BAR is a naturally occurring bioactive anthracycline derived from the leaf exudates of aloe plants, and aloe vera has been used an ingredient in a wide range of dietary and cosmetic products (18). Mounting evidence indicates that aloe exerts a variety of pharmacological activities, including neuroprotective effects (19). For instance, Rathor and colleagues have documented that the extract of aloe vera leaf has anticonvulsant and anti-oxidant activities against acute and chronic epilepsy in mice (20). However, the neuroprotective effects of BAR, the most abundant anthracycline present in aloe vera, have not been reported. In this study, intraperitoneal injection of BAR significantly attenuated MCAO-induced cerebral I/R injury in rats, as shown by improvements in neurological scores and decreases in brain infarct volume. Meanwhile, BAR reduced oxidative stress and iNOS expression and inhibited neuronal apoptosis in cerebral I/R rats. In addition, pretreatment with BAR suppressed LPS-stimulated inflammatory responses, iNOS expression, and NF-κB activation in BV-2 microglial cells.
Aberrant neuronal apoptosis represents a pathological hallmark of many human neurological diseases, such as diabetic encephalopathy, stroke, Alzheimer’s disease, and amyotrophic lateral sclerosis (21, 22). It has been demonstrated that apoptosis is responsible for neuronal death in cortex following cerebral ischemia (23). In this study, the number of TUNEL-positive cells were increased in the ischemic cortex of I/R rats compared with that in the sham rats, and this increase in TUNEL-positive cells was remarkably reversed by BAR treatment. Our findings agree with a previous study showing that BAR inhibits apoptosis in cardiomyocytes and attenuates myocardial I/R injury in rats (15). Caspase-3, an important executioner of programmed cell death, is activated to initiates apoptotic DNA fragmentation in MCAO-induced focal cerebral I/R injury, thereby leading to expansion of infarct volume (24). The Bcl-2 family is consisted of anti-apoptotic (Bcl-2, Bcl-xl, and Bcl-w) and pro‐apoptotic members (Bax, Bak, and Bok) (25). Herein, we observed that BAR administration suppressed activated caspase-3 and Bax protein levels, but increased Bcl-2 protein levels in the ischemic brains, suggesting the mechanisms by which BAR inhibits neuronal apoptosis. However, the mechanisms underlying regulation of Bcl-2 and Bax by BAR need to be further investigated.
Oxidative stress, also termed as excessive reactive oxygen species (ROS) production overwhelming intrinsic antioxidant defense, is linked to the pathophysiology of brain I/R injury (26). High levels of ROS can react with membrane lipids and oxidize membrane proteins to initiate lipid peroxidation, resulting in neuronal apoptosis (27). MDA, a stable final product in the lipid peroxidation process, is commonly determined to indirectly measure ROS content (28). In addition, antioxidant enzymes, such as GSH, GSH-Px and SOD, are a group of ROS scavengers that detoxify ROS to protect against ROS-induced damages (29). Therefore, assessment of these enzyme activities is used to evaluate the antioxidant capacities. Herein, we found that I/R injury resulted in obvious increases in MDA contents and decreases in GSH levels and SOD and GSH-Px activities in the ischemic brains, and BAR administration significantly reversed these changes in brain tissues. Similarly, BAR has also been shown to be potent antioxidant in cardiomyocytes (13, 14). In addition to the antioxidant actions, its ability to restore antioxidant enzymes may also contribute to the therapeutic benefits of BAR. Collectively, our data indicate that BAR protects brains from I/R-induced damages by limiting oxidative stress.
Neuroinflammation is known as a major contributor that results in neuron damage and death following cerebral ischemia (30). NF-κB is a central regulator in inflammatory process (31). In response to ROS, NF-κB is activated in I/R injury to induce excessive release of proinflammatory mediators (e.g. iNOS, TNF-α and IL-6), thereby aggravating brain damages (32). NF-κB inactivation or inhibition of iNOS expression attenuates brain ischemic injury (32, 33). In this study, BAR suppressed I/R-induced iNOS expression in the ischemic cortex and inhibited LPS-stimulated production of proinflammatory cytokines (IL-6, TNF-α and iNOS) and nuclear NF-κB p65 in BV2 cells. Our results are consistent with published literatures reporting that BAR attenuated ulcerative colitis-induced and LPS-induced acute lung injury via suppressing the release of pro-inflammatory cytokines (10, 11). Altogether, it is plausible that BAR exerts the neuroprotective effects in cerebral I/R injury via its anti-inflammatory properties.
There are several limitations to acknowledge. Firstly, the underlying mechanisms by which BAR exerts the anti-oxidative, anti-apoptotic, and anti-inflammatory activities are not investigated. Secondly, the present study treated I/R rats after reperfusion, whether pretreatment of BAR before I/R injury shows similar beneficial effects has not been determined. Thirdly, the animal number in this study is relatively small.