Protective effects of tiger milk mushroom (lignosus rhinocerus) extracts on oxidative stress-induced neurotoxicity and aging in HT22 cells and C. elegans

Abstract

Lignosus rhinocerus (LR) or Tiger Milk Mushroom, a fork medicinal mushroom, has been reported for several pharmacological effects including asthma treatment, anti-inflammatory, anti-proliferative, immuno-modulating effects, promote neurite outgrowth in PC-12 cells, anti-HIV-1 activity, and antioxidants properties. However, the antioxidant properties have only focus on in vitro and no or few studies have reported their protective effects in mouse hippocampal (HT22) cells and Caenorhabditis elegans (C. elegans). This study aims to investigate the neuroprotective effect of three extracts of LR against oxidative stress in both HT22 cells and. C. elegans as well as longevity in C. elegans. In HT22 cells, we assessed the toxicity of three LR extracts (LRE, LRC, and LRH) and their protective activity by MTT assay, Annexin V-FITC/propidium iodide staining, Mitochondrial Membrane Potential (MMP), and assessment of intracellular ROS accumulation. In addition, we determined the antioxidant gene expression by qRT-PCR. In C. elegans, wild-type N2 were determined survival rate under oxidative stress and intracellular ROS. Transgenic strains including TJ356, TJ375, CF1553, CL2166, and LD1 were used to detect DAF-16, HSP-16.2, SOD-3, GST-4, and SKN-1, respectively. Lifespan and aging biomarkers including lipofuscin and pharyngeal pumping rated were also assessed. Furthermore, the neuroprotective effects, such as chemotaxis behavior and PolyQ40 formation were assessed as well. We found that only LRE significantly reduced both apoptotic cells and intracellular ROS level but significantly increased antioxidant genes expression after glutamate-induced oxidative stress in HT-22 cells. However, in C. elegans, all LR extracts decreased intracellular ROS and protected the worms from oxidative stress through DAF-16/FOXO pathway leading to increase SOD-3 and decrease HSP-16.2. On the other hand, the SKN-1 and GST-4 were not changed. All the extracts extended lifespan and reduced lipofuscin, whereas only high concentration improved pharyngeal pumping rate. All the extracts did not alter the body length and the progeny of the worms excluding dietary restriction. In addition, they exhibited the neuroprotective effects by enhancing chemotaxis Index (CI) in Aβ containing worms and decreasing PolyQ40 aggregation. Interestingly, only LRE exerted neuroprotection on both in vitro and in vivo. Therefore, this novel study could suggest that LR extracts, especially LRE, may be an alternative for neuroprotective supplements.