Design of peptides with high affinities for heparin and endothelial cell proteoglycans.

Proteoglycan-binding peptides were designed based on consensus sequences in heparin-binding proteins: XBBXBX and XBBBXXBX, where X and B are hydropathic and basic residues, respectively. Initial peptide constructs included (AKKARA)(n) and (ARKKAAKA)(n) (n = 1-6). Affinity coelectrophoresis revealed that low M(r) peptides (600-1,300) had no affinities for low M(r) heparin, but higher M(r) peptides (2,000-3,500) exhibited significant affinities (K(d) congruent with 50-150 nM), which increased with peptide M(r). Affinity was strongest when sequence arrays were contiguous and alanines and arginines occupied hydropathic and basic positions, but inclusion of prolines was disruptive. A peptide including a single consensus sequence of the serglycin proteoglycan core protein bound heparin strongly (K(d) congruent with 200 nM), likely owing to dimerization through cysteine-cysteine linkages. Circular dichroism showed that high affinity heparin-binding peptides converted from a charged coil to an alpha-helix upon heparin addition, whereas weak heparin-binding peptides did not. Higher M(r) peptides exhibited high affinities for total endothelial cell proteoglycans (K(d) congruent with 300 nM), and approximately 4-fold weaker affinities for their free glycosaminoglycan chains. Thus, peptides including concatamers of heparin-binding consensus sequences may exhibit strong affinities for heparin and proteoglycans. Such peptides may be applicable in promoting cell-substratum adhesion or in the design of drugs targeted to proteoglycan-containing cell surfaces and extracellular matrices.

Heparin sensitive and resistant vascular smooth muscle cells: biology and role in restenosis.

Vascular smooth muscle cells (VSMC)s are characterized by their acute growth inhibition by heparin and heparan sulfates; however, recently the isolation of VSMCs which display greatly diminished sensitivity to the antiproliferative action of heparin have been reported. These heparin resistant (HR) VSMCs have been derived through multiple passage of normal rat VSMCs in culture media containing high heparin doses, by transformation of VSMCs with oncogene-containing vectors, or have been isolated from vascular tissues of spontaneously hypertensive rats, healthy humans, or humans with restenosis where their presence is not limited to sites of injury. Initial characterizations of HR VSMCs are reviewed, and here we propose a definition of HR VSMCs. To date the mechanisms underlying heparin insensitivity remain elusive. Further study of HR VSMCs may expand our understanding of cell growth regulation by heparin, establish whether HR VSMCs contribute to the reported failure of heparin to combat restenosis in humans, and identify cellular mechanisms driving certain vascular proliferative diseases.