Sesaminol prevents Parkinson's disease by activating the Nrf2-ARE signaling pathway

Parkinson’s disease is a neurodegenerative disease caused by the degeneration of substantia nigra neurons. PD is the second most frequent neurodegenerative disease after Alzheimer's disease. Only symptomatic treatment is used for PD, and fundamental treatments or preventive methods are needed.

Oxidative stress is a primary cause of substantia nigra degeneration. The substantia nigra has a high content of oxidizable substances, such as dopamine, and ROS is derived from dopamine, nerve melanin, highly unsaturated fatty acids and iron, and relatively low antioxidant substances. Therefore, oxidative stress easily controls the innate defense mechanism, and the resulting ROS sustains the oxidative stress state and induces the apoptosis of dopaminergic neurons.

Many studies revealed that 6-OHDA induced apoptosis via the application of oxidative stress to nerve cells, which caused mitochondrial damage from the byproducts that accompany autoxidation, such as H2O2. Therefore, ROS production was monitored during the culture time, and ROS production of SH-SY5Y cells increased 3–6 h after the addition of 6-OHDA. Measurement of intracellular ROS production revealed a significant increase in the intracellular ROS amount following the addition of 6-OHDA, and this increase was suppressed to the control level via 2 h of sesaminol pretreatment. The intracellular glutathione ratio (GSSG/GSH), which is an index of oxidative stress, was also measured. The addition of 6-OHDA increased the glutathione ratio, and the addition of sesaminol decreased the ratio. PI staining revealed an abnormal morphology, such as the aggregation of nuclei, which is a characteristic of apoptosis, after the addition of 6-OHDA alone, but not in the sesaminol and 6-OHDA group. These results show that sesaminol prevents apoptotic cell death via the suppression of oxidative stress associated with 6-OHDA-induced ROS production.

The Nrf2-ARE pathway is a potential defense system of cells, and it recent attracted much attention as a therapeutic target for neurodegenerative diseases because it is involved in the expression of numerous antioxidant enzyme systems. The present study suggested that sesaminol promoted the nuclear translocation of the transcription factor Nrf2 and enhanced NQO1 expression via activation of the Nrf2-ARE pathway to enhance the defense system against oxidative stress.

6-Hydroxydopamine (6-OHDA) is an oxidative analogue of dopamine that has a high affinity for catecholamine receptors due to its similar chemical structure to dopamine, and it is taken up into cells via the dopamine transporter (DAT) in neurons. 6-OHDA becomes quinone, which is a reactive oxygen species (ROS), via nonenzymatic autooxidation inside and outside cells, which produces hydrogen peroxide, superoxide radical and hydroxy radicals.

Increased ROS deactivates biopolymers, such as nucleic acids and proteins, in neurons, disrupts organelle functions, such as mitochondria, and induces apoptosis. Human neuroblastoma SH-SY5Y cells are most commonly used for in vitro PD studies. SH-SY5Y cells have nerve cell-like processes and express genes that are characteristic of dopaminergic neurons, such as DAT and vesicular monoamine transporter. To evaluate the protective effect of sesaminol against PD, SH-SY5Y cells were treated with 6-OHDA as an in vitro model of PD.

Rotenone is an active ingredient of the insecticide obtained from the roots of the legume Derris. Rotenone is lipophilic, readily crosses biological membranes, and specifically inhibits complex I in the mitochondrial electron transport chain]. Oral administration of rotenone causes motor and gastrointestinal dysfunction. Because of these properties, rotenone is used to establish PD-like animal models.

Sesaminol (3,4-methylenedioxy phenol)is a sesame lignan found in sesame (Sesamum indicum L.) seeds, and it has a strong antioxidant effect. An anti-cancer effect of sesaminol was also reported. However, most sesaminol exists as a glycoside, and sesaminol remains in the form of glycoside in the defatted residue after the extraction of sesame oil. A recent method for purifying sesaminol from sesaminol glycoside was established, and the effective use of defatted sesame, which was treated as industrial waste, is expected.

Therefore, the present study investigated the preventive effect on PD as a new physiological action of sesaminol produced from sesaminol glycoside using in vitro and in vivo PD models. We also examined the mechanism of sesaminol protection in PD.

We created PD model mice using rotenone and examined the preventive effect of sesaminol in PD. PD patients exhibit motor impairment as a symptom, and mice treated with rotenone exhibit motor impairment. Therefore, a rotor rod test was performed to evaluate motor function. Rotenone significantly reduced rod stay time, and feeding a small amount of sesaminol (0.0008%) significantly restored stay times to the control level.

PD patients complain of constipation 10–20 years before the onset of movement disorder, and the transit time of feces in the intestine is more than twice as long as normal subjects. It is important to measure gastrointestinal motility because constipation adversely affects life and increases the risk of developing PD by a factor of 2.7–4.5. Therefore, Evans blue was orally administered, and the gastrointestinal motility function was evaluated via measuring the migration distance of Evans blue from the pylorus. The intestinal motility function of the rotenone group decreased significantly compared to the control group, but sesaminol restored the intestinal motility.

The first Lewy bodies in PD likely appear in the substantia nigra because neuronal loss in the substantia nigra is observed. However, The various nonmotor symptoms appear before the onset of motor symptoms of PD, and the enteric nervous system (ENS) related to gastrointestinal symptoms in the onset of PD was recently investigated. The dual hit hypothesis proposed by Hawkes et al. suggests that the path starts from the enteric nerve sac of the stomach to the dorsal motor nucleus of the vagus and spreads from the olfactory bulb to the substantia nigra and limbic system routes in the early stages of Lewy body pathology. Therefore, the morphology of the mucosa in the colon was observed using HE staining. Compared to the control group, a shortening of the intestinal mucosal layer and damage to the mucosal surface were observed in the rotenone group, but intestinal mucosal abnormalities were hardly observed in the sesaminol group. These results suggest that sesaminol prevents the progression of PD pathology from the intestine.

The results of the in vivo experimental system of the present study suggest that sesaminol prevents the development of PD pathology from the intestine and reduces α-synuclein expression in the substantia nigra, which suppresses motor dysfunction and the decline of intestinal motor function.

Whether sesaminol is able to cross the blood-brain barrier (BBB) is an important problem. Therefore, we evaluated sesaminol by the following factors that determine the penetrability on the BBB. (1) High lipid solubility (water versus oil partition coefficient) [LogP] < 3, (2) number of hydrogen bonds <8, (3) a neutral charge or low degree of ionization (polar surface area) < 90 Å, (4) a less bulkey (number of rotable bonds) < 5 and (5) smaller molecular size <450 Da. The values of sesaminol for (1), (2), (3), (4) and (5) are 2.14, 7, 75.61 Å, 2 and 370.4, respectively. These values suggest that sesaminol may be able to cross the BBB.

The present study also revealed that sesaminol had a neuroprotective effect in an in vitro experimental system and a PD preventive effect in an in vivo experimental system. Notably, the protective effect was observed with the feeding of a small amount of sesaminol. These results show that sesaminol is very suitable for use as a preventive treatment of PD. Further detailed elucidation of the mechanism of action will be necessary for practical application.

This work was supported by the Japan Society for the Promotion of Science (JSPAS) KAKENHI, Grant Number, JP15K00832 .

The above was an abstract of the report published on Heliyon Cell Press journal from Haruka Kaji, Isao Matsui-Yuasa, Akiko Kojima-Yuasa, Kayo Matsumoto, Ayano Omura, and Kunio Kiyomoto all contributed by performing the experiments, analyzed and interpreted data, contributing reagents, materials, tools and data, as well as writing the paper.