Effects of sodium saccharin and linoleic acid on mRNA levels of Her2/neu and p53 in a human breast epithelial cell line

doi:10.1016/0304-3835(96)04170-5

Cancer Letters

Volume 102, Issues 1–2, 19 April 1996, Pages 91-99

https://doi.org/10.1016/0304-3835(96)04170-5 Get rights and content

Abstract

The effects of two food-related chemicals (sodium saccharin and linoleic acid) on the levels of Her2/neu and p53 mRNA in a non-cancerous human breast epithelial cell line (HBL-100) were tested in comparison with the effects of the known tumor promoter phorbol 12-myristate 13-acetate (TPA). Treatments were made both with and without prior treatment with two well-known tumor initiators, N-nitroso-N-methylurea (NMU) or 7,12-dimethylbenz[a]anthracene (DMBA). The effects in general were small, the greatest being increases of 46–67% in Her2/neu mRNA levels in response to treatments with TPA or sodium saccharin following NMU treatments. These results demonstrate that sodium saccharin following NMU treatments might be involved in transcriptional regulation of Her2/neu in HBL-100 cells and suggest that its effects may not be limited to urinary bladder.

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These results imply that saccharin may affect functions of the molecules involved in neurite elongation. It has been reported that saccharin increases expression of several genes such as Her2/neu (Ogretmen et al., 1996) and proliferin (Parfett, 2003). Saccharin enhances mRNA expression by simultaneous treatment with agents such as phorbol 12-myristate 13-acetate.
In the present study, we found that saccharin, an artificial calorie-free sweetener, promotes neurite extension in the cultured neuronal cells. The purposes of this study are to characterize the effect of saccharine on neurite extension and to determine how saccharin enhances neurite extension.
The analyses were performed using mouse neuroblastoma N1E-115 cells and rat pheochromocytoma PC12 cells. Neurite extension was evaluated by counting the cells bearing neurites and measuring the length of neurites. Formation, severing and transportation of the microtubules were evaluated by immunostaining and western blotting analysis.
Deprivation of glucose increased the number of N1E-115 cells bearing long processes. And the effect was inhibited by addition of glucose. Saccharin increased the number of these cells bearing long processes in a dose-dependent manner and total neurite length and longest neurite length in each cell. Saccharin also had a similar effect on NGF-treated PC12 cells. Saccharin increased the amount of the microtubules reconstructed after treatment with nocodazole, a disruptor of microtubules. The effect of saccharin on microtubule reconstruction was not influenced by dihydrocytochalasin B, an inhibitor of actin polymerization, indicating that saccharin enhances microtubule formation without requiring actin dynamics. In the cells treated with vinblastine, an inhibitor of microtubule polymerization, after microtubule reorganization, filamentous microtubules were observed more distantly from the centrosome in saccharin-treated cells, indicating that saccharin enhances microtubule severing and/or transportation.
These results suggest that saccharin enhances neurite extension by promoting microtubule organization.
Molecular Mechanisms of Ceramide-mediated Telomerase Inhibition in the A549 Human Lung Adenocarcinoma Cell Line
2001, Journal of Biological Chemistry
Citation Excerpt :
The telomerase activity in each sample was quantitated by measuring the ratio of the 36-base pair internal standard to the extended telomerase products as described by the manufacturer using ChemiImager (Alpha Innotech Corp., San Leandro, CA). The mRNA levels of hTERT and hTR were analyzed by RT-PCR and Northern blotting as described previously (40, 41). One µg of total RNA, isolated using a RNA isolation kit (Qiagen), was used in reverse transcription reactions as described by the manufacturer.
This study was aimed at identifying the molecular mechanisms by which ceramide inhibits telomerase activity in the A549 human lung adenocarcinoma cell line. C₆-ceramide (20 µm) caused a significant reduction of telomerase activity at 24 h as detected using the telomeric repeat amplification protocol, and this inhibition correlated with decreased telomerase reverse transcriptase (hTERT) protein. Semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and Northern blot analyses showed that C₆-ceramide significantly decreased hTERT mRNA in a time-dependent manner. Electrophoretic mobility shift and supershift assays demonstrated that the binding activity of c-Myc transcription factor to the E-box sequence on the hTERT promoter was inhibited in response to C₆-ceramide at 24 h. These results were also confirmed by transient transfections of A549 cells with pGL3-Basic plasmid constructs containing the functional hTERT promoter and its E-box deleted sequences cloned upstream of a luciferase reporter gene. Further analysis using RT-PCR and Western blotting showed that c-Myc protein but not its mRNA levels were decreased in response to C₆-ceramide at 24 h. The effects of ceramide on the c-Myc protein were shown to be due to a reduction in half-life via increased ubiquitination. Similar results were obtained by increased endogenous ceramide levels in response to nontoxic concentrations of daunorubicin, resulting in the inhibition of telomerase and c-Myc activities. Furthermore, the elevation of endogenous ceramide by overexpression of bacterial sphingomyelinase after transient transfections also induced the inhibition of telomerase activity with concomitant decreased hTERT and c-Myc protein levels. Taken together, these results show for the first time that both exogenous and endogenous ceramides mediate the modulation of telomerase activity via decreased hTERT promoter activity caused by rapid proteolysis of the ubiquitin-conjugated c-Myc transcription factor.
Capsaicin can alter the expression of tumor forming-related genes which might be followed by induction of apoptosis of a Korean stomach cancer cell line, SNU-1
1997, Cancer Letters
Capsaicin (CAP) has been known to inhibit some tumor development in vivo (J.J. Jang, S.H. Kim, T.K. Yun, Inhibitory effect of capsaicin on mouse lung tumor development, in vivo, J. Korean Med. Sci. 3 (1989) 49–53; J.J. Jang, K.J. Cho, Y.S. Lee, J.H. Bae, Different modifying responses of capsaicin in a wide-spectrum initiation model of F344 rat, J. Korean Med. 6 (1991) 31–36) 1, 2even though its mechanism of action is not well understood. The objectives of this study were to examine the effect of CAP on expression of tumor forming-related genes in a Korean stomach tumor cell, SNU-1. We used slot blot hybridization to investigate its effect on a wide spectrum of proto-oncogenes. It was found that CAP enhanced the transcripts of two proto-oncogenes (c-myc and c-Ha-ras) and tumor suppressor gene p53. While a low concentration of CAP (0.01 μM) did not significantly increase the level of p53 transcript in SNU-1, it did increase it by a factor of 3.5 at a 10 μM dose of CAP. Consequently, SNU-1 cells are sensitive to CAP in the overexpression of tumor suppressor gene, p53 and proto-oncogenes, c-myc and c-Ha-ras, but not those of c-erbB-2, c-jun and bcl-2 genes. Both cell death and DNA fragmentation were shown in SNU-1 cells with treatment of CAP. Our results suggest that CAP induces apoptotic cell death in human gastric cancer cells (SNU-1) in vitro which may be possibly mediated by the overexpression of p53 and/or c-myc genes. Because cell suicide is arguably the most potent natural defense against cancer, the correlation between the induction of apoptosis and the change of tumor forming-related gene expression after CAP treatment should be further studied in detail.
De-regulation of GRP stress protein expression in human breast cancer cell lines
1999, Breast Cancer Research and Treatment

¹: Present address: The University of Chicago, Department of Medicine, Section of Hematology/Oncology, Chicago, IL 60637, USA.

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Effects of sodium saccharin and linoleic acid on mRNA levels of Her2/neu and p53 in a human breast epithelial cell line

Abstract

Biochim. Biophys. Acta

Am. J. Clin. Nutr.

Biochem. Biophys. Res. Commun.

Regul. Toxicol. Pharmacol.

Stimulation of c-myc oncogene expression associated with estrogen induced proliferation of human breast cancer cells

Cancer Res.

Oncogenes, hormones and free radical processes in malignant transformation in vitro

Nature

Studies of the Her2/neu protooncogene in human breast and ovarian cancer

Science

Oncogenes in growth and development

FASEB J.

The neu oncogene: an erb-B-related gene encoding a 185,000-Mr tumour antigen

Nature

p53 mutations in human cancers

Science

p53: a frequent target for genetic abnormalities in lung cancer

Science

The cellular oncogene p53 can be activated by mutagenesis

Nature

Oncogenic forms of p53 inhibit p53-regulated gene expression

Science

Mechanisms of p53 loss in human sarcomas

Increased expression of the putative growth factor receptor p185Her2 causes transformation and tumorigenesis of NIH 3T3 cells

Her2/neu oncogene expression and proliferation in breast cancers

Am. J. Pathol.

Specificity of protooncogene amplification in human malignant diseases

Mol. Biol. Med.

Prognostic significance of erbB-2 oncogene activation in human tumours

Exp. Oncol.

Toward the primary prevention of cancer

Science

Initiation and promotion at different ages and doses in 2200 mice

Br. J. Cancer

Prostaglandins and cancer: a review of tumor initiation through tumor metastasis

Prostaglandins

Dose response of sodium saccharin in induction of urinary bladder hyperplasias in Fischer 344 rats pretreated with BBN

J. Natl. Cancer Inst.

Casarett and Doull's Toxicology

Dietary energy and fat effects on tumor promotion

Cancer Res.

Dietary fat and human cancer

AntiCancer