The selective guanylate cyclase inhibitor ODQ reduces multiple organ injury in rodent models of Gram-positive and Gram-negative shock

Objective, An enhanced formation of endogenous nitric oxide contributes to the circulatory failure caused by endotoxin (lipopolysaccharide). Many of t he biological actions of nitric oxide are mediated by the guanylate cyclase /cyclic guanosine 3 ' ,5 ' -monophosphate system. We recently discovered th at two cell wall components, namely lipoteichoic acid and peptidoglycan of the Gram-positive bacterium Staphylococcus aureus, synergize to cause shock and multiple organ dysfunction syndrome in the rat. Here we investigate th e effects of a selective guanylate cyclase inhibitor, 1H-(1,2,4)oxadiazole( 4,3-alpha )quinoxaline-1 -one (ODQ), on the circulatory failure and multipl e organ dysfunction syndrome (kidney, liver, lung) caused by a) coadministr ation of lipoteichoic acid and peptidoglycan (Grampositive shock) or b) lip opolysaccharide (Gram-negative shock) in the anesthetized rat. Furthermore, we investigated whether ON scavenges superoxide anions and/or hydroxyl rad icals. Design. The in vivo portion of the study was a prospective, randomized, con trolled animal study. The in vitro portion included a) cultured ventricular myoblasts of the rat, H9c2(2-1) cells, and b) a cell free superoxide anion assay system. Setting: University-based research laboratory. Subjects: Seventy-five anesthetized, male Wistar rats were used for the in vivo study. Interventions. For the in vivo portion of the study, after surgical prepara tion, anesthetized rats were observed for 6 hrs. All rats were pretreated a nd received an intravenous infusion of saline (1.5 mL.kg(-1).hr(-1)), which was maintained throughout the experiment. The rats were assigned to nine g roups. Group I contained control rats (sham) subjected to 2 mL/kg saline in traperitoneally, 2 hrs before the experiment (n = 7). Group 2 contained con trol rats (sham) that received 2 mg/kg ODQ intraperitoneally, 2 hrs before the experiment (n = 9). Group 3 contained control rats (sham) that received 2 mL/kg dimethyl sulfoxide, 30% v/v in saline intraperitoneally, as a vehi cle for ODQ, 2 hrs before the experiment (n = 6). In group 4 rats, Gram-pos itive shock was induced by coadministration of lipoteichoic acid (3 mg/kg i ntravenously) and peptidoglycan (10 mg/kg intravenously) (n = 10). In group 5, rats were pretreated with ON (as described previously) before lipoteich oic acid/peptidoglycan (n = 9). In group 6, rats were pretreated with dimet hyl sulfoxide (as described previously) before lipoteichoic acid/peptidogly can (n = 7). In group 7, Gram-negative shock was induced by lipopolysacchar ide (6 mg/kg intravenously) (n = 11). In group 8, rats were pretreated with ODQ (as described previously) before lipopolysaccharide (n = 8). In group 9, rats were pretreated with dimethyl sulfoxide (as described previously) b efore lipopolysaccharide (n = 8). For the in vitro portion of the study, rat cells were preincubated with veh icle (saline and/or dimethyl suffoxide) and ON (0.1 muM to 1 mM) for 2 hrs. The cells then were exposed to H2O2 (1 MM) for 4 hrs at 37 degreesC, after which time cell viability was determined by measuring the mitochondrial-de pendent reduction of 3-(4,5-dimethyliazol-2-yl)-2,5-diphenyltetrazolium bro mide to blue formazan. Next, an aqueous solution was incubated with ODQ (as described previously), and superoxide anions were produced by using a hypo xanthine/xanthine-oxidase assay. The chemiluminescence assay was used to ev aluate any potential antioxidative effects of ODQ. Measurements and Main Results. In vivo, administration of lipoteichoic acid /peptidoglycan or lipopolysaccharide resulted within 6 hrs in hypotension, acute renal dysfunction, hepatocellular injury, and lung injury. Pretreatme nt of rats with ODQ attenuated the renal dysfunction, lung injury, and hepa tocellular injury caused by lipoteichoic acid/peptidoglycan or lipopolysacc haride. In vitro, administration of H2O2 (for 4 hrs) to rat cardiomyoblasts decreased mitochondrial respiration attributable to generation of hydroxyl radicals. Pretreatment of cells with ODQ did not abolish this cell injury. In addition, ODQ did not scavenge superoxide anions. Conclusions., These results imply that ODQ, an inhibitor of guanylate cycla se, reduces the multiple organ injury and dysfunction caused by wall fragme nts of Gram-positive or Gram-negative bacteria in the anesthetized rat. The observed protective effects of ODQ are not attributable to the ability of ON to reduce the formation or the effects of superoxide anions or hydroxyl radicals.