Multi-target neuroprotective effects of notoginsenoside R1 in neurodegenerative diseases: From pharmacokinetics to translational prospects.

For ages, botanical medicine has been used in the treatment of diabetes mellitus (DM). Notoginsenoside R1 (NGR1), a Panax notoginseng (Burkill) F.H.Chen metabolite, has been documented to possess antidiabetic action in vivo. However, its precise molecular mechanism of action is not clear. We evaluated NGR1's effects on blood glucose in vivo and then evaluated in vitro whether NGR1 has effects on insulin secretion and the probable molecular pathways involved in NGR1-induced insulin secretion. Diabetes was induced in mice by streptozotocin. Glucose tolerance test was performed before and after NGR1 was administered intraperitoneally to diabetic animals for 4 weeks. Static and perifusion experiments were performed using isolated female BALB/c mouse islets. Preproinsulin (Ins) mRNA expression was measured using q-PCR. Protein expression of PI3K/Akt pathway was assessed using the fully automated Wes™ capillary-based protein electrophoresis. Treatment of diabetic mice with NGR1 improved their glucose intolerance. In vitro, NGR1 increased insulin secretion in a concentration-dependent manner. NGR1 initiated the secretion of insulin at 2mM glucose and augmented glucose-stimulated insulin secretion which was sustained throughout NGR1 perifusion. NGR1-induced insulin secretion was not altered by a voltage gated calcium channel blocker or protein kinase A inhibitor. NGR1 did not significantly modulate Ins mRNA expression. However, NGR1 significantly increased the levels of phospho-Akt and phopho-p-85. In conclusion, this study has shown that NGR1 ameliorates hyperglycemia in diabetic mice. NGR1 has a direct insulin secretagogue activity on mouse islets, stimulates insulin secretion at both basal and postprandial glucose concentrations, and activates PI3K/Akt pathway to induce insulin secretion. These results suggest that NGR1 may provide an alternative therapy to manage DM.

Background: Alzheimer's disease (AD) has affected numerous elderly individuals worldwide. Panax notoginseng has been shown to ameliorate AD symptoms, and notoginsenoside R2 is a key saponin identified in this plant. Purpose: In the current study, we aimed to explore whether notoginsenoside R2 could improve the prognosis of AD. Methods: Herein, primary rat cortical neurons were isolated and they were treated with amyloid beta-peptide (Aβ) 25-35 oligomers. Cellular apoptosis was examined via flow cytometry and Western blotting. miR-27a and SOX8 mRNA expression levels were quantified by quantitative reverse transcription-polymerase chain reaction. Furthermore, the interaction between miR-27a and SOX8 was investigated by utilizing a dual-luciferase reporter assay. Finally, an AD mouse model was established to validate the in vitro findings. Results: Notoginsenoside R2 alleviated Aβ25-35-triggered neuronal apoptosis and inflammation. During this process, miR-27a expression was decreased by notoginsenoside R2, and miR-27a negatively modulated SOX8 expression. Furthermore, activation of SOX8 upregulated β-catenin expression, thus suppressing apoptosis and neuroinflammation. Conclusions: Our animal experiments revealed that notoginsenoside R2 enhanced the cognitive function of AD mice and inhibited neuronal apoptosis. Notoginsenoside R2 ameliorated AD symptoms by reducing neuronal apoptosis and inflammation, thus suggesting a novel direction for AD pharmacotherapy.

Notoginsenoside R1 (NGR1), a novel phytoestrogen isolated from Panax notoginseng, has antioxidant and anti-apoptotic properties. Oxidative stress plays a pivotal role in neurodegenerative diseases. To mimic oxidative stress in neurons and explore the neuroprotection of NGR1, H2O2-induced neurotoxicity in NGF-induced differentiation of PC12 cells was used. In this study, NGR1 preconditioning provided neuroprotective effects via suppressing H2O2-induced the intracellular ROS accumulation, the increase in the product of lipid peroxidation (MDA), protein oxidation (protein carbonyl), and DNA fragmentation (8-OHdG), and mitochondrial membrane depolarization as well as caspase-3 activation. Moreover, NGR1 treatment alone potently increased the nuclear translocation of Nrf2, augmented ARE enhancer activity, and upregulated the expression and activity of phase II antioxidant enzymes including HO-1, NQO-1, and γ-GCSc. NGR1 could also increase the ERE activity and activate Akt and ERK1/2 pathways. NGR1-mediated activation of Nrf2/ARE signaling and neuroprotection were abolished by genetic silencing of Nrf2 using siRNA or the pharmacological blockade of estrogen receptors using ICI-182780, and partially inhibited by Akt siRNA or ERK siRNA transfection. In addition, the phosphorylation of ERK1/2 mediated by NGR1 was markedly inhibited in PC12 cells transfected with Akt siRNA. On the contrary, ERK1/2 siRNA transfection hardly had any effect on the phosphorylation of Akt mediated by NGR1. NGR1-mediated activation of Akt and ERK1/2 pathways was blocked by ICI-182780. In conclusion, NGR1 provided neuroprotection via inducing an estrogen receptor-dependent crosstalk between Akt and ERK1/2 pathways, subsequently activating Nrf2/ARE signaling and thereby up-regulating phase II antioxidant enzymes.

Our previous studies demonstrated the potential of notoginsenoside R1 (NGR1), a primary bioactive compound from Panax notoginseng, in alleviating diabetic encephalopathy in db/db mice and mitigating amyloid-β (Aβ)-induced neuronal damage. This study aimed to investigate the positive effects of NGR1 against cognitive deficits in a diabetic Alzheimer's disease (AD) mouse model (APP/PS1xdb/db mice). APP/PS1xdb/db mice were intragastrically administrated with NGR1 (40 mg/kg/day) or co-administrated with NGR1 and a selective PPARγ inhibitor GW9662 for 16 weeks. We identified NGR1 as a novel PPARγ agonist through molecular docking, surface plasmon resonance, and dual-luciferase reporter assay. NGR1 treatment significantly promoted the membrane translocation of GLUT4 and enhanced 2-deoxyglucose uptake in primary mouse hippocampal neurons. Furthermore, NGR1 treatment notably mitigated cognitive deficits in APP/PS1xdb/db mice. This treatment correlated with reduced blood glucose levels, lowered blood HbA1c, and decreased serum insulin levels, coupled with enhanced glucose tolerance and insulin sensitivity. Additionally, NGR1 treatment ameliorated Aβ burden, suppressed microglia-induced neuroinflammation, and notably increased cerebral glucose uptake, as demonstrated by 18F-FDG PET scans. NGR1 treatment could upregulate PPARγ and GLUT4 expression and increase phosphorylation of Akt at Ser473 while decreasing phosphorylation of IRS-1 at Ser616 in the hippocampus of APP/PS1xdb/db mice. Crucially, the protective effects of NGR1 were abolished by co-administration with GW9662. NGR1 demonstrated efficacy in enhancing neuronal glucose uptake through the activation of the PPARγ/Akt/GLUT4 signaling pathways in APP/PS1xdb/db mice, positioning it as a promising candidate for diabetic AD treatment.

Notoginsenoside R1 (NTR1) is the main active ingredient of the well-known traditional Chinese herbal medicine Panax notoginseng, the root of Panax notoginseng (Burk.) F. H. Chen. Studies demonstrated that NTR1 may have some neuronal protective effects. Alzheimer's disease (AD) is a neurodegenerative disease characterized by β -amyloid protein (Aβ) deposition, neurofibrillary tangle formation and neuronal loss. This study was designed to explore the protective effect of NTR1 on an APP/PS1 double-transgenic mouse model of AD and investigate the possible mechanism. The 3-month-old mice were fed with 5 mg/(kg•d), 25 mg/(kg•d) NTR1 or vehicle via oral gavage for 3 months and changes in behavior, neuropathology, and amyloid pathology were investigated. The mice with NTR1 treatment showed significant amelioration in the cognitive function and increased choline acetyl transferase expression, as compared to the vehicle treated mice. NTR1 treatment inhibited Aβ accumulation and increased insulin degrading enzyme expression in both APP/PS1 mice and N2a-APP695sw cells, suggesting that of NTR1 may exert its protective effects through the enhancement of the Aβ degradation. Furthermore, our data showed that the increased level of peroxisome proliferator-activated receptor γ (PPARγ) and the up-regulation of insulin degrading enzyme induced by NTR1 were inhibited by administration of GW9662 (a PPARγ antagonist), indicating that the effect of NTR1 was mediated, at least in part, by PPARγ. Thus, our findings provide the evidences that NTR1 has protective effect on AD mouse model and NTR1 may be a potential candidate for AD treatment.

Astragalus and Panax notoginseng are commonly used to treat cardio-cerebrovascular diseases in China and are often combined together to promote curative effect. We speculate that the enhancement of the combination on anticerebral ischemia injury may come from the main active components. The purpose of this work was to probe the effects and mechanisms of Astragaloside IV (the active component of Astragalus) combined with Ginsenoside Rg1, Ginsenoside Rb1, and Notoginsenoside R1 (the active components of P. notoginseng) to antagonize ischemia/reperfusion (I/R) injury via inflammation and apoptosis. C57BL/6 mice were randomly divided into sham, model, Astragaloside IV, Ginsenoside Rg1, Ginsenoside Rb1, Notoginsenoside R1, four active components combination, and Edaravone groups. After administration for 3 days, bilateral common carotid arteries (CCA) were occluded with artery clip for 20[Formula: see text]min followed by reperfusion for 24[Formula: see text]h. Our results showed that the survival rate of nerve cell in hippocampal CA1 decreased while the apoptotic rate increased, and the level of caspase-3 protein in brain tissues was elevated, the expressions of TNF-a, IL-1, and ICAM-1 mRNA as well as phosphorylated nuclear factor kappa B (NF-κB) inhibitor protein α (p-IκBa) in brain tissues were up-regulated, and the nuclear translocation rate of NF-κB was raised. Additionally, the protein expressions of phosphorylated tyrosine kinase 1 (p-JAK1), phosphorylated signal transducer and activator of transcription-1 (p-STAT1), glucose regulated protein 78 (GRP78), caspase-12, and phosphorylated c-Jun N-terminal kinases 1/2 (p-JNK1/2) in brain tissues were also significantly strengthened after I/R for 24 h. All drugs could increase neurocyte survival rate in hippocampal CA1, decrease the apoptotic rate, and inhibit caspase-3 protein expression, in contrast, the effects of four active components combination were better than those of active components alone. In addition, Astragaloside IV and Ginsenoside Rg1 could down-regulate the level of TNF-α, and ICAM-1 mRNA, respectively, Notoginsenoside R1 reduced both TNF-α and ICAM-1 mRNA, and the combination of the 4 effective components had inhibitory effects on the expressions of TNF-α, IL-1β, and ICAM-1 mRNA. Astragaloside IV, Ginsenoside Rg1, Notoginsenoside R1, and 4 effective components combination were able to restrain the phosphorylation of IκBα, and relieve the nuclear translocation rate of NF-κB. Moreover, the effects of the combination are greater than those of active components alone. All drugs could suppress the phosphorylation of JAK1 induced by I/R; meanwhile the expression of p-STAT1 exhibited a decrease in Ginsenoside Rg1 and four active components combination groups. The decreases of p-JAK1 and p-STAT1 in the four active components combination group were more obvious than those in active components alone groups. Astragaloside IV, Ginsenoside Rg1, and Notoginsenoside R1 further augmented GRP78 expression caused by I/R, Notoginsenoside R1 attenuated caspase-12 protein expression, Astragaloside IV and Ginsenoside Rg1 lessened the phosphorylation of JNK1/2, and the four active components combination was capable of up-regulating GRP78 protein while down-regulating the expressions of caspase-12 and p-JNK1/2. Similarly, the effects of the four active components combination were greater than those of effective components alone. These suggested that the combination of the main active components of Astragalus and Panax notoginseng could strengthen protective effects on cerebral ischemia injury via anti-apoptosis and anti-inflammation, and the mechanisms might be associated with restraining the activation of NF-κB and JAK1/STAT1 signal pathways and regulating endoplasmic reticulum stress (ERS) after cerebral ischemia.

Notoginsenoside R1 alleviates blue light-induced corneal injury and wound healing delay by binding to and inhibiting TRIB1.

Notoginsenoside R1 (NGR1) is a novel phytoestrogen that is isolated from Panax notoginseng. We have recently found that NGR1 showed neuroprotection in vitro against oxidative stress through estrogen receptor (ER)-dependent activation of Akt/Nrf2 pathways. However, whether NGR1 has neuroprotective effect against cerebral ischemia–reperfusion (I/R) injury in vivo is unknown. In this study, we used in vivo and in vitro models of cerebral I/R injury that demonstrate middle cerebral artery occlusion and reperfusion in rats, as well as oxygen–glucose deprivation followed by reoxygenation (OGD/R) in primary cortical neurons. These models were used to evaluate NGR1 neuroprotection. Three-day pretreatment with NGR1 (20 mg/kg; i.p.) significantly improved neurologic outcomes and reduced cerebral infarct volume. Pretreatment of primary cortical neurons with NGR1 (25 μM) for 24 h prevented apoptosis and oxidative stress induced by OGD/R. NGR1 inhibited apoptosis by inhibiting mitochondrial membrane potential disruption, caspase-3 activation, and DNA fragmentation. NGR1 prevented oxidative stress by suppressing NADPH oxidase- and mitochondrion-derived superoxide and inhibiting production of malondialdehyde, protein carbonyl, and 8-hydroxydeoxyguanosine in vivo and in vitro. NGR1 induced ER-dependent activation of Akt/Nrf2 pathways by increasing ERα, ERβ, phospho-Akt, phospho-GSK3β, nuclear Nrf2, and HO-1 expression in vivo and in vitro. Pretreatment with ICI-182780, LY294002, or Snpp abolished NGR1-mediated neuroprotection against oxidative stress and apoptosis in vitro. In conclusion, NGR1 showed neuroprotection against cerebral I/R injury in vivo and in vitro. The mechanism of NGR1 neuroprotection involves inhibition of NADPH oxidase activity and mitochondrial dysfunction via ER-dependent activation of Akt/Nrf2 pathways.

Notoginsenoside R2 is a crucial active saponin in Panax notoginseng (Burk.) F. H. Chen, but its natural content is relatively low. In this study, we investigated the biotransformation of notoginsenoside R1 to 20(S/R)-notoginsenoside R2 using Lactiplantibacillus plantarum S165, compared the inhibitory effects on cancer cell proliferation and conducted a mechanistic study. Notoginsenoside R1 was transformed using Lactiplantibacillus plantarum S165 at 37 °C for 21 days. The fermentation products were identified using a combination of HPLC, UPLC-MS/MS, and 13C-NMR methods. The inhibition effects of 20(S/R)-notoginsenoside R2 on H22 hepatoma cells were assessed by CCK-8 and TUNEL assays, and the underlying mechanism was investigated by Western blotting. Lactiplantibacillus plantarum S165 could effectively transform notoginsenoside R1 to 20(S/R)-notoginsenoside R2 with a conversion yield of 82.85%. Our results showed that 20(S/R)-notoginsenoside R2 inhibited H22 hepatoma cells proliferation and promoted apoptosis. The apoptosis of H22 hepatoma cells was promoted by 20(S/R)-notoginsenoside R2 through the blockade of the PI3K/AKT/mTOR signaling pathway. The biotransformation method used in this study resulted in the production of 20(S)-notoginsenoside R2 and 20(R)-notoginsenoside R2 from notoginsenoside R1, and the anti-tumor activity of the transformed substance markedly improved.

Activation of the hepatic stellate cell is implicated in pathological vascularization during development of liver fibrosis. MAPK signaling is involved in the activation of hepatic stellate cell. Oxidative stress and inflammation are also involved in the pathogenesis of liver fibrosis. Notoginsenoside R1 is an effective saponin isolated from the roots of Panax notoginseng (Burk) F. H. Chen and exerts anti-oxidant, anti-inflammatory and anti-fibrotic roles in various diseases. However, the role of Notoginsenoside R1 in liver fibrosis has not been investigated yet. First, a rat model with liver fibrosis was established through oral gavage administration with carbon tetrachloride. Data from hematoxylin and eosin (H&E) and Masson's trichrome stainings showed that carbon tetrachloride induced severe hepatic damages, including inflammatory cell infiltration, lipid droplets deposition in hepatocytes and liver centrilobular necrosis. Meanwhile, the rats were also intraperitoneal injected with different concentrations of Notoginsenoside R1. Results demonstrated that Notoginsenoside R1 treatment suppressed the pathological changes in the livers with enhanced levels of ALB and TP, and reduced levels of ALP, AST and ALT. Second, Notoginsenoside R1 also significantly attenuated carbon tetrachloride-induced decrease in PPAR-[Formula: see text] and increase in Coll-a1, [Formula: see text]-SMA and TIMP1 in liver tissues ([Formula: see text][Formula: see text] 0.001). Third, the decrease in GSH, SOD and GST and increase in MDA, IL-1[Formula: see text], IL-6 and TNF-[Formula: see text] induced by carbon tetrachloride were markedly restored by Notoginsenoside R1 ([Formula: see text][Formula: see text] 0.001). Lastly, Notoginsenoside R1 counteracted with the promotive effects of carbon tetrachloride on levels of proteins involved in MAPK signaling, including phosphorylated p65 (p-p65), p-ERK, p-JNK and p-p38. In conclusion, Notoginsenoside R1 suppressed the activation of hepatic stellate cells and exerted anti- oxidant and anti-inflammatory to attenuate carbon tetrachloride-induced liver fibrosis through inactivation of NF-[Formula: see text]B and MAPK signaling.

Diabetic cardiomyopathy (DCM) leads to heart failure and death in diabetic patients, no effective treatment is available. Notoginsenoside R1 (NGR1) is a novel saponin that is derived from Panax notoginseng and our previous studies have showed cardioprotective and neuroprotective effects of NGR1. However, its role in protecting against DCM remains unexplored. Herein, we examine potential effects of NGR1 on cardiac function of diabetic db/db mice and H9c2 cardiomyocytes treated by advanced glycation end products (AGEs). In vitro experiments revealed that pretreatment with NGR1 significantly decreased AGEs-induced mitochondria injury, limited an increase in ROS, and reduced apoptosis in H9c2 cells. NGR1 eliminated ROS by promoting estrogen receptor α expression, which subsequently activated Akt and Nrf2-mediated anti-oxidant enzymes. In vivo investigation demonstrated that NGR1 significantly reduced serum lipid levels, insulin resistance, the expression of enzymes related to cardiomyopathy, and the expression of apoptotic proteins. Finally, NGR1 improved cardiac dysfunction and attenuated histological abnormalities, as evidenced by elevating ejection fraction and fractional shortening, and reducing cardiac fibrosis. Mechanistically, NGR1 promoted ERα expression, which led to the activation of Akt-Nrf2 signaling and the inhibition of the TGFβ pathway. Collectively, these results strongly indicate that NGR1 exerts cardioprotective effects against DCM through its inhibition of oxidative stress and apoptosis, and eventually suppresses cardiac fibrosis and hypertrophy, which suggests that NGR1 is a potential therapeutic medicine for the treatment of DCM.

Development of paclitaxel-induced neuropathic pain (PINP) during chemotherapy may lead to paclitaxel discontinuation, potentially compromising effective anticancer therapy. PINP can manifest as allodynia. One recently discovered key factor in paclitaxel-induced mechanical allodynia (PIMA) pathogenesis is the elevated activity of monoacylglycerol lipase (MAGL), an enzyme that metabolizes the endocannabinoid 2-arachidonoylglycerol (2-AG). Thus, inhibiting MAGL serves as a potential analgesic target. Notoginsenoside R1 (NGR1), a metabolite of Panax notoginseng, has shown promise in reducing oxidative stress and neuronal apoptosis in nerve injury models. However, its effects on PIMA and MAGL activity have not yet been explored. This study is a proof-of-concept preclinical study investigating the antiallodynic effects of NGR1 on PIMA in female BALB/c mice and also examining its effect on MAGL activity. The effect of treatment of mice with NGR1 intraperitoneally on the development of PIMA was evaluated. Molecular docking using CB-Dock2 compared the binding energies to MAGL of NGR1 and pristimerin, a triterpene MAGL inhibitor. The effects of NGR1 on human recombinant MAGL activity, as well as the MAGL activity in mice paw skin tissues, were assessed using MAGL inhibitor screening and MAGL activity assay kits, respectively. NGR1 prevented the development of PIMA in a dose-dependent manner. The docking scores showed that NGR1 has a good affinity for MAGL (-7.8 kcal/mol, binding energy) but less affinity than pristimerin (-10.3 kcal/mol). NGR1 inhibited the human recombinant MAGL activity in a reversible and concentration-dependent manner, although the inhibition was in a reverse order. Treatment of mice with NGR1 showed a non-significant trend in reducing the paclitaxel-induced increase in MAGL activity in the paw skin. This study shows for the first time that NGR1 prevents the development of PIMA and suggests that NGR1 has affinity for and inhibits human recombinant MAGL activity with a paradoxical inhibition pattern. More mechanistic studies are needed to fully elucidate the molecular mechanisms of NGR1 in preventing PIMA.

The main bioactive compound of Panax notoginseng, notoginsenoside R1, alleviates pulmonary fibrosis via MBD2/SHIP and STAT3 pathway

Notoginsenoside R1 inhibits TNF-α-induced fibronectin production in smooth muscle cells via the ROS/ERK pathway

Previous studies show that notoginsenoside R1 (NG-R1), a novel saponin isolated from Panax notoginseng, protects kidney, intestine, lung, brain and heart from ischemia-reperfusion injury. In this study we investigated the cardioprotective mechanisms of NG-R1 in myocardial ischemia/reperfusion (MI/R) injury in vivo and in vitro. MI/R injury was induced in mice by occluding the left anterior descending coronary artery for 30 min followed by 4 h reperfusion. The mice were treated with NG-R1 (25 mg/kg, i.p.) every 2 h for 3 times starting 30 min prior to ischemic surgery. We showed that NG-R1 administration significantly decreased the myocardial infarction area, alleviated myocardial cell damage and improved cardiac function in MI/R mice. In murine neonatal cardiomyocytes (CMs) subjected to hypoxia/reoxygenation (H/R) in vitro, pretreatment with NG-R1 (25 μM) significantly inhibited apoptosis. We revealed that NG-R1 suppressed the phosphorylation of transforming growth factor β-activated protein kinase 1 (TAK1), JNK and p38 in vivo and in vitro. Pretreatment with JNK agonist anisomycin or p38 agonist P79350 partially abolished the protective effects of NG-R1 in vivo and in vitro. Knockdown of TAK1 greatly ameliorated H/R-induced apoptosis of CMs, and NG-R1 pretreatment did not provide further protection in TAK1-silenced CMs under H/R injury. Overexpression of TAK1 abolished the anti-apoptotic effect of NG-R1 and diminished the inhibition of NG-R1 on JNK/p38 signaling in MI/R mice as well as in H/R-treated CMs. Collectively, NG-R1 alleviates MI/R injury by suppressing the activity of TAK1, subsequently inhibiting JNK/p38 signaling and attenuating cardiomyocyte apoptosis.