Inhibition of c-Jun N-terminal kinase (JNK) by a cell-penetrating, protease-resistant JNK peptide (D-JNKI-1) before hemorrhage and resuscitation (H/R) ameliorated the H/R-induced hepatic injury and blunted the proinflammatory changes. Here we tested the hypothesis if JNK inhibition at a later time point-after hemorrhagic shock but before the onset of resuscitation-in a rat model of H/R also confers protection. Twenty-four male Sprague-Dawley rats (250 - 350 g) were randomly divided into 4 groups: 2 groups of shock animals were hemorrhaged to a MAP of 32 to 37 mmHg for 60 min and randomly received either D-JNKI-1 (11 mg/kg i.p.) or sterile saline as vehicle immediately before the onset of resuscitation. Two groups of sham-operated animals underwent surgical procedures without H/R and were either D-JNKI-1 or vehicle treated. Rats were killed 2 h later. Serum activity of alanine aminotransferase and serum lactate dehydrogenase after H/R increased 3.5-fold in vehicle-treated rats as compared with D-JNKI-1-treated rats. Histopathological analysis revealed that hepatic necrosis and apoptosis (hematoxylin-eosin, TUNEL, and M30, respectively) were significantly inhibited in D-JNKI-1-treated rats after H/R. Hepatic oxidative (4-hydroxynonenal) and nitrosative (3-nitrotyrosine) stress as well as markers of inflammation (hepatic and serum IL-6 levels and hepatic infiltration with polymorphonuclear leukocytes) were also reduced in D-JNKI-1-treated rats. LPS-stimulated TNF-alpha release from whole blood from hemorrhaged and resuscitated animals was higher in vehicle-treated rats as compared with D-JNKI-1-treated rats. c-Jun N-terminal kinase inhibition after hemorrhage before resuscitation resulted in a reduced activation of c-Jun. Taken together, these results indicate that D-JNKI-1 application after hemorrhagic shock before resuscitation blunts hepatic damage and proinflammatory changes during resuscitation. Hence, JNK inhibition is even protective when initiated after blood loss before resuscitation. These experimental results indicate that the JNK pathway may be a possible treatment option for the harmful consequences of H/R.