ABSTRACT
Besides their regulatory role in embryogenesis, homeobox (HOX) genes are expressed in a specific manner in hematopoietic cell lineages, implying a role in the molecular regulation of hematopoiesis. Some HOX C cluster genes are found to be expressed in lymphoid cells of mice and humans. Their function and expression in normal hematopoiesis are still largely unknown. We have studied the mRNA expression of HOXC4, HOXC5, and HOXC6 in several stages of lymphocyte maturation by reverse transcriptase-polymerase chain reaction (RT-PCR) and RNA in situ hybridization (RISH). We examined CD34+/CD38low and CD34+/CD38high cells obtained from normal donor bone marrow (BM), a panel of 19 lymphoid cell lines, several types of leukemias and non-Hodgkin's lymphomas (NHL), and lymphocytes isolated from tonsillar tissue and peripheral blood (PB). HOXC4 and HOXC6 were found to be expressed during maturation in B- and T-lymphoid cells. The expression of each gene was found to be initiated at different cell maturation stages. HOXC4 transcripts were present in CD34+/CD38low cells, which are thought to comprise stem cells and noncommitted progenitor cells, and in subsequent stages to terminally maturated lymphoid cells. HOXC6 expression is initiated in equivalents of prothymocyte and pre-pre-B cell stage and remains present in mature cells. However, HOXC5 is only expressed in neoplastic cell lines and in neoplastic cells of NHL, but not in CD34+ BM cells, nor in resting or activated lymphoid cells isolated from tonsil, PB, or in leukemia cells. In cell lines, weak expression of HOXC5 is initiated in equivalents of pre-B cell and common thymocyte stage and is continuously expressed in mature cell lines. Semi-quantitative RT-PCR showed that expression levels of HOXC5 were much lower than those of HOXC4 and HOXC6; furthermore an increase of expression of HOXC4, HOXC5, and HOXC6 during lymphoid cell differentiation was demonstrated. Thus, mainly mature lymphoid cell lines and neoplastic cells of NHL do express HOXC5, in contrast to the lack of expression in normal lymphoid cells and leukemias. These findings suggest involvement of HOXC5 in lymphomagenesis.
Subject(s)
Gene Expression Regulation , Homeodomain Proteins/biosynthesis , Lymphoid Tissue/metabolism , Base Sequence , Cell Differentiation , Cell Line , Child , Gene Expression Regulation, Leukemic , Gene Expression Regulation, Neoplastic , Homeodomain Proteins/genetics , Humans , Leukemia/genetics , Leukemia/metabolism , Leukemia/pathology , Lymphoid Tissue/cytology , Lymphoma, Non-Hodgkin/genetics , Lymphoma, Non-Hodgkin/metabolism , Lymphoma, Non-Hodgkin/pathology , Molecular Sequence Data , Neoplasm Proteins/biosynthesis , Neoplasm Proteins/genetics , Polymerase Chain Reaction , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Tumor Cells, CulturedABSTRACT
OBJECTIVE: We wanted to test whether the recently described method of using the transferrin receptor system for fluorescence-activated cell-sorter enrichment of nucleated red blood cells can be used for prenatal diagnosis from maternal blood. STUDY DESIGN: Instead of the laborious, expensive fluorescence-activated cell-sorter system, we used the newly described magnetic-activated cell sorter. RESULTS: An effective enrichment could be achieved with separation of lymphocyte subsets. With the transferrin receptor, however, the enrichment was very inefficient because of the poor specificity of the antibody itself. Even in umbilical cord blood only 25% of nucleated red blood cells were labeled as demonstrated by immunogold silver enhancement of transferrin receptor-labeled cells. CONCLUSION: In spite of the availability of a fast and effective separation method (magnetic-activated cell sorter) the use of the transferrin receptor antigen alone is not likely to enable a reliable identification of fetal cells in maternal circulation.
Subject(s)
Cell Separation/methods , Erythrocytes/cytology , Fetal Blood/cytology , Magnetics , Prenatal Diagnosis/methods , Receptors, Transferrin/analysis , Erythrocyte Count , Erythrocytes/chemistry , Female , Flow Cytometry , Fluorescent Antibody Technique , Humans , Leukocyte Count , PregnancyABSTRACT
The alpha 6/beta 4 complex is a member of the integrin family of adhesion receptors. It is found on a variety of epithelial cell types, but is most strongly expressed on stratified squamous epithelia. Fluorescent antibody staining of human epidermis suggests that the beta 4 subunit is strongly localized to the basal region showing a similar distribution to that of the 230-kD bullous pemphigoid antigen. The alpha 6 subunit is also strongly localized to the basal region but in addition is present over the entire surfaces of basal cells and some cells in the immediate suprabasal region. By contrast staining for beta 1, alpha 2, and alpha 3 subunits was very weak basally, but strong on all other surfaces of basal epidermal cells. These results suggest that different integrin complexes play differing roles in cell-cell and cell-matrix adhesion in the epidermis. Immunoelectron microscopy showed that the alpha 6/beta 4 complex at the basal epidermal surface is strongly localized to hemidesmosomes. This result provides the first well-characterized monoclonal antibody markers for hemidesmosomes and suggests that the alpha 6/beta 4 complex plays a major role in epidermal cell-basement membrane adhesion. We suggest that the cytoplasmic domains of these transmembrane glycoproteins may contribute to the structure of hemidesmosomal plaques. Immunoultrastructural localization of the BP antigen suggests that it may be involved in bridging between hemidesmosomal plaques and keratin intermediate filaments of the cytoskeleton.
