Ki67 is a nuclear protein that is tightly linked to the cell cycle. It is a marker of cell proliferation and has been used to stratify good and poor prognostic categories in invasive breast cancer. Its correlation with gene expression patterns has not been fully elucidated. In this study, Ki67 immunohistochemistry using the MIB-1 antibody was performed on sections cut from 21 formalin-fixed, paraffin-embedded invasive breast cancers. Scoring was determined as nil (no immunostaining), low (10% or less immunopositivity) or high (410% immunoreactive cells) respectively. The relationship of Ki67 immunohistochemical detection with clinicopathologic parameters was evaluated. Using Affymetrix U133A GeneChips, expression profiles for these tumors were generated and correlated with Ki67 immunohistochemical findings. Analysis of variance was used to define genes that were differentially regulated between the groups. Real-time polymerase chain reaction (PCR) was used to confirm the presence of a downregulated gene. Our results showed high, low and nil Ki67 immunostaining in nine (43%), six (28.5%) and six (28.5%) invasive breast cancers respectively, with increased Ki67 protein expression correlating with high histologic grade (P ¼ 0.02), mitotic score (P ¼ 0.001) and estrogen receptor immunonegativity (P ¼ 0.002). Expression profiling trends of the Ki67 gene mirrored the observed proportions of immunostained cells when the Ki67 immunoscore was 410%. Genes related to apoptosis and cell death (bcl2, MAP2K4, TNF10) were noted to be downregulated in tumors that disclosed 440% Ki67 immunostaining (Po0.001). Downregulation of the bcl2 gene was confirmed at the RNA level by real-time RT-PCR. Differential regulation of these genes, especially bcl2, may contribute to the biological nature of clinically more aggressive and highly proliferative breast cancers.

Dendritic cells (DCs) exhibit distinct functional properties at immature and mature states. To identify genes preferentially regulated in monocyte-derived immature DCs (imDCs), 13 000-element microarrays were hybridized with RNA isolated from imDCs, mature DCs (mDCs), monocytes, and macrophages and a TGF-␤-induced protein (␤ig-h3) was identified as being most prominently up-regulated in imDCs. By polymerase chain reaction (PCR), little ␤ig-h3 mRNA was detected in monocytes and macrophages, but it was abundant in imDCs. On DC activation with LPS, ␤ig-h3 mRNA became diminished, and in tissues, ␤ig-h3 mRNA was abundantly expressed in lymphoid-rich tissues such as the spleen, bone marrow, small intestines, and colon. ␤ig-h3 was expressed in 293T cells and purified as a 70-kDa protein and, by Western blotting, ␤ig-h3 was predominantly detected in the medium of imDCs. We demonstrate that ␤ig-h3 binds to macrophages and imDCs but not to mDCs and activates the Rac GTPase in macrophages, stimulating macrophage membrane ruffling and enhancing macrophage endocytosis. imDC endocytosis was also inhibited by purified anti-␤ig-h3 antibodies. Therefore, ␤ig-h3 appears to be selectively up-regulated in imDCs to regulate antigen uptake through endocytosis. (Blood. 2006;107:2777-2785)