Neuroprotective Effect for Cerebral Ischemia by Natural Products: A Review

Natural products have a significant role in the prevention of disease and boosting of health in humans and animals. Stroke is a disease with high prevalence and incidence, the pathogenesis is a complex cascade reaction. In recent years, it’s reported that a vast number of natural products have demonstrated beneficial effects on stroke worldwide. Natural products have been discovered to modulate activities with multiple targets and signaling pathways to exert neuroprotection via direct or indirect effects on enzymes, such as kinases, regulatory receptors, and proteins. This review provides a comprehensive summary of the established pharmacological effects and multiple target mechanisms of natural products for cerebral ischemic injury in vitro and in vivo preclinical models, and their potential neuro-therapeutic applications. In addition, the biological activity of natural products is closely related to their structure, and the structure-activity relationship of most natural products in neuroprotection is lacking, which should be further explored in future. Overall, we stress on natural products for their role in neuroprotection, and this wide band of pharmacological or biological activities has made them suitable candidates for the treatment of stroke.


INTRODUCTION
Stroke is a disease with high prevalence and incidence (Benjamin et al., 2018). Strokes can be divided into two categories: ischemic and hemorrhagic stroke, and over 80% are ischemic stroke . Ischemic stroke is a pathological condition characterized by blood vessels occlusion and insufficient of blood supply. Stroke which is one of the most common causes of disability and death worldwide, seriously endangers human health and brings heavy burden to society and family (Donnan et al., 2008;Kalogeris et al., 2016). The pathological mechanism of cerebral ischemia is a complex cascade reaction, and its severity is related to the time of cerebral ischemia and the depth of the ischemic site (Steliga et al., 2020). The occurrence and development of cerebral ischemia included neuron excitotoxicity, mitochondrial dysfunction, neuroinflammatory damage, oxidative stress, etc. It is generally believed that glutamate excitotoxicity, energy metabolism disorder, and Ca 2+ overload happened within 24 h of the onset of stroke, accompanied by the generation of free radicals (Guo, Li and Wang, 2009;Manzanero, Santro, Arumugam, 2013). Apoptosis and necrosis also occurred within a few hours of ischemia . In the subacute phase, brain edema and bloodbrain barrier (BBB) destruction happened. Endothelial cells, pericytes, astrocytes, etc are activated and inflammatory factors are released, accompanied with the proliferation of reactive glial cells (Stanimirovic and Satoh, 2000;Liebner et al., 2018). There exists endogenous repair in the late stage of cerebral ischemia, and neurogenesis, glial scars, angiogenesis could be observed (Steliga et al., 2020). Therefore, the occurrence and development of stroke is complicated, which makes its treatment very difficult. Tissue plasminogen activator (t-PA) is the only therapeutic medicine for ischemic stroke approved by Food and Drug Administration (FDA). However, t-PA has a restricted time window of 6 h, and delayed t-PA infusion increases the risk of hemorrhagic transformation and carries high mortality (Group 1995;Hacke et al., 2008). Despite constantly increasing understanding of ischemic stroke pathophysiology through laboratory and clinical studies, the treatment strategy is still not ideal. Therefore, there is an urgent medical need to identify new molecules that can provide neuroprotection against cerebral ischemic or ischemic reperfusion (I/R) injury.
Natural products obtained from natural sources, including plants, animals, fungi and microorganisms. Natural products have played an important role in ancient traditional medicine systems, such as Unani, Chinese and Ayurveda which are still in common use today. Natural products are known to exert additive, synergistic or antagonistic effects on the body. With the pharmacology technology developing, the administration forms of natural products are various, which has drawn much attention for the treatment of stroke from drug discovery to drug delivery (Hermann et al., 2019;Long et al., 2020;Williams et al., 2020). The pathogenesis of stroke is a complex cascade reaction, including neuro-inflammatory damage, oxidative stress, mitochondrial dysfunction, neurotoxicity, blood-brain barrier (BBB) disruption, neurovascular unit (NVU) damage and other factors. In recent years, it's reported that a vast number of natural products have demonstrated beneficial effects on stroke worldwide (Fei et al., 2020). The neuroprotection of natural products has been discovered to modulate activities via multiple targets and signaling pathways with direct or indirect effects on enzymes, such as kinases, receptors and proteins (Bagli et al., 2016). This wide band of pharmacological or biological activities has made them suitable candidates for the treatment of stroke (Cai et al., 2019;Zhang, C. et al., 2020;Zhong et al., 2020). As a requirement of novel drug development for stroke, there is a focus on the neuroprotection of natural products via multitargets. Hence, we stress on natural products for their role in neuroprotection ( Figure 1). This review provides a comprehensive summary of the pharmacological effects and multiple target mechanisms of natural products for cerebral ischemic or I/R injury.

METHODOLOGY
Database searches using Google scholar, PubMed, and China National Knowledge Internet (CNKI) were conducted until July 2020 to include up to date documented information in the present review. For data mining, the following words were used in the databases mentioned above: neuroprotection, natural product, phytoconstituents, natural products cerebral ischemic injury, natural products cerebral ischemic reperfusion injury, in vivo and in vitro studies for prevention and treatment of stroke. In almost all cases, the original articles or abstracts were obtained and the relevant data was extracted.

Neuroprotective Role of Flavonoids in Ischemic Brain Injury
Flavonoids, a natural bioactive compound found abundant in vegetables, fruits and traditional herbal medicine. Prevention and/or treatment with flavonoids such as baicalin, apigenin, vitexin, quercetin and other flavonoid compounds have shown promising neuroprotective effects against ischemic-induced injury by through increasing neuronal viability, cerebral blood flow and reducing ischemic-related apoptosis (Putteeraj et al., 2018).

Baicalin
Baicalin, mainly derived from the root of Scutellaria baicalensis Georgi, is one kind of a crucial flavonoid (Figure 2A). This medicinal plant is widely distributed in China, Russia, Mongolia, North Korea and Japan, and has the functions of clearing away heat and dampness, purging fire and detoxification . In recent years, baicalin has also been found to have a wide range of neuroprotective effects Sowndhararajan et al., 2018). Middle cerebral artery occlusion (MCAO) induced cerebral ischemic injury in vivo and oxygen-glucose deprivation (OGD) induced hypoxia injury in vitro were implemented. Baicalin at doses ranging from 50 to 200 mg/kg could significantly improve neurological deficit and reduce infarct volume via inhibiting apoptotic and oxidative pathway, including myeloid cell leukemia 1 (Mcl 1), B-cell lymphoma-2 (Bcl 2), superoxide dismutase (SOD), malondialdehyde (MDA), and so on (Cao et al., 2011;Zheng et al., 2015). Neuroprotective activity was also confirmed in another study. Baicalin could decrease nuclear factor kappa B (NF-κB) p 65 to exert neuroprotection (Xue et al., 2010). In addition, baicalin 100 mg/kg inhibits toll like receptor 2/4 (TLR2/ 4) signaling pathway in cerebral ischemia rats (Tu et al., 2011). Baicalin down-regulated nucleotide-binding oligomerization domain protein 2 (NOD2) receptor and tumor necrosis factor α (TNF α) expression of neurons with OGD in vitro and cerebral ischemia-reperfusion in vivo . These findings suggested baicalin could afford neuroprotection through multiple pathways.

Scutellarin and Scutellarein
Scutellarin ( Figure 2B) and scutellarein ( Figure 2C), is the major active component extracted from Erigeron breviscapus (Vant.) Hand-Mazz, a Chinese herbal medicine . Accumulated data have demonstrated the effectiveness and benefits of breviscapine and scutellarin in treating cerebrovascular disease and cardiovascular disease in clinic and in experimental study (Cao et al., 2008;Wang et al., 2015a;Gao et al., 2017). It has demonstrated that scutellarin is benefit to brain injury caused by cerebral ischemia/reperfusion due to its anti-oxidation and anti-inflammatory effects and ability to attenuate neuronal damage . Scutellarin could effectively suppress the inflammatory response in activated microglia/brain macrophage in experimentally induced cerebral ischemia, whose underlying mechanism was associated with suppressing the phosphory c-Jun N-terminal kinase (p-JNK) and p-p38 mitogen-activated protein kinase (MAPK), increasing phosphorylation extracellular regulated kinase 1/2 (p-ERK1/2) (Chen H.-L et al., 2020). In MCAO rats given scutellarin 100 mg/kg treatment, neurological scores were significantly improved coupled with a marked decrease in infarct size, which was found that scutellarin had a neuroprotective effect, amplified TNC1 astrogliosis and activated microglia to remodel injured tissue via notch pathway (Fang et al., 2015;Fang et al., 2016). Pretreated with scutellarin 50 or 75 mg/kg has protective effects for cerebral injury through upregulating endothelial nitric oxide synthase (eNOS) expression and downregulating of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and inducible nitric oxide synthase (iNOS) expression (Hu et al., 2005). Another study also proved scutellarin protected against ischemic injury through regulation of nicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2) and connexin 43 to decrease oxidative damage and apoptotic cell death (Sun, J. B. et al., 2018;Zhang et al., 2009). Scutellarin and scutellarein could improve neuronal injury, and scutellarein had better protective effect than scutellarin through improving the Ca 2+ -ATPase and Na + , K + -ATPase activity in rat cerebral ischemia with bilateral common carotid arteries (BCCAO) (Tang et al., 2014). Scutellarin protects brain from acute ischemic injury probably through its inhibitory effect on the angiotensin-converting enzyme (ACE) and Ang II type 1 receptor (AT1R), also TNF-α, interleukin-6 (IL-6), and interleukin-1β (IL-1β) .

Apigenin and Vitexin
Apigenin ( Figure 2D) is a natural flavonoid and vitexin ( Figure 2E) is an apigenin flavone glycoside, which were found in several dietary plant foods as vegetables and fruits, and had a variety of pharmacological effects (Babaei et al., 2020). It has been reported that apigenin and vitexin played a protective role in diseases associated with the oxidative process, such as cardiovascular and neurological disorders (Ayoobi et al., 2017;Che et al., 2020;Li, S. et al., 2017). Angiogenesis is one of the ways to repair brain injury. It's reported that apigenin could promote cell proliferation, tube formation, and cell migration while inhibiting apoptosis and autophagy by affecting caveolin-1/ VEGF, Bcl-2, caspase-3, Beclin-1, and mechanistic target of rapamycin (mTOR) expression. What's more, apigenin significantly reduced neurobehavioral scores and volume of cerebral infarction while promoting vascular endothelial cell proliferation by upregulating VEGFR2 to affect caveolin-1, VEGF, and eNOS expression in brain tissue of I/R rats (Pang et al., 2018). Vitexin could also play a protective role by suppressing oxidative stress . In addition, vitexin regulated cell apoptosis and autophagy via MAPK and mTOR/Ulk1 pathway to attenuate cerebral I/R injury (Wang et al., 2015b;Jiang et al., 2018). Apigenin also showed long-term therapeutic effect in cognitive impairments after cerebral I/R injury (Tu et al., 2017). Above all, it suggested apigenin and vitexin had neuroprotective effect through inflammation, angiogenesis, apoptosis and autophagy pathways with multitargets.

Icariin
Icariin ( Figure 2F), an active flavonoid extracted from Epimedium brevicornum Maxim, has been proven to possess a wide range of efficacy including anti-tumor, anti-oxidant, antibacterial and anti-inflammatory, etc (Luo et al., 2007;Zhou et al., 2011). It has been also reported to alleviate brain injury and attenuate cognitive deficits Li et al., 2005). Xiong et al. and Deng et al. found that pretreatment with Icariin and Icariside II, a metabolite of icariin, could decrease neurological deficit score, diminish the infarct volume, and reduce IL-1β and transforming growth factor-β (TGF-β1). Moreover, Icariin suppressed inhibitory κB (IκB)-α degradation and NF-κB activation and up-regulated peroxisome proliferator-activated receptor α (PPARα) and peroxisome proliferator activated receptor γ (PPARγ) levels in I/R model Xiong et al., 2016). Moreover, a study by Mo et al. found that Icariin can inhibit apoptosis and inflammatory in neurons after OGD/R through inositol requiring enzyme 1 (IRE1)-X-Box Binding Protein 1 (XBP1) signaling pathway (Mo et al., 2020).

Quercetin
Quercetin ( Figure 2G), is a common flavonoid found in many fruits and vegetables (Lee et al., 2003). It has been reported that quercetin has an anti-oxidative property, anti -inflammation and immunity, anti-cancer, etc (Costa et al., 2016;Shafabakhsh and Asemi 2019;Xu, D. et al., 2019). A study by Jin et al. found that quercetin could increase the expression of ZO-1, Claudin-5, β-catenin, and LEF1, and decrease the expression of MMP-9, GSK-3β and axin to reduce brain edema and BBB leakage and improve BBB dysfunction, which suggested the neuroprotection of quercetin was related to Wnt/β-catenin pathway . Calcium acts as a second messenger that mediates physiologic functions. It's reported that quercetin exerted a preventative effect through attenuation of intracellular calcium overload and restoration of down-regulated hippocalcin expression during ischemic injury (Park et al., 2020). Moreover, quercetin alleviated the increasement of protein tyrosine and serine/threonine phosphatase activity, along with the reduction of ERK and Akt phosphorylation in cerebral I/R injury . In addition, iso-quercetin protected against oxidative stress and neuronal apoptosis via Nrf2-mediated inhibition of the NOX4/ROS/NF-κB signaling pathway in cerebral ischemic stroke .

Calycosin
Calycosin ( Figure 2H), an isoflavone phytoestrogen isolated from Astragalus membranaceus, is found to possess various potential pharmacological activities, such as, anti-tumorigenesis, antioxidation, anti-virus and apoptosis-modulation effects Wang and Zhao, 2016;Zhao, Y. et al., 2016). It was also demonstrated that calycosin 30 mg/kg could ameliorate both the neurological deficit and infarct volume in experimental cerebral ischemia reperfusion injury, and the mechanism might be attributed to its antioxidant effects (Guo et al., 2012). Post-stroke calycosin 30 mg/kg therapy increased brain-derived neurotrophic factor (BDNF)/TrkB to ameliorate the neurological injury due to switching the microglia from the activated ameboid state to the resting ramified state in ischemic stroke rats (Hsu et al., 2020). Calycosin exhibited a downregulation of dexamethasone-induced Ras-related protein 1 (RASD1), and an upregulation of estrogen receptor (ER)-α and Bcl-2 . A study by Wang et al. found that calycosin pretreatment for 14 days dramatically upregulated p62, neighbor of BRCA1 gene 1 (NBR1) and Bcl-2, and downregulated TNF-α to ameliorate neurological scores in cerebral I/R rats .

Neuroprotective Role of Alkaloids in Ischemic Brain Injury
Alkaloids are rated as among the most-investigated plant secondary metabolites with versatile biological activity (Qiu et al., 2014). Recent investigations have pointed out the dynamic role of alkaloids as antibacterial, antidepressant, antioxidant, anti-inflammation (Perviz et al., 2016;Liu et al., 2020;Rosales et al., 2020). Accordingly, alkaloids are emerging as pharmaceutical tools for neuroprotective effects . Alkaloids were investigated on neuroprotective effects including berberine, ligustrazine, tetrahydropalmatine and others.

Berberine
Berberine ( Figure 3A) is an isoquinoline alkaloid isolated from traditional herbal medicine. It is widely present in the roots, rhizomes, stems or bark of various natural plants, such as Coptis chinensis Franch., Phellodendron chinense Schneid., etc. These plants have been used for thousands of years (Wang et al., 2019a;Fan et al., 2019). Studies have shown that berberine has a variety of pharmacological activities, including anti-microbial, antitumor, lowering glucose, lowering cholesterol, antiinflammatory, antioxidant, immunomodulatory and neuroprotective effects (Asemi et al., 2020;Song et al., 2020). So far, the neuroprotective effect of berberine has been involved (Lin and Zhang 2018). Berberine could improve the survival rate in OGD/R cells and improve the neurological deficiency in cerebral ischemic injury, which was involved in reducing the expression of cleave caspase 3 and up-regulating the expression of p-Bad to inhibit cell apoptosis via TrkB-PI3K/Akt pathway to promote neuron growth Chai et al., 2013;Yang et al., 2018a). Other studies also showed berberine 40 mg/kg preconditioning had neuroprotective effects by promoting autophagy, reducing cell apoptosis via PI3K/Akt signaling pathway (Zhang et al., 2016a;Zhang et al., 2016b). In addition, berberine exerted the neuroprotective effects by regulating I/R-induced peripheral lymphocytes early immunoactivation (Song et al., 2012). A study by Zhang et al. found that neuroprotection of early and short-time applying berberine could upregulate p-Akt, p-GSK 3β and phosphor cAMP response element binding protein (p-CREB) and downregulate NF-κB expression to ameliorate BBB permeability in the acute phase of cerebral ischemia . studied and had anti-inflammatory, anti-oxidant and analgesic effect (Kao et al., 2006;Zou et al., 2018). Tetramethylpyrazine treatment with 20 mg/kg suppressed hypoxia inducible factor-1alpha (HIF-1α), TNF-α, and activated caspase-3 expression in MCAO-induced brain ischemia in rats (Chang et al., 2007). Tetramethylpyrazine 20 mg/kg inhibited neutrophil activation via elevating nuclear related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) expression and inhibiting high-mobility group box-1 protein (HMGB1)/toll-like receptor-4 (TLR4), Akt, and ERK pathway following cerebral ischemia rats (Chang et al., 2015). Tetramethylpyrazine analogues were also investigated to induce p-eNOS by activation of PI3K/Akt and TLR-4/NF-κB signaling pathway and regulate the expression of tight junction (TJ) proteins, matrix metalloprotein 9 (MMP-9) and aquaporin 4 (AQP4) under cerebral I/R injury Xu et al., 2017;Zhou et al., 2019). The similar finds of PI3K/ Akt pathway also were shown in another study . What's more, the synergistic effect of drugs has also been shown to have good therapeutic effect. Ligustrazine accompanied with borneol could attenuate I/R injury by downregulating IL-1β, IL-6, and TNF-α, upregulating SOD, GSH-Px and regulating apoptosis and autophagy (Yu, B. et al., 2017;Yu et al., 2016).

Daurinoline
Daurinoline ( Figure 3C) is a kind of dibenzyl tetrahydroisoquinoline alkaloid, isolated from menispermum dauricum DC., which has a variety of pharmacological activities, including neuroprotective effect . Daurinoline 10 mg/kg and 20 mg/kg could enhance of SOD activity and the ability of scavenging oxygen free radicals (Lan and Lianjun 2020). Daurinoline reduced the level of MDA in brain tissue and increase the activity of SOD to prevent and ameliorated energy metabolism disturbance of cortex and mitochondria of brain induced by repeatedly cerebral ischemia reperfusion (Li and Gong 2004). In addition, daurinoline 5 and 10 mg/kg could inhibit cytochrome c (Cyt-C) release and caspase-3 and caspase-9 to inhibit neuronal cells apoptosis in the penumbra after cerebral ischemia (Yang et al., 2009).

Levo-tetrahydropalmatine
Levo-tetrahydropalmatine ( Figure 3D) are a series of alkaloids, isolated from a Chinese analgesic medicine, called Corydalis yanhusuo W. T. Wang. L-Tetrahydropalmatine, one of its main active ingredients, has been demonstrated to have potent analgesic effects and has been used in Chinese clinical practice for this purpose for many years (Chu et al., 2008;Han et al., 2012;Kang et al., 2016). Recent work showed that Ltetrahydropalmatine suppressed the central excitatory effects of amphetamine (Chang and Lin 2001), and played a protective role against neuronal injury in animal models (Liu et al., 2005;Mantsch et al., 2007;Chen et al., 2012). Levotetrahydropalmatine could regulate Src and c-Abl expression to inhibit neuron apoptosis and attenuate BBB injury in cerebral ischemic animals (Mao et al., 2015;Sun, J. B et al., 2018).

Peimine
Peimine ( Figure 3E) is a major biologically active component of Fritillaria ussuriensis. Recent studies have shown that isosteroidal alkaloids in Fritillaria had a wide range of pharmacological activities besides its role in cough remedies Li et al., 2020a;Zhang et al., 2020a;Zhao et al., 2018). It's also reported that peimine had protective effects on cerebral I/R injury in rats. The mechanism of neuroprotective effect might be related to inhibition of apoptosis, oxidative stress and inflammatory response, reduction of pathological damage and neurological dysfunction through regulation of the PI3K/Akt/mTOR pathway Duan et al., 2020).

Neuroprotective Role of Polyphenols and Acids in Ischemic Brain Injury
Polyphenols, a bioactive compound, were found abundant in vegetables, fruits and medical plant. Neuroprotection of polyphenols in medical plants is getting attention in the world. Curcumin, ellagic acid, resveratrol has been extensively studied and show multi-function. They are neuroprotectants, antioxidants, anti-inflammatory and antithrombic agents. It's reported that the neuroprotective efficacy of these compounds was involved in mitigating brain infarction and global ischemia, improving cerebral blood circulation (Lin 2011).

Ellagic Acid
Ellagic acid ( Figure 3H), an important cell protective and antioxidant compound, is a low molecular weight polyphenol derived from several fruits, vegetables and nuts (De Oliveira, 2016). Ellagic acid are recently more taken into accounts since their promising pharmacological effects, such as, antiinflammation, anti-oxidant and anti-cancer (Ceci et al., 2018;Baradaran Rahimi et al., 2020). In recent study, ellagic acid is investigated as a potential endowed with multi-target pharmacological properties on central nervous system (Alfei et al., 2019). BCCAO was carried out to make global cerebral ischemic injury. Ellagic acid pretreatment with 100 mg/kg could reduce MDA level and restore the heart rate to normal level (Nejad et al., 2015). In addition, ellagic acid 100 mg/kg could activate PI3K/Akt/NOS pathway to alleviate brain injury and protect brain tissue in MCAO model (Kaihua and Jiyu 2020). Furthermore, ellagic acid administration with 10-90 mg/kg could improve brain injury outcomes and increase the proliferation of NSCs through the Wnt/β-catenin signaling pathway (Liu, Q. S. et al., 2017). The upregulation of zonula occludens-1 (ZO-1) and down-regulation of AQP 4 and MMP-9 in injured brain tissues after being treated ellagic acid 10-50 mg/kg (Wang et al., 2019b).

Neuroprotective Role of Glycosides in Ischemic Brain Injury
Asiaticoside Asiaticoside ( Figure 4A) is isolated from Centella asiatica (L.), which has been using as a memory enhancing and psychoactive drug for a long time in Asia (Zheng and Qin 2007). Many studies have shown that C. asiatica indeed has many biological activities in central nervous system (Mook-Jung et al., 1999;Zheng and Qin 2007;Dhanasekaran et al., 2009;Orhan 2012). A report demonstrated asiaticoside showed anti-inflammation effect via inhibiting overactivation of p38 MAPK pathway against cerebral I/R mice . Another study found asiaticoside showed a protective effect against cerebral I/R injury via the NOD2/MAPK/NF-kB signaling pathway (Zhang et al., 2020b).

Salidroside
Salidroside ( Figure 4B), the major phenylpropanoid glycoside extract from medicinal Tibetan plant Rhodiola rosea L, has diverse pharmacological activities. Many recent reports and reviews have highlighted that salidroside may exert antiinflammatory, neuroprotective effects and improve cognitive function (Li, M. et al., 2018;Xu, N. et al., 2019). Salidroside may involve the modulation of monoamine metabolism in the striatum and substantia nigra pars compacta (SNpc), which may be related to the function of the dopaminergic system in the cerebral I/R brain . Many studies reported salidroside protected against cerebral I/R injury via PI3K/Akt pathway, accompanying with Nrf2/NF-κB pathway (Wei et al., 2017;Zhang, J. et al., 2019;Zuo et al., 2018). Salidroside exhibited neuroprotective effects against hypoxia/reperfusion injury by activating the SIRT1/FOXO3α pathway (Xu, L. et al., 2018). In addition, salidroside is an effective treatment for ischemic stroke that functions via the fibroblast growth factor 2 (FGF2)-mediated cAMP/PKA/CREB pathway to promote dendritic and synaptic plasticity . Salidroside treatment reduced the expression of M1 microglia/macrophage markers and increased the exdslbpression of M2 microglia/macrophage markers after stroke and induced primary microglia from M1 phenotype to M2 phenotype . What's more, salidroside reduced the markers of endothelial activation and neutrophilic infiltration after I/R injury by inhibition of complement, restoring an anti-inflammatory endothelial phenotype after oxidative stress and inhibiting classical complement activation, in association with anti-apoptotic effects .

Neuroprotective Role of Iridoid Glycosides in Ischemic Brain Injury
Iridoid glycosides are special glycosides. Iridoid glycoside exists broadly in plants of many families and has a wide variety of biological activities including purgative, liver protective, antimicrobial, analgesic, antitumor, sedative and antiinflammatory activities (Ismailoglu et al., 2002). Geniposide, Catalpol and Picroside II were investigated in depth as iridoid glycosides, which had a neuroprotective effect.

Catalpol
Catalpol ( Figure 4D) is the main active component of the radix from Rehmannia glutinosa Libosch, and it belongs to the iridoid monosaccharide glycoside family (Ismailoglu et al., 2002;Zhang et al., 2008), which has pleiotropic protective effects on many diseases, including neurodegenerative diseases (Xia et al., 2012), ischemic stroke (Zhu et al., 2010), metabolic disorders (Zhu et al., 2010) and others. It's reported that the efficacy of catalpol pretreatment on cerebral I/R injury may be attributed to reduction of free radicals and inhibition of lipid peroxidation and endothelin-1 (ET-1) production . Additionally, a study by Li et al. found catalpol also exerted the most significant cytoprotective effect on astrocytes by suppressing the production of free radicals and elevating antioxidant capacity . What's more, catalpol significantly inhibited apoptosis by modulating Bcl-2 and Bax (Li et al., 2006). Catalpol affected angiogenesis via the JAK2/ STAT3 signaling pathway and VEGF expression (Dong, W et al., 2016).

Picroside II
Picrorhiza scrophulariflora belongs to the plant family composed of picroside I, II and III, of which picroside II ( Figure 4E) is one of the most effective components extracted from the dried rhizome and roots of Picrorhiza kurrooa Royle ex Benth and Picrorhiza scrophulariae flora Pennell (Stuppner and Wagner 1989;Wang et al., 1993). Current researches on picroside II are focused on its neuroprotective, anti-apoptotic, anti-cholestatic, antioxidant, anti-inflammation, immunomodulating activities (Cao et al., 2007;Li et al., 2007;He et al., 2009). It has been confirmed that picroside II 25 mg/ml could enhance nerve growth factor -induced PC12 cell axon growth, reduce H 2 O 2 induced PC12 cell damage and improve cell survival in vitro (Cao et al., 2007). Picroside II attenuated cerebral I/R injury via inhibiting apoptosis and inflammation, included COX2, TLR4/NF-κB and MEK-ERK1/2 pathway Wang, F. et al., 2015;Wang, L. Y. et al., 2015). Picroside II could protect BBB possibly through reducing oxidative stress factors (ROS, NOX2 ROCK, MLCK, and MMP-2) and enhancing BBB function factors, claudin-5 . Furthermore, picroside II exerted a neuroprotective effect by inhibiting the mitochondria Cyt C signal pathway and decreasing the permeability of mitochondrial permeability transition pore (mPTP) following I/R injury in rats .

Neuroprotective Role of Saponin in Ischemic Brain Injury
Ginsenoside Rg1 ( Figure 5A) is the representative components in saponin. Ginsenoside Rg1 is one of the main active ingredients of ginseng (Zhang and Zhao 2014;Chuang et al., 2015). It has been shown that as a small molecular substance, ginsenoside Rg1 easily passes through the blood brain barrier. Moreover, ginsenoside Rg1 could promote stem cell orientation transformation, induce stem cell proliferation and played a neuroprotective role in brain repair (Cheng et al., 2005;Tang et al., 2017;Xie, C. J. et al., 2018). It's reported that ginsenoside Rg1 could relieve the I/R injury through multiple pathways. Ginsenoside Rg1 could improve neurological injury, regulate BBB disruption and permeability and downregulate AQP 4 and protease-activated receptor-1 (PAR 1) (Zhou et al., 2014;Xie et al., 2015). Ginsenoside Rg1 alleviated oxidative stress after I/R through inhibiting miR-144 activity and subsequently promoting the Nrf2/ARE pathway . What's more, a study by Li et al. found that ginsenoside Rg1 may exert its neuroprotective action on cerebral I/R injury through the activation of PPARγ signaling . In addition, it's also shown that ginsenoside Rg1 treatment obviously decreased cell apoptosis, while the transplanted cells could be differentiated into neurons and glial cells, which also improved cerebral ischemia (Bao et al., 2015).
Other studies showed that ginsenoside Rg1 40 mg/kg was attributed to a decrease in ubiquitinated aggregates and a suppression of the inflammatory response and increased in the expression of BDNF in the hippocampal CA1 region after I/R insult Zheng et al., 2019).

Ginkgolides
Ginkgolide B ( Figure 5D) were one of ten kinds of diterpene lactone compounds, which were isolated from Ginkgo biloba leaves. It's reported that exact of Ginkgo biloba had a broad range of pharmacological effects, such as anti-inflammation, antioxidant, anti-depressant Hu et al., 2018;Jin, G et al., 2014;Ran et al., 2014;Saini et al., 2014;Wan et al., 2016). Ginkgolides B upregulated the levels of antioxidant proteins through Akt/Nrf2 pathway to protect neurons from oxidative stress injury . In addition, ginkgolide B improved neurological function by promoting the proliferation and differentiation of neural stem cells in rats with cerebral I/R injury (Zheng et al., 2018b).

Borneol
Borneol ( Figure 5E) is a terpene and bicyclic organic compound, a resin from Cinnamomum camphora (L.) Presl, a traditional Chinese medicine Zheng et al., 2018a). As a traditional Chinese medicine, borneol has been used for thousands of years in the treatment of cardiovascular and cerebrovascular diseases. Modern pharmacological studies have shown that borneol had anti-inflammatory, antioxidant stress, anti-apoptosis and other pharmacological effects (Almeida et al., 2013). Borneol protected against cerebral I/R injury through multifunctional cytoprotective pathways, involving in the alleviation of intracellular ROS and iNOS/NO pathway, inhibition of inflammatory factor and depression of caspase-related apoptosis. Additionally, the inhibition of IκBα/ NF-κB pathway might play a significant role in the neuroprotection of borneol (Liu et al., 2011). Another study by Dong et al. found that borneol could alleviate cerebral ischemic injury, most likely executed via anti-apoptosis and anti-inflammation effects and maintenance of the BBB stability and TJs to comprehensively improve NVU function . Additionally, the protection of OGD induced BMECs by tetramethylpyrazine phosphate and borneol combination involved anti-oxidation, apoptosis inhibition, and angiogenesis .

NEUROPROTECTIVE ROLE OF OTHER COMPOUNDS IN ISCHEMIC BRAIN INJURY Emodin
Emodin ( Figure 5F), 1,3,8-trihydroxy-6-methylanthraquinone, is a naturally occurring anthraquinone derivative and an active component from Polygonum multiflorum Thunb. Rheum palmatum L. etc, which have been used widely in Asia in treatment of multiple diseases (Dong, X. et al., 2016). Emodin has been demonstrated to possess a wide spectrum of pharmacological effects, such as anti-viral, anti-bacterial, antiallergic, anti-osteoporotic, immunosuppressive, neuroprotective activities (Dong, W. et al., 2016;Leung et al., 2020;Xue et al., 2020). In fact, the neuroprotective effect of Polygonum multiflorum Thunb was first published in 2000 (Gu et al., 2000) and the neuroprotective effect of emodin was published in 2005 when its ability to interfere with the release of glutamate was identified as a method of neuroprotection (Gu et al., 2005). Additionally, emodin might afford a significant neuroprotective effect against glutamate-induced apoptosis through the critical role including Bcl-2/Bax, active caspase-3, p-Akt, p-CREB, and mature BDNF for potent neuroprotective effects of emodin to subsequently enhance behavioral function in cerebral ischemia (Ahn et al., 2016). Another study by Leung et al. found emodin had neuroprotective effects against I/R or OGD injury both in vitro and in vivo, which may be increase Bcl-2 and glutamate transporter-1 (GLT-l) expression but suppress activated-caspase 3 levels through activating ERK1/2 pathway (Leung et al., 2020).

Cordycepin
Cordycepin ( Figure 5G) is a derivative of nucleoside adenosine and one of the main active components of Cordyceps sinensis, which is mainly distributed in Europe, North America and Asia (Tuli et al., 2013;. Cordycepin has a variety of pharmacological activities, such as anti-inflammatory, antitumor, antioxidant, anti-inflammatory microorganism, antihyperlipidemia, anti-hepatotoxicity, anti-depressant and neuroprotective activities (Ryu et al., 2014;Wang et al., 2015a;Qin et al., 2019). It was shown that cordycepin had an important neuroprotective effect on hypoxic injury by improving the electrophysiological function of neurons (Chen, K. et al., 2017; B et althogenesis of lesions . It is confirmed that cordycepin could significantly reduce glutamic acid and aspartic acid, increase SOD activity and down-regulate MDA and MMP-3 to alleviate cerebral ischemic injury or hypoxia injury, suggesting cordycepin could inhibit oxidative damage to exert neuroprotection (Hwang et al., 2008;Cheng et al., 2011). Cordycepin regulated adenosine A1 receptor to improve long-term potentiation formation and neuronal survival through p38/JNK/ERK pathway in BCCAO model and glutamate-induced HT22 neuronal cell death (Dong et al., 2019;Jin, M. L et al., 2014).

Polysaccharides
Polysaccharides are considered to have a wide range of pharmacological effects, such as scavenging free radicals, immune regulation, anti-tumor, anti-oxidation, anti-viral, antiinflammatory, lowering blood sugar, anti-depression, liver protection, etc (Jin et al., 2012;Kwok et al., 2019;Fang et al., 2020). Panax notoginseng polysaccharide is a kind of heteroglycan derived from the medicinal plant Panax notoginseng, which could increase the ratio of Bcl-2/Bax and reduce caspase-3 in cerebral ischemic brain tissue (Jia et al., 2014). What's more, it could enhance GSH-Px, SOD activity and IL 10 level, while downregulate MDA, TNF-α, IL-1β level to reduce cerebral infarction size and cell apoptosis to afford neuroprotective effect (Jia et al., 2014;Sy et al., 2015). Angelica polysaccharide is the main active ingredient of Angelica sinensis (Oliv.) Diels, which could also enhance the activities of SOD, GSH and GSH-PX, and reduce MDA, IL-1β, TNF-α and NF-κB in cerebral ischemia-reperfusion injury rats (Yan and Xie, 2018;Dai et al., 2020), in addition, enhance angiopoietin 1, angiopoietin 2, VEGF to promote angiogenesis (Hu et al., 2006;Li, J. et al., 2019). Ginkgo biloba polysaccharides could also play a neuroprotective effect by inhibiting oxidative stress and inflammation to improve neurological deficits in cerebral ischemic animals (Yang et al., 2013). Black fungus polysaccharide is the main active ingredient of Auricularia auricula (L.ex Hook.) Underw in China. Studies have found that fungus polysaccharides could reduce the production of ROS, MDA, and NO in rats with acute or chronic cerebral ischemia injury, enhance the activity of SOD and played an anti-oxidant effect. In addition, it can inhibit neuronal apoptosis and improve cerebral ischemia-reperfusion injury. Qian and Min 2010;Ye et al., 2010). Fucose is a marine sulfated polysaccharide extracted from brown algae and some marine invertebrates (Apostolova et al., 2020;Oliveira et al., 2020). Experimental studies have shown that fucose can significantly reduce the levels of pro-inflammatory factors IL-1β, IL-6, MPO and TNF-α in cerebral ischemia model animals, reduce the levels of oxidative stress-related proteins SOD and MDA, and could also regulate cell apoptosis via inhibiting the MAPK pathway to play a neuroprotective effect in cerebral ischemia reperfusion study (Che et al., 2017). Other studies have shown that fucose could also play a neuroprotective effect by inhibiting neuronal apoptosis, glial cell activation and anti-oxidative stress properties Kim et al., 2019). Codonopsis pilosula polysaccharide is a kind of polysaccharide extracted from Codonopsis pilosula. Studies have shown that Codonopsis pilosula polysaccharide could reduce LDH, NO and MDA, increase the content of AChE, SOD and GSH-Px, and inhibit the expression of Beclin-1 to regulate autophagy in the cerebral ischemia-reperfusion injury model Ma et al., 2019). It could also inhibit the expression of GFAP protein . Codonopsis pilosula polysaccharides can also reduce the expression of Bax, upregulate the expression of Bcl-2 and inhibit apoptosis to exert Frontiers in Pharmacology | www.frontiersin.org April 2021 | Volume 12 | Article 607412 neuroprotective effects through mediating the Nrf2/HO-1 pathway . Lycium barbarum polysaccharide, astragalus polysaccharide, aloe polysaccharide, yam polysaccharide, etc. also have a certain protective effect on cerebral ischemia (Ge et al., 2017;Lu et al., 2012;Peng et al., 2019;Wang, Y. et al., 2014;Yan and Zhou, 2012).

Structure-Activity Relationship Between Natural Products and Neuroprotection
Many studies have confirmed the neuroprotective effect of natural products on cerebral ischemia and clarified its preliminary mechanism of action. Different natural products have different mechanisms of anti-cerebral ischemia, which are closely related to the different structures of the compounds. Figure 6 shows the mechanism of cerebral ischemia and the role of different types of natural products in this process. The research on the structure-activity relationship of the biological activity of natural products is mainly related to the basic nucleus of the compound, the number and position of double bonds, and the position of functional groups. They have a great influence on the biological activity of the compound (Aljahdali et al., 2020;Zhou et al., 2021). The flavonoids are based on 2phenylchromone as the skeleton, which has ortho-dihydroxyl in the structure. The hydroxyl substituent on the basic skeleton is the key functional group for scavenging oxygen free radicals. At the same time, a conjugated system is also the key active site to scavenging oxygen free radicals . Therefore, flavonoids have unique advantages in scavenging free radicals and inhibiting lipid peroxidation. In addition, flavonoids had antiinflammatory effects. It is believed that when the C-5 and 7 positions of the A ring of flavonoids have hydroxyl groups at the same time, they will affect the secretion process, mitosis and cell interactions of cells, and then produce strong anti-inflammatory effect (Ying and Chi, 2007).
Alkaloids are quite different in structure. For example, berberine has a methylenedioxy ring structure and is the main functional group for anti-bacterial and hypoglycemic activity Wu and Fang, 2020). This may also be the key structure of its antioxidant effect, which needs further research to confirm. Ligustrazine has a simple structure and a wide range of pharmacological effects. Use it as a lead compound to synthesize various derivatives, which has a variety of biological activities , and the derivative is more stable than ligustrazine, such as ligustrazine hydrochloride injection has better curative effect in the treatment of ischemic cerebrovascular disease You et al., 2020). Dauricine is a kind of bisbenzyltetrahydro isoquinoline alkaloid. It has a phenolic hydroxyl group and a conjugated system that is easy to form hydrogen bonds. It may be similar to flavonoids and is considered to be a key structure for antioxidant effects. Studies have shown that phenolic acids have a strong antioxidant effect, which is due to the existence of carboxyl, carbonyl and other structures (Bialecka-Florjanczyk et al., 2018;. Glycoside components are compounds formed by non-sugar substances and sugars or sugar derivatives, which have the active characteristics both sugar substances and non-sugar substances. For example, flavonoid glycosides and flavonoid aglycones have anti-oxidative biological activity (Xue et al., 2001;Gao et al., 2008;Shao et al., 2013). In addition, most studies believe that glycoside compounds mainly exerted neuroprotective effects by improving the body's immunity (Brito-Arias, 2007). Iridoid glycosides are a special glycoside chemical. The core of iridoid glycosides is hemiacetal-enether cyclopentane, which is an important structural basis. Antioxidant stress is the manifestation of its biological activity (Wang et al., 2019a). Some studies also believe that the double bond on the cyclopentane in iridoid glycosides, the C-11 substituent, and the way of forming the bond after ring opening also have important effects on the anti-inflammatory activity (Mao et al., 2019). For example, loganin, morroniside, and sorgenin all contain several electron-donating groupshydroxyl groups. The hydroxyl groups release hydrogen atoms and combine with free radicals to block the chain reaction mediated by reactive oxygen species and reduce the body's oxidative stress. Stimulus levels, which weaken the phosphorylation of inflammatory signal pathway related factors, thereby inhibiting the inflammatory response, improving insulin resistance and liver damage . Studies believe that tetracyclic triterpene saponins are the leading compounds for the treatment of neurological diseases, and can show strong neurological activity after proper structural modification and modification. Among them, C-7 and C-9 of lanolin triterpenoids The diene bond connected at the position may be the key group for nerve extension promoting activity . The research on the structure-activity relationship of tetracyclic triterpenes is mainly focused on the study of the binding sites related to the action of NMDAR antagonists, indicating that the structural modification and optimization of its aglycon and side chain sugar groups can obtain a new type of neuroactive monomer with high efficiency and low toxicity drug. Tetracyclic triterpene dammarane compounds as NMDAR antagonists will be an effective way to treat neurological diseases (Lee et al., 2006;Ryoo et al., 2020). Studies believe that ginkgolides can improve cerebral blood circulation and energy metabolism through specific inhibition of PAFR, and show a good market prospect in the prevention and treatment of cardiovascular and cerebrovascular diseases. The main differences in their effects are: the number and positions of hydroxyl groups contained in ginkgolides are different, PAF and ginkgolide ligands can be combined, and the introduction of benzyl derivatives at position C-10 is more beneficial for antagonistic activity; In addition, the presence of C, D ring and tert-butyl group also plays an important role in activity (Hui et al., 2013). Some scholars have studied the structureactivity relationship of andrographolide and found that the position of the double bond determines the anti-inflammatory effect of andrographolide and its derivatives, and the antiinflammatory effects of compounds with double bonds in the ring are stronger than those with double bonds outside the ring (Tian et al., 2015). As a natural component of polycarbonyl and polyhydroxyl groups, emodin can coordinate and chelate with the zinc ions of type IV collagenase to inactivate type IV collagenase, thereby preventing type IV collagenase (MMP-2, MMP-9) degradation of basement membrane and extracellular matrix of normal tissue cells, play a role in anti-invasion and antimetastasis (Wang, 2006). As a nucleoside antibiotic, cordycepin has a good application prospect in regulating immunity and scavenging free radicals (Lee et al., 2020).
In summary, the activity of natural products has an important relationship with the structure of the substance itself. The active functional groups of natural products are the key to their functions. Moreover, the bioavailability of many natural products is very low, which limits their use, the structural modifications, changes in dosage forms or improvements in pharmacokinetic parameters can improve their bioavailability. Because the pathogenesis of cerebral ischemia is complex, and cascade reactions occur cross-over. The anti-cerebral ischemia effects of individual natural products are often weak. Therefore, a full understanding of the mechanism of the cerebral ischemia cascade can provide the possibility for the development of specific mechanisms or targeted compounds, and the combined use of different natural products to play a multi-target neuroprotective effect may be a promising therapy. However, it is very important to combine animal models with clinical practice . Choose appropriate experimental models according to actual conditions to provide a solid foundation for the clinical transformation of natural products.
In conclusion, this review summarizes that the facts are comprehensive and deeply informative about the neuroprotective activities of natural products in vitro and in vivo experiment. For prophylactic and therapeutic management of stroke, they are promising candidates. Therefore, this review would provide as a reference for current advances in the study on natural products for neuroprotection.

AUTHOR CONTRIBUTIONS
QX and HL are equally contributed to the article and supervised as a whole and finalized the manuscript. DL, JY, and DG carried out various literature survey studies. RM, JL, and JW prepared the initial draft and graphical representation for figures. MR, HC and YL provided significant input into the chemistry part.   Ma et al., 2019; Frontiers in Pharmacology | www.frontiersin.org April 2021 | Volume 12 | Article 607412