Cancer Res Treat.  2004 Feb;36(1):31-42.

Expression Profiling of the Cellular Processes in Uterine Leiomyomas: Omic Approaches and IGF-2 Association with Leiomyosarcomas

• Bae SM
• Kim YW
• Lee JM
• Namkoong S
• Kim CK
• Ahn WS

Abstract

PURPOSE
This study utilized both cDNA microarray and 2D protein gel electrophoresis technology to investigate the multiple interactions of the genes and proteins involved in the pathophysiology of uterine leiomyomas. Also, Gene Ontology (GO) analysis was used to systematically characterize the global expression profiles, which were found to correlate with the leiomyosarcomas. MATERIALS AND METHODS: The uterine leiomyoma biopsies were obtained from patients in the Department of Obstetrics and Gynecology, The Catholic University of Korea. Differentially expressed transcriptome and proteome, in 6 paired leiomyoma and normal myometrium, were profiled. The total RNAs from the leiomyoma and normal myometrium were labeled with Cy5 and Cy3. All specimens were punch-biopsy-obtained, and frozen in liquid nitrogen. RESULTS: Screening of up to 17, 000 genes identified 71 that were either up-regulated or down-regulated (21 and 50, respectively). The gene expression profiles were classified into 420 mutually dependent functional sets, resulting in 611 cellular processes, according to the gene ontology. Also, the protein analysis, using 2D gel electrophoresis, identified 33 proteins (17 up-regulated and 16 down-regulated) with more than 500 total spots, which were classified into 302 cellular processes. Of these functional profilings, transcriptomes and proteoms down- regulations were shown in the cell adhesion, cell motility, organogenesis, enzyme regulator, structural molecule activity and responses to external stimulus functional activities, which are supposed to play important roles in the pathophysiology. In contrast, up-regulation was only shown in the nucleic acid binding activity. The CDKN2A, ADH1A, DCX, IGF2, CRABP2 and KIF5C were found to increase the reliability of this study, and correlate with the leiomyosarcomas. CONCLUSION: Potentially significant pathogenetic cellular processes showed that down-regulated functional profiling has an important impact on the discovery of the pathogenic pathways in leiomyomas and leiomyosarcomas. GO analysis can also overcome the complexity of the expression profiles of cDNA microarrays and 2D protein analyses, via a cellular process level approach. Thereby, a valuable prognostic candidate gene, with real relevance to disease-specific pathogenesis, can be found at cellular process levels.

Keyword

Uterine leiomyoma; Leiomyosarcoma; cDNA microarray; Proteomics; Gene ontology

MeSH Terms

Animals
Biopsy
Cell Adhesion
Cell Movement
Electrophoresis
Electrophoresis, Gel, Two-Dimensional
Female
Gene Ontology
Gynecology
Humans
Insulin-Like Growth Factor II*
Korea
Leiomyoma*
Leiomyosarcoma*
Mass Screening
Mice
Myometrium
Nitrogen
Obstetrics
Oligonucleotide Array Sequence Analysis
Organogenesis
Proteome
Proteomics
RNA
Social Control, Formal
Transcriptome
Up-Regulation
Insulin-Like Growth Factor II
Nitrogen
Proteome
RNA

Figure

• 

  Fig. 1 Hierarchical cluster analysis of uterine leiomyoma. (A) Type I assay: All the data were median centered; and clustered using a hierarchical clustering. Levels of intensity of red squares correlate with the degree of gene expression; conversely, green squares compare the down-expression, at a scale relative to the color intensity. A cluster image, representing 44 of the cDNAs, is shown. (B) Type II assay (differential expression between uterine leiomyoma and normal myometrium): Type II analysis was used to identify the genes differentially expressed in the uterine leiomyoma, by universal control cell lines, selecting 44 genes. Those genes showing statistically significant differences between the 2 groups are shown as a cluster image. The dendrogram indicates uterine leiomyoma in red, with normal myometrium in green. The normal myometrims cluster together, as do the leiomyomas. The column on the left of the 2-dimensional view indicates the down- regulated genes in the myometrium in green, with those up-regulated in red; conversely, the column on the right indicates the down-regulated genes in the leiomyoma in green, with those up-regulated in red.

• 

  Fig. 2 RT-PCR analysis of selected genes confirmed the differential expression of the cDNA microarray. Total RNAs obtained from normal myometrium (lane 1) and leiomyoma (lane 2) were subjected to RT-PCR assays, as described in Materials and methods.

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  Fig. 3 Uterine leiomyoma protein products on 2D gels. Identification of the proteins in 30 spots, which are indicated by the red arrowheads (A: myometrium), and the blue arrowheads (B: leiomyoma), are shown in Table 2. The 2D gel patterns indicated that of the 30 spots, four were tissue heterogeneity-independent (indicated in bold type).

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