Transcriptome Analysis of mRNA in Uterine Leiomyoma Using Next-generation RNA Sequencing

Author(s): Shadab Anjum, Tahreem Sahar, Aruna Nigam, Saima Wajid*.

Journal Name: Anti-Cancer Agents in Medicinal Chemistry
(Formerly Current Medicinal Chemistry - Anti-Cancer Agents)

Volume 19 , Issue 14 , 2019

Become EABM
Become Reviewer

Graphical Abstract:


Abstract:

Background: Uterine leiomyoma is a benign smooth muscle tumor of monoclonal nature in the female reproductive tract and is one of the major health problems. More than 70% of the female population suffers from uterine leiomyoma in their lifetime and in the advanced condition, it is associated with pregnancy complications and infertility.

Objective: Characterization and relative expression of mRNA transcripts through transcriptome profiling in uterine leiomyoma and adjacent normal myometrium.

Methods: Uterine leiomyoma tissue of an Indian female, age 32 years, with a family history of leiomyoma (evident from mother’s hysterectomy for the same pathology) was used. Patient showed 9 multiple large lesions appearing heterogeneously, deforming the uterine contour and causing distortion and splaying of the endometrial cavity showing disease aggressiveness was taken for Next-generation sequencing (NGS) to develop whole transcriptome profile along with the adjacent normal myometrium as control. The validation of the relative expression of the selective transcripts was done using Real-Time PCR.

Results: The transcriptome profile indicated 128 genes up-regulated and 98 down-regulated, with the Log2 fold change ≥ 2 and P ≤ 0.05, highlighting the molecular network closely associated with focal adhesion, hyaluronan and MAPK-signaling pathways. The mean relative fold change obtained from quantitative PCR as well as the P-values of 10 selected transcripts evaluated from student’s t-test were as follows: BCAN: 7.93 fold (p-value =0.0013); AAK1: 2.2 fold (p-value =0.0036); PCBP3: 3.4 fold (p-value =0.0197); MOV10L1: 3.4 fold (p-value =0.0062); TWISTNB: 1.8 fold (p-value =0.006); TMSB15A: 2.1 fold (p-value =0.0023); SMAD1: 0.8 fold (p-value =0.0112); ANXA1: 0.6 fold (p-value =0.0012); FOS: 0.6 fold (p-value =0.0191); SLFN11: 0.56 fold (p-value =0.0001).

Conclusion: The present study provides a roadmap, towards the analysis of genes and their roles in corresponding pathways throwing light on their possible involvement in the pathology of the disease.

Keywords: Uterine leiomyoma, fibroid, transcriptome, next-generation sequencing, genes, real-time PCR.

[1]
Kempson, R.L.; Hendrickson, M.R. Smooth muscle, endometrial stromal, and mixed Mullerian tumors of the uterus. Mod. Pathol., 2000, 13(3), 328-342.
[2]
Kjerulff, K.H.; Langenberg, P.; Seidman, J.D.; Stolley, P.D.; Guzinski, G.M. Uterine leiomyomas. Racial differences in severity, symptoms and age at diagnosis. J. Reprod. Med., 1996, 41(7), 483-490.
[3]
Othman, E-E.R.; Al-Hendy, A. Molecular genetics and racial disparities of uterine leiomyomas. Best Pract. Res. Clin. Obstet. Gynaecol., 2008, 22(4), 589-601.
[4]
Wegienka, G.; Baird, D.D.; Hertz-Picciotto, I.; Harlow, S.D.; Steege, J.F.; Hill, M.C.; Schectman, J.M.; Hartmann, K.E. Self-reported heavy bleeding associated with uterine leiomyomata. Obstet. Gynecol., 2003, 101(3), 431-437.
[5]
Baird, D.D.; Dunson, D.B.; Hill, M.C.; Cousins, D.; Schectman, J.M. High cumulative incidence of uterine leiomyoma in black and white women: ultrasound evidence. Am. J. Obstet. Gynecol., 2003, 188(1), 100-107.
[6]
Puri, K.; Famuyide, A.O.; Erwin, P.J.; Stewart, E.A.; Laughlin-Tommaso, S.K. Submucosal fibroids and the relation to heavy menstrual bleeding and anemia. Am. J. Obstet. Gynecol., 2014, 210(1), e1-e7.
[7]
Yang, J.H.; Chen, M.J.; Chen, C.D.; Chen, C.L.; Ho, H.N.; Yang, Y.S. Impact of submucous myoma on the severity of anemia. Fertil. Steril., 2011, 95(5), 1769-1772.
[8]
Malik, M.; Norian, J.; McCarthy-Keith, D.; Britten, J.; Catherino, W.H. Why leiomyomas are called fibroids: the central role of extracellular matrix in symptomatic women. Semin. Reprod. Med., 2010, 28(3), 169-179.
[9]
Sandberg, A.A. Updates on the cytogenetics and molecular genetics of bone and soft tissue tumors: leiomyoma. Cancer Genet. Cytogenet., 2005, 158(1), 1-26.
[10]
Cardozo, E.R.; Clark, A.D.; Banks, N.K.; Henne, M.B.; Stegmann, B.J.; Segars, J.H. The estimated annual cost of uterine leiomyomata in the United States. Am. J. Obstet. Gynecol., 2012, 206(3), e1-e9.
[11]
Walker, C.L.; Stewart, E.A. Uterine fibroids: The elephant in the room. Science, 2005, 308(5728), 1589-1592.
[12]
Maruo, T.; Matsuo, H.; Shimomura, Y.; Kurachi, O.; Gao, Z.; Nakago, S.; Yamada, T.; Chen, W.; Wang, J. Effects of progesterone on growth factor expression in human uterine leiomyoma. Steroids, 2003, 68(10-13), 817-824.
[13]
Ishikawa, H.; Ishi, K.; Serna, V.A.; Kakazu, R.; Bulun, S.E.; Kurita, T. Progesterone is essential for maintenance and growth of uterine leiomyoma. Endocrinology, 2010, 151(6), 2433-2442.
[14]
Patel, B.; Elguero, S.; Thakore, S.; Dahoud, W.; Bedaiwy, M.; Mesiano, S. Role of nuclear progesterone receptor isoforms in uterine pathophysiology. Hum. Reprod. Update, 2015, 21(2), 155-173.
[15]
Grings, A.O.; Lora, V.; Ferreira, G.D.; Brum, I.S.; Corleta, H.; Capp, E. Protein expression of estrogen receptors alpha and beta and aromatase in myometrium and uterine leiomyoma. Gynecol. Obstet. Invest., 2012, 73(2), 113-117.
[16]
Ciarmela, P.; Islam, M.S.; Reis, F.M.; Gray, P.C.; Bloise, E.; Petraglia, F.; Vale, W.; Castellucci, M. Growth factors and myometrium: Biological effects in uterine fibroid and possible clinical implications. Hum. Reprod. Update, 2011, 17(6), 772-790.
[17]
Anania, C.A.; Stewart, E.A.; Quade, B.J.; Hill, J.A.; Nowak, R.A. Expression of the fibroblast growth factor receptor in women with leiomyomas and abnormal uterine bleeding. Mol. Hum. Reprod., 1997, 3(8), 685-691.
[18]
Hong, T.; Shimada, Y.; Uchida, S.; Itami, A.; Li, Z.; Ding, Y.; Kaganoi, J.; Komoto, I.; Sakurai, T.; Imamura, M. Expression of angiogenic factors and apoptotic factors in leiomyosarcoma and leiomyoma. Int. J. Mol. Med., 2001, 8(2), 141-148.
[19]
Arita, S.; Kikkawa, F.; Kajiyama, H.; Shibata, K.; Kawai, M.; Mizuno, K.; Nagasaka, T.; Ino, K.; Nomura, S. Prognostic importance of vascular endothelial growth factor and its receptors in the uterine sarcoma. Int. J. Gynecol. Cancer, 2005, 15(2), 329-336.
[20]
Islam, M.S.; Protic, O.; Stortoni, P.; Grechi, G.; Lamanna, P.; Petraglia, F.; Castellucci, M.; Ciarmela, P. Complex networks of multiple factors in the pathogenesis of uterine leiomyoma. Fertil. Steril., 2013, 100(1), 178-193.
[21]
Rossi, M.J.; Chegini, N.; Masterson, B.J. Presence of epidermal growth factor, platelet-derived growth factor, and their receptors in human myometrial tissue and smooth muscle cells: Their action in smooth muscle cells in vitro. Endocrinology, 1992, 130(3), 1716-1727.
[22]
Marshall, L.M.; Spiegelman, D.; Barbieri, R.L.; Goldman, M.B.; Manson, J.E.; Colditz, G.A.; Willett, W.C.; Hunter, D.J. Variation in the incidence of uterine leiomyoma among premenopausal women by age and race. Obstet. Gynecol., 1997, 90(6), 967-973.
[23]
Peddada, S.D.; Laughlin, S.K.; Miner, K.; Guyon, J.P.; Haneke, K.; Vahdat, H.L.; Semelka, R.C.; Kowalik, A.; Armao, D.; Davis, B.; Baird, D.D. Growth of uterine leiomyomata among premenopausal black and white women. Proc. Natl. Acad. Sci. USA, 2008, 105(50), 19887-19892.
[24]
Wise, L.A.; Palmer, J.R.; Cozier, Y.C.; Hunt, M.O.; Stewart, E.A.; Rosenberg, L. Perceived racial discrimination and risk of uterine leiomyomata. Epidemiology, 2007, 18(6), 747-757.
[25]
Wise, L.A.; Ruiz-Narvaez, E.A.; Palmer, J.R.; Cozier, Y.C.; Tandon, A.; Patterson, N.; Radin, R.G.; Rosenberg, L.; Reich, D. African ancestry and genetic risk for uterine leiomyomata. Am. J. Epidemiol., 2012, 176(12), 1159-1168.
[26]
Mittal, P.; Shin, Y.H.; Yatsenko, S.A.; Castro, C.A.; Surti, U.; Rajkovic, A. Med12 gain-of-function mutation causes leiomyomas and genomic instability. J. Clin. Invest., 2015, 125(8), 3280-3284.
[27]
Liegl-Atzwanger, B.; Heitzer, E.; Flicker, K.; Muller, S.; Ulz, P.; Saglam, O.; Tavassoli, F.; Devouassoux-Shisheboran, M.; Geigl, J.; Moinfar, F. Exploring chromosomal abnormalities and genetic changes in uterine smooth muscle tumors. Mod. Pathol., 2016, 29(10), 1262-1277.
[28]
Hayden, M.A.; Ordulu, Z.; Gallagher, C.S.; Quade, B.J.; Anchan, R.M.; Middleton, N.R.; Srouji, S.S.; Stewart, E.A.; Morton, C.C. Clinical, pathologic, cytogenetic, and molecular profiling in self-identified black women with uterine leiomyomata. Cancer Genet., 2018, 222-223, 1-8.
[29]
Wolf, J.B. Principles of transcriptome analysis and gene expression quantification: An RNA-seq tutorial. Mol. Ecol. Resour., 2013, 13(4), 559-572.
[30]
Wang, Z.; Gerstein, M.; Snyder, M. RNA-Seq: A revolutionary tool for transcriptomics. Nat. Rev. Genet., 2009, 10(1), 57-63.
[31]
Islam, M.S.; Ciavattini, A.; Petraglia, F.; Castellucci, M.; Ciarmela, P. Extracellular matrix in uterine leiomyoma pathogenesis: A potential target for future therapeutics. Hum. Reprod. Update, 2018, 24(1), 59-85.
[32]
Dixon, D.; He, H.; Haseman, J.K. Immunohistochemical localization of growth factors and their receptors in uterine leiomyomas and matched myometrium. Environ. Health Perspect., 2000, 108(Suppl. 5), 795-802.
[33]
Sozen, I.; Arici, A. Interactions of cytokines, growth factors, and the extracellular matrix in the cellular biology of uterine leiomyomata. Fertil. Steril., 2002, 78(1), 1-12.
[34]
Norian, J.M.; Owen, C.M.; Taboas, J.; Korecki, C.; Tuan, R.; Malik, M.; Catherino, W.H.; Segars, J.H. Characterization of tissue biomechanics and mechanical signaling in uterine leiomyoma. Matrix Biol., 2012, 31(1), 57-65.
[35]
Kamel, M.; Wagih, M.; Kilic, G.S.; Diaz-Arrastia, C.R.; Baraka, M.A.; Salama, S.A. Overhydroxylation of lysine of collagen increases uterine fibroids proliferation: roles of lysyl hydroxylases, lysyl oxidases, and matrix metalloproteinases. Biomed Res. Int., 2017, 20175316845
[36]
Xu, X.; Kim, J.J.; Li, Y.; Xie, J.; Shao, C.; Wei, J.J. Oxidative stress-induced miRNAs modulate AKT signaling and promote cellular senescence in uterine leiomyoma. J. Mol. Med., 2018, 96(10), 1095-1106.
[37]
Faryna, M.; Konermann, C.; Aulmann, S.; Bermejo, J.L.; Brugger, M.; Diederichs, S.; Rom, J.; Weichenhan, D.; Claus, R.; Rehli, M.; Schirmacher, P.; Sinn, H.P.; Plass, C.; Gerhauser, C. Genome-wide methylation screen in low-grade breast cancer identifies novel epigenetically altered genes as potential biomarkers for tumor diagnosis. FASEB J., 2012, 26(12), 4937-4950.
[38]
Frischknecht, R.; Seidenbecher, C.I. Brevican: A key proteoglycan in the perisynaptic extracellular matrix of the brain. Int. J. Biochem. Cell Biol., 2012, 44(7), 1051-1054.
[39]
Gupta-Rossi, N.; Ortica, S.; Meas-Yedid, V.; Heuss, S.; Moretti, J.; Olivo-Marin, J.C.; Israel, A. The adaptor-associated kinase 1, AAK1, is a positive regulator of the Notch pathway. J. Biol. Chem., 2011, 286(21), 18720-18730.
[40]
Timmerman, L.A.; Grego-Bessa, J.; Raya, A.; Bertran, E.; Perez-Pomares, J.M.; Diez, J.; Aranda, S.; Palomo, S.; McCormick, F.; Izpisua-Belmonte, J.C.; de la Pompa, J.L. Notch promotes epithelial-mesenchymal transition during cardiac development and oncogenic transformation. Genes Dev., 2004, 18(1), 99-115.
[41]
Bisogno, L.S.; Keene, J.D. Analysis of post-transcriptional regulation during cancer progression using a donor-derived isogenic model of tumorigenesis. Methods, 2017, 126, 193-200.
[42]
Pereira, B.; Billaud, M.; Almeida, R. RNA-binding proteins in cancer: Old players and new actors. Trends Cancer, 2017, 3(7), 506-528.
[43]
McStay, B.; Grummt, I. The epigenetics of rRNA genes: From molecular to chromosome biology. Annu. Rev. Cell Dev. Biol., 2008, 24, 131-157.
[44]
Forsburg, S.L. Eukaryotic MCM proteins: Beyond replication initiation. Microbiol. Mol. Biol. Rev., 2004, 68(1), 109-131.
[45]
Lei, M. The MCM complex: Its role in DNA replication and implications for cancer therapy. Curr. Cancer Drug Targets, 2005, 5(5), 365-380.
[46]
Gu, Y.M.; Li, S.Y.; Qiu, X.S.; Wang, E.H. Elevated thymosin beta15 expression is associated with progression and metastasis of non-small cell lung cancer. APMIS, 2008, 116(6), 484-490.
[47]
Pangas, S.A.; Li, X.; Umans, L.; Zwijsen, A.; Huylebroeck, D.; Gutierrez, C.; Wang, D.; Martin, J.F.; Jamin, S.P.; Behringer, R.R.; Robertson, E.J.; Matzuk, M.M. Conditional deletion of Smad1 and Smad5 in somatic cells of male and female gonads leads to metastatic tumor development in mice. Mol. Cell. Biol., 2008, 28(1), 248-257.
[48]
Shen, D.; Nooraie, F.; Elshimali, Y.; Lonsberry, V.; He, J.; Bose, S.; Chia, D.; Seligson, D.; Chang, H.R.; Goodglick, L. Decreased expression of annexin A1 is correlated with breast cancer development and progression as determined by a tissue microarray analysis. Hum. Pathol., 2006, 37(12), 1583-1591.
[49]
Wang, L.P.; Bi, J.; Yao, C.; Xu, X.D.; Li, X.X.; Wang, S.M.; Li, Z.L.; Zhang, D.Y.; Wang, M.; Chang, G.Q. Annexin A1 expression and its prognostic significance in human breast cancer. Neoplasma, 2010, 57(3), 253-259.
[50]
Guo, C.; Liu, S.; Sun, M.Z. Potential role of Anxa1 in cancer. Future Oncol., 2013, 9(11), 1773-1793.
[51]
Boudhraa, Z.; Bouchon, B.; Viallard, C.; D’Incan, M.; Degoul, F. Annexin A1 localization and its relevance to cancer. Clin. Sci. (Lond.), 2016, 130(4), 205-220.
[52]
Smeyne, R.J.; Vendrell, M.; Hayward, M.; Baker, S.J.; Miao, G.G.; Schilling, K.; Robertson, L.M.; Curran, T.; Morgan, J.I. Continuous c-fos expression precedes programmed cell death in vivo. Nature, 1993, 363(6425), 166-169.
[53]
Oliveira-Ferrer, L.; Rößler, K.; Haustein, V.; Schröder, C.; Wicklein, D.; Maltseva, D.; Khaustova, N.; Samatov, T.; Tonevitsky, A.; Mahner, S.; Jänicke, F. c-FOS suppresses ovarian cancer progression by changing adhesion. Br. J. Cancer, 2014, 110(3), 753.
[54]
Chuang, T.D.; Khorram, O. Expression profiling of lncRNAs, miRNAs, and mRNAs and their differential expression in leiomyoma using next-generation RNA sequencing. Reprod. Sci., 2018, 25(2), 246-255.


Rights & PermissionsPrintExport Cite as


Article Details

VOLUME: 19
ISSUE: 14
Year: 2019
Page: [1703 - 1718]
Pages: 16
DOI: 10.2174/1871520619666190409102855
Price: $58

Article Metrics

PDF: 28
HTML: 3
EPUB: 1
PRC: 1