Laminin-8/9 is synthesized by rat glomerular mesangial cells and is required for PDGF-induced mesangial cell migration

Abstract

Background. Laminin (LM), the major glycoprotein component of basement membranes is expressed as multiple isoforms in a developmentally regulated and tissue-specific manner. LM α4 has a limited tissue distribution and is highly expressed in the developing glomerulus. In the present study, we investigate the in vivo and in vitro expression and function of LM α4 in the glomerulus. Methods. LM α4 expression was examined by Northern blot, reverse transcription polymerase chain reaction (RT-PCR), Western blot, and immunofluorescence microscopy. Mesangial cells (MC) were plated on purified LM-1, LM-2, and LM-8/9. Immunofluorescence microscopy was performed to examine the cellular phenotypes induced by LM-1 and LM-8/9. A modified Boyden chamber method was used to assess laminin participation in platelet-derived growth factor (PDGF)-stimulated migration. Results. mRNA for LMα4 is expressed in cultured rat MC, and isolated rat and mouse glomeruli, but not in cultured rat glomerular epithelial cells or glomerular endothelial cells. Using antibodies specific for LM α4, a 240 kD band was detected in MC extract and a slightly smaller band was identified in extracted rat glomeruli. Purified LM-8/9 had MC adhesive activity comparable to LM-1 and LM-2. MC attached to LM-8/9 exhibited a unique phenotype. In contrast to LM-1, attachment of MC to LM-8/9 produced a highly arborized cell morphology with significantly reduced formation of focal contacts or stress fibers. LM α4 is utilized by MC during PDGF-stimulated migration. Conclusion. LM α4 is synthesized by MC and persists in the mature glomerulus. LM-8/9 stimulates a unique cellular morphology, and they are utilized in PDGF-induced migration. These factors suggest that LM α4 plays an important role in MC differentiation and in the maintenance of MC phenotype.