Protection against focal ischemic injury to the brain by trans-sodium crocetinate. Laboratory investigation

Abstract

Object: Ischemic injury is a potential complication in a variety of surgical procedures and is a particular impediment to the success of surgeries involving highly vulnerable neural tissue. One approach to limiting this form of injury is to enhance metabolic supply to the affected tissue. Trans-sodium crocetinate (TSC) is a carotenoid compound that has been shown to increase tissue oxygenation by facilitating the diffusivity of small molecules, such as oxygen and glucose. The present study examined the ability of TSC to modify oxygenation in ischemic neural tissue and tested the potential neuroprotective effects of TSC in permanent and temporary models of focal cerebral ischemia.

Methods: Adult male rats (330–370 g) were subjected to either permanent or temporary focal ischemia by simultaneous occlusion of both common carotid arteries and the left middle cerebral artery (3-vessel occlusion [3-VO]). Using the permanent ischemia paradigm, TSC was administered intravenously beginning 10 minutes after the onset of ischemia at 1 of 8 dosages, ranging from 0.023 to 4.580 mg/kg. Cerebral infarct volume was measured 24 hours after the onset of ischemia. The effect of TSC on infarct volume was also tested after temporary (2-hour) ischemia using a dosage of 0.092 mg/kg. In other animals undergoing temporary ischemia, tissue oxygenation was monitored in the ischemic penumbra using a Licox probe.

Results: Administration of TSC reduced infarct volume in a dose-dependent manner in the permanent ischemia model, achieving statistical significance at dosages ranging from 0.046 to 0.229 mg/kg. The most effective dosage of TSC in the permanent ischemia experiment (0.092 mg/kg) was further tested using a temporary (2-hour) ischemia paradigm. Infarct volume was reduced significantly by TSC in this ischemia-reperfusion model as well. Recordings of oxygen levels in the ischemic penumbra of the temporary ischemia model showed that TSC increased tissue oxygenation during vascular occlusion, but reduced the oxygen overshoot (hyperoxygenation) that occurs upon reperfusion.

Conclusions: The novel carotenoid compound TSC exerts a neuroprotective influence against permanent and temporary ischemic injury when administered soon after the onset of ischemia. The protective mechanism of TSC remains to be confirmed; however, the permissive effect of TSC on the diffusivity of small molecules is a plausible mechanism based on the observed increase in tissue oxygenation in the ischemic penumbra. This represents a form of protection based on “metabolic reflow” that can occur under conditions of partial vascular perfusion. It is particularly noteworthy that TSC could conceivably limit the progression of a wide variety of cellular injury mechanisms by blunting the ischemic challenge to the brain.

Fig. 1

Effect of TSC on cerebral infarct volume in a model of permanent (24-hour) focal ischemia. The bar graph shows group means and SEs for the vehicle- and TSC-treated animals (7 rats per group). The volume of cerebral infarction was reduced by TSC in a dose-dependent manner, characterized by a U-shaped curve. This protective action achieved statistical significance at TSC dosages ranging from 0.023 to 0.229 mg/kg, with a maximal protective effect being achieved at 0.092 mg/kg. * p < 0.05, 1-way ANOVA and the LSD post hoc test.

Fig. 2

Examples of cerebral infarcts in vehicle-treated and TSC-treated animals. Serial coronal sections obtained from a vehicle-treated animal and a TSC-treated animal (0.092 mg/kg) are shown. Sections were stained with tetrazolium chloride for the detection of infarcted tissue. Areas of tissue infarction appear white in these sections, while the healthy tissue appears red. The infarct size is substantially smaller inthe animal from the TSC-treated group.

Fig. 3

Bar graph showing effect of TSC on cerebral infarct volume after transient focal ischemia. The graph shows group means and SEs for cerebral infarct volume in vehicle-treated and TSC-treated (0.092 mg/kg) animals (6 rats per group). The cerebral infarct volume was reduced significantly in the TSC-treated group. ** p < 0.01, Student t-test.

Fig. 4

Graph showing brain tissue partial oxygen levels during ischemia-reperfusion. Values in each animal were normalized to a preischemia baseline and this baseline is expressed as a value of 1.00. Group means and SEs are shown for vehicle-treated animals (7 rats, filled circles) and TSC-treated animals (5 rats, filled triangles). The timings of bolus and infusion treatments of vehicle or TSC are indicated on the graph. Treatment with TSC produced a progressive increase in tissue oxygen levels during ischemia. Upon reperfusion, a substantial hyperoxia response was observed in the vehicle-treated group; this effect was blunted in the TSC-treated group. Note that the y axis is interrupted on the graph above the baseline level to facilitate group comparisons both during and after ischemia. * p < 0.05, compared with corresponding time points for the 2 groups.