Effects of Antioxidants in Human Cancers: Differential Effects on Non-Coding Intronic RNA Expression
Abstract
:1. Introduction
2. Materials and Methods
2.1. RNA-Seq Data Analysis
- Murine lung cancer dataset (Accession: E-GEOD-52594): RNA-Seq data were downloaded from ArrayExpress. The objective of the original study was to study the impact of Vitamin E and NAC supplementation in murine models of KRAS-induced lung cancer [27]. Antioxidants were administered 1 week after the induction of lung cancer, and the mice were euthanized 8 to 10 weeks later. There were 3 experimental groups (untreated, NAC-treated and Vitamin E-treated. Each group consisted of 5 animals, and from each animal two tumor samples were harvested for analysis. A total of 30 samples were profiled by RNA-Seq analyses.
- Human normal and prostate cancer cells dataset (SRP027258): RNA-Seq data were downloaded from NCBI SRA. Normal prostate epithelial cells and androgen-dependent and androgen-independent prostate cancer cells were treated with 15 µM sulforaphane (SFN), a phytochemical derived from cruciferous vegetables, and the transcriptome was determined at 6 and 24 h time points [28].
Antioxidant Supplements Studied | Organism | NCBI/Arrayexpress Accession | Brief Description of The Original Study |
---|---|---|---|
Vitamin E, N-acetyl cysteine (NAC) | Mus musculus (mouse) | E-GEOD-52594 | The original study [27] found that both antioxidants increase tumor cell proliferation by reducing ROS, DNA damage, and p53 expression. |
Sulforaphane (SFN) | Human prostate normal and cancer cell lines | SRP027258 | SFN influenced the expression of genes in functional groups and pathways that are critical in cancer including cell cycle, apoptosis and angiogenesis, but the specific effects of SFN differed depending on the state of cancer progression [28]. |
2.2. Intronic Count Analysis
2.3. In Vitro Analysis on the Effect of Antioxidant Supplements on Superoxide Dismutase Activity on Established Human Cell Lines
2.4. RNA-Isolation and RT-PCR
3. Results
3.1. Murine Lung Cancer Dataset
a. Summary of Results from Murine Model for KRAS-Induced Lung Cancer Data Analysis | ||||||||||
---|---|---|---|---|---|---|---|---|---|---|
Total Number of Genes with Significant Differences in the Intronic Read Counts from Antioxidant Supplement (NAC, Vitamin E) Treated Tumor Tissues Compared to Control Tumor Tissue | ||||||||||
NAC | Vitamin E | |||||||||
Down | Up | Down | Up | |||||||
Number of genes | 459 | 86 | 1143 | 315 | ||||||
b. Summary of Results from Human Prostate Cancer Cells Data Analysis | ||||||||||
Total Number of Genes with Significant Differences in the Intronic Read Counts in Sulforaphane Treated Cells Compared to Untreated Cells | ||||||||||
Normal Prostate Epithelial Cells | LNCAP (Prostate Cancer Cells, Hormone, Dependent) | PC3 (Prostate Cancer Cells, Hormone Independent) | ||||||||
Down | Up | Down | Up | Down | Up | |||||
6 h | 1680 | 1971 | 428 | 575 | 569 | 707 | ||||
24 h | 1522 | 1437 | 1313 | 476 | 496 | 457 |
3.2. Functional Annotation of Genes with Differentially Expressed Intronic RNA
Gene Symbols | ||||
---|---|---|---|---|
ALDOC | F7 * | IL18 | MRC1 * | SOCS3 |
AREG | FCGR2B | ITGAX * | MSR1 | STARD10 |
AXL | GAPDH | ITGB2 | PTGS1 | TNFSF9 |
CAMSAP1 | GJA1 | ITIH4 * | ROS1 * | TYROBP |
CD68 | HDC | KRAS | RPL3 | |
CRLF1 | HK1 * | LCP1 | SERPINE1 | |
CTSK | HK2 * | LRG1 * | SH3RF1 | |
ELL2 | HSPA1B | LRP2 * | SIRPA | |
EPHA7 | HSPA8 | ME1 | SLAIN1 | |
F10 | HSPH1 | MMP12 | SLC38A2 |
For Genes with Increased Expression of Introns |
---|
Fructose and mannose metabolism |
Ensemble of genes encoding ECM-associated proteins including ECM-affiliated proteins, ECM regulators and secreted factors |
Ensemble of genes encoding extracellular matrix and extracellular matrix-associated proteins |
Genes involved in Transmembrane transport of small molecules |
Type II diabetes mellitus |
For Genes with Decreased Expression of Introns |
Drug metabolism—cytochrome P450 |
Genes involved in Biological oxidations |
Genes involved in Muscle contraction |
Metabolism of xenobiotics by cytochrome P450 |
Glutathione metabolism |
Enriched Canonical Pathways |
---|
For genes with Increased Expression of Introns |
Ensemble of genes encoding extracellular matrix and extracellular matrix-associated proteins |
Genes involved in Adaptive Immune System |
Beta2 integrin cell surface interactions |
Ensemble of genes encoding ECM-associated proteins including ECM-affiliated proteins, ECM regulators and secreted factors |
Genes involved in Transmembrane transport of small molecules |
For Genes with Decreased Expression of Introns |
Drug metabolism—cytochrome P450 |
Genes involved in Biological oxidations |
Metabolism of xenobiotics by cytochrome P450 |
Ensemble of genes encoding extracellular matrix and extracellular matrix-associated proteins |
Genes involved in Phase 1—Functionalization of compounds |
3.3. Human Normal and Prostate Cancer Cells Dataset
Enriched Canonical Pathways |
---|
Normal Prostate Epithelial Cells with 6 h of SFN Treatment |
Ensemble of genes encoding extracellular matrix and extracellular matrix-associated proteins |
Genes involved in Developmental Biology |
Genes involved in Transmission across Chemical Synapses |
Genes involved in Immune System |
Genes involved in Axon guidance |
Normal Prostate Epithelial Cells with 24 h of SFN Treatment |
Genes involved in Transmembrane transport of small molecules |
Arrhythmogenic right ventricular cardiomyopathy (ARVC) |
Focal adhesion |
Genes involved in Immune System |
Genes involved in Developmental Biology |
LNCAP Prostate Cancer Cells with 6 h of SFN Treatment |
Caspase cascade in apoptosis |
Endocytosis |
Genes involved in Developmental Biology |
Peroxisome |
Genes involved in Signaling by Rho GTPases |
LNCAP Prostate Cancer cells with 24 h of SFN Treatment |
Genes involved in Collagen formation |
Genes involved in Developmental Biology |
Genes involved in Neuronal System |
Genes involved in Metabolism of lipids and lipoproteins |
Genes involved in Extracellular matrix organization |
PC3 Prostate Cancer Cells with 6 h of SFN Treatment |
Genes involved in Collagen formation |
Genes involved in Extracellular matrix organization |
Regulation of RhoA activity |
Genes involved in Metabolism of lipids and lipoproteins |
Ensemble of genes encoding extracellular matrix and extracellular matrix-associated proteins |
PC3 Prostate Cancer Cells with 24 h of SFN Treatment |
Genes involved in Axon guidance |
Genes involved in Immune System |
Genes involved in NRAGE signals death through JNK |
Genes involved in Signaling by Rho GTPases |
Genes involved in Signaling by NGF |
3.4. Validation of Intronic RNA in DLK1 and LRG1
4. Discussion
5. Conclusions
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Menon, S.; Lu, C.; Menon, R.; Schwartz, J.; Guan, Y. Effects of Antioxidants in Human Cancers: Differential Effects on Non-Coding Intronic RNA Expression. Antioxidants 2016, 5, 1. https://doi.org/10.3390/antiox5010001
Menon S, Lu C, Menon R, Schwartz J, Guan Y. Effects of Antioxidants in Human Cancers: Differential Effects on Non-Coding Intronic RNA Expression. Antioxidants. 2016; 5(1):1. https://doi.org/10.3390/antiox5010001
Chicago/Turabian StyleMenon, Shreya, Chunxia Lu, Rajasree Menon, Jessica Schwartz, and Yuanfang Guan. 2016. "Effects of Antioxidants in Human Cancers: Differential Effects on Non-Coding Intronic RNA Expression" Antioxidants 5, no. 1: 1. https://doi.org/10.3390/antiox5010001
APA StyleMenon, S., Lu, C., Menon, R., Schwartz, J., & Guan, Y. (2016). Effects of Antioxidants in Human Cancers: Differential Effects on Non-Coding Intronic RNA Expression. Antioxidants, 5(1), 1. https://doi.org/10.3390/antiox5010001