Gene expression profiling supports the hypothesis that human ovarian surface epithelia are multipotent and capable of serving as ovarian cancer initiating cells

Abstract

Background: Accumulating evidence suggests that somatic stem cells undergo mutagenic transformation into cancer initiating cells. The serous subtype of ovarian adenocarcinoma in humans has been hypothesized to arise from at least two possible classes of progenitor cells: the ovarian surface epithelia (OSE) and/or an as yet undefined class of progenitor cells residing in the distal end of the fallopian tube.

Methods: Comparative gene expression profiling analyses were carried out on OSE removed from the surface of normal human ovaries and ovarian cancer epithelial cells (CEPI) isolated by laser capture micro-dissection (LCM) from human serous papillary ovarian adenocarcinomas. The results of the gene expression analyses were randomly confirmed in paraffin embedded tissues from ovarian adenocarcinoma of serous subtype and non-neoplastic ovarian tissues using immunohistochemistry. Differentially expressed genes were analyzed using gene ontology, molecular pathway, and gene set enrichment analysis algorithms.

Results: Consistent with multipotent capacity, genes in pathways previously associated with adult stem cell maintenance are highly expressed in ovarian surface epithelia and are not expressed or expressed at very low levels in serous ovarian adenocarcinoma. Among the over 2000 genes that are significantly differentially expressed, a number of pathways and novel pathway interactions are identified that may contribute to ovarian adenocarcinoma development.

Conclusions: Our results are consistent with the hypothesis that human ovarian surface epithelia are multipotent and capable of serving as the origin of ovarian adenocarcinoma. While our findings do not rule out the possibility that ovarian cancers may also arise from other sources, they are inconsistent with claims that ovarian surface epithelia cannot serve as the origin of ovarian cancer initiating cells.

Figure 1

Hierarchical clustering and gene ontology (GO) enrichment of 2320 genes differentially expressed between ovarian surface epithelial cells and ovarian cancer epithelial tissue. The heat map (left) was generated by Z-score normalization of log2 expression values from Affymetrix HGU133 Plus 2.0 3. Displayed are the relative expression levels of genes (rows) differentially expressed (red = relatively over-expressed; green = relatively under-expressed) in 12 ovarian surface epithelial brushings and 12 laser capture microdissected malignant epithelia samples (columns). Unique, enriched GO terms are listed for each set of differentially expressed genes and their statistical significance by false discovery rate (FDR) corrected, hypergeometric distribution p-values. Genes overexpressed in CEPI are labeled as CEPI Genes overexpressed in OSE are labeled OSE Genes.

Figure 2

Cell cycle pathway gene expression. Shown is a GenMAPP http://www.genmapp.org rendering of a modified KEGG (Kyoto Encyclopedia of Genes and Genomes, http://www.genome.jp/kegg/) schematic of cell cycle pathway genes. Genes significantly overexpressed in CEPI relative to OSE are colored red. The execution of the cell cycle is depicted from left to right and individual phases identified below by I- beam brackets. Genes involved in maintaining G1 are generally under-expressed in CEPI while genes involved in G1 to S progression, G2, and M are over-expressed.

Figure 3

Pathway deregulation in ovarian cancer. Individual signaling pathways hypothesized to be deregulated in the oncogenic transformation of ovarian surface epithelia (OSE) into ovarian cancer epithelia (CEPI). An individual OSE is represented on the left with individual pathways (as discussed in the text) labeled adjacent to their respective section of each cell. An individual CEPI with the same signaling pathways is represented on the right as a mirror image of the OSE cell. The legend describes the colored boxes and lines used to represent expression differences and potential interactions among genes. The juxtapositional placement of the two cell halves is meant to emphasize the dichotomous state of signaling between the OSE and CEPI as revealed by our gene expression microarrays.

Figure 4

Immunohistochemistry of OSE gene products. Immunohistochemistry was performed on fresh frozen paraffin embedded OSE and CEPI tissue samples. Staining with primary antibodies against (A) secreted frizzled-related protein 1 (SFRP1), (B) aldehyde dehydrogenase 1 family, member A2 (ALDH1A2), and (C and D) lim homeobox 9 (LXH9) indicated strong protein expression in OSE (labeled arrow) and lower or absent protein expression in CEPI (labeled arrow), consistent with mRNA expression values shown in Additional file 5. Each image is a typical representative from 10 normal and 10 cancer slides. The slides chosen for display contain both CEPI and normal adjacent OSE from the same tissue sample.