Tissue-specific patterns of gene expression in the epithelium and stroma of normal colon in healthy individuals in an aspirin intervention trial

BMC Medical Genetics

John D Potter³  Johanna W Lampe³  Jeannette Bigler⁴  Sanford D Markowitz¹  Min Yan¹  Paul D Lampe³  Rebecca Yvonne Rudolph³  Lisa Levy³  Peiying Yang²  Yingye Zheng³  Lin Li³  Karen W Makar³  Sushma S Thomas³ 

[1] Case Western Reserve University School of Medicine, Cincinnati 44106, OH, USA;M.D. Anderson Cancer Center, Houston 77030, TX, USA;Fred Hutchinson Cancer Research Center, Seattle 98109, WA, USA;Amgen Corporation, Seattle 98119, WA, USA

关键词: Aspirin;    UGT1A6;    Colon biopsy;    Microarray;    Colon epithelium;    Colon stroma;    Gene expression;   

Others  :  1177687
DOI  :  10.1186/s12881-015-0161-6

 received in 2014-10-08, accepted in 2015-02-26,  发布年份 2015

【 摘 要 】

Background

Regular aspirin use reduces colon adenoma and carcinoma incidence. UDP-glucuronosyltransferases (UGT) are involved in aspirin metabolism and clearance, and variant alleles in UGT1A6 have been shown to alter salicylic acid metabolism and risk of colon neoplasia.

Methods

In a randomized, cross-over, placebo-controlled trial of 44 healthy men and women, homozygous for UGT1A6*1 or UGT1A6*2, we explored differences between global epithelial and stromal expression, using Affymetrix U133 + 2.0 microarrays and tested effects of 60-day aspirin supplementation (325 mg/d) on epithelial and stromal gene expression and colon prostaglandin E2 (PGE2) levels.

Results

No statistically significant differences in gene expression were observed in response to aspirin or UGT1A6 genotype, but tissue PGE2 levels were lower with aspirin compared to placebo (p <0.001). Transcripts differentially expressed between epithelium and stroma (N = 4916, P <0.01, false discovery rate <0.001), included a high proportion of genes involved in cell signaling, cellular movement, and cancer. Genes preferentially expressed in epithelium were involved in drug and xenobiotic metabolism, fatty acid and lipid metabolism, apoptosis signaling, and ion transport. Genes preferentially expressed in stroma included those involved in inflammation, cellular adhesion, and extracellular matrix production. Wnt-Tcf4 pathway genes were expressed in both epithelium and stroma but differed by subcellular location.

Conclusions

These results suggest that, in healthy individuals, subtle effects of aspirin on gene expression in normal colon tissue are likely overwhelmed by inter-individual variability in microarray analyses. Differential expression of critical genes between colonic epithelium and stroma suggest that these tissue types need to be considered separately.

【 授权许可】

   
2015 Thomas et al.; licensee BioMed Central.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

【 参考文献 】

• [1]Flossmann E, Rothwell PM: Effect of aspirin on long-term risk of colorectal cancer: consistent evidence from randomised and observational studies. Lancet 2007, 369:1603-13.
• [2]Rothwell PM, Wilson M, Elwin CE, Norrving B, Algra A, Warlow CP, et al.: Long-term effect of aspirin on colorectal cancer incidence and mortality: 20-year follow-up of five randomised trials. Lancet 2010, 376:1741-50.
• [3]Rothwell PM, Price JF, Fowkes FG, Zanchetti A, Roncaglioni MC, Tognoni G, et al.: Short-term effects of daily aspirin on cancer incidence, mortality, and non-vascular death: analysis of the time course of risks and benefits in 51 randomised controlled trials. Lancet 2012, 379:1602-12.
• [4]Chan AT, Arber N, Burn J, Chia WK, Elwood P, Hull MA, et al.: Aspirin in the chemoprevention of colorectal neoplasia: an overview. Cancer Prev Res (Phila) 2012, 5:164-78.
• [5]Taketo MM: Cyclooxygenase-2 inhibitors in tumorigenesis (part I). J Natl Cancer Inst 1998, 90:1529-36.
• [6]Elwood PC, Gallagher AM, Duthie GG, Mur LAJ, Morgan G: Aspirin, salicylates, and cancer. Lancet 2009, 373:1301-9.
• [7]Yin H, Xu H, Zhao Y, Yang W, Cheng J, Zhou Y: Cyclooxygenase-independent effects of aspirin on HT-29 human colon cancer cells, revealed by oligonucleotide microarrays. Biotechnol Lett 2006, 28:1263-70.
• [8]Maglietta R, Liuzzi VC, Cattaneo E, Laczko E, Piepoli A, Panza A, et al.: Molecular pathways undergoing dramatic transcriptomic changes during tumor development in the human colon. BMC Cancer 2012, 12:608. BioMed Central Full Text
• [9]Mariadason JM, Arango D, Corner GA, Aranes MJ, Hotchkiss KA, Yang W, et al.: A gene expression profile that defines colon cell maturation in vitro. Cancer Res 2002, 62:4791-804.
• [10]Halbleib JM, Saaf AM, Brown PO, Nelson WJ: Transcriptional modulation of genes encoding structural characteristics of differentiating enterocytes during development of a polarized epithelium in vitro. Mol Biol Cell 2007, 18:4261-78.
• [11]Sugiyama Y, Farrow B, Murillo C, Li J, Watanabe H, Sugiyama K, et al.: Analysis of differential gene expression patterns in colon cancer and cancer stroma using microdissected tissues. Gastroenterology 2005, 128:480-6.
• [12]Notterman DA, Alon U, Sierk AJ, Levine AJ: Transcriptional gene expression profiles of colorectal adenoma, adenocarcinoma, and normal tissue examined by oligonucleotide arrays. Cancer Res 2001, 61:3124-30.
• [13]St Croix B, Rago C, Velculescu V, Traverso G, Romans KE, Montgomery E, et al.: Genes expressed in human tumor endothelium. Science 2000, 289:1197-202.
• [14]Kitahara O, Furukawa Y, Tanaka T, Kihara C, Ono K, Yanagawa R, et al.: Alterations of gene expression during colorectal carcinogenesis revealed by cDNA microarrays after laser-capture microdissection of tumor tissues and normal epithelia. Cancer Res 2001, 61:3544-9.
• [15]Birkenkamp-Demtroder K, Christensen LL, Olesen SH, Frederiksen CM, Laiho P, Aaltonen LA, et al.: Gene expression in colorectal cancer. Cancer Res 2002, 62:4352-63.
• [16]Mojica W, Hawthorn L: Normal colon epithelium: a dataset for the analysis of gene expression and alternative splicing events in colon disease. BMC Genomics 2010, 11:5. BioMed Central Full Text
• [17]Hutt AJ, Caldwell J, Smith RL: The metabolism of aspirin in man: a population study. Xenobiotica 1986, 16:239-49.
• [18]Kuehl GE, Bigler J, Potter JD, Lampe JW: Glucuronidation of the aspirin metabolite salicylic acid by expressed UDP-glucuronosyltransferases and human liver microsomes. Drug Metab Dispos 2006, 34:199-202.
• [19]Ciotti M, Marrone A, Potter C, Owens IS: Genetic polymorphism in the human UGT1A6 (planar phenol) UDP-glucuronosyltransferase: pharmacological implications. Pharmacogenetics 1997, 7:485-95.
• [20]Chen Y, Kuehl GE, Bigler J, Rimorin CF, Schwarz Y, Shen DD, et al.: UGT1A6 polymorphism and salicylic acid glucuronidation following aspirin. Pharmacogenet Genomics 2007, 17:571-9.
• [21]van Oijen M: Effect of genetic polymorphisms in UDP-glucuronosyltransferase 1A6 (UGT1A6) on acetylsalicylic acid metabolism in healthy female volunteers. Pharmacology 2009, 83:237-42.
• [22]Bigler J, Whitton J, Lampe JW, Fosdick L, Bostick RM, Potter JD: CYP2C9 and UGT1A6 genotypes modulate the protective effect of aspirin on colon adenoma risk. Cancer Res 2001, 61:3566-9.
• [23]Chan AT, Tranah GJ, Giovannucci EL, Hunter DJ, Fuchs CS: Genetic variants in the UGT1A6 enzyme, aspirin use, and the risk of colorectal adenoma. J Natl Cancer Inst 2005, 97:457-60.
• [24]Samowitz WS, Wolff RK, Curtin K, Sweeney C, Ma KN, Andersen K, et al.: Interactions between CYP2C9 and UGT1A6 polymorphisms and nonsteroidal anti-inflammatory drugs in colorectal cancer prevention. Clin Gastroenterol Hepatol 2006, 4:894-901.
• [25]Thompson CL, Plummer SJ, Merkulova A, Cheng I, Tucker TC, Casey G, et al.: No association between cyclooxygenase-2 and uridine diphosphate glucuronosyltransferase 1A6 genetic polymorphisms and colon cancer risk. World J Gastroenterol 2009, 15:2240-4.
• [26]McGreavey LE, Turner F, Smith G, Boylan K, Timothy Bishop D, Forman D, et al.: No evidence that polymorphisms in CYP2C8, CYP2C9, UGT1A6, PPARdelta and PPARgamma act as modifiers of the protective effect of regular NSAID use on the risk of colorectal carcinoma. Pharmacogenet Genomics 2005, 15:713-21.
• [27]Hubner RA, Muir KR, Liu JF, Logan RF, Grainge M, Armitage N, et al.: Genetic variants of UGT1A6 influence risk of colorectal adenoma recurrence. Clin Cancer Res 2006, 12:6585-9.
• [28]Osawa K, Nakarai C, Akiyama M, Hashimoto R, Tsutou A, Takahashi J, et al.: Association between polymorphisms in UDP-glucuronosyltransferase 1A6 and 1A7 and colorectal cancer risk. Asian Pac J Cancer Prev 2012, 13:2311-4.
• [29]Navarro SL, Saracino MR, Makar KW, Thomas SS, Li L, Zheng Y, et al.: Determinants of aspirin metabolism in healthy men and women: Effects of dietary inducers of UGT. J Nutrigenet Nutrigenomics 2011, 4:110-8.
• [30]Lampe JW, Bigler J, Horner NK, Potter JD: UDP-glucuronosyltransferase (UGT1A1*28 and UGT1A6*2) polymorphisms in Caucasians and Asians: relationships to serum bilirubin concentrations. Pharmacogenetics 1999, 9:341-9.
• [31]Schwartz B, Avivi C, Lamprecht SA: Isolation and characterization of normal and neoplastic colonic epithelial cell populations. Gastroenterology 1991, 100:692-702.
• [32]Gene Expression Omnibus. [http://www.ncbi.nlm.nih.gov/geo/]
• [33]Shureiqi I, Chen D, Day RS, Zuo X, Hochman FL, Ross WA, et al.: Profiling lipoxygenase metabolism in specific steps of colorectal tumorigenesis. Cancer Prev Res (Phila) 2010, 3:829-38.
• [34]Wu Z, Irizarry RA: Preprocessing of oligonucleotide array data. Nat Biotechnol 2004, 22:656-8. author reply 658
• [35]Benjamini Y, Hochberg Y: Controlling the false discovery rate - a practical and powerful approach to multiple testing. J Royal Statist Soc Serial B 1995, 57:289-300.
• [36]Eisen MB, Spellman PT, Brown PO, Botstein D: Cluster analysis and display of genome-wide expression patterns. Proc Natl Acad Sci U S A 1998, 95:14863-8.
• [37]Saldanha AJ: Java Treeview–extensible visualization of microarray data. Bioinformatics 2004, 20:3246-8.
• [38]Nagar S, Zalatoris JJ, Blanchard RL: Human UGT1A6 pharmacogenetics: identification of a novel SNP, characterization of allele frequencies and functional analysis of recombinant allozymes in human liver tissue and in cultured cells. Pharmacogenetics 2004, 14:487-99.
• [39]Adegboyega PA, Mifflin RC, DiMari JF, Saada JI, Powell DW: Immunohistochemical study of myofibroblasts in normal colonic mucosa, hyperplastic polyps, and adenomatous colorectal polyps. Arch Pathol Lab Med 2002, 126:829-36.
• [40]Mahida YR, Beltinger J, Makh S, Goke M, Gray T, Podolsky DK, et al.: Adult human colonic subepithelial myofibroblasts express extracellular matrix proteins and cyclooxygenase-1 and -2. Am J Physiol 1997, 273:G1341-8.
• [41]Powell DW, Adegboyega PA, Di Mari JF, Mifflin RC: Epithelial cells and their neighbors I. Role of intestinal myofibroblasts in development, repair, and cancer. Am J Physiol Gastrointest Liver Physiol 2005, 289:G2-7.
• [42]Kosinski C, Li VS, Chan AS, Zhang J, Ho C, Tsui WY, et al.: Gene expression patterns of human colon tops and basal crypts and BMP antagonists as intestinal stem cell niche factors. Proc Natl Acad Sci U S A 2007, 104:15418-23.
• [43]Yan M, Rerko RM, Platzer P, Dawson D, Willis J, Tong M, et al.: 15-Hydroxyprostaglandin dehydrogenase, a COX-2 oncogene antagonist, is a TGF-beta-induced suppressor of human gastrointestinal cancers. Proc Natl Acad Sci U S A 2004, 101:17468-73.
• [44]van de Wetering M, Sancho E, Verweij C, de Lau W, Oving I, Hurlstone A, et al.: The beta-catenin/TCF-4 complex imposes a crypt progenitor phenotype on colorectal cancer cells. Cell 2002, 111:241-50.
• [45]Hulit J, Wang C, Li Z, Albanese C, Rao M, Di Vizio D, et al.: Cyclin D1 genetic heterozygosity regulates colonic epithelial cell differentiation and tumor number in ApcMin mice. Mol Cell Biol 2004, 24:7598-611.
• [46]Flynn C, Montrose DC, Swank DL, Nakanishi M, Ilsley JN, Rosenberg DW: Deoxycholic acid promotes the growth of colonic aberrant crypt foci. Mol Carcinog 2007, 46:60-70.
• [47]Boman BM, Walters R, Fields JZ, Kovatich AJ, Zhang T, Isenberg GA, et al.: Colonic crypt changes during adenoma development in familial adenomatous polyposis: immunohistochemical evidence for expansion of the crypt base cell population. Am J Pathol 2004, 165:1489-98.
• [48]Mikami T, Mitomi H, Hara A, Yanagisawa N, Yoshida T, Tsuruta O, et al.: Decreased expression of CD44, alpha-catenin, and deleted colon carcinoma and altered expression of beta-catenin in ulcerative colitis-associated dysplasia and carcinoma, as compared with sporadic colon neoplasms. Cancer 2000, 89:733-40.
• [49]Jay P, Berta P, Blache P: Expression of the carcinoembryonic antigen gene is inhibited by SOX9 in human colon carcinoma cells. Cancer Res 2005, 65:2193-8.
• [50]Davenport A, Hale RJ, Hunt CR, Bigley G, McMahon RF: Expression of Ki-67 and cytokeratin 20 in hyperplastic polyps of the colorectum. J Clin Pathol 2003, 56:200-4.
• [51]Blache P, van de Wetering M, Duluc I, Domon C, Berta P, Freund JN, et al.: SOX9 is an intestine crypt transcription factor, is regulated by the Wnt pathway, and represses the CDX2 and MUC2 genes. J Cell Biol 2004, 166:37-47.
• [52]Seibert K, Zhang Y, Leahy K, Hauser S, Masferrer J, Isakson P: Distribution of COX-1 and COX-2 in normal and inflamed tissues. Adv Exp Med Biol 1997, 400A:167-70.
• [53]Bond PA, Cundall RL: Properties of monoamine oxidase (MAO) in human blood platelets, plasma, lymphocytes and granulocytes. Clin Chim Acta 1977, 80:317-26.
• [54]Beaulieu JF, Vachon PH: Reciprocal expression of laminin A-chain isoforms along the crypt-villus axis in the human small intestine. Gastroenterology 1994, 106:829-39.
• [55]Clevers H: Wnt/beta-catenin signaling in development and disease. Cell 2006, 127:469-80.
• [56]Cadigan KM, Nusse R: Wnt signaling: a common theme in animal development. Genes Dev 1997, 11:3286-305.
• [57]Bienz M, Clevers H: Linking colorectal cancer to Wnt signaling. Cell 2000, 103:311-20.
• [58]Morin PJ, Sparks AB, Korinek V, Barker N, Clevers H, Vogelstein B, et al.: Activation of beta-catenin-Tcf signaling in colon cancer by mutations in beta-catenin or APC. Science 1997, 275:1787-90.
• [59]Potter JD: Morphogens, morphostats, microarchitecture and malignancy. Nat Rev Cancer 2007, 7:464-74.
• [60]Tlsty TD, Hein PW: Know thy neighbor: stromal cells can contribute oncogenic signals. Curr Opin Genet Dev 2001, 11:54-9.
• [61]Ronnov-Jessen L, Petersen OW, Bissell MJ: Cellular changes involved in conversion of normal to malignant breast: importance of the stromal reaction. Physiol Rev 1996, 76:69-125.
• [62]Elenbaas B, Weinberg RA: Heterotypic signaling between epithelial tumor cells and fibroblasts in carcinoma formation. Exp Cell Res 2001, 264:169-84.
• [63]Lin YM, Furukawa Y, Tsunoda T, Yue CT, Yang KC, Nakamura Y: Molecular diagnosis of colorectal tumors by expression profiles of 50 genes expressed differentially in adenomas and carcinomas. Oncogene 2002, 21:4120-8.
• [64]Perez-Villamil B, Romera-Lopez A, Hernandez-Prieto S, Lopez-Campos G, Calles A, Lopez-Asenjo JA, et al.: Colon cancer molecular subtypes identified by expression profiling and associated to stroma, mucinous type and different clinical behavior. BMC Cancer 2012, 12:260. BioMed Central Full Text
• [65]Vane JR: Inhibition of prostaglandin synthesis as a mechanism of action for aspirin-like drugs. Nat New Biol 1971, 231:232-5.
• [66]Eberhart CE, Coffey RJ, Radhika A, Giardiello FM, Ferrenbach S, DuBois RN: Up-regulation of cyclooxygenase 2 gene expression in human colorectal adenomas and adenocarcinomas. Gastroenterology 1994, 107:1183-8.
• [67]Kunzelmann K, Mall M: Electrolyte transport in the mammalian colon: mechanisms and implications for disease. Physiol Rev 2002, 82:245-89.
• [68]Feske S: Calcium signalling in lymphocyte activation and disease. Nat Rev Immunol 2007, 7:690-702.
• [69]Saaf AM, Halbleib JM, Chen X, Yuen ST, Leung SY, Nelson WJ, et al.: Parallels between global transcriptional programs of polarizing Caco-2 intestinal epithelial cells in vitro and gene expression programs in normal colon and colon cancer. Mol Biol Cell 2007, 18:4245-60.
• [70]Polakis P: Wnt signaling and cancer. Genes Dev 2000, 14:1837-51.
• [71]Sabates-Bellver J, Van der Flier LG, de Palo M, Cattaneo E, Maake C, Rehrauer H, et al.: Transcriptome profile of human colorectal adenomas. Mol Cancer Res 2007, 5:1263-75.
• [72]Fink SP, Yang DH, Barnholtz-Sloan JS, Ryu YM, Mikkola D, Potter JD, et al.: Colonic 15-PGDH Levels are stable across distance and time and are not perturbed by aspirin intervention. Dig Dis Sci 2013, 58:2615-22.
• [73]Chulada PC, Thompson MB, Mahler JF, Doyle CM, Gaul BW, Lee C, et al.: Genetic disruption of Ptgs-1, as well as Ptgs-2, reduces intestinal tumorigenesis in Min mice. Cancer Res 2000, 60:4705-8.
• [74]Tosco P, Lazzarato L: Mechanistic insights into cyclooxygenase irreversible inactivation by aspirin. ChemMedChem 2009, 4:939-45.
• [75]Ruffin MT, Krishnan K, Rock CL, Normolle D, Vaerten MA, Peters-Golden M, et al.: Suppression of human colorectal mucosal prostaglandins: determining the lowest effective aspirin dose. J Natl Cancer Inst 1997, 89:1152-60.
• [76]Bousserouel S, Gosse F, Bouhadjar M, Soler L, Marescaux J, Raul F: Long-term administration of aspirin inhibits tumour formation and triggers anti-neoplastic molecular changes in a pre-clinical model of colon carcinogenesis. Oncol Rep 2010, 23:511-7.
• [77]Agundez JA, Martinez C, Perez-Sala D, Carballo M, Torres MJ, Garcia-Martin E: Pharmacogenomics in aspirin intolerance. Curr Drug Metab 2009, 10:998-1008.