| Abstrakt: |
Tumor cell interactions with fibronectin (FN) are important for the development of secondary tumors inside the bone marrow stroma. We studied and compared thein situdistribution of FN in paraffin-embedded human bone marrow sections and investigated thein vitroregulation of FN assemblage by bone marrow stromal cells (BMSC). Finally, the role of FN in the interaction of BMSC with tumor cells was studied. Fine elongated FN-positive cell extensions, probably of stromal cell origin, were observed as well as a limited amount of extracellular FN deposits in connective tissues around capillaries and sinusoids.In vitrostudies, using the confocal laser scanning microscope, showed that BMSC produced a high amount of FN with a characteristic extracellular matrix formation in an extensive network. FN matrix formation was predominantly detected at contact sites between cultured BMSC. Inin vitrocultures with low cell concentrations andin vivowith a limited number of stromal cell contacts only limited matrix was found. From previous studies it is known that the α5β1 integrin is involved in the regulation of FN assembly. Here the role of the α5-subunit of this integrin was investigated. By using two different monoclonal antibodies (mAb) against the α5-subunit (2H6 and mAb16) the assembly of endogenous FN was completely blocked, indicating that these antibodies are directed against the active epitope. Another mAb (mAb11) against the α5-subunit did not affect the FN assemblage. Codistribution analysis of α5-subunits, αv-subunits, actin, and FN demonstrated that the α5β1 integrin is associated with FN and not with intracellular actin. Integrins αvβ1, αvβ3, and αvβ5, also ligands of FN, did not colocalize with FN. Codistribution of αv with the terminal ends of actin and not with FN indicates that αv-subunits are mainly directed to vitronectin rather than to FN. The dominant role of α5β1 in FN interaction is underlined by effective blocking of tumor cell adhesion with BMSC using anti-α5, anti-β1, and anti-FN antibodies. These results emphasize the important role of α5 integrin subunit in FN matrix assembly in human BMSC and an exclusive role of α5β1 in the anchorage and regulation of FN-mediated adhesion processes in the bone marrow. |
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