MAB3236 Sigma-AldrichAnti-Cytokeratin 18 Antibody, clone RCK106

Previous attempts to isolate pluripotent cell lines from rat preimplantation embryo in mouse embryonic stem (ES) cell culture conditions (serum and LIF) were unsuccessful, however the resulting cells exhibited the expression of such traditional pluripotency markers as SSEA-1 and alkaline phosphatase. We addressed the question, which kind of cell lineages are produced from rat preimplantation embryo under "classical" mouse ES conditions.We characterized two cell lines (C5 and B10) which were obtained from rat blastocysts in medium with serum and LIF. In the B10 cell line we found the expression of genes known to be expressed in trophoblast, Cdx-2, cytokeratin-7, and Hand-1. Also, B10 cells invaded the trophectodermal layer upon injection into rat blastocysts. In contrast to mouse Trophoblast Stem (TS) cells proliferation of B10 cells occurred independently of FGF4. Cells of the C5 line expressed traditional markers of extraembryonic-endoderm (XEN) cells, in particular, GATA-4, but also the pluripotency markers SSEA-1 and Oct-4. C5 cell proliferation exhibited dependence on LIF, which is not known to be required by mouse XEN cells.Our results confirm and extend previous findings about differences between blastocyst-derived cell lines of rat and mice. Our data show, that the B10 cell line represents a population of FGF4-independent rat TS-like cells. C5 cells show features that have recently become known as characteristic of rat XEN cells. Early passages of C5 and B10 cells contained both, TS and XEN cells. We speculate, that mechanisms maintaining self-renewal of cell lineages in rat preimplantation embryo and their in vitro counterparts, including ES, TS and XEN cells are different than in respective mouse lineages.

Using a panel of 21 monoclonal and 2 polyclonal keratin antibodies, capable of detecting separately 11 subtypes of their epithelial intermediate filament proteins at the single cell level, we investigated keratin expression in 16 squamous cell carcinomas, 9 adenocarcinomas, and 3 adenosquamous carcinomas of the human uterine cervix. The keratin phenotype of the keratinizing squamous cell carcinoma was found to be most complex comprising keratins 4, 5, 6, 8, 13, 14, 16, 17, 18, 19, and usually keratin 10. The nonkeratinizing variety of the squamous cell carcinoma expressed keratins 6, 14, 17, and 19 in all cases, usually 4, 5, 7, 8, and 18, and sometimes keratins 10, 13, and 16. Adenocarcinomas displayed a less complex keratin expression pattern comprising keratins 7, 8, 17, 18, and 19, while keratin 14 was often present and keratins 4, 5, 10 and 13 were sporadically found in individual cells in a few cases. These keratin phenotypes may be useful in differential diagnostic considerations when distinguishing between keratinizing and nonkeratinizing carcinomas (using keratin 10, 13, and 16 antibodies), and also in the distinction between nonkeratinizing carcinomas and poorly differentiated adenocarcinomas, which do not express keratins 5 and 6. Keratin 17 may also be useful in distinguishing carcinomas of the cervix from those of the colon and also from mesotheliomas. Furthermore the presence of keratin 17 in a CIN I, II, or III lesion may indicate progressive potential while its absence could be indicative of a regressive behavior. Because most carcinomas express keratins 8, 14, 17, 18, and 19, we propose that this expression pattern reflects the origin of cervical cancer from a common progenitor cell, i.e., the endocervical reserve cell that has been shown to express keratins 5, 8, 14, 17, 18, and 19.

To study the role of the amino-terminal domain of the desmin subunit in intermediate filament (IF) formation, several deletions in the sequence encoding this domain were made. The deleted hamster desmin genes were fused to the RSV promoter. Expression of such constructs in vimentin-free MCF-7 cells as well as in vimentin-containing HeLa cells, resulted in the synthesis of mutant proteins of the expected size. Single- and double-label immunofluorescence assays of transfected cells showed that in the absence of vimentin, desmin subunits missing amino acids 4-13 are still capable of filament formation, although in addition to filaments large numbers of desmin dots are present. Mutant desmin subunits missing larger portions of their amino terminus cannot form filaments on their own. It may be concluded that the amino-terminal region comprising amino acids 7-17 contains residues indispensable for desmin filament formation in vivo. Furthermore it was shown that the endogenous vimentin IF network in HeLa cells masks the effects of mutant desmin on IF assembly. Intact and mutant desmin colocalized completely with endogenous vimentin in HeLa cells. Surprisingly, in these cells endogenous keratin also seemed to colocalize with endogenous vimentin, even if the endogenous vimentin filaments were disturbed after expression of some of the mutant desmin proteins. In MCF-7 cells some overlap between endogenous keratin and intact exogenous desmin filaments was also observed, but mutant desmin proteins did not affect the keratin IF structures. In the absence of vimentin networks (MCF-7 cells), the initiation of desmin filament formation seems to start on the preexisting keratin filaments. However, in the presence of vimentin (HeLa cells) a gradual integration of desmin in the preexisting vimentin filaments apparently takes place.