glycyl-arginyl-glycyl-glutamyl-seryl-proline and arginyl-glycyl-aspartyl-serine

Fibronectin (Fn) is a multifunctional adhesive protein found on cell surfaces as well as in plasma. It is also believed to play an important role in bacterial adherence to host tissues. Molecular analyses of Fn have shown that the amino acid triplet arginine-glycine-aspartic acid (RGD) sequence functions as a binding site. We examined the role of the RGD sequence on bacterial adherence to Fn. The pretreatment of Streptococcus mitis with synthetic RGD-containing peptide reduced the number of bound bacteria to the Fn coated plates by 76%. In contrast, a control peptide containing the RGE sequence showed no inhibition. These data indicate that synthetic RGD peptides may be useful for the inhibition of bacterial adherence to Fn on host cell surfaces.

Topics: Amino Acid Sequence; Bacterial Adhesion; Binding Sites; Fibrinogen; Fibronectins; Oligopeptides; Platelet Aggregation Inhibitors; Receptors, Immunologic; Streptococcus

The vasoocclusive process in patients with sickle cell disease (SCD) is complex and involves interactions among sickle erythrocytes (SS-RBC), vascular endothelium, and plasma and cellular components. The role of neutrophils (PMN) in vasoocclusion has not been examined. Patients with SCD appear to have chronically activated PMN. Because the first step in PMN activation is particle recognition, we explored whether normal PMN recognize SS-RBC and whether this recognition results in PMN monolayers, significantly more SS-RBC adhered to the PMN than did normal erythrocytes (AA-RBC; P < .001). Preincubation of erythrocytes with autologous plasma significantly increased the adherence of SS-RBC to PMN but had no effect on AA-RBC (P < .001). When adhesion of density fractionated SS-RBC was performed, dense SS-RBC showed greater adherence to the PMN monolayers than did light SS-RBC (P < .001). To determine mechanisms of this adhesion, IgG and Arg-Gly-Asp-Ser (RGDS) receptor sites on PMN were saturated. IgG inhibited adherence of dense SS-RBC, whereas RGDS inhibited adherence in both fractions, although to a greater extent in the light fraction. We measured SS-RBC activation of PMN by incubating SS-RBC with 2', 7'-Dichloro-fluroescin Diacetate (DCF)-labeled PMN. Incubation of PMN with SS-RBC resulted in a significant increase in fluorescence compared to AA-RBC. We show here that PMN recognize SS-RBC through multiple mechanisms and that this recognition results in activation of PMN. These findings contribute to the understanding of vasoocclusive crisis in patients with SCD and may have therapeutic implications.

Topics: Amino Acid Sequence; Anemia, Sickle Cell; Blood Flow Velocity; Cell Adhesion; Erythrocytes, Abnormal; Hemoglobin, Sickle; Humans; Immunoglobulin Fc Fragments; Immunoglobulin G; Inflammation; Molecular Sequence Data; Neutrophils; Oligopeptides; Polymers; Receptors, IgG; Respiratory Burst

3H-Labelled kappa-elastin peptides (kE:75 kDa molecular weight) were shown to bind to confluent human skin fibroblast (HSF) cultures in a time-dependent and saturable manner. Scatchard analysis indicated the presence of high affinity binding sites with kD = 2.7 x 10(-10) M and 19,000 sites per cell. Binding of kE to its receptor on HSF accelerates and intensifies the adhesion of insoluble elastin fibres (iE) to confluent HSF. Optimal effect was attained for a kE concentration of 0.3 x 10(-9) M close to kD. This stimulatory effect of kE on the binding of iE to HSF could be inhibited by neomycin, retinal and pertussis toxin, substances which act at different levels of the transduction mechanism following the activation of the receptor and the subsequent triggering of cell biological events (chemotaxis, modification of calcium fluxes). The stimulation of iE adhesion to HSF induced by kE as well as kE binding to the cells could be inhibited by lactose and laminin but not by Arg-Gly-Asp-Ser(RGDS) peptides. This indicates that the elastin peptide receptor on HSF possesses lectin-like properties and shares homology with the laminin receptor as also shown for other cell types. None of the substances tested, that is inhibitors of the transduction mechanism, lactose, laminin and Arg-Gly-Asp-Ser(RGDS) peptides were shown to interfere significantly with the binding of iE (in the absence of added kE) to confluent HSF. The proteins adhering strongly to elastin fibres were isolated by a sequential extraction procedure and the final hydrochloride guanidinium-DTT extract was analysed by SDS-PAGE under reducing conditions, Western blots using specific antibodies against several connective tissue proteins and affinity for [3H]-kE following nitrocellulose electro-transfer of proteins. Fibronectin, vitronectin, tropoelastin(s), and a 120 kDa cysteine rich glycoprotein previously designated as elastonectin were identified. Among these proteins, [3H]-kE was found to bind exclusively to a 65 kDa protein that could be eluted selectively from elastin fibres with a neutral buffer containing 100 mM lactose. Therefore the elastin peptide receptor on human skin fibroblasts shares properties with the elastin receptor characterized from other cell types. Conformational differences between elastin peptides and elastin fibres could explain the differences in the mechanisms of interactions between elastin fibres and elastin peptides with HSF in culture. The stimulatory effect of elastin-de

Topics: Adult; Amino Acid Sequence; Binding Sites; Elastin; Extracellular Matrix; Female; Fibroblasts; Fibronectins; Glycoproteins; Humans; Molecular Sequence Data; Oligopeptides; Platelet Aggregation Inhibitors; Receptors, Cell Surface; Skin; Tropoelastin; Vitronectin