ABSTRACT: Similar to the bone marrow, the mammary gland contains a distinct population of Hoechst-effluxing side population cells, MG-SPs. To better characterize MG-SPs, their microarray gene profiles were compared to the remaining cells, which retain Hoechst dye (MG-NSPs). For analysis, gene ontology (GO) that describes genes in terms of biological processes and ontology traverser (OT) that performs enrichment analysis were utilized. OT showed that MG-SP specific genes were enriched in the GO categories of cell cycle regulation and checkpoints, multi-drug resistant transporters, organogenesis, and vasculogenesis. The MG-NSP upregulated genes were enriched in the GO category of cellular organization and biogenesis which includes basal epithelial markers, p63, smooth muscle actin (SMA), myosin, alpha-6 integrin, cytokeratin (CK) 14, as well as luminal markers, CK8 and CD24. Additional studies showed that a higher percentage of MG-SPs exist in the G1 phase of the cell cycle compared to the MG-NSPs. G1 cell cycle block of MG-SPs may be explained by higher expression of cell cycle negative regulatory genes such as TGF-beta2 (transforming growth factor-beta2), IGFBP-5 (insulin like growth factor binding protein-5), P18 INK4C and Wnt-5a (wingless-5a). Accordingly, a smaller percentage of MG-SPs expressed nuclear b-catenin, possibly as a consequence of the higher expression of Wnt-5a. In conclusion, microarray gene profiling suggests that MG-SPs are a lineage deficient mammary gland sub-population expressing key genes involved in cell cycle regulation, development and angiogenesis. Supplemental File Descriptions: Table 1 is a list of 1632 Genes differentially expressed by MG-SP and MG-NSP; Criteria for comparison included 1.2-fold difference in expression levels, false discovery rate (FDR) 9.4%, P value less than 0.05. Table 2 is a list of 771 Genes differentially expressed by MG-SP and MG-NSP; Criteria for comparison included 1.5-fold difference in expression levels, FDR 3.4%, P value less than 0.05. Table 3 is a list of 335 Genes differentially expressed by MG-SP and MG-NSP; Criteria for comparison included 2-fold difference in expression levels, FDR 0%, P value less than 0.05. Table 4 is a list of 90 Genes differentially expressed by MG-SP and MG-NSP; Criteria for comparison included 2-fold difference in expression levels, FDR 0%, P value less than 0.01. Keywords: Normal C57BL/6 Mammary Gland Epithelial Cells
