Biotransformation, Pharmacokinetics, and Pharmacological Activities of Ginsenoside Rd Against Multiple Diseases.

Panax ginseng C.A. Mey. has a history of more than 4000 years and is widely used in Asian countries. Modern pharmacological studies have proved that ginsenosides and their compounds have a variety of significant biological activities on specific diseases, including neurodegenerative diseases, certain types of cancer, gastrointestinal disease, and metabolic diseases, in which most of the interest has focused on ginsenoside Rd. The evidentiary basis showed that ginsenoside Rd ameliorates ischemic stroke, nerve injury, cancer, and other diseases involved in apoptosis, inflammation, oxidative stress, mitochondrial damage, and autophagy. In this review, we summarized available reports on the molecular biological mechanisms of ginsenoside Rd in neurological diseases, cancer, metabolic diseases, and other diseases. We also discussed the main biotransformation pathways of ginsenoside Rd obtained by fermentation.

Highlights

1) Approximately 120 studies on the use of ginsenoside Rd for the treatment of multiple diseases have been published.

2) This is the first review to report about the biotransformation, pharmacokinetics, and pharmacological effects of ginsenoside Rd.

3) The potential pharmacological mechanisms of ginsenoside Rd have been documented.

4) No specific reviews have been conducted by now.

Introduction

Panax ginseng C.A. Mey. is a well-known herbal medicine widely used in China, Korea, Japan, and other East Asian countries. At present, the ginseng root and its extract are the most widely used herbal medicine. Modern pharmacological studies have proved that ginsenosides are the main active ingredient of ginseng and have a wide range of pharmacological effects, such as anti-inflammatory (Xu et al., 2021; Yi, 2021), anticancer (Zhang et al., 2021a), and anti-viral (Kang et al., 2021), regulate immunity (Kang et al., 2021), metabolism (Wang et al., 2021a), and improve cardiovascular system (Wang et al., 2021b; Sarhene et al., 2021) and nervous system (Brioschi Guevara et al., 2021) function, whereas most attention has been focused on the ginsenoside Rd.

Ginsenoside Rd, a natural compound extracted from the root of Panax ginseng C.A. Mey., is one of the protopanaxadiol (PPD)-type ginsenosides, while the proportion of ginsenoside Rd in ginseng is very low (Liu et al., 2020a). Interestingly, the promising effects of the pretreatment and treatment of ginsenoside Rd on neurological diseases, cancer, gastrointestinal disease, and metabolic diseases have been studied extensively in in vivo and in vitro models (Guo et al., 2021; Chen et al., 2022; Zhou et al., 2022).

Existing studies related to ginsenoside Rd have shown that various ginsenosides, such as Rb1, Rb2, and Rc, can be transformed into ginsenoside Rd after absorption and metabolism in vivo (Park et al., 2010; Shin and Oh, 2016). In addition, Rd can be prepared in a variety of ways based on the in-depth study of biotransformation and the development of modern fermentation technology (He et al., 2019). Based on the above results, we summarized the biotransformation process of other ginsenosides into Rd, thereby hoping to play a positive role in the large-scale industrial production of Rd. In this study, the biotransformation sources, pharmacokinetics, pharmacological effects, and molecular mechanisms of ginsenoside Rd on various systemic diseases in recent years were reviewed, and their therapeutic potential was discussed.

Biotransformation of Ginsenoside Rd

Multiple studies have confirmed that ginsenosides can be transformed into ginsenoside Rd using enzymes and bacterial communities and can promote the transformation of ginsenoside Rd into other metabolites (He et al., 2019). We summarized the precursors, metabolites, and transformation conditions of ginsenoside Rd (Table 1) (Figure 1).

TABLE 1

Summary of the biotransformation of ginsenoside Rd.

References	Conversion	Source	Enzyme	Optimal conditions	Conversion ratio (%)
Akter and Huq, (2018)	Rb1 to Rd	Paenibacillus	MAH-16T	pH 5.0-7.0, 20-40°C
Fang et al. (2020)	Rb1 to Rd		Pectinase	pH 6, 52.5°C	46.15
Renchinkhand et al. (2020)	Rb1 to Rd	Dekkera anomala YAE-1	β-glucosidase	pH 5.0, 40°C, 48 h
Renchinkhand et al. (2017)	Rb1 to Rd	Paenibacillus sp. MBT213	β-glucosidase	pH7.0, 35°C, 14 days
Hong et al. (2012)	Rb1 to Rd	Flavobacterium johnsoniae	β-glucosidase	pH 6.0, 37°C
Quan et al. (2011)	Rb1 to XVII、Rd to F2 to CK	Leuconostoc mesenteroides DC102	Glucosidase	pH6.0–8.0, 30°C, 72 h
Zhong et al. (2016)	Rb1 to Rd	Lactobacillus brevis	β-glucosidase	pH 7.0, 30°C	69
Feng et al. (2016)	Rb1 to Rd	Aspergillus niger	TH-10a	pH 5.0, 32°C, 48 h	86
Ye et al. (2010)	Rb1 to Rd	Paecilomyces bainier 229-7	β-glucosidase	pH5.0, 28°C, 72 h	89-94.9
Ye et al. (2012)	Rb1 to Rd	Paecilomyces bainier 229-7	External calcium regulated β-glucosidase	pH5.0, 28°C, 72 h	92.44
Son et al. (2008)	Rb1 to Rd	Thermus caldophilus	β-glucosidase	pH 5.0, 75°C, 18 h	80
Kim et al. (2013a)	Rb1 to Rd	Microbacterium trichothecenolyticum	M.trichothecenolyticum KCTC 19343	30°C, 24 h
Zhao et al. (2009)	Rb1 to Rd	Cladosporium fulvum	β-glucosidase	pH 5.0, 37°C, 8 days	86
Lin et al. (2015)	Rb1 to Rd	Aspergillus versicolor LFJ1403	β-glucosidase	PH 5.0, 30°C, 96 h	85
Shin et al. (2013)	Rc to Rd	Caldicellulosiruptor saccharolyticus DSM 8903	α-L-arabinofuranosidase	pH 5.5, 80°C, 30 min	100
Xie et al. (2016a)	Rc to Rd	Thermotoga thermarum DSM5069	α-L-arabinofuranosidase	pH 5.0, 85°C, 60 min	99.4
Liu et al. (2013)	Rc to Rd	Leuconostoc sp. strain 22-3	α-L-arabinofuranosidase	pH 6.0, 30°C, 20 min
Zhang et al. (2021b)	Rc to Rd	Bacillus subtilis Str. 168	α-L-arabinofuranosidase	pH 5, 40°C, 24 h	90
Kim et al. (2020)	Rb2 to Rd	Blastococcus saxobsidens	α-L-arabinopyranosidase	pH 7.0, 40°C, 1 h
Jung et al. (2014)	F2 to Rd	Ginseng UDP-glycosyltransferases	UDP-glycosyltransferases 94Q2

FIGURE 1

Biotransformation and pharmacokinetics of ginsenoside Rd in vivo.

Ginsenoside Rd can be synthesized from ginsenoside Rb1 by the hydrolysis of glucose at C-20 (Akter and Huq, 2018). The β-glucosidase produced by pectinase (Fang et al., 2020), Dekkera anomala YAE-1 (Renchinkhand et al., 2020), Paenibacillus sp. MBT213 (Renchinkhand et al., 2017), Flavobacterium johnsoniae (Hong et al., 2012), Leuconostoc mesenteroides DC102 (Quan et al., 2011), and Lactobacillus brevis (Zhong et al., 2016) is able to hydrolyze ginsenoside Rb1 (Rb1) and convert it to ginsenoside Rd during the fermentation of the ginseng. In addition, Aspergillus niger strain TH-10 (Feng et al., 2016), Paecilomyces bainier 229-7 (Ye et al., 2010; Ye et al., 2012), Thermus caldophilus GK24 (Son et al., 2008), Microbacterium trichothecenolyticum (Kim et al., 2013a), Cladosporium fulvum (Zhao et al., 2009), and Aspergillus versicolor (Lin et al., 2015) have shown similar effects as those of hydrolases in Rb1.

The α-L-arabinosidase (AbpBs) from Caldicellulosiruptor saccharolyticus (Shin et al., 2013), Thermotoga thermarum DSM 5069 (Xie et al., 2016a), Leuconostoc sp. 22-3 (Liu et al., 2013), and Bacillus subtilis (Zhang et al., 2021b) converts ginsenoside Rc (Rc) into ginsenoside Rd by attacking the C-20 position of α-linked arabinoside, thereby releasing arabinose (Liu et al., 2013; Zhang et al., 2021b). AbpBs can promote the biotransformation of ginsenoside Rb2 (Rb2) to ginsenoside Rd by attacking C-20, thereby releasing arabinoside (Kim et al., 2020). In addition, enzymes PgUGT74AE2 and PgUGT94Q2, which participate in ginsenoside biosynthesis, transfer two glucose groups from UDP-glucose (UDP-Glc) to the C3 hydroxyl group of ginsenoside compound K (CK) to form ginsenoside Rd (Jung et al., 2014).

β-glucosidase cleaves the glycoside at the C-3 position of ginsenoside Rd and produces the ginsenoside compound CK (Renchinkhand et al., 2020). Ginsenoside M1 is formed by the hydrolysis of the C-3 glucose group in ginsenoside Rd by snailase (Renchinkhand et al., 2017).

Pharmacokinetics

Intestinal flora can promote the metabolic transformation of ginseng extract and Rb1 into ginsenoside Rd in rats and can enter the blood for absorption in rats (Kim et al., 2014a). Ginsenoside Rd is distributed in various organs, with the highest content in the lungs, followed by the liver, kidney, heart, and intestine, and the lowest content in the brain (Sun et al., 2012). After taking urine 0–24 h after oral administration and intravenous administration, liquid chromatography-mass spectrometry (LC-MS) is used to confirm that oxidation and glycosylation (Yang et al., 2006a; Yang et al., 2007a) are the main metabolic pathways of ginsenoside Rd in rats. The absolute bioavailability of Rd in dogs is 0.26% (Wang et al., 2007). As in clinical trials, ginsenoside Rd shows linear pharmacokinetics, is well tolerated in the dose range of 10–75 mg after an intravenous administration, and is slowly cleared from plasma, and the elimination rate does not change after repeated administration (Zeng et al., 2010).

Ginsenoside Rd Targets Multiple Diseases

Ischemic Stroke

In ischemic stroke, ginsenoside Rd plays a neuroprotective role by restoring mitochondrial function, reducing neuronal apoptosis, and eliminating neuroinflammation (Figure 2). As for the therapeutic window study, ginsenoside Rd shows an obvious neuroprotective effect in the middle cerebral artery occlusion (MCAO) model (Ye et al., 2011a). Importantly, the results of a clinical trial showed that ginsenoside Rd has a positive effect on the prognosis of acute ischemic stroke (Liu et al., 2012).

FIGURE 2

Protective effect of ginsenoside Rd on ischemic stroke.

In Ca2+ influx and mitochondrial dysfunction, ginsenoside Rd, a potential Ca2+ channel blocker (Li et al., 2010), significantly reduces the burst of glutamate by increasing the expression of glutamate transporter-1 (GLT-1) and inhibits the channels of Ca2+ influx (Zhang et al., 2013a) to protect the rat hippocampal neurons (Zhang et al., 2012a). Similar to a calcineurin inhibitor, ginsenoside Rd exerts a neuroprotective effect by inhibiting the elevation of N-methyl-D-aspartate (NMDA) receptors and the hyperphosphorylation of the N-methyl-D-aspartate receptor 2B (NR2B) subunit in the MCAO model and oxygen–glucose deprivation (OGD) cultured neurons (Xie et al., 2016b; Zhang et al., 2020a). Ginsenoside Rd pretreatment exerts neuroprotective effects by inhibiting the Ca2+ overload and specificity attenuated the expression of transient receptor potential melastatin (TRPM) 7 and acid-sensing ion channel (ASIC) 1a while promoting ASIC2a expression following focal ischemia (Zhang et al., 2012b). Remarkably, the results of a clinical trial based on Ca2+ disorder and subsequent neurotoxicity induced by acuteischemic stroke, ginsenoside Rd can be considered a calcium channel antagonist and a neuroprotectant (Liu et al., 2009). As for mitochondrial dysfunction, ginsenoside Rd markedly protects the mitochondria, as indicated by regulating enzyme activity, reducing mitochondrial hydrogen peroxide production and depolarizing mitochondrial membrane potential (MMP), decreasing reactive oxygen species (ROS) production in isolated mitochondria from Sprague–Dawley (SD) rats (Ye et al., 2011b), and reducing the mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) damage and cell apoptosis in MCAO-induced ischemic stroke model (Hu et al., 2013; Yang et al., 2016). These findings are also confirmed in primary cultured hippocampal neuron cells (Ye et al., 2009). In addition, in elderly stroke mice, ginsenoside Rd can play an equivalent neuroprotective role in elderly transient focal ischemic mice by regulating lipid peroxide accumulation, mitochondrial complex activity, and MMP (Ye et al., 2011c).

As far as apoptosis is concerned, ginsenoside Rd may reduce cerebral ischemia-induced tau phosphorylation by decreasing the activity of glycogen synthase kinase-3β (GSK-3β) and enhancing the activity of protein kinase B (PKB/AKT) (Zhang et al., 2014). In PC12 cells with OGD/reperfusion (OGD/R) and SD rats with ischemia/reperfusion (I/R) injury, ginsenoside Rd significantly limits the expression of vascular endothelial growth factor (VEGF), brain-derived neurotrophic factor (BDNF), and the phosphatidylinositol 3-kinase (PI3K)/AKT and ERK1/2 pathways (Liu et al., 2015a). As a neuroprotective agent ginsenoside Rd also prevents trimethyltin (TMT)-induced neurotoxicity and significantly reduces neuronal loss in TMT-induced hippocampal dysfunction and active astrocytes via regulation of B-cell lymphoma-2 (Bcl-2), Bcl-2-like protein 4, and caspase-3 (Hou et al., 2017). Taken together, ginsenoside Rd has neuroprotective effects via mitogen-activated protein kinase (MAPK)/ERK-, PI3K/AKT, PI3K/AKT/GSK-3β, and ERK1/2-dependent pathways.

For inflammation, ginsenoside Rd inhibits ischemic stroke-indeced neuronal death and inflammation by inhibiting cleaved poly adenosine diphosphate-ribose polymerase-1(PARP-1) activity, levels of poly (ADP-ribose), sequential apoptosis-inducing factor (AIF) translocation, and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) nuclear accumulation (Hu et al., 2013). Postischemic syntheses of two damaging enzymes, cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), are also significantly inhibited by ginsenoside Rd treatment. Ginsenoside Rd reduces free radical generation during I/R and suppresses oxidative damage and inflammatory injury (Ye et al., 2011d). As a proteasome-related compound, ginsenoside Rd protects against MCAO-induced ischemic brain injury by inhibiting the proteasome activity and NF-κB/matrix metalloproteinase-9 (MMP-9) signal pathway (Zhang et al., 2020b). Ginsenoside Rd inhibits MCAO-induced microglial activation, decreases the expression levels of nuclear factor of kappa light polypeptide gene enhancer in B cell inhibitor, alpha (IκBα) phosphorylation and NF-κB nuclear translocation within a short time, and has fewer side effects than glucocorticoids (Zhang et al., 2016).

Other Nervous System Diseases

Ginsenoside Rd has a significant neuroprotective effect on a variety of neurological diseases, which may be related to its promotion of stem cell proliferation (Shi et al., 2005) and differentiation into astrocytes (Lin et al., 2012). Ginsenoside Rd may promote neurite outgrowth by upregulating growth-associated protein of 43 kDa (GAP-43) expression via ERK- and ARK-dependent signaling pathways in NGF-induced PC12 cells (Wu et al., 2016).

In H2O2-induced PC12 cells, ginsenoside Rd, as a neuroprotective agent, has neuroprotective effects on neurodegenerative diseases (Ye et al., 2008). In the converting monocyte phenotype and macrophages of the Guillain–Barre syndrome (GBS) mouse model, ginsenoside Rd attenuates experimental autoimmune neuritis (Ren et al., 2021). Ginsenoside Rd can regulate MMP by decreasing intracellular ROS and enhancing the activity of antioxidant enzymes and mitochondrial complex, thereby increasing intracellular ATP levels and ultimately reducing 1-methyl-4-phenylpyridinium (MPP+)-induced cell death in Parkinson’s disease (PD) (Liu et al., 2015b). Meanwhile, in the Aβ25–35-induced neuronal damage model, apoptosis and oxidative stress are ameliorated by ginsenoside Rd by regulating antioxidant capacity and the production of apoptotic proteins (Liu et al., 2015c). Learning and memory abilities can be improved in ginsenoside Rd-pretreated APP transgenic mice by significantly suppressing the NF-κB pathway to reduce the generation of proinflammatory factors (Liu et al., 2015d). Ginsenoside Rd-mediated neuroprotective effects against Alzheimer’s disease (AD) progression play a significant role in Neuro2a cells (Kim et al., 2014b). Ginsenoside Rd pretreatment can inhibit tau protein phosphorylation by maintaining a balance of GSK-3β, cyclin-dependent kinase 5 (CDK5/P25), and protein phosphatase 2A (PP-2A) (Li et al., 2013) to inhibit tau phosphorylation of tau protein at Ser199/202, Ser396, or Ser404 in okadaic acid-induced rats, APP transgenic mice, and cortical neurons to increase PP-2A activity for protection against AD (Li et al., 2011a; Li et al., 2021), respectively. Moreover, ginsenoside Rd increases the soluble amyloid-β precursor protein α (sAPPα) level and reduces extracellular Aβ to enhance the cognitive and memory functions of ovariectomy rats (Yan et al., 2017).

In experimental autoimmune encephalomyelitis, ginsenoside Rd exerts a neuroprotective role by regulating the immune response and inflammatory reaction via a signal pathway of IFN-g/IL-4, BDNF/NGF (Zhu et al., 2014), and Foxp3/RORγt/JAK2/STAT3 (Jin et al., 2020a). In spinal cord injury (SCI) models, ginsenoside Rd shows anti-inflammatory effects consistent with dexamethasone that could significantly decrease the biomarkers of apoptosis, inflammation, oxidative damage factor, and repaired damaged mitochondria; particularly, there is no obvious difference in terms of dexamethasone in anti-inflammatory (Zhou et al., 2014; Cong and Chen, 2016), and these effects depended on the ASK1/JNK pathway (Wang et al., 2014). In the pathology of noise-induced hearing loss (NIHL), ginsenoside Rd could alleviate the apoptosis and oxidative stress damage on neuron cells by activating the SIRT1/PGC-1α signaling pathway (Chen et al., 2020). In addition, ginsenoside Rd treatment effectively eliminates the oxidative injury and the production of proinflammatory factors and peroxides in the chronic restraint stress (CRS) paradigm (Wang et al., 2020). Ginsenoside Rd pretreatment may be neuroprotective in old rats following acute Pb exposure through limited microglial activation and maintained neural stem cell proliferation (Wang et al., 2013a).

To summarize, ginsenoside Rd can play a significant role in neuron damage by inhibiting the production of excitatory amino acids, reducing the intracellular Ca2+ influx mediated by the NMDA pathway, changing the neurotoxicity of Ca2+ to mitochondrial function damage, and regulating apoptosis-inducing and neuroinflammatory factors (Table 2).

TABLE 2

Summary of the neuroprotective effects and mechanism of ginsenoside Rd in animal and cell models.

References	Diseases	Inducer	Experimental model	Effects	Mechanism
Zhang et al. (2013a)	Ischemic stroke	MCAO	Male SD rats	GLT-1, PKB/Akt, p-ERK1/2↑	Glutamate metabolism
				Glutamate↓
Zhang et al. (2012a)	Ischemic stroke	Glutamate, NMDA	Primary hippocampal cell cultures from SD rat embryos	TUNEL-positive cells, caspase-3, Ca2+↓	Ca2+, apoptosis
Xie et al. (2016b)	Stroke	OGD/Transient MCAO	Adult male primary cortical neuron cells/SD rats	Infarct volume, NR2B subunit, p-Ser-1303, p-Tyr-1472, p-Tyr-1480↓	Hyperphosphorylation of neurons
Zhang et al. (2020a)	Ischemic stroke	OGD/MCAO, CsA	Primary cortical neurons cells, HEK293 cells/Adult male SD rats	Ca2+, NMDA receptor currents, caspase3↓	Apoptosis
Zhang et al. (2012b)	Ischemic stroke	MCAO	Male SD rats	ASIC2a↑	Ca2+ overload
				TRPM7, ASIC1a↓
Ye et al. (2011b)	Transient ischemic stroke	MCAO	Male SD rats, isolated mitochondria	ETC, aconitase, MMP, Pyruvate↑	Mitochondrial dysfunction, apoptosis
				ROS, Lactate, caspase-3, Cyto C, AIF↓
Yang et al. (2016)	Ischemic stroke	MCAO	Male SD rats	NEIL1, NEIL3↑	mtDNA and nDNA damages, apoptosis
				Cleaved caspase-3↓
Hu et al. (2013)	Cerebral ischemia	MCAO	Adult male SD rats	PARP-1, NF-κB, AIF↓	Apoptosis, inflammation
Ye et al. (2009)	Cerebral ischemic injury	OGD	Primary hippocampal neurons cells	GSH, GPX,SOD,CAT,MMP↑	Oxidative stress, apoptosis
				ROS, MDA,LDH, GSSG↓
Ye et al. (2011c)	Transient focal ischemia in the aged brain	MCAO	Male C57BL/6 mice	Mitochondrial complex, MMP, CAT, SOD, GPX, GST↑	Mitochondrial dysfunction oxidative stress
				MDA, protein carbonyl concentration, ROS, mitochondrial aconitase↓
Zhang et al. (2014)	Ischemic stroke	OGD/MCAO	Primary culture of neurons/Male SD rats	p-AKT, GSK-3β↑	p-tau
				p-tau, S199/202, PHF-1↓
Liu et al. (2015a)	Stroke	OGD/R/Transient MCAO followed by reperfusion	PC12 cells/Male SD rats	p-AKT, p-ERK, VEGF, BDNF↑	Apoptosis
Hou et al. (2017)	TMT intoxication	Trimethyltin	Primary hippocampal neuron/Male ICR mice	Bcl-2↑	Apoptosis
				Bax, caspase-3↓
Ye et al. (2011d)	Transient ischemic stroke	MCAO	Male SD rats	CAT, SOD 1 and 2, GR, GSH/GSSG↑	Oxidative stress, inflammation
				2,3- and 2,5-DHBA, 8-OHdG positive cells, 4-HNE, MDA, AGEs↓
Zhang et al. (2020b)	Transient forebrain ischemia	MCAO	Male SD rats	IκB-α↑	Inflammation
				20S proteasome, NF-κB, p65, matrix MMP-9↓
Zhang et al. (2016)	Ischemic stroke	OGD or LPS/MCAO	BV2 cells/Adult male SD rats	IL-1β, IL-6, TNF-α, IFN-γ, p-IκBα↓	Inflammation
Wu et al. (2016)	Ischemic stroke	NGF	PC12 cells	p-ERK1/2, p-AKT	NGF
				GAP-43↑
Ye et al. (2008)	Oxidative damage	H2O2	PC12 cells	SOD, GPX, MMP↑	Oxidative stress, mitochondrial dysfunction
				LDH, ROS, MDA,↓
Ren et al. (2021)	GBS	Peripheral nerve antigen P0180–199 peptide, Pertussis toxin (PTX)	Male C57 BL/6 mice	Non-classical Ly6Clo monocytes	Immunization, inflammation
				Nr4a1↑
				IL-12, IL-1β, TNF- α, IL-6, CD45+Ly6G +↓
Liu et al. (2015b)	Parkinson disease	MPP +	SH-SY5Y cells/C57BL/6J mice	SOD, GPX, MMP, complex I, ATP, Bcl-2, p-Akt↑	Oxidative stress, mitochondrial dysfunction
				LDH, ROS, MDA, Bax↓
Liu et al. (2015c)	Alzheimer’s disease	Aβ25-35	Primary cultured hippocampal neurons cells	SOD, GSH-Px, Bcl-2 mRNA↑	Oxidative stress, Neuronal apoptosis
				ROS, Bax mRNA, Caspase-3, Cyt C mRNA↓
Liu et al. (2015d)	Alzheimer’s disease		APP transgenic mice	IL-1β, IL-6, TNF-α, S100β mRNA, NF-κB p65↓	Inflammation
				IL-10↑
Kim et al. (2014b)	Neurodegenerative diseases		Neuro2a cells	ChAT, VAChT, ACh, MAP-2, p75, p21, TrkA↑	Cholinergic markers
Li et al. (2013)	Alzheimer’s disease		APP transgenic mice	Ser9, PP-2A↑	p-tau
				GSK-3β, Tyr216↓
Li et al. (2011a)	Alzheimer’s disease	Okadaic Acid	Adult male SD rats/Cortical neurons cells	PP-2A↑	Tau
				Tau↓
Li et al. (2021)	Alzheimer’s disease		APP transgenic mice	P35↑	p-tau
				Tau, P25↓
Yan et al. (2017)	Alzheimer’s disease	Ovariectomy/Inhibitor	Adult female rats/HT22 hippocampal neuronal cells	BACE1, Aβ↓ sAPPα, ADAM↑	Activating estrogen-like activity
Zhu et al. (2014)	Multiple sclerosis	Experimental autoimmune encephalomyelitis	6-8 weeks female C57 BL/6 mice	IL-4, BDNF, NGF↑	Blood–brain barrier, inflammation
				IFN-γ↓
Jin et al. (2020a)	Multiple sclerosis	Experimental autoimmune encephalomyelitis	Splenocyte/6-8 weeks C57BL/6 mice	TGF-β, IL-10, Treg, Foxp3↑	Inflammation, autoimmunity
				IL-6, IL-17, RORγt, Jak1, Jak2,STAT↓
Cong and Chen, (2016)	Spinal cord injury	T8 laminectomy and a spinal contusion injury	Adult female SD rats	MDA, TNF-α, IL-1β, IL-6, Bax, GSK, SOD, Bcl-2↑	Oxidative stress, inflammation, apoptosis
				cleaved-caspase 3, p-ERK, p-JNK, p-p38↓
Zhou et al. (2014)	Paraplegia	Ca2+	Isolated spinal cord mitochondria/Male C57BL/6J mice	p-AKT, p-ERK↑	Mitochondrial dysfunction
				Cyto C↓
Wang et al. (2014)	Delayed paralysis	Occlusion of the abdominal aorta for 1 h	Female SD rats	Caspase 3, ASK1, JNK↓	Apoptosis
Wang et al. (2020)	Cognitive impairment	Respiration in a transparent plexiglas restrainer with many air holes to for 10 h	Male C57BL/6J mice	SOD, CAT, GSH, GPX, p-PI3K, p-CREB, BDNF, TrkB↑	Oxidative stress, inflammation, neurotrophic factors
				TNF-α, IL-6, p-AKT↓
Wang et al. (2013a)		Lead (Pb) exposure	Retired breeder SD rats	IL-1β, IL-6, TNF-α↓	Inflammation

Abbreviations: CsA, cyclosporin A; ETC, mitochondrial electron transport chain; CAT, catalase; SOD, superoxide dismutase; GPX, glutathione peroxidase; GR, glutathione reductase; GSH, glutathione; GSSG, glutathione disulfide; 8-OHdG, 8-hydroxy-deoxyguanosine; 4-HNE, 4-hydroxynonenal; MDA, malondialdehyde; AGEs, advanced glycosylation end products; NGF, nerve growth factor; PTX, pertussis toxin; Nr4a1, nuclear receptor subfamily 4 group A member 1; ChAT, choline acetyltransferase; VAChT, vesicular acetylcholine transporter; ACh, acetylcholine.

Cancer

As indicated in Table 3 and Figure 3, ginsenoside Rd can inhibit the proliferation of various cancer cells by participating in the apoptotic pathway. As a potential therapeutic and specific 26S proteasome inhibitor, ginsenoside Rd plays an important role in anticancer therapy by targeting 26S proteasome (Chang et al., 2008).

TABLE 3

Summary of the effects and mechanisms of ginsenoside Rd on cell and animal models of multiple cancers.

References	Diseases	Experimental model	Effects	Mechanism
Tian et al. (2020)	Gastric cancer	MKN-45, SGC-7901 cells	Caspase-3, caspase-9↑	Apoptosis
			Cyclin D1↓
Kim et al. (2013b)	Gastric cancer	AGS cells	Caspase-3, caspase8, PARP↑	Apoptosis
Chian et al. (2019)	NSCLC	A549 NSCLC cells	NRF2↓	Proliferation
Gu et al. (2019)	Glioblastoma	U251 cells	Caspase‐3↑	Apoptosis
			Bcl‐2, hTERT↓
Liu et al. (2020b)	Glioblastoma	U251 cells, H4 (HTB148) cells, U87 MG cells	miR-144-5p, TLR2↑	Proliferation
			Toll-like receptor 2↓
Phi et al. (2019)	Colorectal cancer	Human CRC cell, HT29 cells/SW620, NSG mice	Smad2↓	Apoptosis
Zhang et al. (2017)	Breast cancer	HUVECs, MDA-MB-231 cells/Athymic nude mice	Bax, caspase-3, HIF1-α↑	Apoptosis
			Bcl-2↓
Kim, (2013)	Breast cancer	AGS cells, MCF-7 cells	Caspase-3↑	Apoptosis
Wang et al. (2016)	Breast cancer	4T1 cells, MDA-MB-231cells/Female BALB/c mice	Smad2↑	Attenuates metastasis
			miR-18a↓
Pokharel et al. (2010)	Breast cancer	MCF-7/ADR cells	MDR1↓	Resistance
Yang et al. (2006b)	Cervical cancer	HeLa cells	Bax↑	Apoptosis
			Bcl-2↓
Yang et al. (2021a)	Hepatocellular carcinoma	HepG2 cells/Male BALB/c nude mice		Proliferation, apoptosis
Yoon et al. (2012)	Hepatocellular carcinoma	HepG2 cells	MMP-1, MMP-2, MMP-7↓	Blocking MAPK signaling and inducing the formation of focal adhesions

FIGURE 3

Molecular mechanism of ginsenoside Rd on anticancer activity in multiple cancers.

Ginsenoside Rd can appreciably inhibit the proliferation of gastric cancer cells and can stimulate apoptosis by downregulating cyclin D1, thereby inducing cell cycle arrest in the G0/G1 phase and enhancing the expression of caspase-3 and caspase-9 and the ratio of Bax/Bcl-2 (Tian et al., 2020). After heat processing, the anticancer activity of deglycosylated Rd could be improved via the apoptotic pathway for AGS cells (Kim et al., 2013b).

In non-small-cell lung cancer (NSCLC), ginsenoside Rd, as a therapeutic drug, inhibits the nuclear factor erythroid 2-associated factor 2 (NRF2) pathway, and the synergistic effect of ginsenoside Rd in A549 and cisplatin (DDP)-resistant A549 cell lines (A549/DDP) can be weakened by knocking out NRF2 (Chian et al., 2019). As for glioblastoma, ginsenoside Rd decreases the proliferation of human glioma U251 cells and promotes apoptosis by downregulating the expression of hTERT and Bcl-2, upregulating the expression of the caspase-3 level, and inhibiting the telomerase activity of U251 cells (Gu et al., 2019). Ginsenoside Rd inhibits the proliferation and migration of glioblastoma cells by decreasing the expression of tumor-suppressor Mir-144-5p and promoting the expression of the target of Mir-144-5p toll-like receptor 2 (Liu et al., 2020b). In colorectal cancer cells, ginsenoside Rd, a therapeutic agent, targets epidermal growth factor receptor (EGFR)/SOX2 signaling (Phi et al., 2019).

Ginsenoside Rd also plays a crucial role in breast cancer. In MDA-MB-231 cell xenografted mice, ginsenoside Rd treatment inhibits the activation of PI3K, AKT, mammalian target of rapamycin (mTOR), and p70S6K in cells and decreases the expression of hypoxia-inducible factor 1-α (HIF1-α) (Zhang et al., 2017). In MCF-7 cells, ginsenoside Rd inhibits the proliferation of MCF-7 cells by enhancing caspase-3 activity, mitochondrial depolarization, and sub-G1 populations (Kim, 2013). In 4T1 cells, the expression of Mir-18a and Smad2 decreases with ginsenoside Rd treatment (Wang et al., 2016). Furthermore, ginsenoside Rd promotes the ubiquitination of MDR1 and inhibits doxorubicin resistance in MCF-7/ADR cells (Pokharel et al., 2010). In cervical cancer, ginsenoside Rd treatment in HeLa cells upregulates Bax expression, downregulates Bcl-2 expression, decreases the mitochondrial transmembrane potential, activates the caspase-3 pathway, significantly inhibits proliferation, and induces apoptosis (Yang et al., 2006b).

Finally, in HepG2 cells and the HepG2 cell-injected nude mice-induced hepatocellular carcinoma model, the combination of CA4P and ginsenoside Rd has synergistic antitumor effects via the PI3K/AKT/mTOR signaling pathway-related inhibition of HIF-1α (Yang et al., 2021a). HepG2 cells treated with ginsenoside Rd noticeably promoted matrix metalloproteinases’ (MMPs) activation, and MAPK signaling pathways were involved in cancer cell migration, thereby suggesting that ginsenoside Rd inhibits the activity of HepG2 cells in a dose-dependent and time-dependent manner (Yoon et al., 2012).

Gastric and Gut

In a sodium dextran sulfate (DSS)-induced colitis model, ginsenoside Rd reduces DSS-induced colonic pathology via the adenosine 5′-monophosphate-activated protein kinase/Unc-51 like autophagy activating kinase 1 (AMPK/ULK1)-induced autophagy signaling pathway and the inhibition of the production of proinflammatory cytokines (IL-1β, TNF-α, and IL-6) in serum and colon tissues (Liu et al., 2018). In irradiation-induced intestinal epithelial cells, ginsenoside Rd reduces apoptosis by activating a pathway of PI3K/AKT, inactivates MEK, and inhibits a mitochondria/caspase pathway (Tamura et al., 2008). Meanwhile, in 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced ulcerative colitis model, ginsenoside Rd showed obvious anti-inflammatory activity by inhibiting neutrophil infiltration, regulating apoptosis signal and oxidative stress (Yang et al., 2012a), reduced the accumulation of leukocytes, and downregulated multiple proinflammatory cytokines (Yang et al., 2012b).

Metabolic Diseases

Laboratory data of ginsenoside Rd suggest that it has effects on multiple metabolic diseases. The browning of white adipose tissue induced by cold stress and cAMP levels are increased by ginsenoside Rd. In particular, Rd alleviates obesity and insulin resistance by upregulating thermogenesis through the cAMP/protein kinase A (PKA) signaling pathway (Yao et al., 2020). In fast-food diet-induced non-alcoholic fatty liver disease (NAFLD), fermented ginsenoside Rd with Cordyceps militaris regulates lipid metabolism and the inflammatory response via mTORC1 signaling (Choi et al., 2019). Ginsenoside Rd inhibits the progress of the death of islet transplantation by decreasing the apoptosis of the islet cells (Kaviani et al., 2019). In the atherosclerosis process, ginsenoside Rd decreases oxidized low-density lipoprotein (Ox-LDL) and cholesterol by inhibiting Ca2+ influx (Li et al., 2011b). In diabetic db/db mice and mesangial cells, pectin-lyase-modified ginsenoside Rd relieves diabetic nephropathy via alleviated ROS production (Jung et al., 2021).

Other Diseases

Ginsenoside Rd has positive effects on skin injury, osteoporosis, kidney injury, vessel injury, heart injury, lung injury, aging, and inflammation. In animal wound models, ginsenoside Rd significantly increases wound healing by promoting the proliferation and migration level of keratinocyte progenitor cells (KPCs) and human dermal fibroblasts (HDFs) (Kim et al., 2013c). Ginsenoside Rd also has a positive effect on rejection caused by a transplant skin allograft (Wang et al., 2012a). Beyond that, ginsenoside Rd, as an antiosteoporotic agent, promotes differentiation and mineralization in osteoblastic MC3T3-E1 cells (Kim et al., 2012). In animal models of renal I/R injury and cultured proximal tubule cells, ginsenoside Rd has a protective effect by inhibiting inflammation and regulating biochemical indexes of renal function (Yokozawa et al., 1998; Ren et al., 2016). In addition, ginsenoside Rd downregulates NF-κB and the expression of iNOS and COX-2 in lipopolysaccharide (LPS)-induced Institute of Cancer Research (ICR) mice, and RAW264.7 cells were suppressed (Kim et al., 2013d). In the nicotine-induced vascular endothelial injury model, ginsenoside Rd plays an important role in the prevention of cardiovascular diseases via participation in NO signaling and regulates platelet and vascular function (Zhang et al., 2020c). Ginsenoside Rd upregulates Cyto C release and caspase-9/caspase-3 activation and decreases the MMP and the ratio of Bcl-2/Bax via the mitochondria-dependent pathway in H2O2-induced apoptosis in basilar artery smooth muscle cells (BASMCs) (Li et al., 2012). Furthermore, ginsenoside Rd could relieve the cisplatin-induced kidney injury (Yokozawa and Liu, 2000; Yokozawa and Dong, 2001) and kidney proximal tubules cephaloridine injury under cephaloridine treatment (Yokozawa and Dong, 2001). In an adrenocorticotrophic hormone (ACTH)–induced corticosterone secretion cell model, ginsenoside Rd inhibits ACTH-induced corticosterone production by inhibiting the MC2R-cAMP/PKA/cyclic AMP response element binding (CREB) pathway in adrenocortical cells (Jin et al., 2020b). In myocardial I/R-induced rats and simulated I/R-induced primary neonatal rat cardiomyocyte models, ginsenoside Rd promotes cardioprotection via the activation of AKT/GSK-3b signaling (Wang et al., 2013b). In addition, ginsenoside Rd can protect against LPS-induced acute lung injury by inhibiting the PI3K/AKT signaling pathway (Yang et al., 2021b). Other studies have indicated that ginsenoside Rd can significantly enhance the survival time of Caenorhabditis elegans via lipid metabolism and the activation of the stress response signaling pathway (Yu et al., 2021) and can alleviate the oxidative damage caused by aging in senescence-accelerated mice (Yokozawa et al., 2004). Finally, the anti-inflammatory activity of ginsenoside Rd is well documented, is considered to be associated with its antioxidant effects (Kim et al., 2007; Zhang et al., 2013b), and selectively produces prostaglandin E2 (PGE2) by activating the CCAAT/enhancer binding protein (C/EBP) and CREB to express COX-2 (Jeong et al., 2007). Ginsenoside Rd exerts anti-inflammation effects in carrageenan-induced inflammation rats via the inhibition of the NF-κB signaling pathway (Wang et al., 2012b) and in ovalbumin-induced allergic rhinitis mice by regulating multiple inflammatory factors (Kim et al., 2019) and elicits a Th1 and Th2 immune responses (Yang et al., 2007b). Ginsenoside Rd enhances the Th1 response to surface mannan extract in mice, which protects mice from disseminated candida infection by stimulating higher titers of Th1 antibodies and a Th1-dominated immune response (Han and Rhew, 2013).

Conclusion and Perspective

As a widely used herbal medicine, ginseng appears in the form of dietary supplements nowadays. Available evidence suggests that the antiapoptotic, antioxidant, and anti-inflammatory activities, which suppress the calcium influx of ginsenoside Rd, may have an important role in the neuroprotective and anticancer effects. Ginsenoside Rd play a crucial role in neuroprotective, anticancer effects, metabolism, and other diseases by regulating PI3K/AKT, inhibiting Cyto C released and caspase activation, and regulating the release of inflammatory factors, which play a crucial role in neuroprotective, anticancer effects, metabolism, and other diseases.

In addition, ginsenoside Rd has potential therapeutic effects on regulating metabolism and in multiorgan protection. However, attributable to the shortage of clinical studies on ginsenoside Rd, it is difficult to make a clear decision. In addition to exploring its various activities, it is suggested to verify existing activities in a deeper mechanism, design clinical trials to prove its safety and effectiveness, and obtain a more extensive clinical application.