N-cadherin is developmentally regulated and functionally involved in early hematopoietic cell differentiation.

The cadherins, an important family of cell adhesion molecules, are known to play major roles during embryonic development and in the maintenance of solid tissue architecture. In the hematopoietic system, however, little is known of the role of this cell adhesion family. By RT-PCR, western blot analysis and immunofluorescence staining we show that N-cadherin, a classical type I cadherin mainly expressed on neuronal, endothelial and muscle cells, is expressed on the cell surface of resident bone marrow stromal cells. FACS analysis of bone marrow mononuclear cells revealed that N-cadherin is also expressed on a subpopulation of early hematopoietic progenitor cells. Triple-color FACS analysis defined a new CD34(+) CD19(+) N-cadherin(+) progenitor cell population. During further differentiation, however, N-cadherin expression is lost. Treatment of CD34(+) progenitor cells with function-perturbing N-cadherin antibodies drastically diminished colony formation, indicating a direct involvement of N-cadherin in the differentiation program of early hematopoietic progenitors. N-cadherin can also mediate adhesive interactions within the bone marrow as demonstrated by inhibition of homotypic interactions of bone-marrow-derived cells with N-cadherin antibodies. Together, these data strongly suggest that N-cadherin is involved in the development and retention of early hematopoietic progenitors within the bone marrow microenvironment.

Adhesive interactions of human multiple myeloma cell lines with different extracellular matrix molecules.

Multiple myeloma represents a human B cell malignancy which is characterized by a predominant localization of the malignant cell clone within the bone marrow. With the exception of the terminal stage of the disease the myeloma tumor cells do not circulate in the peripheral blood. The bone marrow microenvironment is believed to play an important role in homing, proliferation and terminal differentiation of myeloma cells. Here we have studied the expression of several extracellular matrix (ECM) molecules in the bone marrow of multiple myeloma patients and analyzed their adhesive capacities with four different human myeloma-derived cell lines. All ECM molecules analyzed (tenascin, laminin, fibronectin, collagen types I, III, V and VI) could be detected in bone marrow cryostat sections of multiple myeloma patients. Adhesion assays showed that only laminin, the microfibrillar collagen type VI and fibronectin were strong adhesive components for the myeloma cell lines U266, IM-9, OPM-2 and NCI-H929. Tenascin and collagen type I were only weak adhesive substrates for these myeloma cells. Adhesion to laminin and fibronectin was beta 1-integrin-mediated since addition of anti-beta 1-integrin antibodies could inhibit the binding of the four different cell types to both matrix molecules. In contrast, integrins do not seem to be involved in binding of the myeloma cells to collagen type VI. Instead, inhibition of binding by heparin suggested that membrane-bound heparan sulfate proteoglycans are responsible ligands for binding to collagen type VI. Adhesion assays with several B-cell lines resembling earlier differentiation stages revealed only weak interactions with tenascin and no interactions with collagen type VI, laminin or fibronectin. In summary, the interactions of human myeloma cells with the extracellular matrix may explain the specific retention of the plasma cells within the bone marrow.

Cell binding properties of collagen type XIV for human hematopoietic cells.

Collagen XIV, which belongs to the subclass of fibril-associated collagens with interrupted triple helices (FACITs), is a homotrimeric molecule consisting of three alpha 1 (XIV) chains. Collagen type XIV is strongly expressed in the native human bone marrow, as shown by immunofluorescence staining and immunoblotting with an affinity-purified antibody. Hematopoietic cell lines of myeloid (KG1a, U937, K562) and lymphoid (U266, IM-9) origin were able to attach firmly to purified human collagen XIV preparations. Attachment of these cells was shown to be concentration-dependent. However, other hematopoietic cell lines tested were unable to adhere to collagen XIV, indicating restriction of this cellular interaction. The cellular receptors involved in cell binding to collagen type XIV are probably membrane-bound heparansulfate proteoglycans, since only the the addition of heparin inhibited attachment of the hematopoietic cells to collagen XIV in a concentration-dependent manner. Antibodies against the beta 1-integrin subunit could not interfere with binding to collagen type XIV. Using purified fragments of collagen XIV, it could be demonstrated that at least two different heparin-sensitive adhesion sites are present in the N-terminal globular domain and in the triple-helical domain. These data indicate that collagen XIV represents another collagen type expressed in human bone marrow with strong cell binding properties for defined populations of hematopoietic cells.