Leiomyomas are benign uterine tumors which develop from transformation of myometrial cells without a known etiology. Through genomic and proteomic analysis we have identified a number of differentially expressed and regulated genes in leiomyomas as compared to myometrium, with diverse biological functions, including cellular transformation, proliferation, and pro-inflammatory/pro-fibrotic activities such as IL-13 and TGF-2. ZEB1, a transcription factor and major repressor of E-cadherin, which is lost during cellular transition into mesenchymal phenotype, is overexpressed in leiomyomas as compared to myometrium. Based on differential expression, hormonal regulation and regulation by TGF-2 and IL-13, we propose that ZEB1 and ZEB2 also serve as key regulators of myometrial phenotypic transformation into leiomyomas. We further propose that expression and stability of ZEBs are subject to epigenetic and microRNA regulation, respectively. To test the hypothesis that the expression, regulation and function of ZEBs result in development and growth of leiomyoma, we propose the following specific aims.
Aim#1 will test the hypothesis that ZEBs and E-cadherin are differentially expressed and regulated in leiomyomas as compared to myometrium, and their expression directly/inversely correlates with the expression of TGF-2 and IL-13 and altered in patients who receive hormonal therapies to suppress leiomyoma growth. Furthermore, the expression profiles of HDAC's 2, 4, and EZH2 in leiomyoma and myometrium from the above cohorts correlate with their role as epigenetic regulators of the expression of ZEBs and miRNAs that target their expression, and are differentially expressed in tissues from African Americans as compared to Caucasians.
Aim#2 will test the hypothesis that pro-inflammatory/pro-fibrotic molecules, such as IL-13 and TGF-2 directly regulate the expression of ZEBs in myometrial smooth muscle cells (MSMC) resulting in phenotypic transformation as a consequence of loss of E-cadherin, and their actions are prevented following treatments with GnRHa, RU-486, Depo-Provera and Tranilast, an inhibitor of pro-fibrotic/inflammatory mediators. Further functional activity of ZEBs is altered through treatments with siRNAs and miR-200b/c in MSMC and LSMC thus their downstream target genes, including E-cadherin.
Aim#3 will test the hypothesis that ZEBs expression is regulated via epigenetic modifications and by the action of miRNAs in myometrial cells, and alteration in this mechanism results in transformation into leiomyoma cells. This will be tested by assessing the patterns of expression of HDACs and EZH2, as well as miRNAs that target their expression in myometrial and leiomyoma cells;following treatments with IL-13 and TGF-2, alone and in combination, and following treatment with Tranilast, miRNAs gain-of functions and inhibitors of HDACs and CpG methylation, respectively. To achieve these Aims, we will use leiomyoma and matched myometrium and utilize a combination of biochemical, molecular and cell biological approaches. We anticipate that this proposed research will lead to further identification of a direct role of pro-fibrotic/inflammatory derived mediators in myometrial cellular transformation into leiomyoma cells and help to work toward the overall development of a novel therapeutic approach to prevent the growth and specifically treat leiomyomas.

Public Health Relevance

Fibroids are benign uterine tumors estimated to develop in 70% of women, more specifically among African Americans during their reproductive years. Symptomatic fibroids cause chronic pelvic pain and abnormal uterine bleeding which are the main indications for women undergoing hysterectomy. This proposal will investigate how fibroids are developed and grow, and ways to prevent them, including their symptoms.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD037432-11
Application #
8146143
Study Section
Integrative and Clinical Endocrinology and Reproduction Study Section (ICER)
Program Officer
De Paolo, Louis V
Project Start
2001-03-06
Project End
2013-03-31
Budget Start
2011-06-01
Budget End
2013-03-31
Support Year
11
Fiscal Year
2011
Total Cost
$142,013
Indirect Cost
Name
University of Florida
Department
Obstetrics & Gynecology
Type
Schools of Medicine
DUNS #
969663814
City
Gainesville
State
FL
Country
United States
Zip Code
32611
Chuang, Tsai-Der; Luo, Xiaoping; Panda, Harekrushna et al. (2012) miR-93/106b and their host gene, MCM7, are differentially expressed in leiomyomas and functionally target F3 and IL-8. Mol Endocrinol 26:1028-42
Chuang, Tsai-Der; Panda, Harekrushna; Luo, Xiaoping et al. (2012) miR-200c is aberrantly expressed in leiomyomas in an ethnic-dependent manner and targets ZEBs, VEGFA, TIMP2, and FBLN5. Endocr Relat Cancer 19:541-56
Panda, Harekrushna; Pelakh, Leslie; Chuang, Tsai-Der et al. (2012) Endometrial miR-200c is altered during transformation into cancerous states and targets the expression of ZEBs, VEGFA, FLT1, IKK?, KLF9, and FBLN5. Reprod Sci 19:786-96
Panda, Harekrushna; Chuang, Tsai-Der; Luo, Xiaoping et al. (2012) Endometrial miR-181a and miR-98 expression is altered during transition from normal into cancerous state and target PGR, PGRMC1, CYP19A1, DDX3X, and TIMP3. J Clin Endocrinol Metab 97:E1316-26
Chegini, Nasser (2010) Proinflammatory and profibrotic mediators: principal effectors of leiomyoma development as a fibrotic disorder. Semin Reprod Med 28:180-203
Pan, Qun; Luo, Xiaoping; Chegini, Nasser (2010) microRNA 21: response to hormonal therapies and regulatory function in leiomyoma, transformed leiomyoma and leiomyosarcoma cells. Mol Hum Reprod 16:215-27
Chegini, Nasser (2010) Uterine microRNA signature and consequence of their dysregulation in uterine disorders. Anim Reprod 7:117-128
Luo, Xiaoping; Chegini, Nasser (2008) The expression and potential regulatory function of microRNAs in the pathogenesis of leiomyoma. Semin Reprod Med 26:500-14
Pan, Qun; Chegini, Nasser (2008) MicroRNA signature and regulatory functions in the endometrium during normal and disease states. Semin Reprod Med 26:479-93
Pan, Qun; Luo, Xiaoping; Chegini, Nasser (2008) Differential expression of microRNAs in myometrium and leiomyomas and regulation by ovarian steroids. J Cell Mol Med 12:227-40

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