Gamma Frequency Inhibits the Secretion and Aggregation of Amyloid-β and Decreases the Phosphorylation of mTOR and Tau Proteins in vitro

Abstract

Background:

Alzheimer’s disease (AD) was the main cause of dementia in an aging society; unfortunately, there is no effective treatment for AD now. Meditation has been reported to thicken the cerebral cortex, and gamma wave at a frequency of 40 hertz (Hz) was recorded during the meditation process from the brain. Previous study showed that non-invasive scintillation gamma frequency oscillation increased the space in recognition and memory of auditory cortex hippocampal gyrus in AD mice model. However, the AD-related molecular change by exposure of 40 Hz gamma frequency in brain cells was still unclear.

Objective:

We investigated the AD-related molecular change by exposure of 40 Hz gamma frequency in SH-SY5Y cells.

Methods:

We designed the light and sound generators at 40 Hz gamma frequency for this study. SH-SY5Y cells were exposed to sound or light of 40 Hz gamma frequency, respectively. The concentrations of amyloid-β₄₀ (Aβ₄₀) and amyloid-β₄₂ (Aβ₄₂) were quantified by enzyme-linked immunosorbent assay. The protein levels were examined by Western blotting. The aggregation of Aβ₄₂ was examined by thioflavin T assay.

Results:

Our results showed that the secretion of Aβ, phosphorylation of AKT, mTOR, and tau, and aggregation of Aβ₄₂ were significantly inhibited by 40 Hz gamma frequency in SH-SY5Y cells. The phosphorylation of 4E-BP1, downstream of mTOR, was induced by 40 Hz gamma frequency in SH-SY5Y cells.

Conclusion:

Our study showed 40 Hz gamma frequency involved in the inhibition of secretion and aggregation of Aβ and inhibition of p-Tau protein expression through the mTOR/4E-BP1/Tau signaling pathway.

Keywords

Alzheimer disease amyloid-β gamma frequency mTOR tau

INTRODUCTION

Alzheimer’s disease (AD) is the main cause of dementia in an aging society. The 40–60% of AD patients can improve symptom from acetyl-cholinesterase inhibitor or N-methyl-D-aspartic acid antagonist. Currently, there are no effective drugs for the treatment of AD. Therefore, early diagnosis and treatment may improve clinical outcomes of AD patients [1, 2]. The accumulation of plaques in brain were derived from the amyloid-β protein precursor (AβPP) cleaved and aggregated to amyloid-β (Aβ) plaques, and the high level of tau phosphorylation to aggregate into neurofibrillary tangles [3, 4]. Recent studies showed overexpression of abnormal Aβ induced the expression of inflammatory factors to inhibit the clearance of Aβ in neuron cells by autophagy [5]. Hyperphosphorylation of tau protein induced destruction and degradation in microtubule and then phosphorylated tau was aggregated into neurofibrillary tangles. This progression of plaque accumulation in brain mediated gradually synaptic dysfunction and loss of neuronal function or cell death [6 –8]. Current studies indicated abnormal neural activity in molecular signaling [6 –8] and the state of dysbiosis [9] aggravated the pathological features of AD. Therefore, induction of brain neural activity can be adjusted to delay the progression of AD.

Recently, meditation was conceptualized as a series of complex emotional and attentional regulatory training regimes for various ends [10]. Previous study showed the participants with more meditation experience were recorded gamma waves at a frequency of 40 Hz in meditation process by electroencephalography (EEG) [11]. Kang et al. (2013) showed that long-term meditation thickened the cerebral cortex, especially the frontal and temporal lobes by magnetic resonance imaging (MRI) [6] and increased blood flow in prefrontal and posterior cingulate cortex, and retard cognitive decline in patients with mild dementia [12 –14]. Meanwhile, Basar et al. (2016) showed the AD patients have a delayed response to EEG gamma frequency and decreased blood flow in posterior cingulate cortex [15]. Based on these studies, meditation may potentially stimulate brain gamma frequency and increase posterior cingulate cortical blood. Iaccarino et al. (2016) and Martorell et al. (2019) reported that the short-term memory and spatial cognition performance was re-activated by non-invasive scintillation oscillation at gamma frequency (gamma entrainment using sensory stimulus) in AD mouse models [16, 17]. They found the expression of Aβ₄₀ and Aβ₄₂ were inhibited, and the expression of microglia morphology transformation-related genes and microglia to remove Aβ ability were induced by exposure of oscillation at gamma frequency in AD mouse model. In particular, oscillation at gamma frequency in auditory reduced phosphorylation of tau in hippocampus of AD mouse model [16, 17]. These studies showed that gamma frequency can be achieved using different sensory stimuli and has a wide range of effects in multiple brain regions to improve cognitive function and neural activity.

However, the molecular mechanism of 40 Hz gamma frequency inhibited the expression of Aβ₄₀ and Aβ₄₂ in neural cells was still unclear. Therefore, the purpose of this study was to investigate the influence of 40 Hz gamma frequency on the AD-related signaling pathway and the molecular mechanism in cell model.

MATERIALS AND METHODS

40 Hz generator

Similarly to the 40 Hz auditory tones used by Martorell et al. (2019) [17], the fundamental frequency of the periodic audio signal for this study was 40 Hz. For each of the 25 ms periods of the signal, with a duty cycle of 4%, the first 1 ms was a 10 kHz sinusoidal signal and the remaining 24 ms was a silent signal. The auditory signal was generated by a soundbar speaker (Mi soundbar, Xiaomi, Beijing, China) with a bandwidth of 20,000 Hz. The sound pressure level received by the cell was about 70 dB. A 12.5 ms light-on and 12.5 ms light-off period was used and the 40 Hz visual flicker was generated using a 12 voltage (V) RGB light-emitting diode (LED) strip (WS2812 5050) that was controlled by an Arduino microcontroller. Measured by a lux-meter, the intensity of the light signal received by the cells is about 885–928 lumens. The bandwidth of the LED is well over 1 megahertz.

Chemicals and reagents

Aβ₄₂ peptide was purchased from Apex Biotechnolog, TX, USA, and the purity was 98.83%. Thioflavin T (ThT) was purchased from Sigma-Aldrich, MA, USA. Aβ₄₀ and Aβ₄₂ ELISA kits were purchased from Invitrogen, MA, USA. The following primary antibodies were used for Western blot analysis: anti-phospho-Tau (Thr 181) (Invitrogen, MA, USA), anti-Tau (Total) (Dako, Glostrup, Denmark), anti-phospho-AKT (Ser 473), anti-phospho-AKT (Thr 308), anti-AKT (Santa Cruz Biotechnology, CA, USA), anti-phospho-mTOR (Ser 2448), anti-mTOR, anti-phospho-eukaryotic translation initiation factor 4E-binding protein 1 (p-4E-BP1) (T37/46), anti-eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) (Cell Signaling Technology, MA, USA), and anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (Gene Tex Biotechnology, CA, USA).

Cell culture

Human neuroblastoma cell line SH-SY5Y was purchased from ATCC. Cells were cultured in Dulbecco’s Modified Eagle Medium:F12 (DMEM:F12) supplemented with 10% fetal bovine serum (FBS), 1% penicillin/streptomycin, 1% L-glutamine, and 1% non-essential amino acids in humidified 5% CO₂/95% humidified air incubator at 37°C.

Western blotting analysis

Cells (1×10⁵) were seeded in 6-cm dish for 24 h and were exposed to sound or light of 40 Hz for 0, 15, 30, 45, and 60 min respectively. Then cells were scraped from 6 cm dish and resuspended by 100 μL of cell lysis buffer (0.5 % sodium deoxycholate, 1 % NP-40, 150 mM NaCl, 10 mM EDTA, 50 mM Tris-HCl (pH 7.5), 1 mM sodium ovanadate, 0.1 % sodium dodecyl sulfate (SDS), 10 μg/mL aprotinin, 1 mM phenylmethanesulfonyl fluoride, and 10 μg/mL leupeptin). The lysate was centrifuged (13,000×g at 4°C for 30 min) to collect supernatant and was used a protein assay dye for quantitation of protein concentration (Bio-Red, CA, US). Protein extracts (20 μg/mL) were separated by 10% SDS-polyacrylamide gels and transferred to polyvinylidene difluoride membranes. The membranes were blocked with 5% non-fat milk for 1 h then incubated with specific primary antibodies at 4°C overnight. The membranes were washed for five times with tris-buffered saline with 0.1% Tween (TBST) and were incubated with horseradish peroxidase-conjugated goat anti-rabbit, anti-mouse or anti-goat secondary antibodies for 1 h at room temperature. The proteins were detected using Western blot luminol reagent (Advansta, CA, US).

ELISA assay for Aβ₄₀ and Aβ₄₂

Cells (2×10⁴) were seeded in 6-cm dish for 24 h and were exposed to sound or light of 40 Hz for 0, 15, 30, 45, and 60 min respectively, and incubated for 24 h. The cell supernatants were harvested for further assay. The Aβ₄₀ and Aβ₄₂ levels in culture medium were measured, respectively using Aβ₄₀ and Aβ₄₂ ELISA kit (Invitrogen, CA, US), according to the manufacturer’s instructions. The standard curve sensitivity range of Aβ₄₀ was 0–500 pg/ml and Aβ₄₂ was 0–1,000 pg/ml.

Aggregation inhibitory assay (ThT assay)

Aβ₄₂ lyophilized peptide was dissolved in 10% dimethyl sulfoxide (dimethyl sulfoxide diluted in 10 mM Tris-HCl buffer having pH 7.4), underwent vortexing for 1 min and sonicated for 5 min to Aβ₄₂ peptide stock solution (62.5 μM). The Aβ₄₂ peptide stock solution was stored at –80°C. The 20 μL Aβ₄₂ peptide (25 μM) from the stock solution was incubated for 7 days to grow the pre-formed Aβ₄₂ fibrils at the room temperature, and pre-formed Aβ₄₂ fibrils was exposed of 40 Hz gamma frequency for 0, 15, 30, 45, and 60 min. We were performed using a ThT assay method was described elsewhere [18, 19]. The 5 μM ThT stock solution (ThT diluted in 10 mM Tris-HCl buffer, pH 7.4) was stored in an aluminum foil and the vial was wrapped to prevent photo-oxidation. A black 96-well plate was used to incubate the ThT solution stock with an equal volume (20 μL) of pre-formed Aβ₄₂ fibril, and the volume was made up to 200 μL with 1 mM Tris-HCl buffer at room temperature for 1 h. The absorbance was measured at an excitation of 450 nm and an emission of 480 nm.

Statistical analysis

The all figures were showed from one representative experiment of three independent experiments and were presented as the mean±standard deviation (SD). Significant differences between groups were analyzed by student’s t test. All tests were two-sided with p < 0.05 being statistically significant. Statistical analysis was performed by SigmaPlotsoftware.

RESULTS

The secretion of Aβ₄₀ and Aβ₄₂ were inhibited by 40 Hz gamma frequency in SH-SY5Y cells

Light and sound generators at 40 Hz gamma frequency were used for this study. SH-SY5Y cells were exposed to sound or light of 40 Hz gamma frequency for 15, 30, 45, and 60 min respectively. Our results showed the secretion concentration of Aβ₄₀ was 35.52 ± 0.54 pg/ml by exposure of sound of 40 Hz gamma frequency for 15 min, and the control was 42.19 ± 2.04 pg/ml. We indicated the secretion of Aβ₄₀ was significantly inhibited by exposure to sound of 40 Hz gamma frequency for 15 minutes in SH-SY5Y cells (Fig. 1A). However, the secretion of Aβ₄₂ was not significantly inhibited in SH-SY5Y cells under the same experimental condition (Fig. 1B).

Fig. 1

The secretion of Aβ₄₀ in SH-SY5Y cells was inhibited by sound of 40 Hz gamma frequency. SH-SY5Y cells were exposed to sound of 40 Hz gamma frequency for 15, 30, 45, and 60 min respectively, and incubated for 24 h. The secretion of (A) Aβ₄₀ and (B) Aβ₄₂ were detected by ELISA. The data was showed from representative of three independent experiments and was expressed as the mean±SD, **p < 0.01 (t test).

Similarly, the secretion concentration of Aβ₄₂ was 18.56±0.38 pg/ml by exposure of light of 40 Hz gamma frequency for 60 min, and the control was 13±0.58 pg/ml. We found the secretion of Aβ₄₂ was significantly inhibited by exposure to light of 40 Hz gamma frequency for 60 min in SH-SY5Y cells (Fig. 2D). The secretion of Aβ₄₀, however, was not significantly inhibited in SH-SY5Y cells under the same experimental condition (Fig. 2 C). However, our results indicated the secretion of Aβ₄₀ and Aβ₄₂ was not inhibited by the exposure to light of 30 Hz or 50 Hz gamma frequency, respectively in SH-SY5Y cells (Fig. 2A, B, E, and F).

Fig. 2

The secretion of Aβ₄₂ in SH-SY5Y cells was inhibited by light of 40 Hz gamma frequency. SH-SY5Y cells were exposed to light of (A, B) 30, (C, D) 40, or (E, F) 50 Hz gamma frequency for 15, 30, 45, and 60 min respectively, and incubated for 24 h. The secretion of (A, C, E) Aβ₄₀ and (B, D, F) Aβ₄₂ were detected by ELISA. The data was showed from representative of three independent experiments and was expressed as the mean±SD, ***p < 0.001 (t test).

The phosphorylation of tau protein was inhibited by 40 Hz gamma frequency

Thijssen et al. (2020) demonstrated the higher level of p-Tau (T181) was detected in plasma of AD and other neurodegenerative disease patients. The elevated level of plasma p-Tau (T181) protein may be a potential biomarker for the detection of AD [20]. Our study was the first research study, to our knowledge, to show that the expression of p-Tau (T181) protein was significantly inhibited by light or sound of 40 Hz gamma frequency, respectively in SH-SY5Y cells (Fig. 3).

Fig. 3

The phosphorylation of tau was inhibited by 40 Hz gamma frequency. SH-SY5Y cells were exposed by 40 Hz gamma frequency for 15, 30, 45, and 60 min respectively. A, B) The expression of total tau, p-Tau (T181) and GAPDH proteins were determined by Western blotting. Similar results were obtained from three independent experiments. C-F) The expression of total tau and p-Tau (T181) proteins were quantified by densitometry using Image J software and were normalized with the corresponding GAPDH. These data was expressed as the mean±SD, *p < 0.05, **p < 0.01, ***p < 0.001 (t test).

The phosphorylation of mTOR (S2448) protein was inhibited, and the phosphorylation of 4E-BP1 (T37/46) protein was induced by 40 Hz gamma frequency

SH-SY5Y cells were exposed to sound or light of 40 Hz gamma frequency for 15, 30, 45, and 60 min respectively. Interestingly, our results showed the expression of p-mTOR (S2448) protein was significantly inhibited by exposure to light or sound of 40 Hz gamma frequency, respectively in SH-SY5Y cells (Fig. 4A, B, G, and H).

We found light of 40 Hz gamma frequency significantly inhibited the expression of p-AKT (S473) protein for 15 and 30 min in SH-SY5Y cells (Fig. 4A, C). Similarly, sound of 40 Hz gamma frequency significantly inhibited the expression of p-AKT (S473) protein for 30 min in SH-SY5Y cells (Fig. 4B, D). However, both light and sound of 40 Hz gamma frequency did not inhibit the expression of p-AKT (T308) and p-ERK (T202/Y204) in our study (data not shown).

Fig. 4

The phosphorylation of AKT and mTOR were inhibited and phosphorylation of 4E-BP1 was induced by 40 Hz gamma frequency. SH-SY5Y cells were exposed by 40 Hz gamma frequency for 15, 30, 45 and 60 minutes respectively. A, B) The expression of p-AKT (S473), AKT, p-mTOR (S2448), mTOR, p-4E-BP1 (T37/46), 4E-BP1 and GAPDH proteins were determined by Western blotting. Similar results were obtained from three independent experiments. C-N) The expression of p-AKT (S473), AKT, p-mTOR (S2448), mTOR, p-4E-BP1 (T37/46), and 4E-BP1 proteins were quantified by densitometry using Image J software, and normalized with the corresponding GAPDH. These data were expressed as the mean±SD, *p < 0.05, **p < 0.01, ***p < 0.001 (t test).

It was a novel finding that both light and sound of 40 Hz gamma frequency significantly induced the expression of p-4E-BP1 (T37/46) protein in SH-SY5Y cells (Fig. 4A, B, K, and L). However, there were no significant difference in the expression of AKT, mTOR, and 4E-BP1 total proteins by exposure of light and sound of 40 Hz gamma frequency in SH-SY5Y cells (Fig. 4A, B, E, F, I, J, M, and N).

Aβ₄₂ fibril aggregation was inhibited by 40 Hz gamma frequency in vitro

ThT was a common probe to monitor the formation of amyloid fibril. ThT displayed a strong fluorescence signal at approximately 480 nm when excited at 450 nm if there was binding to amyloid fibrils [21, 22]. Obviously, we found the Aβ₄₂ fibril aggregation was significantly inhibited by exposure to light and sound of 40 Hz gamma frequency in vitro assay (Fig. 5).

Fig. 5

The aggregation of Aβ₄₂ in vitro was inhibited by 40 Hz gamma frequency. Aβ₄₂ fibril was exposed by (A) light and (B) sound of 40 Hz gamma frequency for 0, 15, 30, 45, and 60 min respectively. The aggregation rate for Aβ₄₂ was determined by ThT assay. The data was showed from representative of three independent experiments and was expressed as the mean±SD, *p < 0.05, **p < 0.01, ***p < 0.001 (t test).

DISCUSSION

Our study has demonstrated the changes of mTOR, tau, and Aβ after the intervention of light with 40 Hz gamma frequency. Previous studies demonstrated the hyperactive mTOR increased the brain levels of total and phosphorylated tau in animal model. The suppression of mTOR signaling has a beneficial effect on tau pathology [23, 24]. Expressive level of mTOR may be a potential predictor for the development of tau pathology through the induction of tau protein translation initiated by the suppression of 4E-BP1 in neuronal cells [25 –27]. Recent studies demonstrated rapamycin or temsirolimus significantly inhibited mTOR activity and these can decrease Aβ deposition, ameliorate AD-related pathology and cognitive deficits in AD mouse model [28, 29]. The activation and expression of AKT [30, 31] and mTOR [23, 24] were involved in progression of AD, and these molecules were also associated with molecular pathway of tau and Aβ expression. In this study, we found light and sound of 40 Hz gamma frequency significantly inhibited the phosphorylation of mTOR (S2448) and tau (T181) proteins in SH-SY5Y cells. In particular, our results showed the secretion of Aβ₄₂ at 60 min and the expression of p-AKT (S473) at 15 and 30 min were inhibited by light of 40 Hz gamma frequency. Similarly, the secretion of Aβ₄₀ only at 15 min and the expression of p-AKT (S473) at 15 min were inhibited by sound of 40 Hz gamma frequency. Although the differences of Aβ₄₀ and Aβ₄₂ levels between exposure and non-exposure groups were small, but they did reach the significant difference by statistical analysis, and the data was from three independent experiments. Based on this evidence, we demonstrated that Aβ₄₀ and Aβ₄₂ were significantly inhibited by 40 Hz gamma frequency in SH-SY5Y cells. The exact mechanisms of how the exposure of 40 Hz gamma frequency will modulate the cell function, especially in the adjusting the dementia-related proteins were still to be determined. We supposed the energy accumulation of 40 Hz gamma frequency at specific timing induced the various signaling pathway for the cellular responses. Based on results of this study, we found similar features that 40 Hz gamma frequency obviously induced the expression of AD-related molecular events at specific timing. However, the molecular mechanism of 40 Hz gamma frequency induced the specific response in cells remained unknown. In sum, inhibition of AKT/mTOR/Tau signaling pathway by 40 Hz gamma frequency may attenuate the AD progression. Therefore, it may be useful to apply on clinical treatment of AD patients due to possibly modify the formation of amyloid and tau protein, which has been conducted with promising findings [32].

Argentati et al. (2019) showed the cell proliferation, growth, migration, and differentiation were regulated by cell microenvironment [33]. The senses of physical cues and mechanical forces from the cell microenvironment were translated to biochemical signals of ion channels, receptors and cytoskeletons in cells [34 –36]. Tan et al. (2016) showed the novel heat-sensitive transient receptor potential channels (TRPM2) was a thermal sensor in somatosensory neurons [37]. Recent research also showed the focused ultrasound of 1 MHz frequency induced signaling pathway of ion channels in cells. Data accumulated suggested the exposure of cells to different physical effects induced various signaling pathways for cellular response [36, 38]. Previous study showed that the exposure of low-power laser irradiation were attenuated Aβ-induced cell apoptosis by AKT/GSK3β/β-catenin pathway in SH-SY5Y cells [39]. These studies demonstrated that the cellular responses in cells can induce by senses of physical cues and mechanical forces in microenvironment. In our study, the 40 Hz gamma frequency was a physical oscillation energy, which was usually found in inter-neuron for learning function [40] and will be applied through light with 885-928 lumens and sound around 70 dB to SH-SY5Y cells. To our knowledge, our study was innovative to report that the protein secretion and molecular signaling in SH-SY5Y cells were induced by 40 Hz gamma frequency. Based on our findings, we supposed the diverse cellular responses were induced by the light and sound of 40 Hz gamma frequency at specific energy level and timing. As expected, we found the secretion of Aβ and expression of p-AKT (S473) were significantly inhibited by 40 Hz gamma frequency at specific energy level and timing.

Recent studies showed the neurotoxicity of Aβ was correlated with the level of Aβ aggregation [41]. The well-known technique to identify the presence of Aβ₄₂ aggregation used ThT assay. ThT interacted with the four consecutive β-strand and binding in Aβ₄₂ fibril surface. ThT only recognized the bind to Aβ₄₂ fibril but not Aβ₄₂ single structure [21, 42]. In a similar study, Omar et al. (2018) found olive biophenols potential inhibited the Aβ₄₂ aggregation by ThT assay [18]. Interestingly, we also found the Aβ₄₂ aggregation was inhibited by 40 Hz gamma frequency in vitro.

Our results of exposure to light and sound 40 Hz gamma frequency novelty demonstrated that the secretion of Aβ and the phosphorylation of AKT (S473), mTOR (S2448), and Tau (T181) proteins were inhibited, the phosphorylation of 4E-BP1 (T37/46) protein was induced in SH-SY5Y cells, and the aggregation of Aβ₄₂ was inhibited in vitro (Fig. 6).

Fig. 6

The hypothesis of this study showed the signaling pathway of AKT/mTOR/4E-BP1/Tau was activated in SH-SY5Y cells by 40 Hz gamma frequency. Dashed lines represent predicated pathway.

Based on our study, we suggested the repetitive transcranial magnetic stimulation machine can be modified to a 40 Hz gamma frequency stimulation machine for clinical treatment of AD patients. Our study may provide the novelty and feasibility of AD symptoms improved by 40 Hz gamma frequency clinically, and the possible efforts from other frequency, if any, could be examined in coming studies.

CONCLUSION

Our hypothesis (as shown in Fig. 6) first demonstrated the secretion of Aβ and the phosphorylation of mTOR (S2448), AKT (S473), and tau (T181) were inhibited, the phosphorylation of 4E-BP1 (T37/46) was induced in SH-SY5Y cells, and the aggregation of Aβ₄₂ was inhibited by 40 Hz gamma frequency in vitro.

Footnotes

ACKNOWLEDGMENTS

This study was supported by grants from Kaohsiung Medical University Research Center Grant (KMU-TC111B02), Kaohsiung Municipal Ta-Tung Hospital, Kaohsiung, Taiwan (KMTTH-108-R010), Kaohsiung Medical University Research Foundation, Taiwan (KMU-096011, KMU-M098006 and NSYSUKMU108-I003), the Ministry of Science and Technology, Executive Yuan, Taiwan (MOST 105-2314-B-037-041), National Health Research Institutes (NHRI-11A1-CG-CO-06-2225-1).

Authors’ disclosures available online ().

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Gamma Frequency Inhibits the Secretion and Aggregation of Amyloid-β and Decreases the Phosphorylation of mTOR and Tau Proteins in vitro

Abstract

Background:

Objective:

Methods:

Results:

Conclusion:

Keywords

INTRODUCTION

MATERIALS AND METHODS

40 Hz generator

Chemicals and reagents

Cell culture

Western blotting analysis

ELISA assay for Aβ40 and Aβ42

Aggregation inhibitory assay (ThT assay)

Statistical analysis

RESULTS

The secretion of Aβ40 and Aβ42 were inhibited by 40 Hz gamma frequency in SH-SY5Y cells

The phosphorylation of tau protein was inhibited by 40 Hz gamma frequency

The phosphorylation of mTOR (S2448) protein was inhibited, and the phosphorylation of 4E-BP1 (T37/46) protein was induced by 40 Hz gamma frequency

Aβ42 fibril aggregation was inhibited by 40 Hz gamma frequency in vitro

DISCUSSION

CONCLUSION

Footnotes

ACKNOWLEDGMENTS

References

ELISA assay for Aβ₄₀ and Aβ₄₂

The secretion of Aβ₄₀ and Aβ₄₂ were inhibited by 40 Hz gamma frequency in SH-SY5Y cells

Aβ₄₂ fibril aggregation was inhibited by 40 Hz gamma frequency in vitro