Feature Articles Inosine improves gut permeability and vascular reactivity in endotoxic shock Francisco Garcia Soriano, MD, PhD; Lucas Liaudet, MD; Anita Marton, MD; György Haskó, MD, PhD; Clara Batista Lorigados, MD; Edwin A. Deitch, MD; Csaba Szabó, MD, PhD I t is well recognized that certain naturally occurring purines can exert powerful effects on the im- mune system. The nucleoside adenosine is the best characterized of these purines; both extracellular and in- tracellular adenosine have been shown to affect almost all aspects of an immune response (1–3). Adenosine and its analogs can alter the course of a variety of im- mune-mediated/inflammatory diseases, such as endotoxin shock (4, 5), rheuma- toid arthritis (6, 7), pleural inflammation (8), nephritis (9), and uveitis (10). Aden- osine is also recognized as one of the most important endogenous molecules able to prevent tissue injury in ischemia- reperfusion (11). Its effect is partly medi- ated by the inhibition of deleterious im- mune-mediated processes, including the release of proinflammatory cytokines and free radicals (11). Inosine is another endogenous purine nucleoside, which is formed during the breakdown of adenosine by adenosine deaminase (12). This molecule is released into the extracellular space from cells upon metabolic stress (13–15) or from the sympathetic nervous system (16). In ischemic tissues, inosine concentration can increase to levels as high as 1–3 mM (17), and increased tissue inosine levels are present in various inflammatory states (17–20). In a recent study, we showed that— contrary to the previous, widely held belief that inosine is an inert metabolite without biological effects— inosine inhibits proinflammatory cyto- kine and chemokine production in vitro and in vivo, enhances the production of the anti-inflammatory interleukin-10 via a posttranscriptional mechanism, and re- duces endotoxin-induced mortality in mice (21). In the present study, we investigated whether inosine can affect the deteriora- tion in vascular reactivity, intestinal function, neutrophil infiltration, and in- traorgan oxidant production, pivotal events of the septic multiple organ dys- function syndrome. Furthermore, we tested whether inosine maintains its pro- tective effects in the posttreatment regi- men. Our hypothesis was that inosine suppresses intraorgan oxidant production and neutrophil accumulation in endotox- emia and prevents the deterioration of vascular responsiveness. Furthermore, because of the posttranscriptional mech- anism of its anti-inflammatory action, we hypothesized that some of the protective effects of inosine may be sustained, even From Inotek Corporation (Drs. Soriano, Liaudet, Marton, Haskó, and Szabó), Beverly, MA; the Depart- ment of Surgery (Drs. Soriano, Liaudet, Haskó, Deitch, and Szabó), New Jersey Medical School, UMDNJ, Newark, NJ; and the Hospital das Clinicas FMUSP (Drs. Soriano and Lorigados), Sao Paulo, SP- Brazil. Supported, in part, by grants from the National Institutes of Health (R43GM59560 and R29GM54773) to Dr. Szabó. Dr. Liaudet is on leave from the Critical Care Division, Department of Internal Medicine, Uni- versity Hospital, Lausanne, Switzerland, and is sup- ported by a grant from the ADUMED foundation, Swit- zerland. Dr. Soriano is on leave from the Department of Critical Care Medicine, Hospital das Clinicas da Uni- versidade de Sao Paulo, Brasil, and is supported by a fellowship from FAPESP (Brazil). Address requests for reprints to: Csaba Szabó, MD, PhD, Inotek Corporation, Suite 419E, 100 Cummings Center, Bev- erly, MA 01915. E-mail: szabocsaba@aol.com Copyright © 2001 by Lippincott Williams & Wilkins Objective: To investigate the effects of inosine administration on vascular reactivity, gut permeability, neutrophil accumulation and lipid peroxidation in tissues in murine endotoxin shock. Design: Randomized, prospective laboratory study. Setting: Research laboratory. Subjects: BALB/c mice 6 – 8 wks age. Interventions: BALB/c mice were randomly assigned to one of five groups: a) vehicle controls, which received saline intraperi- toneally; b) inosine controls, which received inosine alone (100 mg/kg, ip); c) lipopolysaccharide (LPS)-treated animals, which received LPS (40 and 100 mg/kg, ip, depending on the experi- mental protocol); d) inosine pretreatment group, which received inosine (100 mg/kg, ip) 30 mins before LPS; and finally, e) inosine posttreatment group, which received inosine (100 mg/kg, ip) 60 mins after LPS. Measurements and Main Results: The passage of fluorescein isothiocyanate-conjugated dextran (4 kDa, FD4) was analyzed in everted gut ileal sacs incubated ex vivo as an index of gut permeability. LPS induced a significant intestinal hyperpermeabil- ity, and inosine exerted protective effects both in pre- and post- treatment regimens. Myeloperoxidase and malondialdehyde were also measured to study neutrophil accumulation and lipid peroxi- dation in selected tissues. Inosine, both in pre- and posttreatment regimens ameliorated the increases in myeloperoxidase and mal- ondialdehyde in the lung and gut. LPS-treated animals showed decreased contractile and relaxant responses, and inosine pre- treatment (but not posttreatment) partially improved these re- sponses. Conclusions: Taken together, inosine has organ protective effects during shock. A significant portion of its protective action is maintained even in the posttreatment scenario. (Crit Care Med 2001; 29:703–708) KEY WORDS: inosine; shock; endotoxin; gut; vascular; endothe- lial; permeability; reactivity; lung; liver; myeloperoxidase; malon- dialdehyde 703 Crit Care Med 2001 Vol. 29, No. 4