The human placenta utilizes both active and passive mechanisms to evade rejection by the maternal immune system. We investigated the pattern of expression of the B7 family of immunomodulatory molecules B7-H1 (PD-L1), B7-2 (CD86), and B7-1 (CD80) at the term maternal-fetal interface. Northern blot and reverse transcription-polymerase chain reaction (RT-PCR) analyses showed that B7-H1 mRNA is abundant in term placenta and that cytotrophoblasts are sources of this message. Immunohistochemistry demonstrated that B7-H1 is constitutively expressed by the syncytiotrophoblast and by extravillous cytotrophoblasts, both of which are juxtaposed to maternal blood and tissue. By contrast, placental stromal cells, including macrophages, lacked the protein. Expression of B7-H1 protein was low in first-trimester placenta compared to second- and third-trimester tissue (P < 0.05) and was enhanced in cultured cytotrophoblasts by treatment with either interferon-gamma or epidermal growth factor (P < 0.05), suggesting that one or both of these mediators regulates B7-H1 expression in the placenta. RT-PCR and immunofluorescence analysis of term placental tissue revealed different patterns of expression of the immunostimulatory protein, B7-2. In contrast to B7-H1, B7-2 mRNA and protein were absent in cytotrophoblast cells but present in maternal macrophages and some fetal macrophages. The B7-1 mRNA and protein were absent at the maternal-fetal interface. These studies document expression of the B7 family proteins at the maternal-fetal interface and demonstrate that B7-H1 is positioned such that it could facilitate protection of fetal cells against activated maternal leukocytes. Conversely, B7-2 was absent on trophoblasts and was appropriately localized to fetal and maternal macrophages, which may participate in antigen presentation.

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The HLA-G message is alternatively spliced into multiple transcripts, two of which encode soluble isoforms. To initiate studies on the specific functions of the soluble isoforms, we produced soluble rHLA-G1 (rsG1) and rsG2 in human embryonic kidney 293 cells and characterized the proteins. Both isoforms were glycosylated and formed disulfide-bonded oligomers. Recombinant sG1 associated with β2-microglobulin, whereas rsG2 did not. Mouse mAb generated to rsG1 (1-2C3), which identified exclusively sG1, and mAb generated to rsG2 (26-2H11), which identified both soluble and membrane G2 (m/sG2), were used for immunohistochemical isoform mapping studies on placental tissue sections. Soluble G1 protein was abundant in many subpopulations of trophoblast cells, whereas m/sG2 protein was present exclusively in extravillous cytotrophoblast cells. Although both isolated placental villous cytotrophoblast cells and chorion membrane extravillous cytotrophoblast cells contained mRNAs encoding sG1 and sG2, protein expression was as predicted from the immunostains with m/sG2 present only in the invasive trophoblast subpopulation. Analysis of function by Northern and Western blotting demonstrated that both rsG1 and rsG2 inhibit CD8α expression on PBMC without changing CD3δ expression or causing apoptotic cell death. Collectively, the studies indicate that: 1) both sG1 and m/sG2 are produced in placentas; 2) transcription and translation are linked for sG1, but not G2; 3) expression of G2 is exclusively associated with the invasive phenotype; and 4) the two isoforms of sG may promote semiallogeneic pregnancy by reducing expression of CD8, a molecule required for functional activation of CTL.

Although previous in vitro studies predicted that CCN5/ WISP-2 may act as an anti-invasive gene in breast cancer, the distribution pattern of CCN5 in breast cancer samples is conflicting. Thus, we systematically investigated the CCN5 expression profile in noninvasive and invasive breast tumor samples and its functional relevance in breast cancer progression. The studies showed that CCN5 expression is biphasic, such that in normal samples CCN5 expression is undetectable, whereas its expression is markedly increased in noninvasive breast lesions, including atypical ductal hyperplasia and ductal carcinoma in situ. Further, CCN5 mRNA and protein levels are significantly reduced as the cancer progresses from a noninvasive to invasive type. Additionally, we showed that CCN5 mRNA and protein level was almost undetectable in poorly differentiated cancers compared with the moderately or well-differentiated samples and its expression inversely correlated with lymph node positivity. The result was further supported by evaluating the RNA expression profile in microdissected sections using real-time PCR analysis. Therefore, our data suggest a protective function of CCN5 in noninvasive breast tumor cells. This hypothesis was further supported by our in vitro studies illuminating that CCN5 is a negative regulator of migration and invasion of breast cancer cells, and these events could be regulated by CCN5 through the modulation of the expression of genes essential for an invasive front. These include Snail-E-cadherin signaling and matrix metalloproteinase (MMP)-9 and MMP-2. Collectively, these studies suggest that the protective effect of CCN5 in breast cancer progression may have important therapeutic implications. [Cancer Res 2008;68(18):7606-12]

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