heparitin-sulfate and Edema

Ischemia-reperfusion (I/R) injury is a major clinical problem linked to vascular surgery. Currently, no drugs to prevent or to treat I/R injury are approved for clinical use. C1 inhibitor (C1 INH) is known to reduce activation of the plasma cascade systems that are involved in the pathophysiologic process of I/R injury. The aim of this study was therefore to investigate the effect of C1 INH on complement deposition and endothelial cell activation in a rat model of hind limb I/R injury.. Male Wistar rats (wild type, bred at the central animal facility, University of Bern), weighing 250 to 320 g, were used. The rats underwent 2-hour ischemia and 24-hour reperfusion by unilateral clamping of the femoral artery and additional use of a tourniquet. Five groups were divided according to intravenous treatment 5 minutes before ischemia: 50 IU/kg C1 INH (n = 5); 100 IU/kg C1 INH (n = 7); vehicle control (n = 5); nontreated control (n = 7); and normal, healthy control without intervention (n = 4). At the end, muscle edema, tissue viability, and histologic features were assessed. Deposition of immunoglobulin M, C1r, C4d, and fibrin and expression of plasminogen activator inhibitor 1, heparan sulfate (HS), E-selectin, and vascular cell adhesion molecule 1 were evaluated by fluorescence staining. In addition, high-mobility group box 1 protein was measured in plasma.. Edema formation was reduced by C1 INH at two dosages, mirrored by improved histologic injury scores and preserved muscle viability. Deposition of immunoglobulin M, C4d, and fibrin was significantly decreased by 100 IU/kg C1 INH compared with nontreated controls. Pretreatment with 100 IU/kg C1 INH also significantly reduced HS shedding and expression of plasminogen activator inhibitor 1 as well as plasma levels of high-mobility group box 1 protein.. Pretreatment with both 50 and 100 IU/kg C1 INH attenuated reperfusion injury of rat hind limbs. Pretreatment with 100 IU/kg also preserved the endothelial HS layer as well as the natural, profibrinolytic phenotype of the endothelium. Prevention of endothelial cell activation by C1 INH may therefore be a promising strategy to prevent I/R injury in the clinical setting of peripheral vascular diseases and elective surgery on extremities.

Topics: Animals; Complement Activation; Complement C1 Inhibitor Protein; Complement C1r; Complement C4b; Complement Inactivating Agents; Disease Models, Animal; E-Selectin; Edema; Endothelial Cells; Fibrin; Heparitin Sulfate; Hindlimb; HMGB1 Protein; Immunoglobulin M; Male; Muscle, Skeletal; Peptide Fragments; Plasminogen Activator Inhibitor 1; Rats, Wistar; Reperfusion Injury; Tissue Survival; Vascular Cell Adhesion Molecule-1

Small molecule inhibitors of biologically important protein-glycosaminoglycan (GAG) interactions have yet to be identified.. Compound libraries were screened in an assay of L-selectin-IgG binding to heparin (a species of heparan sulfate [HS-GAG]). Hits were validated, IC-50s established and direct binding of hits to HS-GAGs was investigated by incubating compounds alone with heparin. Selectivity of inhibitors was assessed in 11 different protein-GAG binding assays. Anti-inflammatory activity of selected compounds was evaluated in animal models.. Screening identified a number of structurally-diverse planar aromatic cationic amines. Scaffolds similar to known GAG binders, chloroquine and tilorone, were also identified. Inhibitors displayed activity also against bovine kidney heparan sulfate. Direct binding of compounds to GAGs was verified by incubating compounds with heparin alone. Selectivity of inhibitors was demonstrated in a panel of 11 heparin binding proteins, including selectins, chemokines (IL-8, IP-10), Beta Amyloid and cytokines (VEGF, IL-6). A number of selected lead compounds showed dose-dependent efficacy in peritonitis, paw edema and delayed type hypersensitivity.. A new class of compounds, SMIGs, inhibits protein-GAG interaction by direct binding to GAGs. Although their IC-50s were in the low micro-molar range, SMIGs binding to HS-GAGs appeared to be stable in physiological conditions, indicating high avidity binding. SMIGs may interfere with major checkpoints for inflammatory and autoimmune events.. SMIGs are a class of structurally-diverse planar aromatic cationic amines that have an unusual mode of action - inhibiting protein-GAG interactions via direct and stable binding to GAGs. SMIGs may have therapeutic potential in inflammatory and autoimmune disorders.

Topics: Animals; Anti-Inflammatory Agents; Carrageenan; Cattle; Chemokines; Cytokines; Edema; Glycosaminoglycans; Heparin; Heparitin Sulfate; High-Throughput Screening Assays; Humans; Hypersensitivity, Delayed; Immunoglobulin G; Inflammation; L-Selectin; Leukocytes; Mice; Mice, Inbred BALB C; Mice, Inbred ICR; Molecular Structure; Neutrophils; Protein Binding; Protein Interaction Domains and Motifs; Small Molecule Libraries

Adhesion of polymorphonuclear neutrophils and platelets to the vessel wall contributes to generating ischemia-reperfusion injury. Endothelial adhesion molecules are harbored within the glycocalyx, which covers every healthy vascular endothelium but is deteriorated by ischemia-reperfusion. Pretreating the heart with volatile anesthetics reduces myocardial infarct size and protects against ischemia-reperfusion injury. The authors analyzed a possible protective effect of sevoflurane on the glycocalyx and implications for postischemic cell adhesion.. Isolated guinea pig hearts were perfused with crystalloid buffer and subjected to 20 min of global warm ischemia and 10 min of reperfusion. An intracoronary bolus of 3 x 10(6) polymorphonuclear neutrophilic leukocytes or 1 x 10(9) platelets of human origin was applied after reperfusion, either with or without pretreating with 0.5 or 1 minimal alveolar concentration sevoflurane. The number of sequestered cells was calculated from the difference between coronary input and output. Coronary effluent was collected throughout reperfusion to measure shedding of the glycocalyx.. Ischemia-reperfusion induced a significant increase in median (interquartile range) adhesion versus control nonischemic hearts of both leukocytes (38.9 (36.3-42.9) vs. 14.5 (13.1-16.0)%) and platelets (25.0 (22.5-27.1) vs. 9.4 (8.4-10.7)%). Shedding was evidenced by eightfold increases in washout of syndecan-1 and heparan sulfate versus basal. Sevoflurane reduced cell adhesion to near basal at 1 minimal alveolar concentration (leukocytes: 21.2% (19.2-23.9%), platelets: 11.5% (10.4-12.0%). Shedding measurements and electron microscopy demonstrated that sevoflurane-treated hearts retained much of their 200 nm-thick glycocalyx.. Sevoflurane reduces glycocalyx shedding in the postischemic coronary bed, maintaining the natural cover for endothelial adhesion molecules and, thus, reducing cell adhesion. This may explain beneficial outcomes linked to clinical use of volatile anesthetics after ischemia-reperfusion.

Topics: Anesthetics, Inhalation; Animals; Cell Adhesion; Coronary Circulation; Edema; Endothelium; Flow Cytometry; Glycocalyx; Guinea Pigs; Heparitin Sulfate; Humans; In Vitro Techniques; Methyl Ethers; Microscopy, Electron; Neutrophils; Platelet Adhesiveness; Reperfusion Injury; Sevoflurane; Syndecan-1

The overgrowth-affected gingiva of patients treated with cyclosporin A after kidney transplant was examined with ultrastructural and histochemical methods to evaluate the involvement of connective tissue. Gingival overgrowth has the same clinical signs as local edema. The ultrastructural study showed that the dimensional increase was largely due to increased production of amorphous ground substance by fibroblasts, possibly resulting from an increased release of histamine by mast cells. The histochemical data revealed that the affected tissues contained higher levels of glycosaminoglycans and that cyclosporin A induced comparably high levels of glycosaminoglycans in in vitro cultures of fibroblasts obtained from normal gingiva. The combination of ultrastructural and histochemical data, therefore, strongly suggests that the response of the connective tissue in gingival overgrowth cannot be ignored and may be the main cause of the observed pathological condition.

Topics: Alcian Blue; Capillaries; Chondroitin Sulfates; Coloring Agents; Connective Tissue; Cyclosporine; Dermatan Sulfate; Edema; Epithelium; Fibroblasts; Gingiva; Gingival Hyperplasia; Glycosaminoglycans; Heparin; Heparitin Sulfate; Histamine Release; Humans; Immunosuppressive Agents; Keratan Sulfate; Kidney Transplantation; Mast Cells; Tetrapyrroles