Abstract
The laminin (LN) family of large heterotrimeric extracellular matrix glycoproteins has multiple functions: LNs take part in the regulation of processes such as cell migration, differentiation, and proliferation, in addition to contributing to the structure of basement membranes. LN-10, composed of alpha5, beta1, and gamma1 chains, is widely distributed in most basement membranes of both epithelia and endothelia. We determined the complete human cDNA sequence for the LN alpha5 chain and produced recombinant human LN-10 (rLN-10) in HEK293 cells by triple transfection of full-length cDNAs encoding the human LN alpha5, beta1, and gamma1 chains. The rLN-10 was purified using affinity chromatography and had an apparent molecular mass of approximately 800 kDa in SDS-PAGE and a native domain structure in rotary shadowing electron microscopy. By using function-blocking monoclonal antibodies, integrin alpha(3)beta(1) was found to be a major mediator of adhesion of HT-1080 and human saphenous vein endothelial cells. Human saphenous vein endothelial cells adhered more strongly to rLN-10 than to LN-1 and LN-8 and showed better migration on rLN-10, compared with several other matrices. Considering the cell adhesive and migration-promoting properties of rLN-10 on endothelial cells, this molecule could be useful in improving the biocompatibility and endothelialization of vascular grafts.
Highlights
The laminin (LN) family of large heterotrimeric extracellular matrix glycoproteins has multiple functions: LNs take part in the regulation of processes such as cell migration, differentiation, and proliferation, in addition to contributing to the structure of basement membranes
A partial sequence covering 953 amino acids in the C terminus of human LN ␣5 has been reported previously [12], and this contains no discrepancies compared with our sequence
The quantity of recombinant human LN-10 (rLN-10) produced by the HEK293 cells in monolayer cultures was quite high, considering the size and complexity of the protein
Summary
The laminin (LN) family of large heterotrimeric extracellular matrix glycoproteins has multiple functions: LNs take part in the regulation of processes such as cell migration, differentiation, and proliferation, in addition to contributing to the structure of basement membranes. Have been identified [1, 2] These LN isoforms are expressed in tissue-specific and developmentally regulated patterns, and they play significant roles in the adhesion, migration, proliferation, and differentiation of many cell types [3,4,5]. LN-8 seems to be an important component of the newly formed endothelial basement membranes because mice lacking the ␣4 chain exhibit neonatal hemorrhages due to unstable microvessels [9]. LN ␣5 chain-containing isoforms may be important in placental endothelial cell migration and blood vessel branching and in formation of proper basal laminae
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