Assessment of Zymosan-Induced Leukocyte Influx in a Rat Model using Sulfated Polysaccharides

Abstract

Fucoidan, a sulfated polysaccharide from the brown algae Fucus vesiculosus, has diverse biological properties, including anti-inflammatory, anticoagulant and antithrombotic activity. This study analyzed the therapeutic activity of total fucoidan (TF) from F. vesiculosus and that of purified fractions (F1 and F2) on zymosan-induced arthritis. Arthritis was induced by injecting zymosan into the knee joint. Thus, three fucoidan fractions were obtained by acetone fractionation. Due to the yield obtained from F3, we used only fucoidans F1 and F2 in the induced inflammation tests. Chemical analyses and electrophoretic characterization of these fractions demonstrated that they contain polysaccharides, sulfate ester and very low protein levels. The fucoidans obtained from TF showed only an electrophoretic band in agarose gel with much lower polydispersion. The F2 fraction showed a migration between fucoidans F1 and F3. We administered TF (15, 30, 50 mg/kg i. p.), F1 or F2 (10, 25 and 50 mg/kg i. p.), diclofenac sodium (10 mg/kg i. p.), lumiracoxib (5 mg/kg o. a.) or L-NAME (30 mg/kg i. p.), 1 hour after induction of articular inflammation. We analyzed cell influx and nitrite levels in addition to performing histopathological analysis. TF (total fucoidan) at 15, 30, 50 mg/kg i. p. and its fractions (F1 and F2 at concentrations of 25 and 50 mg/kg i. p.) significantly reduced cellular influx and nitric oxide concentration. Moreover, the articular inflammation in zymosan-induced arthritis caused a progressive loss in glycosaminoglycan content. This loss decreased when TF (30 mg/kg) was administered. These data suggest that fucoidan exerts anti-inflammatory action in a zymosan-induced model of acute inflammation in rats. Taken together with the fact that these natural compounds have minimal toxicity, this may have important therapeutic implications.

Fucus vesiculosus - Fucaceae - fucoidan - sulfated polysaccharides - brown algae - arthritis - zymosan

References

• 1 Leite E L, Medeiros M G L, Rocha H A O, Farias G G M, da Silva L F, Chavante S F, de Abreu L D, Dietrich C P, Nader H B. Structure and pharmacological activities of a sulfated xylofucanglucuronan from the algae Spatoglossum scroederi.  Plant Sci. 1998;  14 215-228
• 2 Patankar M S, Oehninger S, Barnett T, Williams R L, Clark G F. A revised structure for fucoidan may explain some of its biological activities.  J Biol Chem. 1993;  268 21770-21776
• 3 Derbocio A M, Kelmer-Bracht A M, Bracht L, Bracht A, Ishii-Iwamoto E L. The hemodynamic effects of zymosan in the perfused rat liver.  Vasc Pharmacol. 2005;  43 75-85
• 4 Bezerra M M, Brain S D, Greenacre S, Jerônimo S M B, Melo L B, Keeble J, Rocha F A C. Reactive nitrogen species scavenging, rather than nitric oxide inhibition, protects from articular cartilage damage in rat zymosan-induced arthritis.  Br J Pharmacol. 2004;  141 172-182
• 5 Kannan K, Ortmann R A, Kimpel D. Animal models of rheumatoid arthritis and their relevance to human disease.  Pathophysiology. 2005;  12 167-181
• 6 Hardy M M, Seibert K, Manning P T, Currie M G, Woerner B M, Edwards D, Koki A, Tripp C S. Cyclooxygenase 2-dependent prostaglandin E2 modulates cartilage proteoglycan degradation in human osteoarthritis explants.  Arthritis Rheum. 2002;  46 789-803
• 7 Dietrich C P, Dietrich S M C. Electrophoretic behavior of acidic mucopolysaccharides by agarose gel electrophoresis.  J Chromatogr. 1977;  130 299-304
• 8 Dubois M, Gilles K A, Hamilton J K, Rebers P A, Smith F. Colorimetric method for determination of sugars, and related substances.  Anal Chem. 1956;  28 350-356
• 9 Spector J. Refinement of the coomassie blue method of protein quantification. A simple and linear spectrophotometric assay of 0.5 to 50 µg of protein.  Anal Biochem. 1978;  86 142-143
• 10 Dodgson K S, Price R G. A note on the determination of the ester sulphate content of sulphated polysaccharides.  Biochem J. 1962;  84 106-110
• 11 Granger D L, Tainitor R R, Boockvar K S, Hibbs J B. Measurement of nitrate and nitrite in biological samples using nitrate reductase and Griess reaction.  Methods Enzymol. 1996;  268 142-145
• 12 Dietrich C P, Farias G G M, Abreu L R D, Leite E L, Silva L F, Nader H B. A new approach for the characterization of polysaccharides from algae: presence of four main acidic polysaccharides in three species of the class Phaeophyceae.  Plant Sci. 1995;  108 143-153
• 13 Silva T M A, Chavante S F, Queiroz K C S, Marques C T, Rocha H A O, Leite E L. Characterization partial and anticoagulant activity of a heterofucan from the brown seaweed Padina gymnospora.  Braz J Med Biol Res. 2005;  38 523-533
• 14 Gadó K, Gliger G. Zymosan inflammation: a new method suitable for evaluating new anti-inflammatory drugs.  Agents Actions. 1991;  32 119-121
• 15 Capone M L, Tacconelli S, Sciulli M G, Patrignani P. Clinical pharmacology of selective COX-2 inhibitors.  Int J Immunopathol Pharmacol. 2003;  16 49-58
• 16 Salvemini D, Misko T P, Masferrer J, Seibert K, Currie M G. Nitric oxide activates cyclooxygenase enzymes.  Proc Natl Acad Sci USA. 1993;  90 7240-7244
• 17 Bachelet L, Bertholon I, Lavigne D, Vassy R, Jandrot-Perrus M, Chaubet F, Letourneur D. Affinity of low molecular weight fucoidan for P-selectin triggers its binding to activated human platelets.  Biochim Biophys Acta. 2009;  1790 141-146
• 18 Angstwurm K, Weber J R, Segert A, Burger W, Weih M, Freyer D, Einhaupl K M, Dirnagl U. Fucoidan, a polysaccharide inhibiting leukocyte rolling, attenuates inflammatory responses in experimental pneumococcal meningitis in rats.  Neurosci Lett. 1995;  191 1-4
• 19 Woodman R C, Teoh D, Payne D, Kubes P. Thrombin and leukocyte recruitment in endotoxemia.  Am J Physiol Heart Circ Physiol. 2000;  279 338-345
• 20 McCartney-Francis N L, Song X-Y, Mizel D E, Wahl S M. Selective inhibition of inducible nitric oxide synthase exacerbates erosive joint disease.  J Immunol. 2001;  166 2734-2740