Bruno Palmier

Minocycline effects on cerebral edema: relations with inflammatory and oxidative stress markers following traumatic brain injury in mice.

One of the severe complications following traumatic brain injury (TBI) is cerebral edema and its effective treatment is of great interest to prevent further brain damage. This study investigated the effects of minocycline, known for its anti-inflammatory properties, on cerebral edema and its respective inflammatory markers by comparing different dose regimens, on oxidative stress and on neurological dysfunction following TBI. The weight drop model was used to induce TBI in mice. The brain water content was measured to evaluate cerebral edema. Inflammatory markers were detected by ELISA (IL-1beta), zymography and Western blot (MMP-9). The oxidative stress marker (glutathione levels) and neurological function were measured by Griffith technique and string test, respectively. Minocycline was administered i.p. once (5 min), twice (5 min and 3 h) or triple (5 min, 3 h and 9 h) following TBI. The first dose of minocycline only varied (45 or 90 mg/kg), whereas the following doses were all at 45 mg/kg. The single and double administrations of minocycline reduced the increase of inflammatory markers at 6 h post-TBI. Minocycline also reduced cerebral edema at this time point, only after double administration and at the high dose regimen, although with no effect on the TBI-induced oxidized glutathione increase. The anti-edematous effect of minocycline persisted up to 24 h, upon a triple administration, and accompanied by a neurological recovery. In conclusion, we reported an anti-edematous effect of minocycline after TBI in mice according to a specific treatment regimen. These findings emphasize that the beneficial effects of minocycline depend on the treatment regimen following a brain injury.

Selective inhibition of inducible nitric oxide synthase reduces neurological deficit but not cerebral edema following traumatic brain injury.

The role of inducible nitric oxide synthase (iNOS) in cerebral edema and neurological deficit following traumatic brain injury (TBI) is not yet clear-cut. Therefore, the aim of this study was to investigate the effect of three different iNOS inhibitors on cerebral edema and functional outcome after TBI. First, the time courses of blood--brain barrier (BBB) breakdown, cerebral edema, and neurological deficit were studied in a rat model of fluid percussion-induced TBI. The permeability of BBB to Evans blue was increased from 1 h to 24 h after TBI. Consistently, a significant increase in brain water content (BWC) was observed at 6 and 24 h post-TBI. A deficit in sensorimotor neurological functions was also observed from 6 h to 7 days with a maximum 24 h after TBI. Second, a single dose of aminoguanidine (AG; 100 mg/kg, i.p.), L-N-iminoethyl-lysine (L-NIL; 20 mg/kg, i.p.), or N-[3-(aminomethyl)benzyl]acetamide (1400W; 20 mg/kg, s.c.) was administered at 6 h post-TBI. Treatment with AG reduced by 71% the increase in BWC evaluated at 24 h, while L-NIL and 1400W had no effect. In contrast, the three iNOS inhibitors reduced the neurological deficit from 30% to 40%. Third, 1400W (20 mg/kg, s.c.) was administered at 5 min, 8 and 16 h post-TBI. Although this treatment paradigm had no effect on cerebral edema evaluated at 24 h, it significantly reduced the neurological deficit and iNOS activity. In conclusion, iNOS contributes to post-TBI neurological deficit but not to cerebral edema. The beneficial effect of iNOS inhibitors is not due to their anti-edematous effect, and the reduction of cerebral edema by AG is unlikely related to iNOS inhibition. The 6 h therapeutic window of iNOS inhibitors could allow their use in the treatment of functional deficit at the acute phase of TBI.

Changes in oxidative stress, iNOS activity and neutrophil infiltration in severe transient focal cerebral ischemia in rats

Oxidative stress, inducible nitric oxide synthase (iNOS) and neutrophils all contribute to post-ischemic brain damage. This study has determined the time courses of these three phenomena after ischemia in parallel with histological and functional outcomes. Ischemia was produced in rats by occluding the left middle cerebral artery and both common carotid arteries for 20 min. Regional cerebral blood flow (rCBF) rapidly decreased to 20% of its preischemic value during occlusion and stabilized at 60% following reperfusion. The striatal infarction was maximal 15 h after reperfusion (50+/-3 mm(3)), whereas the cortical infarction reached its maximum at 48 h (183+/-10 mm(3)). This drastic decrease in rCBF followed by incomplete reperfusion and massive infarction is, thus, extremely severe. The cortical infarction was strongly correlated with the neurologic deficit and loss of body weight. Oxidative stress, evaluated by the decrease in glutathione concentrations, appeared in the striatum at 6 h after reperfusion and in the cortex at 15 h. Calcium-independent NOS activity, considered as inducible NOS activity, was significantly enhanced at 24 h in the striatum and at 48 h in the cortex. Myeloperoxidase activity, a marker of neutrophil infiltration, was significantly increased at 48 h in both the striatum and cortex. These time courses show that the delayed iNOS activity and neutrophil infiltration that occur after the maturation of infarction in severe ischemia may not contribute to ischemic brain damage. By contrast, early oxidative stress may well be implicated in cerebral injury.

Static pressure-volume curves and effect of positive end-expiratory pressure on gas exchange in adult respiratory distress syndrome.

Fifteen patients with adult respiratory distress syndrome (ARDS) were studied: 11 in the early stage of ARDS (group 1); 4 in the late stage (group 2). The inspiratory and expiratory static pressure-volume (P-V) curves of the respiratory system were compared to the pulmonary shunt (Qsp/Qt) when PEEP was increased; cardiac output was kept constant. In group 1 patients, we found that a concavity on the P-V curves was associated with an abrupt decrease in Qsp/Qt when PEEP was increased; the concavity on the expiratory curve was correlated with the change in Qsp/Qt but not the concavity on the inspiratory curve. In group 2 patients, the P-V curves were found rectilinear and Qsp/Qt was not abruptly decreased when PEEP was increased. Expiratory P-V curves can be used to determine: first, whether a patient should be ventilated with PEEP; second, the PEEP level which can be set on the respirator. In group 1 patients, when PEEP was set to a value corresponding to the inflexion point, i.e., the point of departure from the exponential shape (mean value 14.6 ± 2.8 cm H2O), Qsp/Qt compared to zero PEEP was abruptly decreased to 87.6 ± 6%; further increase in PEEP had little advantage.

Percutaneous tracheostomy : prospective comparison of the translaryngeal technique versus the forceps-dilational technique in 100 critically ill adults

Objective To compare two different techniques of percutaneous tracheostomy: Griggs’ forceps-dilational technique and Fantoni’s translaryngeal technique, both performed with the manufacturer’s basic kit and with bronchoscopic guidance. Design A prospective, randomized trial was designed to compare the two tracheostomy techniques. Critically ill patients requiring elective tracheostomy for long-term ventilation were randomized for translaryngeal tracheostomy or forceps-dilational tracheostomy. Setting Intensive care unit of a military teaching hospital. Patients A total of 100 adult patients in the intensive care unit who were mechanically ventilated. Procedures All tracheostomy procedures were performed at the bedside by using a commercially available set. The procedures were performed by two surgeons, one for bronchoscopic guidance and management of the airway and one for the tracheostomy. Measurements and Main Results The measurements were divided into procedure-related variables (duration, technical difficulties, oxygenation): major and minor complications. The procedure was longer in the translaryngeal technique group (12.9 vs. 6.9 mins, p = .0018). Technical difficulties occurred in 11 patients in the translaryngeal technique group. Uneventful forceps dilational tracheostomy was performed instead. There has been no mortality associated with either technique. Serious complications occurred in one patient in the forceps-dilational technique group (one posterior tracheal wall injury) and in four patients in the translaryngeal technique group (one with a posterior tracheal wall injury and three with severe hypoxia). Significant hypercarbia and acidosis occurred in both the translaryngeal technique group and the forceps-dilational technique group. A significant decrease in Pao2 was observed in the translaryngeal technique group (311 to 261, p = .0069). No bleeding requiring intervention occurred. Conclusions Serious complications related to percutaneous tracheostomy occurred in 8.5% and 1.8% of the cases in the translaryngeal technique and the forceps-dilational technique group, respectively (p < .001). Technical difficulties were not rare when using the translaryngeal technique (23%). On the basis of our results, we concluded that the forceps-dilation technique is superior to the translaryngeal technique, with fewer technical difficulties and fewer complications for critically ill patients.

Detrimental Role of Bradykinin B2 Receptor in a Murine Model of Diffuse Brain Injury

Inhibition of the bradykinin B2 receptor type (B2R) has been shown to improve neurological outcome in models of focal traumatic brain injury. However, the involvement of B2R in trauma-induced diffuse injury has not yet been explored. This is an important point, since in humans a pattern of diffuse injury is commonly found in severely injured patients and has been associated with a poor neurological outcome and prognosis. Using the non-peptide B2R antagonist LF 16-0687 Ms and B2R null (B2R-/-) mice, we investigated the role of B2R in a model of closed head trauma (CHT). LF 16-0687 Ms given 30 min after injury reduced the neurological deficit by 26% and the cerebral edema by 22% when evaluated 4 h after CHT. Neurological function after CHT was improved in B2R-/- mice compared to B2R+/+ mice, although there was no difference in the development of brain edema. Treatment with LF 16-0687 Ms and B(2)R gene deletion decreased the accumulation of neutrophils at 24 h after CHT (50% and 36%, respectively). In addition, the inducible NO synthase (iNOS) mRNA level increased markedly, and this was reduced by LF 16-0687 Ms. Taken together, these data support a detrimental role of B2R in the development of the neurological deficit and of the inflammatory secondary damage resulting from diffuse traumatic brain injury. Therefore, blockade of bradykinin B2 receptors might represent an attractive therapeutic approach in the pharmacological treatment of traumatic brain injury.

Acute systemic inflammation induces central mitochondrial damage and mnesic deficit in adult Swiss mice

The aim of this study was to investigate how the brain is affected during systemic inflammation. For this purpose, Swiss mice were challenged with a single intraperitoneal dose of lipopolysaccharide (LPS; 250microg/mouse) to mimic aspects of systemic infection. Spatial learning in Y-maze test demonstrated a differential learning profile during the training test between control and LPS-treated mice, with an alteration in the latter group. We show that systemic LPS-induced inflammation and oxidative injury as assessed by reactive oxygen species (ROS) and nitrites/nitrates (NOx) production associated with reduced glutathione (GSH) depletion, cyclooxygenase-2 (COX-2) expression, and lipid peroxidation. LPS also induced a loss in mitochondrial integrity as shown by a significant decrease in membrane potential and impairment in mitochondrial redox activity. Thus, peripheral inflammation by producing brain inflammation and oxidative injury causes mnesic deficits. It remains to determine whether such events can induce neuronal dysfunction/degeneration and, with time, lead to cholinergic deficiency, amyloid deposits and cognitive impairments as they occur in Alzheimers disease.

LF 16-0687 Ms, a bradykinin B2 receptor antagonist, reduces ischemic brain injury in a murine model of transient focal cerebral ischemia.

Bradykinin promotes neuronal damage and brain edema through the activation of the B2 receptor. The neuroprotective effect of LF 16‐0687 Ms, a B2 receptor antagonist, has been described when given prior to induction of transient focal cerebral ischemia in rat, but there are no data regarding the consequence of a treatment when given after injury. Therefore, in a murine model of transient middle cerebral artery occlusion (MCAO), we evaluated the effect of LF 16‐0687 Ms given prior to and/or after the onset of ischemia on neurological deficit, infarct volume and inflammatory responses including cerebral edema, blood–brain barrier (BBB) disruption and neutrophil accumulation. LF 16‐0687 Ms (1, 2 and 4 mg kg−1) administered 0.5 h before and, 1.25 and 6 h after MCAO, decreased the infarct volume by a maximum of 33% and significantly improved the neurological recovery. When given at 0.25 and 6.25 h after MCAO, LF 16‐0687 Ms (1.5, 3 and 6 mg kg−1) decreased the infarct volume by a maximum of 25% and improved the neurological score. Post‐treatment with LF 16‐0687 Ms (1.5 mg kg−1) significantly decreased brain edema (−28%), BBB disruption (−60%) and neutrophil accumulation (−65%) induced by ischemia. Physiological parameters were not modified by LF 16‐0687 Ms. These data emphasize the role of bradykinin B2 receptor in the development of infarct lesion, neurological deficit and inflammatory responses resulting from transient focal cerebral ischemia. Therefore, B2 receptor antagonist might represent a new therapeutic approach in the pharmacological treatment of stroke.

Neutrophils do not contribute to infarction, oxidative stress, and NO synthase activity in severe brain ischemia

Polymorphonuclear leukocytes (PMNs) were reported to contribute to ischemia-reperfusion-induced brain damage. The present work examined whether PMN infiltration is deleterious in a severe model of transient focal cerebral ischemia and in which part PMNs contribute to oxidative stress and nitric oxide (NO) production. A 20-min occlusion of the left middle cerebral artery and both common carotid arteries was performed in rats. Infarction was maximal 24 h after reperfusion, while accumulation of PMNs in infarcted tissue was not significant before 48 h. Moreover, neutropenia induced by vinblastine (0.5 mg/kg iv) significantly decreased by 60-80% PMN infiltration 48 h after reperfusion but did not reduce the infarct volume. Thus PMNs do not contribute to cerebral injury in our model. Furthermore, decreased PMN infiltration modified neither oxidative stress evaluated by glutathione concentrations nor NO synthase activities 48 h after reperfusion. In conclusion, our results suggest that PMNs are not involved in severe cerebral ischemia and that anti-PMN strategies may be inefficient in some pathological conditions.

Reduction of hemorrhagic transformation by PJ34, a poly(ADP-ribose)polymerase inhibitor, after permanent focal cerebral ischemia in mice.

Hemorrhagic transformation is an aggravating event that occurs in 15 to 43% of patients suffering from ischemic stroke. This phenomenon due to blood-brain barrier breakdown appears to be mediated in part by matrix metalloproteinases (MMPs) among which MMP-2 and MMP-9 could be particularly involved. Recent experimental studies demonstrated that post-ischemic MMP-9 overexpression is regulated by poly(ADP-ribose)polymerase (PARP). In this context, our study aimed to evaluate the effect of PJ34 (N-(6-oxo-5,6-dihydrophenanthridin-2-yl)-2-(N,N-dimethylamino)acetamide), a potent PARP inhibitor, on MMP-2 and MMP-9 levels and on hemorrhagic transformations in a model of permanent focal cerebral ischemia in mice. PJ34 (6.25-12.5 mg/kg, i.p.) was given at the time of ischemia onset and 4 h later. Hemorrhagic transformations, divided into microscopic and macroscopic hemorrhages, were counted 48 h after ischemia on 12 coronal brain slices. Microscopic and macroscopic hemorrhages were respectively reduced by 38% and 69% with 6.25 mg/kg PJ34. The anti-hemorrhagic effect of PJ34 was associated with a 57% decrease in MMP-9 overexpression assessed by gelatin zymography. No increase in MMP-2 activity was observed after ischemia in our model. The vascular protection achieved by PJ34 was associated with a reduction in the motor deficit (P<0.05) and in infarct volume (-31%, P<0.01). In conclusion, our study demonstrates for the first time that PJ34 reduces hemorrhagic transformations after cerebral ischemia. Thus this PARP inhibitor exhibits both anti-hemorrhagic and neuroprotective effects that may be of valuable interest for the treatment of stroke.