A Novel Supplement Attenuates Oxidative Stress-Induced TDP-43-Related Pathogenesis in TDP-43-Expressed Cells.

Eun Jin Yang,Rajeev K Singla

doi:10.1155/2021/6773260

Abstract

Amyotrophic lateral sclerosis (ALS) is caused by selective the loss of spinal motor neurons by multifactorial pathological mechanisms and results in muscle atrophy. Incidence rates of ALS are increasing over time, but there are no effective treatments at present due to limitations on approved therapies (riluzole and edaravone). Therefore, this study investigated whether combined treatment with Bojungikgi-tang and riluzole could act synergistically in transactive response DNA-binding protein 43 (TDP-43) stress granule cells. To examine the effect of combined treatment on oxidative stress-induced cell death, the CCK8 assay was performed for the detection of cell viability. The expression of oxidative stress-induced proteins was determined by Western blot. Quantification of sodium arsenite-induced reactive oxygen species (ROS) was measured in TDP-43 stress granular cells using 2,7-diacetyl dichlorofluorescein diacetate. To investigate the effect of combined treatment on TDP-43 aggregation, immunofluorescence and immunoblotting were performed in TDP-43 stress granular cells. This combined treatment alleviated oxidative stress-induced cell death by increasing the expression levels of antioxidation proteins, such as heme oxygenase-1 and B cell lymphoma-2-associated X protein. Furthermore, it reduced oxidative stress-induced TDP-43 aggregates and lowered the levels of autophagy-related proteins, including p62, light chain 3b, and ATG8, in TDP-43-expressing cells. Our results suggest that this combined treatment could be helpful for autophagy regulation in other neurodegenerative diseases.

Highlights

Amyotrophic lateral sclerosis (ALS) is caused by selective loss of spinal motor neurons by multifactorial pathological mechanisms and results in muscle atrophy [1]
We have investigated the effects of combined treatment with BJIGT and riluzole in transactive response DNA-binding protein 43 (TDP-43) stress granular cells to examine the synergistic effect of combined treatment. e combined treatment reduced oxidative stress-induced cell death and regulated autophagy dysfunction compared with sodium arsenite (SA) only-treated TDP-43 cells
We evaluated the supernatants with the bicinchoninic acid (BCA) protein assay to determine the protein concentration of the lysates. e radioimmunoprecipitation assay (RIPA)-insoluble pellets were lysed with 10% SDS buffer, sonicated, and supplemented with SDSpolyacrylamide gel electrophoresis (PAGE). e soluble and insoluble proteins were separated on 4–12% SD-PAGE gels and transferred to PVDF membranes

Summary

Introduction

Amyotrophic lateral sclerosis (ALS) is caused by selective loss of spinal motor neurons by multifactorial pathological mechanisms and results in muscle atrophy [1]. E pathological mechanisms of ALS include excitotoxicity, apoptosis, inflammation, and oxidative stress at the cellular level [2]. TDP-43 is involved in frontotemporal lobar degeneration (FTLD), and ALS and its pathological mechanisms include oxidative stress [3], neuroinflammation [4], and mitochondrial dysfunction [5]. TDP-43 has been associated with the RNA-related metabolism, stress granule formation, the ubiquitin-proteasome system (UPS), autophagy, and mitochondrial dysfunction [7]. Xu et al and Wegorzewska et al have shown that mutant TDP43 protein is toxic to neurons in several TDP-43 animal models [11, 12]. Vander Broeck et al suggested that the loss of TDP-43 induces TDP-43-related proteinopathies rather than toxic effects by aggregates [13]. Riluzole does not improve neuropathology in TDP-43 expressed animal models [14, 15]

Methods

Results

Conclusion