Regulation of arterial smooth muscle cell proliferation by cell-surface heparan sulfate proteoglycan

Tuesday II October 1994: Poster Abstracts Pathobiology

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Ile-Pro-Asp. Biglycans are shown to have the ability to bind and neutralize the biologic activity of growth factors such as transforming growth factor/l that are relevant to atherogenesis. 1 m,

Regulation of arterial smooth muscle cell proliferation by cell-surface heparan suRate proteoglycan Edwards IJ, Glory D, Bowman Gray Sch. of Med.,

Wake Forest Univ., Winston-Salem, NC 27157, USA

A key event in the progression of atherosclerosis and in restenosis following angioplasty is the proliferation of smooth muscle cells. Cell surface heparan sulfate proteoglycan (HSPG) is proposed to be a primary regulator of cell proliferation. To investigate genetic determinants of this regulation we have conducted studies with cultured arterial smooth muscle cells from atherosclerosissusceptible White Cameau (WC) and -resistant Show Racer (SR) pigeons. Arterial cells from WC pigeons have increased growth rates and reduced cell surface HSPG compared to cells from SR pigeons. Current studies were designed; (1) to determine if WC cells respond to growth inhibition by heparin thus demonstrating a pathway for an antiproliferative effect, (2) to determine if cell surface HSPG of WC is defective in antiproliferation potential, or (3) whether enhanced proliferation in WC is primarily due to low levels of HSPG. Purified cell-surface HSPG was used in a crossover experiment to test antiproliferative effect on WC and SR cells in the log phase of growth. WC HSPG had a lower potential to inhibit cell proliferation of both WC and SR cells. The concentration required to yield 50% growth inhibition was significantly greater than SR-derived HSPG, regardless of cell type (6.4,@ml for WC HSPG vs 0.8pg/ml SR HSPG on SR cells), (1 lO,@ml for WC HSPG vs 9.5 &ml SR HSPG on WC cells). In comparison to HSPG both cell types responded to heparin with growth inhibition. The results indicate that the major determining factor in growth regulation resides in the structural properties of the HSPG rather than the level on the cell surface, but WC cells in addition have a defective cellular response to cell surface HSPG. Lipoprotein lipase provides an internalization pathway for lipoproteins via heparan sulfate proteogly cans in human fibroblasts Femandez-Borja M, Bellido D, Vilella E, IQ&& Olivecrona G, Vilar6 S, Unit of Cell Biol., Dept. Biochem. and Physiol., Univ. of

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Barcelona, E-08071 Barcelona, Spain

Lipoprotein lipase (LPL) is the rate-limiting enzyme for hydrolysis of plasma lipoprotein triglycerides. Recent studies have shown that the lipase enhances the binding and uptake of all classes of lipoproteins by cultured cells in a non-enzymatic way. It is widely accepted that the LPL enhanced binding of lipoprotein to cell surfaces is mediated through its binding to heparan sulfate proteoglycans (HSPG). However, there is some controversy about the mechanism by which LPL increases the uptake of lipoproteins. Some authors have proposed that internalization occurs exclusively via receptor-mediated endocytosis by the LDL receptor or

the LRP, a mechanism where HSPG patticipates by presenting the bound LPL-lipoprotein complexes to these receptors. Other authors have asserted that LPL-mediated uptake occurs via HSPG endocytosis. In view of these conflicting data, we sought to perform morphological studies of the effects of LPL on lipoprotein binding and uptake by human skin tibroblasts. Using fluorescence and immunoelectron microscopy we showed that LPL mediates the binding of LDL, chylomicron and chylomicron remnants to the whole surface of the fibroblast cell. The ligands completely colocalize, which indicates that LPL acts as a bridge between the cell surface and the lipoproteins. The binding sites correspond to HSPG, as shown by heparin competition and heparinase treatment. Furthermore, LPL and lipoproteins are internalized together, at least in part, by a pathway clearly distinct from that mediated by specific receptors such as the LDL receptor or LRP. We conclude that, through its bridging between lipoproteins and HSPG. LPL orovides high-caoacitv binding sites for lioonroteins and .an additional in;ema&ibn pathiay mediated- by HSPG. This could be physiologically relevant since this pathway is not under regulation by intracellular sterol content. Displacement of tbe %bFGF from rat aortlc smooth muscle cells by a series of glycosaminoglycans. A structure-activity relatlouship study Giorgini L, Musant R, Naggi A, mG. Pharmacia Farmita-

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lia Cado Erba, Via Giovanni XXIII, Nerviano 20014, Italy The proliferation of smooth muscle cells (SMC) of the arterial wall is a key event in the development of atherosclerosis. The locally formed mitogen bFGF is likely to play an important role in initiating and in maintaining this proliferative status. A distinguishing characteristic of bFGF is its strong interaction with the sulfated glycosaminoglycans (GAGS) present in the extracellular matrix, which accounts for the low-affinity component of the ‘*%bFGF binding to the cells. In this study the ability of a series of chemically modified GAGS to displace l*?-bFGF from the extracellular matrix of rat aortic SMC has been investigated. Cells between the 8th and 18th passage were used when they had reached confluence. Low-aftinitv bindine of ‘*%bFGF to the cells were assessed as described by Moicatelli (J Cell Physiol 1987; 131: 123-130). When unmodified heparin (Hep), heparin sulfate (HS), chondroitin Csulfate (C4-S) and dermatan sulfate (DS) were tested, the order of potency was: Hep > HS > > C4-S, DS. Supersulfated Hep and HS had the same activity as the parent compounds. Splitting of the uranic acid ring of Hep and HS did not affect their potency. Within the modified Hep series, the order of potency was: Hep, O-sulfated > 6-O desulfated > 2’-0desulfated >> N-acetylated. N-acetylated HS had no activity. Activity of C4-S and DS could only be increased by supersulfation. No correlation could be found between the molecular weight of the GAGS investigated and their ability to displace the mitogen. Present results are consistent with the concept that spccific structural corn nents are involved in the interaction of RASMC GAGS with’?-bFGF.

PATHOBIOLOGY Presence of serum amyloid P component in the neerotic core of human atherosclerotic plaque: a novel mediator of atherosclerosis? &&a~&&*, Li XA*, Yutani C**, Masuda K**, Yamamoto A*,

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Depts. of *Etiology/Pathophysiology and **Pathology, National Cardiovascular Center, 5-7-I Fujishiro-dai, Suita City, Osaka 565, Japan

Serum amyloid P component (SAP) is found in all forms of amy-

loid deposits. SAP is present in normal human serum and is also a normal tissue matrix constituent associated with elastic fibers and the glomerular basement membrane. The physiological role of SAP is unclear, but the association of SAP with many ligands and its stable evolutionary conservation imply that it has important functions. We report here the presence of SAP in human atheromatous plaque. Serial sections of formalin-fixed, paraffin-embedded human atherosclerotic abdominal aorta derived from autopsy specimens

Atherosclerosis X, Montreal, October 1994