Pyridoxine (VB6 ) restores the down-regulation of serine palmitoyltransferase mRNA expression in keratinocytes cultured in highly oxidative conditions through enhancement of the intracellular antioxidant system

This study aimed to investigate the anti-cancer effect of lobetyolin on breast cancer cells. Lobetyolin was incubated with MDA-MB-231 and MDA-MB-468 breast cancer cells for 24 h. Glucose uptake and the mRNA expression of GLUT4 ( SLC2A4), HK2 and PKM2 were detected to assess the effect of lobetyolin on glucose metabolism. Glutamine uptake and the mRNA expression of ASCT2 ( SLC1A5), GLS1, GDH and GLUL were measured to assess the effect of lobetyolin on glutamine metabolism. Annexin V/PI double staining and Hoechst 33342 staining were used to investigate the effect of lobetyolin on cell apoptosis. Immunoblot was employed to estimate the effect of lobetyolin on the expression of proliferation-related markers and apoptosis-related markers. SLC1A5 knockdown with specific siRNA was performed to study the role of ASCT2 played in the anti-cancer effect of lobetyolin on MDA-MB-231 and MDA-MB-468 breast cancer cells. C-MYC knockdown with specific siRNA was performed to study the role of c-Myc played in lobetyolin-induced ASCT2 down-regulation. Myr-AKT overexpression was performed to investigate the role of AKT/GSK3β signaling played in lobetyolin-induced down-regulation of c-Myc and ASCT2. The results showed that lobetyolin inhibited the proliferation of both MDA-MB-231 and MDA-MB-468 breast cancer cells. Lobetyolin disrupted glutamine uptake via down-regulating ASCT2. SLC1A5 knockdown attenuated the anti-cancer effect of lobetyolin. C-MYC knockdown attenuated lobetyolin-caused down-regulation of ASCT2 and Myr-AKT overexpression reversed lobetyolin-caused down-regulation of both c-Myc and ASCT2. In conclusion, the present work suggested that lobetyolin exerted anti-cancer effect via ASCT2 down-regulation-induced apoptosis in breast cancer cells.

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Lower antioxidant capacity in the prefrontal cortex of individuals with schizophrenia

Australian & New Zealand Journal of Psychiatry ◽

10.1177/0004867417728805 ◽

2017 ◽

Vol 52 (7) ◽

pp. 690-698 ◽

Cited By ~ 8

Author(s):

Yiru Zhang ◽

Vibeke Sørensen Catts ◽

Cynthia Shannon Weickert

Keyword(s):

Prefrontal Cortex ◽

Antioxidant Capacity ◽

Mrna Expression ◽

Antioxidant System ◽

Messenger Rna ◽

Total Glutathione ◽

Protein Levels ◽

Significant Difference ◽

Dorsolateral Prefrontal ◽

And Control

Objective: The glutathione (GSH) pathway is the main antioxidant system to protect against oxidative stress in the human brain. In this study, we tested whether molecular components of the GSH antioxidant system are changed in dorsolateral prefrontal cortex tissue from people with schizophrenia compared to controls. Method: The levels of total glutathione and reduced GSH were determined by fluorometric assay via quantifying thiols in extracts from frontal cortex of 68 people. Immunoblotting was used to measure levels of enzymes responsible for maintaining GSH, the glutamyl-cysteine ligase (GCL) catalytic subunit (GCLC) and the GSH peroxidase (GPx)-like protein ( n = 74). Quantitative reverse transcription polymerase chain reaction (RT-PCR) was used to measure GCLC messenger RNA (mRNA) expression. Results: Both total glutathione ( t(66) = 2.467, p = 0.016) and reduced GSH ( t(66) = 3.001, p = 0.004) levels were significantly less in people with schizophrenia than in controls. However, there were no significant differences in either GCLC-like protein ( t(72) = −1.077, p = 0.285) or GCLC mRNA expression ( t(71) = −0.376, p = 0.708) between people with schizophrenia and control subjects. There was also no significant difference of GPx-like protein levels between schizophrenia and controls ( t(72) = −0.060, p = 0.952). Moreover, no significant correlations of putative confounding factors with GSH changes were detected. Discussion: These results suggest that people with schizophrenia have impaired GSH antioxidant capacity, alongside normal levels of key regulatory proteins.

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Lysophosphatidylcholine Decreases the Synthesis of Tissue Factor Pathway Inhibitor in Human Umbilical Vein Endothelial Cells

Thrombosis and Haemostasis ◽

10.1055/s-0037-1614242 ◽

1998 ◽

Vol 79 (01) ◽

pp. 217-221 ◽

Cited By ~ 13

Author(s):

Koichi Kokame ◽

Toshiyuki Miyata ◽

Naoaki Sato ◽

Hisao Kato

Keyword(s):

Endothelial Cells ◽

Mrna Expression ◽

Tissue Factor ◽

Tissue Factor Pathway Inhibitor ◽

Umbilical Vein ◽

Mrna Levels ◽

Down Regulation ◽

Human Umbilical Vein ◽

Tissue Factor Pathway ◽

Umbilical Vein Endothelial Cells

SummaryThrombotic complications are frequently associated with atherosclerosis. Lysophosphatidylcholine (LPC), a component accumulated in oxidatively modified LDL (ox-LDL), is known to play a crucial role in the initiation and progression of atherosclerotic vascular lesions. Since a vascular anticoagulant, tissue factor pathway inhibitor (TFPI), has the function of regulating the initial reaction of tissue factor (TF)-induced coagulation, we investigated the effect of LPC on TFPI synthesis in cultured human umbilical vein endothelial cells (HUVEC). The treatment of HUVEC with LPC for 24 h decreased TFPI antigen levels in both the culture medium and the cell lysate in a dose-dependent manner. Northern blot analysis revealed that LPC caused a time-dependent decrease in the TFPI mRNA levels. The levels of TFPI antigen and mRNA were decreased to 72% and 38%, respectively, by the incubation with 50 μM LPC for 24 h. The down-regulation by LPC of TFPI mRNA expression was not observed in the presence of cycloheximide, suggesting that protein synthesis was involved in the suppression of TFPI mRNA expression. The TFPI mRNA levels in actinomycin D-treated cells were relatively stable, indicating that the down-regulation of TFPI mRNA by LPC would be partly explained by the enhanced mRNA destabilization. In contrast to the significant down-regulatory effects of LPC on TFPI expression, LPC did not induce TF mRNA expression in HUVEC. These results indicate that LPC accumulated in the atherosclerotic vascular wall would suppress endothelial TFPI synthesis, reducing the antithrombotic property of endothelial cells.

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Parallel Down-Regulation of FOXO1, PPAR? and Adiponectin mRNA Expression in Visceral Adipose Tissue of Class III Obese Individuals

Obesity Facts ◽

10.1159/000339574 ◽

2012 ◽

Vol 5 (3) ◽

pp. 452-459 ◽

Cited By ~ 11

Author(s):

Thais Ortiz Hammes ◽

Cíntia dos Santos Costa ◽

Francieli Rohden ◽

Rogério Margis ◽

Jussara Carnevale de Almeida ◽

...

Keyword(s):

Adipose Tissue ◽

Mrna Expression ◽

Visceral Adipose Tissue ◽

Class Iii ◽

Down Regulation ◽

Adiponectin Mrna ◽

Visceral Adipose

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Aberrant CpG Island Hypermethylation and Down-Regulation of Oct-6 mRNA Expression in Human Hepatocellular Carcinoma

Digestive Diseases and Sciences ◽

10.1007/s10620-011-1686-y ◽

2011 ◽

Vol 56 (10) ◽

pp. 3072-3077 ◽

Cited By ~ 2

Author(s):

Jing-zhe Sun ◽

Xue-xi Yang ◽

Xiang-hong Li ◽

Wei-wen Xu ◽

Ying Wang ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Mrna Expression ◽

Cpg Island ◽

Human Hepatocellular Carcinoma ◽

Down Regulation

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The Regulation of Telomere Binding Proteins on Telomerase during Differentiation of ATRA Responsive and Resistant NB4 Cells.

Blood ◽

10.1182/blood.v104.11.3390.3390 ◽

2004 ◽

Vol 104 (11) ◽

pp. 3390-3390

Author(s):

He Huang ◽

Jie Sun ◽

Yuan Yuan Zhu ◽

Jian Ping Lan ◽

Xiao Yu Lai

Keyword(s):

Protein Expression ◽

Mrna Expression ◽

Peak Level ◽

Telomerase Activity ◽

Expression Level ◽

Htert Mrna ◽

Down Regulation ◽

Nb4 Cells ◽

Significant Change ◽

Htert Mrna Expression

Abstract It has been reported that the down-regulation of telomerase activity associated with maturation of APL cells is not mechanistically linked to cell maturation, and requires only a RAR, but not a RXR-dependent pathway. However, it is not clear whether and how telomeric proteins respond to the retinoid treatment. Using maturation-sensitive and resistant APL cell lines NB4, NB4-R1 and NB4-R2 cells, we analyzed a panel of telomeric proteins using western blotting analyses in addition to temporal profile of corresponding mRNA during the course of retinoid-induced differentiation. Our analyses show hTERTmRNA expression decreased rapidly during differentiation of NB4 and NB4-R1 cells, telomerase activity also declined. But in NB4-R2 cells, hTERT mRNA was initially decreased to 38.2% on day1 (P<0.05) and then increased to 80.0% on day 3 (P<0.05). Telomerase activity remained unchanged overtime (P>0.05), which may be caused by the increasing of hTERT mRNA expression during its later period of differentiation. TRF1 mRNA and protein expression have no significant change during differentiation in NB4 and NB4-R1 cells but has a small increase in NB4-R2 cells. The TRF1 mRNA expression level has no significant change during differentiation of NB4 and NB4-R1 cell line cells. However, it was increased to 235% on day 2 (P<0.05) and remains at this level until day 3 during the differentiation of NB4-R2 cells. TRF1 protein expression level also remains stable during differentiation of NB4 and NB4-R1 cells, but has a little increase in NB4-R2 cells. This indicates TRF1 has different regulation in RARα dependent or RXRα dependent pathways. Pinx1 mRNA expression decreased during the differentiation of NB4 and NB4-R1. But during the differentiation of NB4-R2, Pinx1mRNA expressions level was initially decreased to 34.3% (P<0.05) on day 1 then increased to 64.5% (P<0.05 compared to day1) on day2. The change of Pinx1 mRNA expression and hTERT mRNA expression in NB4(r=0.902, P=0.036), NB4-R1(r=1.00, P<0.001), and NB4-R2(r=0.880, P=0.049) cells are positive correlated. Pinx1 is the only telomere binding protein that can bind to hTERT directly, it might be responsible for the different regulation of telomerase activity through RARα dependent or RXRα dependent pathways. During NB4 cell differentiation, TANK1 mRNA expression decreased gradually to 31.6% (P<0.05) on day1, and remained this level until day3. In NB4-R1 cells, TANK1 mRNA expression increased initially to 197% at 12h(P<0.05), and then decreased gradually to 111% (P<0.05) on day 3. During the differentiation of NB4-R2 cells, TANK1 mRNA expression was initially increased to 204% at 12h(P<0.01), and then decreased gradually to 96.9% on day3 (P<0.01). Its protein expression initially increased and reached a peak level at day 1 and then decreased in the later period of differentiation of all three NB4 cells. Both TANK1 mRNA expression and its protein expression were down-regulated at the later period of differentiation in all three NB4 cells. It seems that TANK1 may act as a positive regulator on telomerase activity during differentiation. TANK2 mRNA expression remained no change during differentiation of three NB4 cells. As results show, Pinx1 and TANK1 may interfere in the regulation of telomerase. The decrease of TANK1 may be the cause of the down-regulation of telomerase activity. Further studies will focus on the mechanism of their regulation on telomerase.

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Integrating microRNA and mRNA expression profiling using a novel algorithm identified a small set of unique genes upregulated in malignant pleural mesothelioma (MPM)

Journal of Clinical Oncology ◽

10.1200/jco.2009.27.15_suppl.e22111 ◽

2009 ◽

Vol 27 (15_suppl) ◽

pp. e22111-e22111

Author(s):

M. Suraokar ◽

A. Corvalan ◽

C. Chow ◽

A. Gazdar ◽

C. Moran ◽

...

Keyword(s):

Mrna Expression ◽

Cell Lines ◽

Expression Profiling ◽

Tumor Tissue ◽

Control Cell ◽

Seed Region ◽

Down Regulation ◽

Total Rna ◽

Tissue Specimens ◽

Unique Genes

e22111 Background: We employed a global profiling strategy using miRNA microarrays in MPM cell lines and archival tumor tissue. Methods: We isolated total RNA from 4 MPM cell lines, 2 control cell lines, and 16 tissue specimens from patients with resected MPM (n=8) and normal counterpart (n=8) patients as controls. Total RNA was labeled with Cyanine 3, then hybridized with Agilent human miRNA microarray v1 slides. Results: Preliminary miRNA profiles show up-regulation of 44 versus down-regulation of 29 miRNA's in MSTO-211H cancer cells compared to HCT-4012 (pleural telomerase-transformed control). Profiling of 16 tissue specimens (8 normal vs 8 MPM) revealed down-regulation of 11 miRNA's in MPM tumor tissue. To focus on relevant miRNA that regulate genes involved in carcinogenesis and progression, we identified > 1000 unique genes using the online targetscan 4.2 program ( http://www.targetscan.org ), which predicts biological targets of miRNAs by identifying the presence of conserved 8-mer and 7-mer sites that match the seed region of each miRNA. We then explored a novel screening strategy, which combines mRNA expression dataset with the miRNA dataset, to narrow the list of relevant miRNA's. We conducted gene expression profiling on the cell lines and MPM tissue samples with Affymetrix U133 plus 2.0 chips. Bioinformatic analysis was conducted with MeV: MultiExperiment Viewer software, data reduction techniques (Correspondance Analysis), hierarchical clustering methods, and Serial Analysis for Microarray (SAM), and showed up-regulation of ∼300 genes in MPM compared to normal tissues. We then computed of the ∼300 mRNA's up-regulated in MPM only 32 are recognized by the 11 down-regulated miRNA's using the targetscan 4.2 algorithm. Most of the miRNA's regulate single messages while ∼20 % of the messages are regulated by more than 1 miRNA's. Some of these targets include Ets variant 1 and Protein kinase C - epsilon. Conclusions: This innovative approach of selecting highly relevant miRNA is feasible and enables discovery of novel genes based on their ability to be bound by single or multiple miRNA's. Validation of our profiling studies using real-time PCR and protein analysis methods will be presented. No significant financial relationships to disclose.

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Aminoglutethimide suppresses adrenocorticotropin receptor expression in the NCI-h295 adrenocortical tumor cell line

Journal of Endocrinology ◽

10.1677/joe.0.1590035 ◽

1998 ◽

Vol 159 (1) ◽

pp. 35-42 ◽

Cited By ~ 18

Author(s):

M Fassnacht ◽

F Beuschlein ◽

S Vay ◽

P Mora ◽

B Allolio ◽

...

Keyword(s):

Mrna Expression ◽

Cell Line ◽

Cdna Probe ◽

Tumor Cell Line ◽

Receptor Expression ◽

Intracellular Camp ◽

Camp Accumulation ◽

Transcript Accumulation ◽

Down Regulation ◽

Stimulation Of

The adrenostatic compound aminoglutethimide (AG), a potent inhibitor of the P450 side chain cleavage enzyme, is used in the treatment of ACTH-dependent or adrenal Cushing's syndrome. Recently, AG has been shown to inhibit ACTH receptor (ACTH-R) mRNA expression in ovine adrenocortical cells in a time-dependent fashion. To investigate whether ACTH-R down-regulation will also be induced in tumor cells, we studied the effect of AG on ACTH-R expression in the human NCI-h295 adrenocortical carcinoma cell line, which expresses functional ACTH receptors and produces steroids of the glucocorticoid, mineralocorticoid and androgen pathway. The cells were incubated in triplicate with increasing doses of AG (3, 30, 300 microM) which suppressed steroid secretion dose-dependently. After 48 h, cells were harvested, and total RNA was extracted, electrophoresed, blotted and hybridized with a human ACTH-R cDNA probe. In parallel experiments, after preincubation with AG the cells were stimulated with ACTH (10 nM) for 10 min and the intracellular cAMP accumulation was determined by RIA. AG significantly suppressed the baseline ACTH-R mRNA expression in a dose-dependent fashion (300 microM AG, 5+/-1%; 30 microM AG, 64+/-1%; 3 microM AG, 108+/-19% compared with control cells, 100+/-11%). The reduced ACTH-R mRNA expression was paralleled by low ACTH-induced cAMP accumulation indicating reduced expression of the ACTH-R protein. The adrenostatic compound metyrapone, an inhibitor of 11beta-hydroxylase activity, also suppressed ACTH-R mRNA expression in a similar fashion. Stimulation of the protein kinase A pathway by simultaneous incubation of ACTH (10 nM) or forskolin (10 microM) together with AG was not able to overcome the steroid biosynthesis blockade, but reversed the inhibitory effects of AG on the ACTH-R mRNA expression. Also, cortisol (12 microM) reversed the AG-induced ACTH-R mRNA expression. We conclude that AG induces profound ACTH-R down-regulation in the NCI-h295 cell line either by affecting the gene expression or by decreasing transcript accumulation via an effect on RNA stability. This novel action of AG can be reversed by stimulation of the cAMP pathway and of the glucocorticoid-mediated signal transduction cascade. As the down-regulation occurs in vitro at concentrations which are reached during treatment with AG in humans it may contribute to its therapeutic activity in adrenal disease.

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Inhibition of Uncoupling Protein Expression during Lactation: Role of Leptin

Endocrinology ◽

10.1210/en.2003-0836 ◽

2004 ◽

Vol 145 (2) ◽

pp. 830-838 ◽

Cited By ~ 48

Author(s):

Xiao Qiu Xiao ◽

Kevin L. Grove ◽

Bernadette E. Grayson ◽

M. Susan Smith

Keyword(s):

Skeletal Muscle ◽

Protein Expression ◽

Mrna Expression ◽

Uncoupling Protein ◽

Atp Synthesis ◽

Negative Energy ◽

Down Regulation ◽

Serum Free Fatty Acid ◽

Serum Leptin ◽

Brown Adipose

Abstract Uncoupling proteins (UCPs) are mitochondrial proteins that play a role in regulation of energy expenditure by uncoupling respiration from ATP synthesis. Lactation is a physiological condition characterized by negative energy balance due to the loss of energy sources to the production of milk. The objective of the current study was to investigate whether UCP mRNA and protein expressions were altered during lactation compared with those after 48 h of fasting. Lactation significantly reduced serum leptin levels, and removal of pups for 48 h increased serum leptin to higher levels than those observed in control rats. Compared with control rats, mRNA expression of UCP1 and UCP3 in brown adipose tissue (BAT) was dramatically reduced during lactation and fasting. The reduction in mRNAs was reflected by a lowered UCP1 protein level, and to some extent, UCP3 protein. Treatment of lactating rats with exogenous leptin (3 mg/kg) or removal of pups for 48 h completely reversed the down-regulation of UCP1 and UCP3 mRNA expression in BAT, and pup removal led to a recovery of protein expression. In contrast to BAT, UCP3 expression in skeletal muscle was increased in fasted rats and decreased during lactation. Similar changes were observed in serum free fatty acid levels. These changes are consistent with the idea that the utilization of free fatty acids as a fuel source is spared during lactation. As in BAT, leptin treatment and removal of pups were able to restore changes in mRNA expression of UCP3 in skeletal muscle during lactation. The present results suggest that the inhibition of leptin secretion during lactation is involved in the down-regulation of UCP expression in BAT and skeletal muscle, which, in turn, is responsible for the decrease in metabolic fuel oxidation and thermogenesis.

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