# Cytoprotective-Antioxidant Effect of _Brunfelsia grandiflora_ Extract on Neuron-like Cells
## 4 Discussion
grandiflora_ extract concentrations used in the present study should be considered realistic and comparable to those of previous studies focused on neuroprotection with natural compounds. Indeed, when SH-SYSY cells were exposed to _B. grandiflora_ extracts at the selected range of doses, there was a dose-dependent reduction in cytotoxicity induced by t-BOOH, indicating that the stressed cells' integrity was significantly protected against the oxidative insult. Although the chemoprotective effect of the _B. grandiflora_ extract seemed to be slightly more prominent in the pretreatment condition, an important quantity of the bioactive phenolic compounds in the extract [9] had enough capacity for fractional but substantial dose-dependent cell protection, comparable to that previously reported for other phenolic extracts in similar cell culture conditions [5,6]. Recently, the higher adequacy of differentiated neuroblastoma as a neuron-like cell culture model has been reported [37, 38, 39], and, indeed, we have previously used SH-SYSY differentiated with retinoic acid submitted to stress conditions similar to those utilized in the present study [5]. Interestingly, in from that study that were obtained simultaneously from undifferentiated SH-SYSY cells, very similar results regarding ROS concentration (determined via two methods, flow cytometry and in situ fluorescence with a microplate reader) were observed. Furthermore, the present study was carried out in parallel to a similar approach that was used for testing the cytoprotective effect of an extract from another Peruvian medicinal plant, _Desmodium tortuosum_, where the same cell line, undifferentiated SH-SYSY, was used, in order to facilitate stimulating comparisons [6]. As stated above, a substantial overproduction of ROS that might be responsible for the increased cell death was observed when t-BOOH was added to SH-SYSY cells in both treatment conditions. Both co- and pretreatment of stressed cells with the _B. grandiflora_ extracts evoked a remarkable dose-dependent inhibition of ROS, which plainly confirms the antioxidant capacity of the phenolic compounds in the extract and offers a first reason for the reported cell protection. This chemoprotective capacity of _B. grandiflora_ does not seem specific to this particular neuron-like cell type since we have observed a comparable ROS-quenching capacity of this plan in cultured endothelial cells [31]. In addition, we have recently reported a similar protective effect of extracts from _D. tortuosum_ in both endothelial and neuron-like cells [6], as well as of phenolic extracts from other medicinal plants in hepatic cells [13; 14]. All these results clearly indicate that the chemoprotective effect is the result of the antioxidant capacity of natural compounds. In addition to their ROS-quenching capacity, plant extracts rich in bioactive phenolics may balance cell redox status by regulating antioxidant defenses such as GSH, the major reducing power in eukaryotic cells. GSH concentration is strongly regulated within the cell and its diminution denotes an increase in intracellular oxidation and hazardous oxidative status, whereas a stable GSH level permits the cell to handle in good condition an impending oxidative challenge [36]. As recently reported in our model of neuron-like cells, the addition of t-BOOH to any treatment condition resulted in a substantial decrease in GSH concentration, which is indicative of oxidative stress that might provoke severe damage to cellular macromolecules [6]. SH-SYSY cells pretreated with _B. grandiflora_ extracts prior to the stress prevented this dangerous situation in a dose-dependent pattern, whereas only 200 ug/mL of _B. grandiflora_ caused a significant recovery of GSH concentration in cotreated stressed cells. As in the case of cell viability protection, the condition of pretreatment with extracts prior to the stress seems to be more effective than that of cotreatment with both prooxidants and extracts.