Combined zinc supplementation with proinsulin C-peptide treatment decreases the inflammatory response and mortality in murine polymicrobial sepsis.

Zinc is a trace element vital for immune function during host response to infection. The proinsulin C-peptide has been shown to exert beneficial effects through activation of the anti-inflammatory peroxisome proliferator-activated receptor γ (PPARγ) in experimental endotoxemia. Some in vitro activities of C-peptide appear dependent on the presence of zinc. We investigated the effect of zinc supplementation before onset of sepsis on the anti-inflammatory properties of C-peptide. Male C57BL/6 mice were subjected to polymicrobial sepsis by cecal ligation and puncture (CLP). Mice received zinc gluconate (1.3 mg/kg) intraperitoneally (i.p.) for 3 days before CLP. One hour after CLP, animals received C-peptide (280 nmol/kg i.p.) or the antimicrobial agent imipenem (25 mg/kg i.p.). Cecal ligation and puncture was associated with an 11% survival rate, pulmonary leukosequestration, and liver injury. Molecular analysis in lungs of septic mice showed increased nuclear activation of the proinflammatory extracellular signal-regulated kinases 1 and 2 and nuclear factor κB, but decreased PPARγ expression, when compared with sham animals. Combination of zinc supplementation with C-peptide posttreatment significantly improved survival rate (61%) similarly to antibiotic treatment (60%), ameliorated lung architecture and liver function, reduced tissue neutrophil infiltration, and increased bacterial clearance when compared with vehicle, C-peptide, or zinc treatment alone. These beneficial effects were associated with restored lung nuclear expression of PPARγ and reduction of phosphorylated extracellular signal-regulated kinases 1 and 2 and nuclear factor κB activities in comparison to vehicle or single treatment protocols. Our data demonstrate that short-term zinc prophylaxis before the infectious insult is a requisite for the anti-inflammatory properties of C-peptide by facilitating modulation of inflammatory pathways.

Hypoxic stress exacerbates hyperoxia-induced lung injury in a neonatal mouse model of bronchopulmonary dysplasia.

BACKGROUND: Premature infants with lung injury often experience intermittent episodes of hypoxemia. OBJECTIVE: This study investigates whether intermittent hypoxemia exacerbates oxidative stress and lung injury in neonatal mice in a hyperoxia-induced model of bronchopulmonary dysplasia (BPD). METHODS: For the BPD model, 3-day-old C57Bl/6J mice were exposed to hyperoxia (65% O(2)) for 4 weeks (O(2) group) or to hyperoxia and intermittent (10 min daily) hypoxia (O(2) + H group). Upon completion of O(2) or O(2) + H exposure, the degree of pulmonary alveolarization and granulocytic infiltration were examined. The severity of oxidative injury in lungs was defined by tissue glutathione and protein carbonyl content. Data were compared to those in naïve mice and mice subjected only to intermittent hypoxia. RESULTS: Hyperoxia-exposed mice exhibited a dramatic (p < 0.0001) decrease of alveolarization, significantly increased granulocytic infiltration (p < 0.0001) and increased protein carbonyl content (p = 0.04) compared to naïve mice. However, O(2) + H mice demonstrated significantly (p = 0.03) fewer alveoli compared to their O(2) counterparts. This was associated with a significantly (p = 0.02) decreased pulmonary total/oxidized glutathione ratio and a significant (p = 0.03) elevation of protein carbonyl content. CONCLUSIONS: Thus, intermittent hypoxic stress during hyperoxic induction of BPD in mice potentiates oxidative stress in lung tissue and exacerbates alveolar developmental arrest.

Systemic hyperthermia induces ischemic brain injury in neonatal mice with ligated carotid artery and jugular vein.

Postnatal d 7 (p7) or p12 mice had their right carotid artery (CA) and jugular vein (JV) ligated to mimic veno-arterial (VA) access for extracorporeal membrane oxygenation (ECMO). At p9-11 (early) or p19-21 (late) mice were exposed to hyperthermia or normothermia followed by assessment of neuropathological injury score. In separate cohorts of mice, cerebral and peripheral blood flow (CBF, PBF) and cerebral ATP content was measured. Hyperthermia resulted in ischemic brain injury in 57% and 77% of mice subjected to early or late hyperthermia, respectively. Isolated CA+JV ligation induced minimal injury (score 0.47 +/- 0.34) in 2/8 mice from the late normothermia group. No cerebral injury was detected in mice subjected to early normothermia. In 3/19 shams (2/10 early, 1/9 late) hyperthermia induced a subtle (score, 0.6 +/- 0.27) injury in the ipsilateral to the site of surgery cortex. CBF and PBF increased in response to hyperthermia in all mice. The rise in CBF was significantly attenuated in the "ligated" versus intact hemisphere, which was associated with a profound depletion of ATP content. Systemic hyperthermia induces ischemic brain injury in mice with ligated CA+JV. We speculate that hyperthermia/fever can be a potential risk factor for brain injury in infants treated with VA ECMO.

The contribution of intermittent hypoxemia to late neurological handicap in mice with hyperoxia-induced lung injury.

Bronchopulmonary dysplasia (BPD) is considered by many to be an independent risk factor for poor neurodevelopment in premature infants. However, infants with BPD experience intermittent hypoxic episodes. This study was undertaken to determine whether intermittent hypoxic stress associated with BPD contributes to the development of neurological deficit. The model of BPD was produced in neonatal mice by exposure to hyperoxia (65% O(2)) for 4 weeks. Arterial blood gases, pulmonary mechanics, and histopathology were used to define the degree of lung injury. The mice were subjected to brief (10 min/day) and intermittent (10 days) hypoxic stress (8% O(2)) at different stages of the development of hyperoxia-induced lung injury. At 8 weeks of life, the neurofunction was assessed by water maze and rota-rod tests followed by cerebral morphological analysis using Nissl, bromodeoxyuridine, and caspase-3 immunostaining. Data were compared to naïve normoxic littermates and those mice that were exposed only to hyperoxia or intermittent hypoxia alone. Mice with BPD subjected to brief/intermittent hypoxia demonstrated a significantly poorer navigational memory performance as compared with normoxic mice and mice with BPD that were not subjected to intermittent hypoxia. The neurofunctional handicap in these mice was associated with significantly decreased brain weight and increased cerebral expression of caspase-3. Our results suggest that intermittent hypoxia associated with hyperoxia-induced lung injury, but not lung injury itself, results in significant neurological handicap in neonatal mice with BPD.

Reoxygenation with 100% oxygen versus room air: late neuroanatomical and neurofunctional outcome in neonatal mice with hypoxic-ischemic brain injury.

Study investigated neuroutcome in mice subjected at 7-8 d of life to hypoxic-ischemic brain injury (HI) followed by 30 min of reoxygenation with 100% O(2) (Re-O(2)) or room air (Re-Air). At 24 h of recovery, mouse reflexes were tested. At 7 wks after HI spatial orientation and memory were assessed in the same mice. Mortality rate was recorded at 24 h and at 7 wks of recovery. In separate cohort of mice, changes in cerebral blood flow (CBF) during HI-insult and reoxygenation were recorded. Re-O(2)versus Re-Air mice exhibited significantly delayed geotaxis reflex. Adult Re-O(2)versus Re-Air mice exhibited significantly better spatial learning and orientation with strong tendency toward better preserved memory. Histopathology revealed significantly less hippocampal atrophy in Re-O(2)versus Re-Air mice. Following a hypoxia-induced hypoperfusion, Re-O(2) re-established CBF in the ipsilateral side to the prehypoxic level significantly faster than Re-Air. The mortality was higher among Re-O2 versus Re-Air mice, although, it did not reach statistical significance. Re-O(2)versus Re-Air restores CBF significantly faster and results in better late neuroutcome. However, greater early motor deficit and higher mortality rate among Re-O(2)versus Re-Air mice suggest that Re-O(2) may be deleterious at the early stage of recovery.