Perlecan Mediates the Antiproliferative Effect of Apolipoprotein E on Smooth Muscle Cells

Abstract

Apolipoprotein E (apoE) is known to inhibit cell proliferation; however, the mechanism of this inhibition is not clear. We recently showed that apoE stimulates endothelial production of heparan sulfate (HS) enriched in heparin-like sequences. Because heparin and HS are potent inhibitors of smooth muscle cell (SMC) proliferation, in this study we determined apoE effects on SMC HS production and cell growth. In confluent SMCs, apoE (10 g/ml) increased 35 SO 4 incorporation into PG in media by 25-30%. The increase in the medium was exclusively due to an increase in HSPGs (2.2-fold), and apoE did not alter chondroitin and dermatan sulfate proteoglycans. In proliferating SMCs, apoE inhibited [ 3 H]thymidine incorporation into DNA by 50%; however, despite decreasing cell number, apoE increased the ratio of 35 SO 4 to [ 3 H]thymidine from 2 to 3.6, suggesting increased HS per cell. Purified HSPGs from apoE-stimulated cells inhibited cell proliferation in the absence of apoE. ApoE did not inhibit proliferation of endothelial cells, which are resistant to heparin inhibition. Analysis of the conditioned medium from apoE-stimulated cells revealed that the HSPG increase was in perlecan and that apoE also stimulated perlecan mRNA expression by >2-fold. The ability of apoE isoforms to inhibit cell proliferation correlated with their ability to stimulate perlecan expression. An anti-perlecan antibody completely abrogated the antiproliferative effect of apoE. Thus, these data show that perlecan is a potent inhibitor of SMC proliferation and is required to mediate the antiproliferative effect of apoE. Because other growth modulators also regulate perlecan expression, this may be a key pathway in the regulation of SMC growth.

Negative and independent influence of apolipoprotein E on C-reactive protein (CRP) concentration in obese adults. Potential anti-inflammatory role of apoE in vivo

“…It is not possible to conclude if it is a reduced apoE level in obese adults which leads in part to low grade inflammation, expressed by increased CRP level or, inversely, if it is low grade systemic inflammation which leads to reduced apoE level. However, in vitro cellular studies have suggested an anti-proliferative role of apoE on T lymphocytes, 45,55 smooth muscle cells 44,46 and microglia. 47 Indirect in vivo anti-inflammatory role of apoE has also been suggested in apoE knock-out mice model.…”

A Protective Role for Kidney Apolipoprotein E

“…ApoE effects on proteoglycans in different pools, pericellular (trypsinreleasable representing cell surface and extracellular proteoglycans), cellular (Triton X-100/NH 4 releasable), and matrix (guanidine hydrochloride extractable following Triton X-100 treatment), were determined as described previously (18). Pericellular proteoglycans minus matrix proteoglycans was represented as cell surface proteoglycans.…”

“…Cell Proliferation-Cell proliferation was assessed by [ 3 H]thymidine incorporation as described previously (18). Mesangial cells were seeded at a density of 2 ϫ 10 4 /well in a 48-well plate, and experiments were performed the following day (40 -50% confluency).…”

“…Metabolic Labeling and Determination of HSPG-Confluent mesangial cells were incubated in culture medium containing either [ 3 H]glucosamine or 35 SO 4 with or without 10 g/ml apoE for 24 h. Heparan sulfate proteoglycans associated with cells and those secreted into medium were determined as described previously following purification by DEAE-cellulose chromatography and digestion with heparinase (18). ApoE effects on proteoglycans in different pools, pericellular (trypsinreleasable representing cell surface and extracellular proteoglycans), cellular (Triton X-100/NH 4 releasable), and matrix (guanidine hydrochloride extractable following Triton X-100 treatment), were determined as described previously (18).…”

The cell cycle: A critical therapeutic target to prevent vascular proliferative disease

“…One potential growth suppressor is perlecan, a large heparan sulfate proteoglycan that is expressed in most extracellular matrixes and basement membranes (148). Perlecan has been shown to inhibit in vitro SMC proliferation (149,150), adhesion (151) and migration (152). In vivo, perlecan has been shown to inhibit thrombosis and intimal hyperplasia after arterial injury (153).…”