Contribution of mast cells and snake venom metalloproteinases to the hyperalgesia induced by Bothrops jararaca venom in rats.
ABSTRACT Bothrops jararaca venom (Bjv) is known to induce local inflammation and severe pain. Since, mast cells are able to secrete mediators involved in algesic processes, in this study we examined the putative role of these cells in the hyperalgesia triggered by Bjv in the rat paw. We noted that treatment with mast cell stabilizer sodium cromoglicate as well as with histamine and 5-hydroxytriptamine receptor antagonists meclizine and methysergide, respectively, inhibited the Bjv-induced hyperalgesia. In addition, we showed that stimulation of isolated rat peritoneal mast cells with Bjv in vitro resulted in the release of stored and neo-generated inflammatory mediators such as histamine and leukotriene C(4), respectively. Bjv-induced histamine secretion was clearly sensitive to treatment with sodium cromoglicate and sodium nedocromil. We further observed that metalloproteinase inhibitors 1,10-phenantroline and DM43 inhibited mast cell degranulation in vitro, under conditions where inhibitors of phospholipase A(2) as well as of serine- and cysteine-proteinases were inactive. Altogether, our findings indicate that mast cells seem to contribute to the hyperalgesia caused by Bjv in the rat paw, and also provide evidence that this response might be dependent on the ability of the Bjv to activate directly mast cells.
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ABSTRACT: In Brazil, the genus Crotalus is responsible for approximately 1500 cases of snakebite annually. The most common complication in the lethal cases is acute renal failure, although the mechanisms of the damaging effects are not totally understood. In this work, we have examined the renal effects caused by a supernatant of macrophages stimulated by Crotalus durissus cascavella venom as well the potential role of phospholipase A2 and cyclo-oxygenase. Rat peritoneal macrophages were collected and placed in a RPMI medium and stimulated by crude Crotalus durissus cascavella venom (1, 3 or 10 microg/ml) for 1 hr. They were then washed and kept in a culture for 2 hr. The supernatant (1 ml) was tested in an isolated perfused rat kidney. The first 30 min. of each experiment were used as an internal control, and the supernatant was added to the system after this period. All experiments lasted 120 min. A study of toxic effect on perfusion pressure, glomerular filtration rate, urinary flow percent of sodium tubular transport and percent of proximal tubular sodium transport was made. The lowest concentration of venom (1 microg/ml) was not statistically different from the control values. The most intense effects were seen at 10 microg/ml for all renal parameters. The infusion of the supernatant of macrophages stimulated with crude venom (3 or 10 microg/ml) increased the perfusion pressure, glomerular filtration rate and urinary flow, decreased the percent of sodium tubular transport and percent of proximal tubular sodium transport. Dexamethasone (10 microM) and quinacrine (10 microM) provided protection against the effect of the venom on glomerular filtration rate, urinary flow, percent of sodium tubular transport, percent of proximal tubular sodium transport and perfusion pressure. Indomethacin (10 microM) and nordiidroguaretic acid (1 microM) reversed almost all functional changes, except those of the perfusion pressure. These results suggest that macrophages stimulated with Crotalus durissus cascavella venom release mediators capable of promoting nephrotoxicity in vitro. Moreover, phospholipase A2 and cyclooxygenase products are involved in these biologic effects.Pharmacology & Toxicology 02/2003; 92(1):14-20.
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ABSTRACT: The reprolysin subfamily of metalloproteinases includes snake venom metalloproteinases (SVMP) and mammalian disintegrin/metalloproteinase. These proteins are synthesized as zymogens and undergo proteolytic processing resulting in a variety of multifunctional proteins. Jararhagin is a P-III SVMP isolated from the venom of Bothrops jararaca. In crude venom, two forms of jararhagin are typically found, full-length jararhagin and jararhagin-C, a proteolytically processed form of jararhagin that is composed of the disintegrin-like and cysteine-rich domains of jararhagin. To better understand the structural and mechanistic bases for these forms of jararhagin in the venom of B. jararaca and the source of venom complexity in general, we have examined the jararhagin forms isolated from venom and the autolysis of isolated jararhagin under the conditions of varying pH, calcium ion concentration, and reducing agents. From our results, jararhagin isolated from venom appears as two forms: a predominant form that is stable to in vitro autolysis and a minor form that is susceptible to autolysis under a variety of conditions including alkaline pH, low calcium ion concentrations, or reducing agent. The autolysis site for production of jararhagin-C from isolated jararhagin was different from that observed for jararhagin-C as isolated from crude venom. Taken together, these data lead us to the conclusion that during the biosynthesis of jararhagin in the venom gland at least three forms are present: one form which is rapidly processed to give rise to jararhagin-C, one form which is resistant to processing in the venom and autolysis in vitro, and one minor form which is susceptible to autolysis under conditions that promote destabilization of its structure. The presence of these different forms of jararhagin contributes to greater structural and functional complexity of the venom and may be a common feature among all snake venoms. The biological and biochemical features in the venom gland responsible for these jararhagin isoforms are currently under investigation.Archives of Biochemistry and Biophysics 02/2003; 409(2):395-401. · 3.37 Impact Factor
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ABSTRACT: Neutralization of hyperalgesia induced by Bothrops jararaca and B. asper venoms was studied in rats using bothropic antivenom produced at Instituto Butantan (AVIB, 1 ml neutralizes 5 mg B. jararaca venom) and polyvalent antivenom produced at Instituto Clodomiro Picado (AVCP, 1 ml neutralizes 2.5 mg B. aspar venom). The intraplantar injection of B. jararaca and B. asper venoms caused hyperalgesia, which peaked 1 and 2 h after injection, respectively. Both venoms also induced edema with a similar time course. When neutralization assays involving the independent injection of venom and antivenom were performed, the hyperalgesia induced by B. jararaca venom was neutralized only when bothropic antivenom was administered iv 15 min before venom injection, whereas edema was neutralized when antivenom was injected 15 min or immediately before venom injection. On the other hand, polyvalent antivenom did not interfere with hyperalgesia or edema induced by B. asper venom, even when administered prior to envenomation. The lack of neutralization of hyperalgesia and edema induced by B. asper venom is not attributable to the absence of neutralizing antibodies in the antivenom, since neutralization was achieved in assays involving preincubation of venom and antivenom. Cross-neutralization of AVCP or AVIB against B. jararaca and B. asper venoms, respectively, was also evaluated. Only bothropic antivenom partially neutralized hyperalgesia induced by B. asper venom in preincubation experiments. The present data suggest that hyperalgesia and edema induced by Bothrops venoms are poorly neutralized by commercial antivenoms even when antibodies are administered immediately after envenomation.Brazilian Journal of Medical and Biological Research 11/2002; 35(10):1221-8. · 1.14 Impact Factor