batimastat and Hemorrhage

The ability of two peptidomimetic hydroxamate metalloproteinase inhibitors, Batimastat and Marimastat, to abrogate toxic and proteinase activities of the venom of Echis ocellatus from Cameroon and Ghana was assessed. Since this venom largely relies for its toxicity on the action of zinc-dependent metalloproteinases (SVMPs), the hypothesis was raised that toxicity could be largely eliminated by using SVMP inhibitors. Both hydroxamate molecules inhibited local and pulmonary hemorrhagic, in vitro coagulant, defibrinogenating, and proteinase activities of the venoms in conditions in which venom and inhibitors were incubated prior to the test. In addition, the inhibitors prolonged the time of death of mice receiving 4 LD

Topics: Animals; Cameroon; Dose-Response Relationship, Drug; Ghana; Hemorrhage; Hydroxamic Acids; Lung; Metalloproteases; Mice; Peptidomimetics; Phenylalanine; Snake Bites; Thiophenes; Viper Venoms; Viperidae

Previously, we established delayed jellyfish envenomation syndrome (DJES) models and proposed that the hemorrhagic toxins in jellyfish tentacle extracts (TE) play a significant role in the liver and kidney injuries of the experimental model. Further, we also demonstrated that metalloproteinases are the central toxic components of the jellyfish Cyanea capillata (C. capillata), which may be responsible for the hemorrhagic effects. Thus, metalloproteinase inhibitors appear to be a promising therapeutic alternative for the treatment of hemorrhagic injuries in DJES. In this study, we examined the metalloproteinase activity of TE from the jellyfish C. capillata using zymography analyses. Our results confirmed that TE possessed a metalloproteinase activity, which was also sensitive to heat. Then, we tested the effect of metalloproteinase inhibitor batimastat (BB-94) on TE-induced hemorrhagic injuries in DJES models. Firstly, using SR-based X-ray microangiography, we found that BB-94 significantly improved TE-induced hepatic and renal microvasculature alterations in DJES mouse model. Secondly, under synchrotron radiation micro-computed tomography (SR-μCT), we also confirmed that BB-94 reduced TE-induced hepatic and renal microvasculature changes in DJES rat model. In addition, being consistent with the imaging results, histopathological and terminal deoxynucleotidyl transferase-mediated UTP end labeling (TUNEL)-like staining observations also clearly corroborated this hypothesis, as BB-94 was highly effective in neutralizing TE-induced extensive hemorrhage and necrosis in DJES rat model. Although it may require further clinical studies in the near future, the current study opens up the possibilities for the use of the metalloproteinase inhibitor, BB-94, in the treatment of multiple organ hemorrhagic injuries in DJES.

Topics: Angiography; Animals; Bites and Stings; Cnidarian Venoms; Hemorrhage; In Situ Nick-End Labeling; Kidney; Liver; Male; Metalloproteases; Mice; Necrosis; Phenylalanine; Protective Agents; Rats; Rats, Sprague-Dawley; Scyphozoa; Syndrome; Thiophenes; X-Ray Microtomography

The peptidomimetic hydroxamate metalloproteinase inhibitor batimastat (BB-94) was assessed for its ability to neutralize the systemic effects (lethality, hemorrhage and coagulopathy) induced by the venom of Bothrops asper, the most important snake from a medical standpoint in Central America. Batimastat inhibited lethality when a venom challenge dose of two LD(50)s was used by intraperitoneal and intravenous routes, with ED(50)s of 250 and 22 microM, respectively. With a challenge dose of three LD(50)s, lethality was not abrogated, but a conspicuous and dose-dependent delay in the time of death was observed in mice injected with mixtures of venom plus batimastat. Upon incubation with 500 microM batimastat, venom LD(50) increased 2.86-fold (intraperitoneal route) and 2.37-fold (intravenous route), when compared with LD(50) of venom alone. Batimastat also inhibited the hemorrhagic effect induced by venom in the lungs after intravenous injection. Moreover, batimastat exerted a significant inhibition of in vitro coagulant and in vivo defibrinogenating effects of venom, evidencing that metalloproteinases play a key role in the coagulopathy characteristic of B. asper envenomation. The remaining uninhibited coagulant effect is due to serine proteinases, i.e. thrombin-like enzymes, since this effect was completely abrogated by the combination of batimastat and PMSF. Our results stress the view that metalloproteinases play a relevant role in the systemic pathophysiology of B. asper envenomation and that metalloproteinase inhibitors may become a therapeutic alternative in this pathology.

Topics: Animals; Blood Coagulation; Bothrops; Costa Rica; Crotalid Venoms; Dose-Response Relationship, Drug; Drug Interactions; Factor X; Hemorrhage; Histological Techniques; Lethal Dose 50; Lung; Metalloendopeptidases; Mice; Phenylalanine; Prothrombin; Snake Bites; Thiophenes; Time Factors

Jararhagin is the most important hemorrhagic component in the venom of the snake Bothrops jararaca, a species of medical importance in South America. It is a P-III zinc-dependent metalloproteinase comprising catalytic, disintegrin-like, and cysteine-rich domains. Jararhagin injected intravenously into mice induced rapid and prominent bleeding in the lungs, whereas other organs were devoid of overt hemorrhagic manifestations. This action depends on the proteolytic activity of jararhagin, since it was abrogated by the synthetic inhibitor batimastat. There were conspicuous ultrastructural alterations in cells at the alveolo-capillary unit, i.e., capillary endothelial cells and type I pneumocytes, with a characteristic pattern of "regional alveolar damage" associated with extravasation. These pathological effects were observed under conditions in which the whole blood clotting time, bleeding time, and fibrinogen levels were not affected. 125I-labeled jararhagin is concentrated mainly in liver and kidneys after iv injection, with little radioactivity observed in the lungs, thereby indicating that the predominance of pulmonary microvascular damage is not due to a preferential concentration of this enzyme in the lungs. Despite the fact that jararhagin is complexed by plasma proteins after iv injection, its hemorrhagic activity was not inhibited by the plasma proteinase inhibitor alpha(2)-macroglobulin, and was only partially reduced by normal mouse serum, suggesting that resistance to inhibition may contribute to its ability to cause pulmonary hemorrhage.

Topics: alpha-Macroglobulins; Animals; Blood Coagulation; Bothrops; Bothrops jararaca Venom; Crotalid Venoms; Hemorrhage; Lung Diseases; Metalloendopeptidases; Mice; Microscopy, Electron; Phenylalanine; Platelet Aggregation Inhibitors; Protease Inhibitors; Thiophenes; Tissue Distribution

Batimastat (BB-94), a synthetic hydroxamate peptidomimetic matrix metalloproteinase inhibitor, was tested for its ability to inhibit proteolytic and toxic effects induced by BaP1, a 24-kDa hemorrhagic metalloproteinase isolated from the venom of Bothrops asper, the medically most important snake species in Central America and southern Mexico. Batimastat inhibited proteolytic activity on biotinylated casein, with anIC(50) of 80 nM. In addition, batimastat was effective in inhibiting hemorrhagic, dermonecrotic, and edema-forming activities of this metalloproteinase if incubated with the enzyme prior to the assays. When the inhibitor was administered i.m. at the site of the toxin injection without preincubation, rapidly after metalloproteinase administration, it totally abrogated the hemorrhagic and dermonecrotic effects of BaP1. Inhibition was less effective as the time lapse between toxin and batimastat injection increased, due to the extremely rapid development of BaP1-induced local tissue damage in this experimental model. On the other hand, batimastat was ineffective if administered by the i.p. route immediately after toxin injection. It is concluded that batimastat, and probably other synthetic metalloproteinase inhibitors, may become useful therapeutic tools aimed at the in situ inhibition of venom metalloproteinases, when injected at the site of the bite rapidly after envenomation.

Topics: Animals; Bothrops; Carbon-Oxygen Lyases; Crotalid Venoms; Deoxyribonuclease IV (Phage T4-Induced); DNA-(Apurinic or Apyrimidinic Site) Lyase; Drug Interactions; Edema; Hemorrhage; Matrix Metalloproteinase Inhibitors; Metalloendopeptidases; Mice; Phenylalanine; Protease Inhibitors; Thiophenes