a potential therapeutic target for treatment of obesity and metabolic disorders.

ERK/MAPK Is Essential for Endogenous Neuroprotection in SCN2.2 Cells

Background Glutamate (Glu) is essential to central nervous system function; however excessive Glu release leads to neurodegenerative disease. Strategies to protect neurons are underdeveloped, in part due to a limited understanding of natural neuroprotective mechanisms, such as those present in the suprachiasmatic nucleus (SCN). This study tests the hypothesis that activation of ERK/MAPK provides essential protection to the SCN after exposure to excessive Glu using the SCN2.2 cells as a model. Methodology Immortalized SCN2.2 cells (derived from SCN) and GT1-7 cells (neurons from the neighboring hypothalamus) were treated with 10 mM Glu in the presence or absence of the ERK/MAPK inhibitor PD98059. Cell death was assessed by Live/Dead assay, MTS assay and TUNEL. Caspase 3 activity was also measured. Activation of MAPK family members was determined by immunoblot. Bcl2, neuritin and Bid mRNA (by quantitative-PCR) and protein levels (by immunoblot) were also measured. Principal Findings As expected Glu treatment increased caspase 3 activity and cell death in the GT1-7 cells, but Glu alone did not induce cell death or affect caspase 3 activity in the SCN2.2 cells. However, pretreatment with PD98059 increased caspase 3 activity and resulted in cell death after Glu treatment in SCN2.2 cells. This effect was dependent on NMDA receptor activation. Glu treatment in the SCN2.2 cells resulted in sustained activation of the anti-apoptotic pERK/MAPK, without affecting the pro-apoptotic p-p38/MAPK. In contrast, Glu exposure in GT1-7 cells caused an increase in p-p38/MAPK and a decrease in pERK/MAPK. Bcl2-protein increased in SCN2.2 cells following Glu treatment, but not in GT1-7 cells; bid mRNA and cleaved-Bid protein increased in GT1-7, but not SCN2.2 cells. Conclusions Facilitation of ERK activation and inhibition of caspase activation promotes resistance to Glu excitotoxicity in SCN2.2 cells. Significance Further research will explore ERK/MAPK as a key molecule in the prevention of neurodegenerative processes.

Aryl Hydrocarbon Receptor Activation Attenuates Per1 Gene Induction and Influences Circadian Clock Resetting

Light-stimulated adjustment of the circadian clock is an important adaptive physiological response that allows maintenance of behavioral synchrony with solar time. Our previous studies indicate that the aryl hydrocarbon receptor (AhR) agonist 2,3,7,8- tetrachlorodibenzo-p-dioxin attenuates light-induced phase resetting in early night. However, the mechanism of inhibition remains unclear. In this study, we showed that another potent AhR agonist-β-naphthoflavone (BNF)-significantly decreased light-induced phase shifts in wild-type (WT) mice, whereas AhR knockout mice had an enhanced response to light that was unaffected by BNF. Mechanistically, BNF blocked light induction of the Per1 transcript in suprachiasmatic nucleus and liver in WT mice, and BNF blocked forskolin (FSK)-induced Per1 transcripts in Hepa-1c1c7 (c7) cells. An E-box decoy did not affect BNF inhibition of FSK-induced Per1 transcripts in c7 cells. cAMP-response element (CRE)-dependent induction of Per1 promoter activity in response to FSK in combination with phorbol 12-tetradecanoate 13-acetate was suppressed in cells that expressed high levels of AhR (c7) compared with cells lacking functional AhR activity (c12). In addition, the inhibitory effect of BNF on FSK-induced Per1 was dependent on phosphorylation of JNK. Together, these results suggest that AhR activation inhibits light-induced phase resetting through the activation of JNK, negative regulation of CREs in the Per1 promoter, and suppression of Per1.

Beta-naphthoflavone (DB06732) mediates estrogen receptor-positive breast cancer cell cycle arrest through AhR-dependent regulation of PI3K/AKT and MAPK/ERK signaling

Beta-naphthoflavone (BNF, DB06732) is an agonist of aryl hydrocarbon receptor (AhR) and a putative chemotherapeutic agent that has antitumor activity against mammary carcinomas in vivo. However, the mechanism by which BNF exerts this antitumor effect remains unclear. Thus, we explored mechanisms of BNFs antitumor effects in human breast cancer cells. This study showed that BNF suppressed cell proliferation and induced cell cycle arrest in the G0/G1 phase with downregulation of cyclin D1/D3 and CDK4 and upregulation of p21(Cip1/Waf1), leading to a senescence-like phenotype in estrogen receptor (ER)-positive MCF-7 cells, but not in ER-negative MDA-MB-231 cells. In addition, BNF inhibited PI3K/AKT signaling, and the PI3K inhibitor, LY294,002, exhibited the same inhibitory effects on cyclinD1/D3, CDK4 and the cell cycle as BNF. Interestingly, BNF activated mitogen-activated protein kinase-extracellular signal-regulated kinase (MAPK-ERK) signaling, and more notably, MEK inhibitor PD98059 significantly blocked the BNF-induced cell cycle arrest and upregulation of p21(Cip1/Waf1). Furthermore, specific ERα and AhR siRNA studies indicate that ERα is required in BNF-induced p21(Cip1/Waf1) expression, and BNF-mediated cell cycle arrest and modulation of AKT and ERK signaling is AhR-dependent. Taken together, AhR-dependent inhibition of the PI3K/AKT pathway, activation of MAPK/ERK and modulation of ERα is a novel mechanism underlying BNF-mediated antitumor effects in breast cancer, which may represent a promising strategy to be exploited in future clinical trials.

Suprachiasmatic nucleus neurons display endogenous resistance to excitotoxicity

A comprehensive understanding of neuroprotective pathways is essential to progress in the battle against numerous neurodegenerative conditions. The hypothalamic suprachiasmatic nucleus (SCN) is endogenously resistant to glutamate (Glu) excitotoxicity in vivo. This study was designed to determine whether immortalized SCN neurons (SCN2.2 cells) retain this characteristic. We first established that SCN2.2 cells retained the ability to respond to Glu. SCN2.2 cells expressed N-methyl-d-aspartate (NMDA) receptor subtypes NR1 and NR2A/2B, suggesting the presence of functional receptors. mRNA for the NMDA receptor subunits NR2A and NR2B were higher in the SCN2.2 than in the control hypothalamic neurons (GT1-7). Specific NMDA receptor antagonists (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate and d-(–)-2-amino-5-phosphonovaleric acid blocked Glu-induced activation of gene expression. SCN2.2 cells were resistant to Glu excitotoxicity compared with GT1-7 neurons as assessed with a mitochondrial function assay, cell death by trypan blue exclusion and apoptosis by terminal deoxynucleotidyl transferase dUTP nick end labeling. SCN2.2 resistance to Glu excitoxicity was retained in the presence of the broad spectrum Glu transport inhibitor, l-trans-pyrrolidine-2,4 dicarboxylate, excluding glial Glu uptake as a major neuroprotective mechanism. Collectively, these observations demonstrate endogenous neuroprotection in SCN2.2 cells; this cell line is resistant to excitotoxicity under conditions that are toxic to other immortalized cell lines. Thus, the SCN2.2 cell line may provide insights into the molecular mechanisms that confer endogenous neuroprotection in the SCN.

Suppression of Catatonia-Like Signs by Lorazepam in Nonconvulsive Status Epilepticus Without Seizure Termination

Nonconvulsive status epilepticus (NCSE) cannot be reliably distinguished from catatonia by signs or symptoms. The authors report on a 68-year-old man with endogenous major depression who displayed catatonia-like psychopathology, which temporarily disappeared with lorazepam. An EEG then revealed NCSE. Presumably, lorazepam suppressed seizure in areas where it had caused clouding of consciousness, but it did not suppress all seizure activity. When lorazepam was stopped, the catatonia-like delirium returned; it disappeared again with valproate administration. This case illustrates that the reduction of signs consistent with catatonia by benzodiazepines does not by itself confirm catatonia, even in patients with endogenous depression.

Transformations of reconstituted high-density lipoprotein subclasses as a function of temperature or LDL concentration

The objectives of this study were to determine the structural changes in defined, reconstituted high density lipoproteins (rHDL) resulting from spontaneous phospholipid depletion in the presence or absence of low-density lipoproteins (LDL), to establish the precursor-product relationships among the rHDL particles and to assess the differences in behavior of rHDL particles containing apo A-I or apo A-II. The rHDL particles were prepared by the sodium cholate dialysis method, and were incubated in buffer at 50 degrees C, or in buffer containing different concentrations of LDL at 37 degrees C, for up to 24 h. The changes in the rHDL particle distributions with time were followed by non-denaturing gradient gel electrophoresis, and the rHDL were isolated at various time points for chemical analysis. We found that rHDL particles containing apo A-I or apo A-II lose phospholipid and gain cholesterol when incubated with LDL. Increasing LDL concentrations remove increasingly larger amounts of phospholipid. With phospholipid loss the apo A-I containing particles undergo major structural rearrangements that give rise to 78 A and 106 A particles from 86 A and 94 A precursors. The 78 A products appear to be the most stable, lipid-poor species. Reconstituted HDL particles prepared with apo A-II (94 and 101 A in diameter) are more resistant to structural rearrangements than the apo A-I counterparts under similar reaction conditions.

Environmental factors act through aryl hydrocarbon receptor activation and circadian rhythm disruption to regulate energy metabolism

php and incorporate the Creative Commons Attribution – Non Commercial (unported, v3.0) License (http://creativecommons.org/licenses/by-nc/3.0/). By accessing the work you hereby accept the Terms. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed. For permission for commercial use of this work, please see paragraphs 4.2 and 5 of our Terms (https://www.dovepress.com/terms.php). Journal of Receptor, Ligand and Channel Research 2018:10 13–24 Journal of Receptor, Ligand and Channel Research Dovepress