Omar Elmadhoun

Ethanol and Normobaric Oxygen: Novel Approach in Modulating Pyruvate Dehydrogenase Complex After Severe Transient and Permanent Ischemic Stroke

Background and Purpose— Ischemic stroke induces metabolic disarray. A central regulatory site, pyruvate dehydrogeanse complex (PDHC) sits at the cross-roads of 2 fundamental metabolic pathways: aerobic and anaerobic. In this study, we combined ethanol (EtOH) and normobaric oxygen (NBO) to develop a novel treatment to modulate PDHC and its regulatory proteins, namely pyruvate dehydrogenase phosphatase and pyruvate dehydrogenase kinase, leading to improved metabolism and reduced oxidative damage. Methods— Sprague–Dawley rats were subjected to transient (2, 3, or 4 hours) middle cerebral artery occlusion followed by 3- or 24-hour reperfusion, or permanent (28 hours) middle cerebral artery occlusion without reperfusion. At 2 hours after the onset of ischemia, rats received either an intraperitoneal injection of saline, 1 dose of EtOH (1.5 g/kg) for 2- and 3-hour middle cerebral artery occlusion, 2 doses of EtOH (1.5 g/kg followed by 1.0 g/kg in 2 hours) in 4 hours or permanent middle cerebral artery occlusion, and EtOH+95% NBO (at 2 hours after the onset of ischemia for 6 hours) in permanent stroke. Infarct volumes and neurological deficits were examined. Oxidative metabolism and stress were determined by measuring ADP/ATP ratio and reactive oxygen species levels. Protein levels of PDHC, pyruvate dehydrogenase kinase, and pyruvate dehydrogenase phosphatase were assessed. Results— EtOH induced dose-dependent neuroprotection in transient ischemia. Compared to EtOH or NBO alone, NBO+EtOH produced the best outcomes in permanent ischemia. These therapies improved brain oxidative metabolism by decreasing ADP/ATP ratios and reactive oxygen species levels, in association with significantly raised levels of PDHC and pyruvate dehydrogenase phosphatase, as well as decreased pyruvate dehydrogenase kinase. Conclusions— Both EtOH and EtOH+NBO treatments conferred neuroprotection in severe stroke by affecting brain metabolism. The treatment may modulate the damaging cascade of metabolic events by bringing the PDHC activity back to normal metabolic levels.

Reduced Apoptosis by Ethanol and Its Association with PKC-δ and Akt Signaling in Ischemic Stroke

Along with thrombolytic therapy, which has a number of limitations, stroke outcome may be improved with neuroprotective therapies that disrupt ischemic cell death. Recent research has shown a neuroprotective role of ethanol administration during ischemic stroke, such as its ability to reduce infarct volume and neurologic deficit. In order to investigate this further, we assessed the hypothesis that ethanols neuroprotective effect is through reduction of apoptosis and the modulation of the important apoptotic PKC-δ and Akt signaling pathway. Ethanol (1.5 g/kg) was given by intraperitoneal injections to 54 Sprague-Dawley rats after 2 hours of middle cerebral artery (MCA) occlusion, followed by 3 or 24 hours of reperfusion. We measured apoptotic cell death, PKC-δ, and Akt mRNA and protein expressions in each of ischemic groups with or without ethanol treatment using ELISA, real-time PCR and Western blot analysis. Our results showed that cell death was significantly increased in rats following 2 hour MCA occlusion and 24 hour reperfusion. Subsequently, cell death was significantly reduced by an administration of ethanol. We further found that ethanol administration, prior to either 3 or 24 hours of reperfusion, significantly decreased the expression of PKC-δ while simultaneously increasing the expression Akt at both mRNA and protein levels at the two points. In conclusion, our study suggests that ethanol administration following ischemic stroke modulates the gene and protein profile in such a way that it increased expression of anti-apoptotic Akt and decreased the pro-apoptotic PKC-δ. This ultimately results in a decrease in neuronal apoptosis, thus conferring neuroprotection.

Dihydrocapsaicin (DHC) enhances the hypothermia-induced neuroprotection following ischemic stroke via PI3K/Akt regulation in rat

OBJECTIVE Hypothermia has demonstrated neuroprotection following ischemia in preclinical studies while its clinical application is still very limited. The aim of this study was to explore whether combining local hypothermia in ischemic territory achieved by intra-arterial cold infusions (IACIs) with pharmacologically induced hypothermia enhances therapeutic outcomes, as well as the underlying mechanism. METHODS Sprague-Dawley rats were subjected to right middle cerebral artery occlusion (MCAO) for 2h using intraluminal hollow filament. The ischemic rats were randomized to receive: 1) pharmacological hypothermia by intraperitoneal (i.p.) injection of dihydrocapsaicin (DHC); 2) physical hypothermia by IACIs for 10min; or 3) the combined treatments. Extent of brain injury was determined by neurological deficit, infarct volume, and apoptotic cell death at 24h and/or 7d following reperfusion. ATP and ROS levels were measured. Expression of p-Akt, cleaved Caspase-3, pro-apoptotic (AIF, Bax) and anti-apoptotic proteins (Bcl-2, Bcl-xL) was evaluated at 24h. Finally, PI3K inhibitor was used to determine the effect of p-Akt. RESULTS DHC or IACIs each exhibited hypothermic effect and neuroprotection in rat MCAO models. The combination of pharmacological and physical approaches led to a faster and sustained reduction in brain temperatures and improved ischemia-induced injury than either alone (P<0.01). Furthermore, the combination treatment favorably increased the expression of anti-apoptotic proteins and decreased pro-apoptotic protein levels (P<0.01 or 0.05). This neuroprotective effect was largely blocked by p-Akt inhibition, indicating a potential role of Akt pathway in this mechanism (P<0.01 or 0.05). CONCLUSIONS The combination approach is able to enhance the efficiency of hypothermia and efficacy of hypothermia-induced neuroprotection following ischemic stroke. The findings here move us a step closer towards translating this long recognized TH from bench to bedside.

Protocol-directed weaning from mechanical ventilation in neurological patients: a randomised controlled trial and subgroup analyses based on consciousness

Abstract Objectives: To assess whether a weaning protocol reduces the mechanical ventilation (MV) duration compared to physicians judgement-based weaning in neurological patients and to determine whether patient consciousness influences this reduction. Methods: A randomised controlled trial was conducted in a neurological intensive care unit (NCU) of a tertiary hospital; 144 patients requiring MV for more than 24 hours were randomly allocated to protocol-directed (intervention) (n = 71) or physician-directed (control) group (n = 73). Results: The intervention group displayed a significantly shorter median weaning time than the control group (2.00 vs 5.07 days, P < 0.05). The median MV duration tended to be shorter in the intervention group (10.8 vs 14.2 days, P = 0.106). The median length of NCU stay was 19.0 and 26.1 days in the intervention and control groups, respectively (P = 0.063). The median NCU cost was 9.26 × 104 and 12.24 × 104 ¥ in the intervention and control groups, respectively (P = 0.059). The unsuccessful weaning, ventilator-associated pneumonia (VAP) and mortality rates were similar between the groups. Among conscious patients, the median weaning time (2.00 vs 7.00 days, P < 0.05) and the median MV duration (8.8 vs 18.0 days, P = 0.017) were significantly reduced in the intervention group. Among unconscious patients, the intervention group displayed a reduced median weaning time (1.00 vs 3.10 days, P < 0.05), but not median MV duration (11.6 vs 11.1 days, P = 0.702), compared to the control group. Conclusion: Protocol-directed weaning reduces weaning time, MV duration, length of NCU stay and NCU cost in neurological patients, and these effects are more significant in conscious patients than in unconscious patients.

Synergistically Induced Hypothermia and Enhanced Neuroprotection by Pharmacological and Physical Approaches in Stroke

Hypothermia is considered as a promising neuroprotective treatment for ischemic stroke but with many limitations. To expand its clinical relevance, this study evaluated the combination of physical (ice pad) and pharmacological [transient receptor potential vanilloid channel 1 (TRPV1) receptor agonist, dihydrocapsaicin (DHC)] approaches for faster cooling and stronger neuroprotection. A total of 144 male Sprague Dawley rats were randomized to 7 groups: sham (n=16), stroke only (n=24), stroke with physical hypothermia at 31ºC for 3 h after the onset of reperfusion (n=24), high-dose DHC (H-DHC)(1.5 mg/kg, n=24), low-dose DHC (L-DHC)(0.5 mg/kg, n=32) with (n=8) or without (n=24) external body temperature control at ~38 ºC (L-DHC, 38 ºC), and combination therapy (L-DHC+ ice pad, n=24). Rats were subjected to middle cerebral artery occlusion (MCAO) for 2 h. Infarct volume, neurological deficits and apoptotic cell death were determined at 24 h after reperfusion. Expression of pro- and anti-apoptotic proteins was evaluated by Western blot. ATP and reactive oxygen species (ROS) were detected by biochemical assays at 6 and 24 h after reperfusion. Combination therapy of L-DHC and ice pad significantly improved every measured outcome compared to monotherapies. Combination therapy achieved hypothermia faster by 28.6% than ice pad, 350% than L-DHC and 200% than H-DHC alone. Combination therapy reduced (p<0.05) neurological deficits by 63% vs. 26% with L-DHC. No effect was observed when using ice pad or H-DHC alone. L-DHC and ice pad combination improved brain oxidative metabolism by reducing (p<0.05) ROS at 6 and 24 h after reperfusion and increasing ATP levels by 42.9% compared to 25% elevation with L-DHC alone. Finally, combination therapy decreased apoptotic cell death by 48.5% vs. 24.9% with L-DHC, associated with increased anti-apoptotic protein and reduced pro-apoptotic protein levels (p<0.001). Our study has demonstrated that combining physical and pharmacological hypothermia is a promising therapeutic approach in ischemic stroke, and warrants further translational investigations.

Phenothiazines Enhance Mild Hypothermia-induced Neuroprotection via PI3K/Akt Regulation in Experimental Stroke

Physical hypothermia has long been considered a promising neuroprotective treatment of ischemic stroke, but the treatment’s various complications along with the impractical duration and depth of therapy significantly narrow its clinical scope. In the present study, the model of reversible right middle cerebral artery occlusion (MCAO) for 2 h was used. We combined hypothermia (33–35 °C for 1 h) with phenothiazine neuroleptics (chlorpromazine & promethazine) as additive neuroprotectants, with the aim of augmenting its efficacy while only using mild temperatures. We also investigated its therapeutic effects on the Phosphatidylinositol 3 kinase/Protein kinase B (PI3K/Akt) apoptotic pathway. The combination treatment achieved reduction in ischemic rat temperatures in the rectum, cortex and striatum significantly (P < 0.01) faster than hypothermia alone, accompanied by more obvious (P < 0.01) reduction of brain infarct volume and neurological deficits. The combination treatment remarkably (P < 0.05) increased expression of p-Akt and anti-apoptotic proteins (Bcl-2 and Bcl-xL), while reduced expression of pro-apoptotic proteins (AIF and Bax). Finally, the treatment’s neuroprotective effects were blocked by a p-Akt inhibitor. By combining hypothermia with phenothiazines, we significantly enhanced the neuroprotective effects of mild hypothermia. This study also sheds light on the possible molecular mechanism for these effects which involves the PI3K/Akt signaling and apoptotic pathway.

Aetiological diagnosis of middle-aged and elderly cryptogenic ischaemic cerebral vascular disease

Abstract Although tremendous efforts have been made to explore the potential aetiologies of cryptogenic ischaemic cerebral vascular disease (CICVD), it remains a great challenge for neurologists to get a comprehensive picture of CICVD across the world. Part of the reason why is that the vast majority of studies have focussed on CICVD in young stroke patients while the underlying causes of CICVD in middle-aged or elderly stroke population have not been fully investigated. The focus of this paper has been dedicated to review the different studies that explore the aetiologies of CICVD cases in this patient population. While there is a set of heterogeneous causes that can lead to CICVD in middle-aged and elderly patients, our review reveals that emboli originated from or across occult places within the heart or produced by transient arrhythmias could possibly be the main culprit. Dislodged aortic plaques might also account for certain CICVD cases and in fewer cases, hereditary arteriopathy and thrombophilia can also play a role. The aforementioned factors have similar roles in middle-aged and elderly CICVD patients as in their younger counterparts. However, more studies are needed to explore the role of these factors in older patients.