NaturalEssential Oils: A Promising Therapy Way for Treating Ischemic Stroke

Abstract

Stroke is an acute cerebrovascular disease with high morbidity, mortality, and disability, making it the second leading cause of death worldwide. Ischemic stroke (IS) accounts for the majority of strokes, and its pathogenesis is complex, often involving complications and sequelae. Currently, conventional clinical approaches are ineffective, with few drugs available for intravenous thrombolysis and mechanical thrombolysis limited by a short time window. With the poor efficacy of monotherapy, the search for new complementary or alternative therapies has become the focus of researchers. In traditional medicine, Chinese aromatherapy has a long history of using aromatic medicines to treat IS. Natural essential oils (EOs), as the main pharmacological substances in aromatic drugs, are composed of different ratios of active metabolites with multi-targets and multi-components, which makes EOs have a wide range of pharmacological effects. Modern studies have also shown that EOs extracts and isolated monomers are beneficial for pathologically complex CIS. Therefore, this paper summarizes the EOs and monomers obtained from EOs that can prevent and treat IS in the last 20 years, and finds that EOs exert their anti-CIS effects mainly through anti-oxidative stress, anti-inflammation, anti-apoptosis, and inhibition of excitotoxicity. The amelioration of IS complications by natural EOs and their active monomer components for the treatment of IS are further discussed.

1 Introduction

Stroke is a cerebrovascular disease that seriously jeopardizes human health. According to The Global Burden of Disease Study 2019, as many as 655 million people worldwide died from stroke in 2019, making it the second leading cause of death worldwide^1）. After a stroke, common sequelae such as, hemiparesis, speech disorders, etc. seriously affect the quality of life of the patient’s life. Insufficient blood supply to the brain due to clots or other particles blocking the arteries, resulting in impaired neurological function is Ischemic stroke (IS) , which accounts for about 62.4% of all stroke types^2）. Currently, intravenous thrombolysis and mechanical thrombolysis are recognized as effective protocols for IS, restoring blood and significantly improving patient prognosis. However, available clinical trials have shown that the time window for intravenous thrombolysis is only 6 h, and mechanical thrombolysis requires a patient assessment of the pros and cons before proceeding. This leaves only a small percentage of patients to be treated with thrombolysis or thrombus retrieval within the specified time window^3）. In addition, there is a risk of tissue edema, hemorrhage, and delayed neuronal necrosis after reperfusion. Therefore, it is important to focus on the treatment of IS and find new and safe complementary therapies.

Aromatherapy is considered a safe and effective natural therapy^4）. As an important material basis for the efficacy of aromatherapy, essential oils (EOs) are mixtures of secondary metabolites extracted from aromatic plants^5）. It has a wide range of pharmacological effects, including antibacterial, antiviral, insecticidal, anti-inflammatory, antioxidant, antiulcerative, etc^6）. The use of EOs for stroke treatment in humans is well documented. In China, Angong Niuhuang Wan, a formula from the Qing Dynasty’s 〝Item Differentiation Of Warm Febrile Diseases〟, has been used for stroke for more than 200 years. Its modern dosage form, Qingkailing Injection, is the only Chinese medicine injection approved for stroke use in China. Despite the different dosage forms, the basic composition of the prescription includes aromatic medicines such as borneolum and moschus^4）. With the increased interest in aromatic medicines, more and more preclinical studies have shown that EOs exert pharmacological effects such as antioxidant, anti-inflammatory, anti-apoptotic, and inhibition of excitotoxicity, and are able to reduce infarct size in mice/rats with middle cerebral artery occlusion (MCAO) model^7）. EOs have significant advantages over chemicals, and they are generally recognized as natural, environmentally friendly, and safe. Products derived from more than 150 aromatic plants (e.g., EOs, extracts, oleic acid) are recognized by the U.S. Food and Drug Administration as safe for human consumption^8）. In addition, the fat solubility and volatility of essential oils are good, and it is easier to pass through the blood-brain barrier (BBB) and reach the brain lesion site to exert the medicinal effect. EOs, as a class of mixed bioactive metabolites, are multi-component and multi-targeted, and can modulate multiple targets while ameliorating the complications of IS. Therefore, natural EOs are valuable for the treatment of IS and improvement of prognosis.

In this article, we begin by discussing the types of stroke, the possible pharmacological actions and mechanisms by which essential oils and monomers derived from essential oils are able to prevent and treat IS and IS complications are then summarized, providing the composition of these EOs, as well as the structures of the classical monomers. We hope that this paper will provide a scientific basis for the development of anti-IS drugs with good efficacy, low side effects and good prognosis. In future studies, we also hope that researchers will be able to find more suitable and safer natural medicines based on the medicinal properties of EOs.

2 Classification and Structure of Essential Oils

EOs are volatile complex mixtures extracted and processed from aromatic plants or animals, soluble in alcohols, ethers and fixed oils, but insoluble in water and with a special odor. Due to volatility, thermal instability and good fat solubility, EOs are commonly extracted by traditional extraction methods including, water vapor distillation, cold pressing and solvent extraction. In recent years, the rise of many new extraction and separation techniques has greatly increased the efficiency of essential oil extraction, such as supercritical fluid extraction, subcritical extraction, solvent-free microwave-assisted extraction. Garcez et al. compared the effects of different extraction methods on the extract and extraction efficiency of Anethum graveolens L. EOs and showed that supercritical fluid extraction was 2.6 times more efficient than hydrodistillation and there are unique components of the extract^9）. The pharmacological activity of EOs is a synergistic effect of different ratios of active monomers. Currently, researchers mainly determine the components of EOs by gas chromatography-mass spectrometry, and most EOs are able to detect more than 20 components^10）. With the development of essential oil extraction technology, separation and purification technology, structure identification technology, extensive pharmacological studies, more and more new conformational compounds with better pharmacological activities have been discovered. These single compounds derived from EOs have been validated in extensive preclinical experiments and are expected to be the lead compounds for anti-IS. We therefore summarize the plant EOs and monomer compounds with anti-IS properties reported in the literature, as shown in Fig. 1 and list the structures and classifications of these monomers.

Fig.1

Structureand classification of monomers from essential oils.

As can be seen from Table 1, there are more than 21 types of plant EOs currently used for anti-IS, most of which are derived from the Umbelliferae, Rutaceae, and Zingiberaceae, with a few from the Leguminosae, Santalaceae, and Labiatae. As is known, Umbelliferae and Rutaceae are important sources of aromatic medicines, and these EOs from the same families may have the same components. For example, Angelica sinensis (Oliv.) Diels and Ligusticum chuanxiong Hort., both in the Umbelliferae, have ligustilide. In addition, certain components are prevalent in EOs, only in different proportions. For example, linalool contains about 7% of Lavandula angustifolia EO^11）, 1.1% of Pimpinella anisum L. EO^12）, and 3.31% of Gleditsia sinensis Lam. fruit EO^13）. In summary, most of these EOs consist of one or two major components in high concentrations and multiple trace components that synergize to exert anti-IS effects. For example, Pomelo Peel EO and Bergamot EO both contain D-limonene as the highest component, but they are very different in terms of the composition and proportion of trace components. Citrus medica L. var. sarcodactylis Swingle EO is mainly composed of D-limonene (34.69%) , _γ-Terpinene (20.42%) , β-Bisabolene (4.59%) ^14）. The composition and proportions of EOs are the basis for their efficacy, and therefore the two EOs are not identical in their anti-IS mechanisms. Citrus maxima (Burm.) Merr. EO exhibited anti-inflammatory, antioxidant, and anti-apoptotic effects, while Citrus medica L. var. sarcodactylis Swingle EO exhibited inhibition of excitotoxicity. Overall, these compounds synergize with each other or play major/minor roles, fully reflecting the multi-component, multi-target model of EOs exerting a holistic modulatory effect on IS.

Table1

Essentialoils isolated from natural products against ischemic stroke.

Table1

Essentialoils isolated from natural products against ischemic stroke.

Table1

Essentialoils isolated from natural products against ischemic stroke.

Volatile terpenoids are the main components of the aromatic properties of aromatic plants, in addition to a number of aromatic compounds that also play an important role. From the chemical point of view, these terpenes can be categorized as monoterpenes, monoterpene alcohols, monoterpene ketones, sesquiterpene ketones, and sesquiterpene alkenes. The general ability of these aromatic plants to make large amounts of terpenes is due to the presence of a large number of genes already involved in terpene biosynthesis in the genomes of their species^15）. As can be seen in Fig. 1 and Table 2, there are 26 monomers with 13 terpenoids, 2 polyphenols, 2 flavonoids and other types of compounds. Understanding the structures of these compounds is necessary to provide possible drug core structures for future development of anti-IS drugs.

3 Anti-ischemic Stroke Essential Oils

3.1 For oxidative stress

Brain tissue is more sensitive to oxidative stress due to the highest metabolic activity per unit weight of brain, low antioxidant enzyme content, and high membrane content. Oxidative stress is the overproduction of free radicals beyond the scavenging capacity of one’s own antioxidant defense system. Free radicals are categorized into reactive oxygen species (ROS) and reactive nitrogen species (RNS) ^112）. ROS are mainly superoxide anion (O^2－) , hydroxyl radical (OH^－) , lipid radical (ROO^－) , and hydrogen peroxide (H₂O₂) , and are mainly derived from xanthine oxidoreductase, NADPH oxidase, 5-lipoxygenase, and mitochondria^113）. In addition, nitric oxide (NO) is synthesized through nitric oxide synthase (NOS) -catalyzed synthesis and may react with O^2－ to generate the highly reactive free radical peroxynitrite anion (ONOO^－) ^114）. During cerebral ischemia-reperfusion, the glutathione peroxidase free radical scavenging system decreases, while lipid peroxides such as malondialdehyde (MDA) and lactate dehydrogenase (LDH) all rise significantly, leading to an oxidative-antioxidative imbalance that allows free radicals to accumulate and accumulate^115）. Excess free radicals damage proteins, peroxidize lipids and damage DNA, as well as affect intercellular signaling cascades, ultimately leading to cellular damage^116）. A large amount of research evidence suggests that oxidative stress is an important mechanism involved in ischemia-induced neuronal injury and is directly related to the degree of neurological deficit and clinical prognosis. Preventing the action of these free radicals has been shown to be an effective measure to limit the expansion of ischemic lesions. For example, the antioxidant edaravone scavenges hydroxyl, peroxyl, and superoxide radicals, inhibits lipid peroxidation, reduces the size of the ischemic hemidiaphragm that develops or infarcts, and inhibits delayed neuronal death, improving the prognosis of IS, and it is currently in use in China and Japan^117）. Overall, overproduction against ROS and RNS may provide satisfactory results in the treatment of IS.

As can be seen from the Table 1 and Table 2, most of the EOs and monomers are associated with the inhibition of oxidative stress. Various EOs inhibit oxidative stress by inhibiting lipid peroxidation and increasing antioxidant enzyme activity. For example, Citrus aurantium L. EO significantly reduced the levels of lipid peroxides MDA in brain tissue and serum of rats after cerebral ischemia-reperfusion^23）. Lavandula angustifolia EO has been reported to significantly increase superoxide dismutase (SOD) , GPX, and catalase (CAT) activities and reduce MDA production in brain tissues after cerebral ischemia and reperfusion^118）. Notably, linalool, the shared active ingredient of the two EOs, has been shown to upregulate SOD and catalase activities in an OGD/R-induced cortical neuronal cells cell model, which may be one of the main active ingredients of the two EOs in inhibiting oxidative stress^65）. Carvacrol is mostly found in the EO of Origanum Linn. and is able to alleviate cerebral ischemic oxidative stress^63）. Its derivative Carvacryl acetate was able to activate the nuclear factor erythroid2-related factor 2 (Nrf2) signaling pathway to alleviate CIRI-induced oxidative stress injury in MCAO-induced rat and OGD/R-induced PC12 cell models^64）. In addition, there are EOs that inhibit the production of RNS. Curcuma longa L. EO has been found to reduce infarct volume, neurologic scores, and cerebral water content when administered either pre-MCAO treatment or post-MCAO^37）. It may exert neuroprotective effects by inhibiting NO-induced peroxynitrite formation. Angelica sinensis (Oliv.) Diels EO significantly reduced NO content and NOS activity after cerebral ischemia and reperfusion, suggesting that the underlying mechanism is related to the inhibition of NO neurotoxicity^47）.

Table2

Monomers isolated from natural essential oils against ischemic stroke.

Table2

Monomers isolated from natural essential oils against ischemic stroke.

Table2

Monomers isolated from natural essential oils against ischemic stroke.

3.2 For inflammatory

Immune-mediated inflammatory response is an important pathological mechanism of brain injury after IS. The inflammatory response after stroke is a complex pathological process involving intrinsic cells in the brain, peripheral immune cells, inflammatory mediators and their signaling^115）. After cerebral ischemia, dying and necrotic cells release danger-associated molecular patterns that bind to receptors on microglia, astrocytes, and other cells, activating downstream pathways. Inflammatory mediators, trend factors, and adhesion factors released by these cells prompt a variety of immune cells in the circulation to gather and infiltrate into the focal area, further secrete a variety of pro-inflammatory mediators, and exacerbate the cerebral inflammatory response^119）. At the same time, the aggregated immune cells release oxygen free radicals, protein hydrolyzing enzymes and cellular agonists, which directly damage the endothelial cells and further destroy the BBB^120）. In addition, inflammation is an important factor in the course and prognosis of IS. Numerous studies have shown that the inflammatory markers interleukin 6 (IL-6) and C-reactive protein are associated with neurologic deficits and are proportional to infarct size^121）.

Treatment with Citrus maxima (Burm.) Merr. EO (30 mg/kg) has been found to significantly reduce infarct volume and improve neurologic dysfunction and pathologic changes. It inhibits neutrophil infiltration and levels of pro-inflammatory factors. These mechanisms are associated with inhibition of the Toll-like receptor 4 (TLR4) / nuclear factor kappa-B (NF-κB) signaling pathway^16）. Reduction of Serum levels of tumor necrosis factor-α (TNF-a) and interleukin－1β (IL-1β) and increased serum levels of IL-10 in HFD and MCAO rats after intranasal administration of a high dose of Gleditsia sinensis Lam. fruit EO (0.308 mg/kg) ^13）. In addition, Ligusticum chuanxiong Hort. EO may exert a neuroprotective effect by attenuating the post-IR inflammatory response^44）. Z-Ligustilide, a major constituent of Ligusticum chuanxiong Hort. EO, also reduced tumor necrosis factor-α (TNF-α) levels in ischemic brain tissue^122）. β-Caryophyllene, a common component of several EOs, was able to down-regulate the expression of inducible NO synthase, interleukin-1β, IL-6 and cyclooxygenase 2 in microglia in a dose-dependent manner^76）. NBP is found in the EOs of many Apiaceae Lindl., such as Ligusticum chuanxiong Hort. EO, Angelica sinensis (Oliv.) Diels EO, and Apium graveolens EO. NBP softgels are used as a treatment for IS in China. In MCAO rats model and lipopolysaccharide (LPS) -induced astrocytes, it has been demonstrated that NBP inhibits the TLR4/NF-κB pathway and reduces the release of the pro-inflammatory factors IL-6, IL-1β, and TNFα by upregulating the expression of hepatocyte growth factor (HGF) ^97）.

3.3 For apoptosis

After ischemia, tissue necrosis in the core is severe and irreversible, but brain tissue in the penumbra is salvageable. In the ischemic penumbra, apoptosis is the main mode of cell death^123）. Apoptosis induced by cerebral ischemia is generally of two pathways. One is an exogenous pathway triggered by death receptors: death receptors such as Fas bind to ligands and recruit FADD and caspase-8 zymogen to form death-inducing signaling complexes. Caspase-8 is activated, which in turn activates caspase-3. The other is the mitochondria-triggered endogenous pathway: after the cell is subjected to an endogenous apoptotic signal, it releases cytochrome c (Cyt-c) , which forms an apoptotic complex with APAF1 and caspase-9 zymogen. Activated caspase-9, which further activates caspase-3 and caspase-7, triggers apoptosis. The endogenous and exogenous pathways of apoptosis are interconnected. For example, activated caspase-8 cleaves the signaling molecule Bid, triggering endogenous apoptosis. Studies have shown that inhibition of apoptosis reduces ischemic injury, such as inhibition of cysteine 3, gene deletion of Bid, and the use of viral vector-mediated gene transfer of B-cell lymphoma-2 (Bcl-2) and Bcl-XL. Mu-Xiang-You-Formulation EO showed significant protective effect on MCAO animal model.

Neurologic severity scores, infarct volume, and expression of apoptosis-related proteins and their messenger RNA (mRNA) collectively demonstrated this effect. Neurologic severity scores, infarct volume, and expression of apoptosis-related proteins and their mRNAs collectively demonstrated this effect^54）. The neuroprotective effects of Mu-Xiang-You-Formulation EO are related to its components methyleugenol, beta-caryophyllene, arachidonic acid, piperine and sesquiterpenes. All of the above components showed anti-IS effects in animal models. Furthermore, in an OGD/R-induced rat cortical neuronal cell model, different doses of Liquidambar orientalis Mill. EO have been shown to have neuroprotective effects attributed in part to apoptosis-inhibiting activity. Inhibition of the toll-like receptor 9 (TLR9) signaling pathway is the primary mechanism^29）. Preeti Dohare et al. found that Curcuma longa L. EO (250 mg/kg) significantly reduced the number of apoptotic cells when administered in MCAO mice, and its main component, Germacrone, also attenuated cerebral ischemia-reperfusion injury through anti-apoptosis^37）,71）.

3.4 For glutamate excitotoxicity

Under physiological conditions, glutamate is maintained at low extracellular levels and is involved in signaling as an excitatory neurotransmitter^124）. Presynaptic Ca²⁺ overload after cerebral ischemia leads to calcium-dependent glutamate cytosolic release^125）. At the same time, excitatory glutamate transporters on astroglial thin membranes are dysfunctional, with a reduced capacity for glutamate clearance or even reverse release^{126）,127）}. Eventually excess glutamate accumulates extracellularly, causing sustained activation of glutamate receptors on the postsynaptic membrane, which triggers a series of cascading reactions leading to neuronal damage, including collapse of electrochemical gradients, activation of proteases and phospholipases, etc., degradation of vital substances, and increased ROS^128）. N-methyl-D-aspartic acid receptor (NMDAR) has a high affinity for glutamate and play a major role in glutamate-mediated excitotoxicity. Over the years, a large number of antagonists have been developed against NMDAR, e.g., ketamine, memantine, MK-801, etc. However, complete antagonism of glutamate receptors, at the same time, interferes with the normal transmission of nerve signals. Therefore how to i.e. maintain the normal activity of NMDAR and selectively inhibit the over-activation of NMDAR is the goal of developing glutamate inhibitors in the future.

β-Asarone ether is the active ingredient in Acorus tatarinowii EO. It has been shown that beta-asarone, borneolum, musk ketone, and Liquidambar orientalis Mill. EO can treat IS by inhibiting excitatory amino acid toxicity. The four drugs could reduce the level of aspartic acid (Asp) and glutamate (Glu) in the brain tissue of rats with different degrees of injury (p<0.05) , and the strengths of their effects were as follows: Liquidambar orientalis Mill. EO>beta-asarone>borneolum>musk ketone group^27）. Fariba Karimzadeh et al. initially evaluated the effects of Pimpinella anisum L. EO on hypoxia-induced neurons and found that the neuroprotective effects of Pimpinella anisum L. EO were associated with inhibition of N-methyl-D-aspartic acid (NMDA) receptors^12）.

3.5 Other

The pathology involved in IS and its complexity, in addition to oxidative stress, inflammation, apoptosis, excitotoxicity, but also iron death, impaired energy metabolism, autophagy, impaired BBB and other mechanisms, and each other crosstalk intervention, as shown in Fig. 2. The BBB is an important physical and biochemical barrier that maintains central nervous system homeostasis and protects brain tissue^129）. After stroke, BBB impairment is associated with poor patient outcomes, hemorrhagic transformation^{130）,131）}. Early post-stroke events include decreased ATP synthesis, dysregulation of ionic homeostasis, structural disruption of tight junction proteins, and increased paracellular permeability, leading to cerebral edema as well as impairment of the BBB. In addition, the inflammatory cascade response and the aggregation of immune cells in the brain exacerbate the destruction of the BBB^131）. Autophagy acts as a cellular survival mechanism that degrades unwanted or dysfunctional components^132）. Several studies have shown that autophagy plays a dual role in cerebral ischemia-reperfusion. On the one hand, autophagy exerts neuroprotective effects by removing damaged organelles and proteins; on the other hand, over-activated autophagy promotes damage to normal organelles and proteins and aggravates cell death. Autophagy is tightly regulated and can be induced in cerebral ischemia/reperfusion via phosphoinositide 3-kinase (PI3K) / protein kinase B (Akt) / mammalian target of rapamycin (mTOR) , AMPK, and MAPK-mTOR pathways.

Fig.2

Pharmacologicmechanism of essential oils anti-ischemic stroke.BBB: blood-brain barrier; Glu: glutamate; NMDAR: N-methyl-D-aspartic acid receptor triphospha; NOS: nitric oxide synthase; NO: nitric oxide; ONOO^–:peroxynitrite anion; H₂O₂:hydrogen peroxide; OH^–:hydroxyl radical; SOD: superoxide dismutase; O^2..:superoxide anion; RNS: reactive nitrogen species; Bax: Bcl2-associated X; Bcl-2: B-cell lymphoma-2; Cyt-c: cytochrome c; DAMP: danger-associated molecular patterns; ROS: reactive oxygen species

Several extracted parts of Zingiber officinale Rosc., including Zingiber officinale Rosc. EO, alcoholic extracts, and aqueous extracts, have been shown to be protective against ischemic brain tissue injury^133）. Notably, Zingiber officinale Rosc. EO improved brain tissue energy metabolism and significantly increased Na⁺, K⁺-ATPase, Ca⁺- ATPase activities in brain tissue in a dose-dependent manner^50）. β-Caryophyllene, a bicyclic sesquiterpene widely found in EOs, has been extensively studied for its pharmacological effects on IS, including antioxidant, anti-inflammatory and anti-apoptotic effects. A recent study showed that beta-caryophyllene preconditioning protected rats from ischemia-reperfusion injury, and neurological scores, cerebral infarct volume, and HE staining collectively demonstrated this effect. In the MCAO rats model and OGD/R astrocytes, there was decreased expression of the important regulatory proteins of iron death: GPX4 and increased expression of ACSL4 and COX-2; lipid peroxidation and disruption of mitochondrial structure, and beta-caryophyllene could reverse these pathological changes^72）. These evidences suggest that beta-caryophyllene can modulate iron death to exert neuroprotective effects, and further studies have shown a close correlation with pathways activating the Nrf2/heme oxygenase-1 (HO-1) axis. Ligustilide is one of the main active components of the EOs of Angelica sinensis (Oliv.) Diels EO and Ligusticum chuanxiong Hort. Juan Li et al. pretreated ligustilide (15 mg/kg) by nasal drip for 3 days and then established the MCAO rat model^94）. Researchers found that ligustilide was able to attenuate infarct volume, neurological dysfunction, BBB permeability, and cerebral edema after cerebral ischemic injury by reducing the loss of Vascular Basement Membrane Protein and Tight Junction Proteins after cerebral ischemia and significantly lowering the levels of matrix metalloproteinases (MMP) -2 and MMP-9, which is related to the possible mechanisms behind Nrf2 and heat shock protein 70 (HSP70) signaling pathways. β-asarone is the main effective component of Acorus tatarinowii EO. In the PC12 model of OGD/R, it showed pharmacological effects to reduce Beclin-1 dependent autophagy.

4 Improvement of Ischemic Stroke Comorbidities

Clinical studies have found that comorbidities in IS patients greatly affect their prognosis and mortality rates^134）. In other words, the presence of underlying diseases and comorbidities predisposes to a vicious cycle of progressive IS. Risk factors for IS have been extensively studied. A recent study showed that the top five risk factors for IS are high systolic blood pressure, high body mass index, high fasting blood sugar, environmental particulate pollution and smoking^1）. In addition, risk factors were associated with the type of stroke. A large case-control study in 2016 reported 10 risk factors associated with stroke, with seven risk factors for ischemic and hemorrhagic stroke, and a greater correlation between hypertension and cerebral hemorrhage, with smoking, diabetes mellitus, apolipoproteins, and cardiac causes now even more highly correlated with IS^135）. Patients with stroke co-morbidities are common in clinical practice after stroke due to various factors such as ischemia, immunosuppression, and infection^136）. Common complications include cerebral edema, pneumonia, hemorrhagic transformation, seizures, impaired consciousness, dementia, and depression. These complications are not only detrimental to the patient’s prognosis, but also contribute to the high mortality and disability rates in IS patients^{134）,137）,138）}. Aromatherapy, represented by essential oils, is widely used as an intervention treatment for IS complications, especially to relieve patients’ moods such as insomnia and depression.

We summarized the combined application of EOs and its monomers in the treatment of IS. 3-n-Butylphthalide (NBP) significantly alleviated learning and memory deficits as well as pathological changes in the hippocampal region after RCIR. NBP exerted antioxidant, anti-neuroinflammatory and neuroprotective effects in a mouse model of RCIR-induced vascular dementia. The mechanism involved activation of the BDNF/TrkB signaling pathway and Nrf2 and inhibition of the TLR4/MyD88/NF-κB signaling pathway^139）. In addition, NBP had been shown to improve cognitive deficits after cerebral ischemia-reperfusion in animal models^{140）,141）,142）}. Studies had shown that Lavandula angustifolia EO treatment, especially at a dose of 200 mg/kg, significantly reduces infarct size (TTC staining) , cerebral edema (wet and dry gravimetry) , and neurological function scores. Further studies revealed that its pharmacological effects were associated with enhancement of endogenous antioxidant defense, inhibition of oxidative stress, and increased vascular endothelial growth factor (VEGF) expression. Alpinia zerumbet EO improved post-stroke skeletal muscle spasms in patients by relaxing muscles. Brain injury is the main cause of coma and death in patients undergoing the cardiac arrest/cerebral ischemia reperfusion model (CA/CPR) . EO from Citrus maxima (Burm.) Merr. peel improved survival and neurological deficit scores in rats 24 h after CA/CPR. Citrus maxima (Burm.) Merr. EO inhibited the expression of TNF-α, RIPK1, RIPK3, and p-MLKL/MLKL proteins, suggesting that it exerts its neuroprotective effects by inhibiting TNF-α-induced necrotic apoptosis. In particular, a medium dose of Citrus maxima (Burm.) Merr. EO (20 mg/kg) was the most efficacious^19）. Hypertension is a common comorbidity and risk factor for IS^{143）,144）}. Linalyl acetate monomer from finger Citrus medica L. var. sarcodactylis Swingle EO and Lavandula angustifolia EO prevents hypertension-associated ischemic injury. Yu et al. established a hypertension-ischemic injury model by inducing hypertension in rats in vivo and taking aortic OGD/R to induce ischemia after linalyl acetate injection. Linalyl acetate pretreatment significantly reduced systolic blood pressure in vivo and led to a reduction in ROS production through anti-p47phox expression, thereby attenuating ROS-induced LDH release as well as endothelial nitric oxide synthase (eNOS) inhibition. These results suggest that linalyl acetate can prevent hypertension-related ischemic injury^145）. In addition, D-limonene reduced systolic blood pressure and cerebral infarct size in post-stroke SHRsp rats, which is thought to be associated with anti-inflammatory, vascular remodeling, and antioxidant activities^146）.

5 Conclusion

In summary, there has been a growing body of evidence over the past 20 years regarding the treatment of IS with EOs or their main components. They exert multi-targeted pharmacological effects by responding to the pathological mechanisms of IS-oxidative stress, inflammation, apoptosis, and excitotoxicity. Not only do these natural EOs and monomers act synergistically by combining them with other drugs, both chemical and botanical extracts, to improve the efficacy of individual drugs, but they also have a positive preventive and therapeutic effect on the complications of IS. This evidence suggests the potential of natural EOs and their active constituents for the prevention and treatment of IS.

The use of natural EOs in IS patients is still in its infancy. On the one hand, in most preclinical experiments, researchers have mostly focused on the histologic and morphological improvements of natural EOs on IS model animals and pathology-related biochemical indices. There is less research on how natural EOs work by affecting genes, proteins, and metabolic levels. In order to further translate EOs to the clinic, further exploration of the targets and mechanisms of action by which the drugs exert their anti-IS effects is needed. One can explore the pharmacodynamic basis of EOs in conjunction with network pharmacology, genomics, proteomics, and metabolomics. On the other hand, the consistent quality of EOs is the basis for good medicinal effects. In order to ensure industrial production and clinical use, it is necessary to establish a standardized plant growing base for raw materials, develop quality standards for EOs of origin, and adopt advanced extraction methods such as supercritical extraction to improve. In addition, the use of pharmacological means facilitates the retention and slow release of the active ingredients of EOs. It is worth noting that natural EOs are fat-soluble, volatile and other special properties, easy to volatilize into the nasal cavity, penetrate through the nasal mucosa, through the olfactory region delivered to the brain. This 〝nose-to-brain〟 mode of transportation is able to bypass the BBB and rapidly deliver the EOs to the IS site. Therefore, when designing routes of administration and dosage forms of natural EOs, due consideration should be given to the characteristics and advantages of the drugs. In conclusion, only by ensuring the stable quality of the EO, exploring the material basis and mechanism of action of its anti-IS, having the support of relevant clinical trial data, and pharmacological means to improve the performance of playing the oil preparation, can we provide more possibilities for the future research of new drugs for IS.

Funding

This work was supported by the Xinglin Scholar Discipline Promotion Talent Program of Chengdu University of Traditional Chinese Medicine [grant numbers: QJJJ2022014] , the Xinglin Scholar Discipline Promotion Talent Program of Chengdu University of Traditional Chinese Medicine [grant numbers: XCZX2022007] , Project of the Science and Technology Department of Sichuan Province [grant numbers: 2022NSFSC1406] .

AuthorContributions

Yuanyuan Wu: writing-original draft; writing-review and editing. Leying Gao: writing-review and editing. Yue Hu: writing-original draft. Xiaofang He: writing-review and editing. Wenli Ye: writing-original draft. Yu Long: writing-review and editing. Xiaoqiu Li: writing-review and editing. Jie Deng: visualization. Yin Ma: Supervision. Huiyi Feng: visualization. Haolin Liu: writing-review and editing. Qianqian Wu: conceptualization Nan Li: supervision.

Declarationof Competing Interest

There are no conflicts to declare.

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