CBL194F Sigma-AldrichAnti-Cytokeratin 7 Antibody, clone LP5K, FITC conjugated

In early pregnancy, human extravillous trophoblasts (EVTs) invade and remodel maternal arteries. We have previously demonstrated that CCR1 is expressed on perivascular/endovascular trophoblasts and that CCR1 ligands promote EVT migration. In this study, we examined the physiologic roles of platelet-derived chemoattractants on EVT invasion. By immunohistochemistry, maternal platelets were localized among endovascular trophoblasts within the lumen of spiral arteries. Extracellular matrices (ECMs) were also detected among endovascular trophoblasts and platelets, suggesting that the platelets in these arteries were activated by ECMs. In vitro, platelets attached to EVTs isolated from human villous explant cultures and expressed P-selectin on the cell surface. Platelets significantly enhanced migration of EVTs without affecting proliferation of EVTs or secretion of MMP-2 or MMP-9. The invasion-enhancing effect of platelet-derived culture medium on EVTs was neutralized by anti-CCR1 antibody. Heat treatment completely abrogated the invasion-promoting effects of platelet-derived culture medium, but charcoal stripping did not. Platelets also induced endovascular trophoblast-like morphologic changes and integrin alpha1 expression in EVTs during 48-hour culture. These findings suggest that maternal platelets activated in the spiral arteries can regulate trophoblastic vascular infiltration and differentiation by releasing various soluble factors.

At the human feto-maternal interface, trophoblasts differentiate towards extravillous trophoblasts (EVTs) and form the cell column. EVTs acquire invasive activity in the distal part of the cell column and begin to migrate into the maternal tissue. We previously reported that dipeptidyl peptidase IV (DPPIV) is expressed on EVTs in the proximal part of cell column and is involved in the inhibition of their migration. Because DPPIV has been shown to degrade several chemokines, we examined possible roles of chemokines in EVT migration. Immunohistochemistry demonstrated that C-C chemokine receptor 1 (CCR1) was hardly detected on cytotrophoblasts and syncytiotrophoblast but was expressed on EVTs in the cell column. In vitro, CCR1 protein was also present on the surface of EVTs that grew out from chorionic villous explants cultured under 20% O2. Chemokines that can bind to CCR1 (CCR1 ligands), such as regulated on activation, normal T cell expressed and secreted (RANTES) and macrophage inflammatory protein-1alpha (MIP-1alpha), were confirmed in the decidual tissues by RT-PCR and immunohistochemistry. These CCR1 ligands promoted the migration of the EVTs that were isolated from the explant cultures in vitro. These results indicate that CCR1 is expressed on trophoblasts as they differentiate to EVTs and that CCR1 ligands produced from the decidual tissue induce EVT migration. By contrast, CCR1 was scarcely expressed on EVTs that grew out from villous explants cultured in 1% O2, indicating that a relatively high oxygenic environment is needed to induce CCR1 expression. Moreover, CCR1 expression on the isolated EVTs was significantly reduced in the presence of decidua-conditioned medium. Such regulation of CCR1 by surrounding oxygenic and decidual environments supports a close correlation between EVT invasion and their expression of CCR1. This study demonstrates that trophoblasts acquire CCR1 as they differentiate to an invasive phenotype at the villus-anchoring sites and indicates a novel role for the chemokine-CCR1 system in the initial step of trophoblastic invasion towards the maternal tissue.

Reactivity patterns of seven mouse monoclonal antibodies to human keratin 7 were compared using immunoblotting and immunohistochemistry on cultured cells and normal human and animal tissues. Differences in keratin specificities as determined by two-dimensional immunoblots and interspecies cross-reactivity data on 8 mammalian species suggest that at least six nonidentical epitopes of the keratin 7 molecule are recognized by this panel of reagents. Immunohistochemical examination of a panel of various human neoplasms with monoclonal antibodies monospecific for keratins 7, 18 and 19 revealed potential value of keratin subtyping in differential diagnosis of tumors in general and in subclassification of carcinomas in particular.