Frances Kern

Density- and proliferation status-dependent expression of T-cadherin, a novel lipoprotein-binding glycoprotein: a function in negative regulation of smooth muscle cell growth?

The atypical low density lipoprotein (LDL) binding proteins (M r 105 and 130 kDa; p105 and p130) in human aortic medial membranes and cultured human and rat aortic smooth muscle cells (SMC) have recently been identified as the cell adhesion glycoprotein T‐cadherin. Although cadherins are generally recognized to be important regulators of morphogenesis, the function of T‐cadherin in the vasculature is poorly understood. This study has examined the relationship between expression of T‐cadherin and the density and proliferation status of SMC. T‐cadherin (p105 and p130) levels in SMC lysates were measured on Western blots using ligand‐binding techniques. T‐cadherin expression was dependent upon cell density, and maximal levels were achieved at confluency. T‐cadherin levels were reversibly modulated by switching cultures between serum‐free (upmodulation) and serum‐containing (downmodulation) conditions. Platelet‐derived growth factor (PDGF)‐BB, epidermal growth factor (EGF) or insulin‐like growth factor (IGF) elicited a dose‐ and time‐dependent downmodulation that was reversible after transfer of SMC to growth factor‐free medium. Our results support the hypothesis that T‐cadherin may function as a negative determinant of cell growth.

Functional Aspects of Vascular Tenascin-C Expression

The arterial tenascin C expression in vivo and in vitro has been studied using immunohistochemistry. The functional relevance of localized tenascin C expression was assessed in vitro using various human cell types involved in the progression of vascular disease. Normotensive and hypertensive rats exhibited age-dependent patterns of vascular (aorta) tenascin expression, but the lumen-to-media-directed progression of tenascin induction was accelerated in hypertensive rats. Tenascin-rich neointimal lesions (spontaneous) were observed at branching sites of aorta from aged (80 weeks) hypertensive rats. Subendothelial tenascin foci contained lipid-laden smooth muscle cells and monocytes/macrophages. Medial tenascin foci encaged smooth muscle cells which synthesized DNA. Tenascin was expressed both in vivo and in vitro by endothelial and smooth muscle cells but not by monocytes/macrophages; angiotensin II, oxidized-low density lipoprotein and transforming growth factor beta 1 induced expression of tenascin transcripts and glycoprotein in vitro. Endothelial and smooth muscle cells, but not monocytes, adhered to tenascin substrata. Tenascin reduced focal adhesion integrity in confluent endothelial and smooth muscle cell cultures. Angiotensin II-induced migration of endothelial and smooth muscle cells was accompanied by tenascin deposition within extracellular matrix migration trails. Tenascin may function both as a defense against monocyte invasion and medial smooth muscle replication, as well as a substratum for directed endothelial and smooth muscle cell migration.

Peptide vasoconstrictors, vessel structure, and vascular smooth-muscle proliferation

The peptide vasoconstrictors angiotensin II (Ang II) and endothelin-1 (ET-1), originally thought to derive exclusively from the plasma renin-angiotensin system and vascular endothelium, respectively, have been demonstrated to be produced independently of such sources. Local tissue angiotensin-generating systems are well documented, and endothelin production has been demonstrated for a variety of nonendothelial cells, including vascular smooth-muscle cells (VSMC). There is increasing evidence from in vitro studies that local production of these vasoconstrictor peptides may contribute to blood vessel homeostasis and the development of vascular pathologies. Results obtained from pharmaceutical intervention in humans and animals of these systems strongly support this hypothesis. In addition to their vasoconstrictor properties, Ang II and ET-1 act as potent biological effectors. In vitro, both vasoconstrictor peptides appear to modulate the activity of autocrine feedback loops in VSMC. The activity of these feedback loops in vivo may represent a central mechanism for regulation and phenotypic differentiation of this cell type. The best-recognized autocrine feedback loops of VSMC are constituted by platelet-derived growth factor and transforming growth factor-beta, both of which are influenced by the action of Ang II and ET-1. Because both vasoconstrictors (via their induction of autocrine growth modulators) may influence the composition of the extracellular matrix of VSMC, the effects of the peptide vasoconstrictors on the (auto-) regulated feedback loops are of long-term structural importance. Ang II and ET-1 promote the synthesis and secretion of the glycoproteins thrombospondin, fibronectin, and tenascin.(ABSTRACT TRUNCATED AT 250 WORDS)

LDL binds to surface-expressed human T-cadherin in transfected HEK293 cells and influences homophilic adhesive interactions.

T‐cadherin (T‐cad) is an unusual glycosylphosphatidylinositol‐anchored member of the cadherin family of cell adhesion molecules. Binding of low density lipoproteins (LDLs) to T‐cad can be demonstrated on Western blots of smooth muscle cell lysates, membranes and purified proteins. Using HEK293 cells transfected with human T‐cad cDNA (T‐cad+), we have investigated the adhesion properties of expressed mature and precursor proteins and examined the postulate that LDL represents a physiologically relevant ligand for T‐cad. T‐cad+ exhibits an increased Ca2+‐dependent aggregation (vs. control) that was reduced by selective proteolytic cleavage of precursor T‐cad and abolished after either proteolytic or phosphatidylinositol‐specific phospholipase C (PI‐PLC) cleavage of both mature and precursor proteins, indicating that both proteins function in intercellular adhesion. T‐cad+ exhibited a significantly increased specific cell surface‐binding of [125I]‐LDL that was sensitive to PI‐PLC pre‐treatment of cells. Ca2+‐dependent intercellular adhesion of T‐cad+ was significantly inhibited by LDL. Our results support the suggestion that LDL is a physiologically relevant ligand for T‐cad.

Effects of endothelin-1 on vascular smooth muscle cell phenotypic differentiation

Summary: Endothelin‐1 (ET‐1) produced by vascular endothelial cells has been proposed to act in a paracrine manner on adjacent smooth muscle cells (SMCs) in vivo, exerting a variety of short‐ and long‐term effects. Although some of the in vitro ET‐1‐mediated effects are related to growth‐promoting events, the physiological significance of these observations remains to be clarified. Reported discrepancies of the mitogenic potential of ET‐1 may relate to differences in culturing conditions (submitogenic levels of serum in combination with ET‐1). Because ET‐1 has been implicated in proliferation of vascular SMCs (VSMCs) at sites of vascular injury, as well as pathological events during atherogenesis, a clarification of the mitogenic effects of ET‐1 is important. This study demonstrates the possible autocrine role for ET‐1 in the regulation of the vasculature, its influence on VSMC cell cycle, and autocrine and phenotypic regulation of VSMCs. Stimulation of quiescent VSMCs with a variety of peptides resulted in the secretion of biologically active ET‐1 by VSMCs. In contrast to previous reports, long‐term exposure (12‐15 days) of VSMCs to ET‐1 in nonmitogenic medium did not promote cycling of cells. On the contrary, ET‐1 attenuated the cycling of VSMCs in the S and G2/M phases and interrupted progression through the cell cycle at late G1/early S phase. Subsequent to ET‐1 exposure, VSMCs expressed increased levels of smooth muscle‐specific &agr;‐actin. Therefore, autocrineproduced ET‐1 may contribute to phenotypic differentiation of VSMCs.