Enhancing the Neuroprotection Potential of Edaravone in Transient Global Ischemia Treatment with Glutathione- (GSH-) Conjugated Poly(methacrylic acid) Nanogel as a Promising Carrier for Targeted Brain Drug Delivery

Abstract

Ischemic stroke is the most common among various stroke types and the second leading cause of death, worldwide. Edaravone (EDV) is one of the cardinal antioxidants that is capable of scavenging reactive oxygen species, especially hydroxyl molecules, and has been already used for ischemic stroke treatment. However, poor water solubility, low stability, and bioavailability in aqueous media are major EDV drawbacks. Thus, to overcome the aforementioned drawbacks, nanogel was exploited as a drug carrier of EDV. Furthermore, decorating the nanogel surface with glutathione as targeting ligands would potentiate the therapeutic efficacy. Nanovehicle characterization was assessed with various analytical techniques. Size (199 nm, hydrodynamic diameter) and zeta potential (-25 mV) of optimum formulation were assessed. The outcome demonstrated a diameter of around 100 nm, sphere shape, and homogenous morphology. Encapsulation efficiency and drug loading were determined to be 99.9% and 37.5%, respectively. In vitro drug release profile depicted a sustained release process. EDV and glutathione presence in one vehicle simultaneously made the possibility of antioxidant effects on the brain in specific doses, which resulted in elevated spatial memory and learning along with cognitive function in Wistar rats. In addition, significantly lower MDA and PCO and higher levels of neural GSH and antioxidant levels were observed, while histopathological improvement was approved. The developed nanogel can be a suited vehicle for drug delivery of EDV to the brain and improve ischemia-induced oxidative stress cell damage.

Figure 1

Timeline of procedure, treatment, and evaluation of behavioral and biochemical analyses.

Figure 2

FTIR spectra of Bis AM, MAA, PMAA, GSH, and GSH-PMAA nanoparticles.

Figure 3

HNMR spectra of PMAA, GSH, and GSH-PMAA.

Figure 4

Particle size and morphology of GSH-PMAA using TEM at different magnification scale bars.

Figure 5

AFM image and morphology of GSH-PMAA.

Figure 6

In vitro drug release profile of EDV and GSH-PMAA-EDV nanogel.

Figure 7

Size (a) and zeta potential (b) of GSH-PMAA nanogel.

Figure 8

The effects of several doses of GSH-PMAA-EDV nanogel (5, 20, 40, and 250 μg/kg) on behavioral function. (a) Discrimination ratio as an output of novel object recognition test. (b) Tendency of the rats to choose one arm in 6 trials in T-maze test. (c) Choosing correct arm by rat in T-maze test. Values are expressed as the mean ± SD and were analyzed using one-way ANOVA followed by Bonferroni's multiple comparison test (n = 7 − 8). ^∗P < 0.05, ^∗∗P < 0.01, and ^∗∗∗P < 0.001 compared with the control group. ^#P < 0.05, ^##P < 0.01, and ^###P < 0.001 compared with stroke rats (n = 7 − 8).

Figure 9

The effects of several doses of GSH-PMAA-EDV nanogel (5, 20, 40, and 250 μg/kg) on biochemical alteration. (a) Reduced glutathione level, (b) FRAP level, (c) MDA level, and (d) protein carbonyl amount. Values are expressed as the mean ± SD and were analyzed using one-way ANOVA followed by Bonferroni's multiple comparison test. ^∗P < 0.05, ^∗∗P < 0.01, and ^∗∗∗P < 0.001 compared with the control group. ^#P < 0.05, ^##P < 0.01, and ^###P < 0.001 compared with stroke rats, and in case of GSH and FRAP levels, ^$P < 0.05, ^$$P < 0.01, and ^$$$P < 0.001 were compared with the stroke-GSH-PMAA group (n = 3 − 4).

Figure 10

Histological sections of experimental groups' brain. H&E staining, ×100. (a, b) Normal saline and GSH-PMAA group, respectively: histological structure was normal. (c) GSH-PMAA-EDV 250 μg/kg group: microglial nodule in the brain was observed (arrow). (d, e) TGI group: vacuolation and degeneration (arrows in (d)) and basophilic neuronal necrosis (arrows in (e)) were observed. (f–i) Received GSH-PMAA-EDV 5, 20, 40, and 250 μg/kg groups, respectively: histological structure was normal.