27 - 30 May 2005

GENE PROFILING EXPERIMENTS REVEAL A DISORDERED EXTRACELLULAR MATRIX IN UTERINE LEIOMYOMA James Segars MD, William Catherino MD PhD, Phyllis Leppert, MD PhD Reproductive Biology and Medicine Branch National Institute of Child Health and Human Development National Institutes of Health Bethesda, Maryland USA

Leiomyoma (uterine fibroids) are a prevalent disorder of the reproductive tract that affects millions of women leading to bleeding, pelvic pain, infertility, pre-term birth, and pregnancy loss. The cause of these common benign growths remains unknown. Our group has used global gene expression profiling to study uterine leiomyoma (fibroids) and ask the question: what are the genetic features that characterize the leiomyoma cell?

To address this question we compared surgically-obtained tissue from normal myometrium with fibroid tumors using AffymetrixTM U133 A&B chips, which contain transcripts from up to 33,000 genes. We confirmed differences in gene expression using RT-PCR, real-time PCR, immunohistochemistry, or other approaches. Results revealed that approximately 1% of genes differed greater than two-fold between normal myometrium and tissues harvested from uterine fibroids. An unexpected observation was that there were rather marked differences between arrays from different core facilities using different AffymetrixTM platforms.

We interpret the differences to variation in procedure, but concluded it was essential to confirm results using an ancillary approach. Another unexpected result was that genes involved in sex steroid action were not featured as differentially expressed genes. Instead, genes involved in formation of extracellular matrix (ECM) were differentially expressed. We noted remarkable changes in expression of genes encoding collagen and ultrastructural studies revealed that collagen fibrils were more abundant, loosely packed and arrayed in a non-parallel manner in leiomyoma, compared to tight bundles in normal myometrium.

In addition, analysis of mature collagen fibrils in myometrium revealed an ordered, barbed structure at 64,000 X magnification, whereas collagen fibrils in leiomyoma lacked the barbed structure. Furthermore, when our list of differentially expressed genes was compared with published lists obtained using a similar methodology, six transcripts were consistently identified. One transcript encoded dermatopontin, a 22kd extracellular protein known to bind the collagen-binding protein decorin, as well as TGF-beta. Reverse-transcriptase real-time PCR, RT-PCR, and immunohistochemical experiments confirmed the reduction in dermatopontin in fibroids. Interestingly, reductions in dermatopontin were previously recognised in hypertrophic scar and keloid, two disorders of tissue remodeling in skin. Next, we compared our list of genes with a recently reported list for keloid tissues where a similar approach had been used.

The similarities in gene expression noted raise the possibility that disordered tissue remodeling may contribute to leiomyoma growth. In conclusion, global gene profiling experiments suggested the possibility that leiomyoma may arise from normal uterine cells that undergo alteration in response to disordered extracellular signals. We are now examining the hypothesis that abnormal tissue repair may contribute to fibroid development.

List of abstracts from the 3rd International Conference on the Female Reproductive Tract