Cloning of the Mouse Laminin α4 cDNA. Expression in a Subset of Endothelium

Abstract

Endothelial cells express a 400-kDa or 240-kDa laminin a chain, depending on their tissue of origin or physiological state [l, 21. Using differential display and subsequent screening of a mouse endothelial cell cDNA library we here identify the gene coding for the 240-kDa laminin chain as the laminin a4 gene. The complete mouse laminin a4 cDNA sequence is reported and compared with other laminin a chains. In situ hybridization of embryonic and new born mouse tissues revealed expression of laminin a4 mRNA in a subset of endothelium, in particular aortic endothelium, endocardium and endothelium of blood vessels in the skin and in the brain. Strong laminin a4 expression by aortic endothelia was confirmed by data obtained from cultured bovine aortic endothelial cells (BAEC). Isolation of laminin from BAEC conditioned medium revealed a Y-shaped molecule in rotary shadowing. Subsequent sequencing of BAEC laminin resulted in laminin a4, pl and y l amino acid sequences, confirming that laminin a4 is one of the major laminin a chains expressed by aortic endothelium not only in the mouse. In addition, strong laminin a4 mRNA expression occurred in peripheral nerves, cardiac muscle, fat, the dermis of the skin and lung stroma of mouse tissues. The data demonstrate a cytokine and progesterone-regulated differential expression of laminin a4 mRNA in mouse endothelium, suggesting a distinct functional role for this laminin chain in endothelium.

A High Endothelial Venule Secretory Protein, Mac25/Angiomodulin, Interacts with Multiple High Endothelial Venule-Associated Molecules Including Chemokines

“…such as fibronectin and vitronectin. In addition, it failed to bind various LN isoforms that are also expressed in the blood vascular basal lamina, including LN-8 and -10/11 (32)(33)(34). Thus, these results extend previous observations (11) and indicate that mac25/ AGM interacts with restricted members of the ECM proteins that are expressed in the basal lamina of HEVs.…”

Complex interactions between the laminin α4 subunit and integrins regulate endothelial cell behavior in vitro and angiogenesis in vivo

“…Compared with ␣1, ␣2, and ␣5, the ␣4 subunit present in laminins 8 and 9 contains a truncated N terminus (8)(9)(10). In this regard, it is similar to the ␣3 subunit present in laminins 5, 6, and 7.…”

Venular basement membranes contain specific matrix protein low expression regions that act as exit points for emigrating neutrophils

“…Total RNA was reverse transcribed using SuperScript II Reverse Transcriptase (Invitrogen) and cDNAs of interest were amplifi ed with BIOTaq DNA (Bioline) in a PTC-100 Thermocycler (Bio-Rad Laboratories), using specifi c primers (MWG) for LN-α5 chain (sense: 5′-A C G G C T C A G A A G G T T-T C C C G -3′; antisense: 5′-C T T C A G A C A G C C G C T G A A C C -3′) (27) and LN-α4 chain (sense: 5′-G T C A G C T A G C G G A T G C G C C T T C A T G G G -3′; antisense: 5′-C A G T C G G C C G C G G C T G T G G G A C A G G A G T T G -3′) (13) or β-actin (sense: 5′-G T G G G C C G C C C T A G G C A C C A G -3′; antisense:…”

“…To investigate the expression of venular BM constituents, cremaster muscles were immunostained for laminin 8 (α4β1γ1 chains) and 10 (α5β1γ1 chains), the principal vascular laminin isoforms (6,(12)(13), in parallel with other key basement membrane components, collagen type IV (25), perlecan (26), and nidogen-2 (27), using a panel of wellcharacterized anti-mouse antibodies (28). Immunostaining of unstimulated whole cremasteric muscles with a rabbit anti-laminin α5 (LN-α5) chain polyclonal antibody (405), detecting the laminin 10 isoform, consistently indicated a dis continuous expression profi le of this matrix protein in venules but not arterioles (both at 20-40 μm in diameter).…”

“…The major architectural feature of this structure is two independent protein networks formed by laminins (laminin 8 [composed of α4β1γ1 chains] and laminin 10 [composed of α5β1γ1 chains]) (12,13) and collagen type IV that are largely interconnected with other components of the BM, such as nidogen-2 and perlecan (14). The perivascular BM envelops pericytes that are long cells, ‫-051ف‬ 200 μm in length and ‫52-01ف‬ μm wide, which form a discontinuous layer within the wall of all postcapillary venules (15).…”