Ferroptosis is a unique iron-dependent form of regulated cell death (Dixon et al., 2012). The accumulation of lipid peroxidation products and lethal reactive oxygen species (ROS) is the main characteristic of ferroptosis (Xie et al., 2016). Ferroptosis, as a way to promote cell death, may be implicated in the occurrence and development of many diseases. Studies have shown the importance of ferroptosis in many diseases of the central nervous system, such as Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), amyotrophic lateral sclerosis (ALS), and traumatic brain injury (TBI; Wu et al., 2018).

Gastrodiae Rhizoma (Tianma), a notable Chinese herb, is dry tubers of Gastrodia elata Blume which belongs to Orchidaceae. Gastrodiae Rhizoma is considered a top-grade medicine described to treat the hypertension of liver-yang hyperactivity in the tradition Chinese medicine. Studies have shown that the application of gastrodia elatahas biological activities of anticonvulsion, antioxidation, neuroprotection, anti-denguevirus, anti-cardio-cerebral-vascular diseases, and anti-inflammation (Ahn et al., 2007; Han et al., 2014; Zhan et al., 2016). The major active component and material basis of Gastrodia elata are Gastrodin (GAS). GAS, a chemical compound that known as 4-hydroxybenzyl alcohol-4O-β-D-glucopyranoside, is isolated from the rhizome of Gastrodia elata. Furthermore, the molecular formula of GAS is C₁₃H₁₈O₇, and its chemical structural formula is shown in Figure 1. GAS has numerous pharmacological activities including analgesic (Guo et al., 2013), antidepressant (Chen and Sheen, 2011), anxiolytic (Peng et al., 2013), anti-inflammatory (Kim et al., 2012), antiobesity (Park et al., 2011), and memory and retrieval improvements (Wu et al., 1996; Hsieh et al., 1997). Among them, analgesic, antioxidant, anti-inflammatory, and neuroprotective effects are the main research hotspots in recent years. Recent findings suggest that GAS exerts a protective effect on primary neural progenitor cells (NPCs) by resisting amyloidβ (Aβ; 1–42)-induced neurotoxicity (Li and Qian, 2016). In the meantime, GAS increased the expression of HO-1, Nrf2, and GPX4 protein in Rat Glioma Cell Line C6, which protected Rat Glioma Cell Line C6 from ferroptosis induced by H₂O₂ (Jiang et al., 2020). In recent years, several neuroprotective mechanisms of GAS have been fund. However, the study regarding to the effects of Gas on ferroptosis is rare. GAS was also reported to display powerful anti-inflammation properties. Based on the above research progress and analysis, it is speculated that GAS might be a potential therapeutic for the inhibition of ferroptosis. This study was designed to explore the analgesic, anti-inflammatory, and anxiolytic effects of GAS. We also examined whether GAS can exhibit neuroprotective effect through inhibition of ferroptosis, as well as its relation with intestinal microbiota.

Acute inflammatory pain induced by injection of CFA. In this process, rats were allergic to mechanical allodynia and thermal hyperalgesia, and the pain-induced anxiogenic effect lasted for more than 14 days (Nagakura et al., 2003). Clinically, it has been reported that chronic pain leads to mental problems such as anxiety and depression, which seriously reduces the quality of life of patients and hinders their normal life (Gallagher et al., 1995). GAS is a phenolic glucoside with significant analgesic and anti-inflammatory effects. In the CFA-induced chronic inflammatory pain model, we found that mechanical and thermal pain threshold were increased with treatment of GAS in a dose-dependent pattern. In addition, the number of entries in open arms and retention times of open arms were increased by GAS. These studies further confirmed that GAS has powerful analgesic, anti-inflammatory, and anti-anxiety effects in the chronic inflammatory pain model of mice. GAS exerted analgesic and anti-inflammatory effects by decreasing the activation of astrocyte and microglia and the induction of TNF-α and IL-6 in the ACC (Sun et al., 2016). In a mouse model of chemotherapeutic agent-induced neuropathic pain, 5-HT 1A receptor can mediate the powerful antinociceptive of GAS (Guo et al., 2013).

Inflammatory disease (ID) is a series of diseases characterized by inflammatory response, and ferroptosis is closely related to inflammatory response (Andersen et al., 2020). There are some inflammatory factors related to the metabolism of peroxides and arachidonic acid in ferroptosis tissues (Stockwell et al., 2020). Studies have shown that both ferroptosis and inflammatory diseases have the depletion of Gx4 and GSH, the increase of lipid peroxidation products, and the interruption of iron metabolism (Mao et al., 2020). At present, although a variety of molecular mechanisms and signaling pathways can lead to ferroptosis, iron metabolism and lipid peroxidation signaling are the main way to regulate ferroptosis (Dixon et al., 2015). During iron metabolism, excessive iron leads to ferroptosis by producing ROS. Ferritin heavy chain 1 (FTH1), as an iron storage protein complex, is involved in the uptake of excessive iron (Xie et al., 2016). We found that GAS increased the expression of FTH1 and thus balanced intracellular iron levels. The heme oxygenase-1 (HO-1), a major intracellular source of iron (Kwon et al., 2015), plays an important role in ferroptosis and inflammation. It was reported that p38 MAPK phosphorylation could mediate the protective effect of GAS on H₂O₂-induced oxidative stress (Zhang et al., 2018). GAS could ameliorate MPP+-induced oxidative stress by regulating the expression of HO-1 in human dopaminergic cells (Jiang et al., 2014). We also demonstrated that GAS increases HO-1 expression, which accelerates the decomposition of heme and inhibits inflammation. In addition, the expressions of glutathione peroxidase4 (GPX4) and prostaglandin-endoperoxide synthase2 (PTGS2) are also important for the induction of ferroptosis.

In our experiments, GAS significantly upregulates the expression of FTH1 and GPX4, decreases PTGS2 expression, and suggests that GAS against ferroptosis by reducing lipid peroxidation. CFA-induced chronic inflammatory pain is accompanied by the ferroptosis of neuronal cells, and GAS has an inhibitory effect on ferroptosis, which is one of the possible mechanisms to protect neuronal cells.

The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.

All experimental procedures were approved by the Animal Ethics Committee of Southwest Jiaotong University and were conducted in accordance with the university’s animal experiment guidelines.

JW and ZH: data collection. XW and XL: conceive and design the study. JW and XL: statistical analysis. XC, JW, and ZH: drafting the manuscript. XC and HL: critical revision of the manuscript. All authors contributed to the article and approved the submitted version.

This work was supported by the Science and Technology Department of Sichuan Province (project no.: 2018FZ0096) and the Fundamental Research Funds for the Central Universities (project no.: A0920502052001-10).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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