Recombinant human laminin-10 (alpha5beta1gamma1). Production, purification, and migration-promoting activity on vascular endothelial cells.
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.
Doi
Masayuki
M
Division of Matrix Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Scheeles vag 2, 17177 Stockholm, Sweden.
Thyboll
Jill
J
Kortesmaa
Jarkko
J
Jansson
Katarina
K
Iivanainen
Antti
A
Parvardeh
Masomeh
M
Timpl
Rupert
R
Hedin
Ulf
U
Swedenborg
Jesper
J
Tryggvason
Karl
K
D001483:Base Sequence; D002448:Cell Adhesion / Q000502:physiology; D002460:Cell Line; D002465:Cell Movement / Q000502:physiology; D002846:Chromatography, Affinity / Q000379:methods; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary; D006801:Humans; D007797:Laminin / Q000235:genetics* / Q000302:isolation & purification / Q000502:physiology / Q000648:ultrastructure; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000648:ultrastructure
D017931:DNA Primers; D018076:DNA, Complementary; D007797:Laminin; D011994:Recombinant Proteins; C119013:laminin 10