Efficacy of Novel Carbon Nanoparticle Antioxidant Therapy in a Severe Model of Reversible Middle Cerebral Artery Stroke in Acutely Hyperglycemic Rats.

Roderic H Fabian,Lizanne G Nilewski,Paul J Derry,Harriett Charmaine Rea,James M Tour,William V Dalmeida,Thomas A Kent,Pitchaiah Mandava,Vladimir Berka,Ah-Lim Tsai,Kimberly Mendoza,William K A Sikkema

doi:10.3389/fneur.2018.00199

Abstract

While oxidative stress can be measured during transient cerebral ischemia, antioxidant therapies for ischemic stroke have been clinically unsuccessful. Many antioxidants are limited in their range and/or capacity for quenching radicals and can generate toxic intermediates overwhelming depleted endogenous protection. We developed a new antioxidant class, 40 nm × 2 nm carbon nanoparticles, hydrophilic carbon clusters, conjugated to poly(ethylene glycol) termed PEG-HCCs. These particles are high-capacity superoxide dismutase mimics, are effective against hydroxyl radical, and restore the balance between nitric oxide and superoxide in the vasculature. Here, we report the effects of PEG-HCCs administered during reperfusion after transient middle cerebral artery occlusion (tMCAO) by suture in the rat under hyperglycemic conditions. Hyperglycemia occurs in one-third of stroke patients and worsens clinical outcome. In animal models, this worsening occurs largely by accelerating elaboration of reactive oxygen species (ROS) during reperfusion. PEG-HCCs were studied for their protective ability against hydrogen peroxide in b.End3 brain endothelial cell line and E17 primary cortical neuron cultures. In vivo, hyperglycemia was induced by streptozotocin injection 2 days before tMCAO. 58 Male Sprague-Dawley rats were analyzed. They were injected IV with PBS or PEG-HCCs (4 mg/kg 2×) at the time of recanalization after either 90- or 120-min occlusion. Rats were survived for up to 3 days, and infarct volume characteristics and neurological functional outcome (modified Bederson Score) were assessed. PEG-HCCs were protective against hydrogen peroxide in both culture models. In vivo improvement was found after PEG-HCCs with 90-min ischemia with reduction in infarct size (42%), hemisphere swelling (46%), hemorrhage score (53%), and improvement in Bederson score (70%) (p = 0.068-0.001). Early high mortality in the 2-h in the PBS control group precluded detailed analysis, but a trend was found in improvement in all factors, e.g., reduction in infarct volume (48%; p = 0.034) and a 56% improvement in Bederson score (p = 0.055) with PEG-HCCs. This nano-antioxidant showed some improvement in several outcome measures in a severe model of tMCAO when administered at a clinically relevant time point. Long-term studies and additional models are required to assess potential for clinical use, especially for patients hyperglycemic at the time of their stroke, as these patients have the worst outcomes.

Highlights

While oxidative stress can be measured during transient cerebral ischemia, antioxidant therapies for ischemic stroke have been clinically unsuccessful
We demonstrated their superior efficacy to two clinical failed antioxidants, poly(ethylene glycol) (PEG)-superoxide dismutase (SOD) and the precursor to NXY-059 [4], phenyl butyl nitrone, PBN [41] in culture by their ability to reduce the damaging effect of the mitochondrial toxin, antimycin A, when given AFTER the toxin, while pretreatment was needed for the other agents
In E17 neurons, we found that 100 μM H2O2 was more lethal in neurons than b.End3 cells, partial restoration was achieved with posttreatment with PEG-hydrophilic carbon clusters (HCCs) (Figure 2)

Summary

Introduction

While oxidative stress can be measured during transient cerebral ischemia, antioxidant therapies for ischemic stroke have been clinically unsuccessful. We believe this failure is due to two major factors: [1] There are severe limitations in currently available antioxidants that hinder their effectiveness when employed following ischemia as opposed to pretreatment [5] and [2] oxidative stress injury is quantitatively more important under specific clinical circumstances, so a benefit might be missed if it is not tested under the most relevant conditions In stroke, those conditions are typically those that have the worst outcomes such as hyperglycemia at the time of stroke when treated with recanalization therapy [6]. These mechanisms consist of enzymes and other proteins that modify the radical species in a series of steps leading to water

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