Senegenin Attenuates Pulmonary Fibrosis by Inhibiting Oxidative-Stress-Induced Epithelial Cell Senescence through Activation of the Sirt1/Pgc-1α Signaling Pathway.

Idiopathic pulmonary fibrosis is a fatal interstitial lung disease for which effective drug therapies are lacking. Senegenin, an effective active compound from the traditional Chinese herb Polygala tenuifolia Willd, has been shown to have a wide range of pharmacological effects. In this study, we investigated the therapeutic effects of senegenin on pulmonary fibrosis and their associated mechanisms of action. We found that senegenin inhibited the senescence of epithelial cells and thus exerted anti-pulmonary-fibrosis effects by inhibiting oxidative stress. In addition, we found that senegenin promoted the expression of Sirt1 and Pgc-1α and that the antioxidative and antisenescent effects of senegenin were suppressed by specific silencing of the Sirt1 and Pgc-1α genes, respectively. Moreover, the senegenin-induced effects of antioxidation, antisenescence of epithelial cells, and antifibrosis were inhibited by treatment with Sirt1 inhibitors in vivo. Thus, the Sirt1/Pgc-1α pathway exerts its antifibrotic effect on lung fibrosis by mediating the antioxidative and antisenescent effects of senegenin.

Senegenin Rescues PC12 Cells with Oxidative Damage Through Inhibition of Ferroptosis.

Oxidative stress is one of the pathological mechanisms of Alzheimer's disease (AD), and ferroptosis has been determined to be involved in neurodegenerative diseases such as AD. Senegenin (Sen) prevents oxidative damage in nerve cells via a mechanism that may be highly related to ferroptosis. However, the mechanism of ferroptosis pathway involvement in AD is unclear. In this study, we established a model of PC12 cytotoxic injury induced by Aß25-35, and we detected the level of oxidative damage, MMP, and ferroptosis-related protein expression. The results showed that, compared with control group, the level of ROS increased, GPX activities decreased, and MDA levels increased in Aß25-35 group. Aß25-35 could induce mitochondrial depolarization in PC12 cells and Fer-1 could not reverse this damage. WB revealed that Aß25-35 group had increased ACSL4 and PEBP1 proteins, and decreased GPX4 protein. After adding Sen in the model, the level of oxidative damage was reduced, and mitochondrial depolarization was reversed compared with Aß25-35 group. WB suggested that the expression of ACSL4 and PEBP1 proteins decreased, and the expression of GPX4 protein increased by Sen treatment. In conclusion, we found that Sen exhibits strong neuroprotective activity against Aß25-35 induced oxidative damage and lipid metabolic associated with ferroptosis. Inhibiting nerve cell ferroptosis might facilitate the future development of strategies to AD.

Neuroprotective Effects and Mechanisms of Senegenin, an Effective Compound Originated From the Roots of Polygala Tenuifolia.

Senegenin is the main bioactive ingredient isolated from the dried roots of Polygala tenuifolia Willd. In recent years, senegenin has been proved to possess a variety of pharmacological activities, such as anti-oxidation, anti-inflammation, anti-apoptosis, enhancement of cognitive function. Besides, it has a good development prospect for the treatment of neurodegenerative diseases, depression, osteoporosis, cognitive dysfunction, ischemia-reperfusion injury and other diseases. However, there is no systematic literature that fully demonstrates the pharmacological effects of senegenin. In order to meet the needs of new drug research and precise medication, this review summarized the neuroprotective effects, mechanisms and gastrointestinal toxicity of senegenin based on the literatures published from the past 2 decades. In addition, an in-depth analysis of the existing problems in the current research as well as the future research directions have been conducted in order to provide a basis for the clinical application of this important plant extract.

Senegenin alleviates Aß1-42 induced cell damage through triggering mitophagy.

ETHNOPHARMACOLOGICAL RELEVANCE: Senegenin (SEN), an active compound extracted from the traditional Chinese herb Polygala tenuifolia Willd. (a species in the genus Polygala, family Polygalaceae), could nourish neurons and resist neuronal damage in mouse models of Alzheimer's disease (AD). Amyloid-ß (Aß) depositions in neuronal cells may cause pathological changes such as oxidative stress which one return could cause severe damage to mitochondria in AD patients or animal models. Mitophagy is an important mechanism to selectively remove damaged mitochondria. In neurons, this process is mainly mediated by PTEN-induced putative kinase 1 (PINK1)/Parkin pathway. Previous studies have shown that SEN could reduce mitochondrial damage and inhibit apoptosis in neurons. Therefore, this study speculated that SEN might activate mitophagy to clear damaged mitochondria, thereby mitigating Aß-induced cell damage in neuronal cells. AIM OF THE STUDY: This study aimed to determine the effects of SEN on Aß-induced cell damage, and further to explore whether SEN could induce mitophagy. Moreover, the regulatory role of mitophagy in the neuroptrotective effect of SEN would be elucidated. MATERIALS AND METHODS: This study established an in vitro cell damage model using Aß1-42 to treat mouse hippocampal neuron HT22 cells. The effects of SEN on cell damage were determined by MTT assay and lactate dehydrogenase (LDH) release assay. Reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were detected by Cytation™5 cell imaging microplate detection system. The apoptotic rate was analyzed by flow cytometry. The effects of SEN on mitophagy were detected by transmission electron microscope, immunofluorescence and immunoblotting. RESULTS: Firstly, HT22 cells were treated with 30 µM Aß1-42 for 24 h to establish the damage model. It was found that 30 µM Aß1-42 caused neuronal damages as evidenced by reduced cell viability, increased LDH release and ROS, collapsed MMP and elevated apoptosis. Secondly, Aß1-42-incubated cells were treated with 10, 20, 40 and 60 µM SEN for 24 h. SEN significantly reduced the damage of Aß1-42-incubated cells as shown by recovered cell viability and MMP, reduced apoptosis and ROS. Notably, SEN induced the formation of mitophagosomes and mitolysosomes, and elevated the conversion of LC3 I to LC3 II. Moreover, SEN down-regulated the expression of p62, promoted the accumulation of full-length PINK1 and the translocation of Parkin to mitochondria, decreased the expression of mitochondrial matrix protein HSP60, thus activating the PINK1/Parkin-mediated mitophagy. However, when cells were pretreated with 5 µM CsA (Cyclosporine A, a mitophagy inhibitor) for 2 h and then co-treated with 20 and 40 µM SEN for 24 h, the protective effects of SEN were compromised. CONCLUSIONS: The present study demonstrated that SEN could alleviate Aß1-42-induced cell damage through PINK1/Parkin-mediated mitophagy. Our findings justify the traditional use of P. tenuifolia in China with anti-aging or anti-neurodegenerative effects.

Senegenin Inhibits Aß1-42-Induced PC12 Cells Apoptosis and Oxidative Stress via Activation of the PI3K/Akt Signaling Pathway.

Background/Aim: Apoptosis and oxidative stress have been considered as key events in the pathogenesis of Alzheimer's disease (AD). Senegenin (Sen), the major and most effective ingredient of Radix Polygalae, which has anti-apoptotic and anti-oxidative effects. The aim of this study was to investigate the anti-apoptotic and anti-oxidant effects of Sen on Aß1-42-induced PC12 cells apoptosis and oxidative stress as well as its possible signaling pathway. Methods: Rat pheochromocytoma (PC12) cells were treated by 20 µM Aß1-42 and then divided into 5 different treatment groups (Control; Aß1-42 20 µM; Aß1-42 20 µM + Sen 10 µM; Aß1-42 20 µM + Sen 30 µM; Aß1-42 20µM + Sen 60 µM). PC12 cells activity was detected by MTT assay. Colony formation assay was performed to assess the clonogenic ability of cells. The cell apoptosis was detected by Annexin-V/PI staining. The pro-apoptotic protein (Bax), anti-apoptotic protein (Bcl-2), anti-oxidative stress factor (HO-1, Nuclear Nrf2, Total Nrf2) and pathway-related protein (Akt, P-Akt, PI3K, P-PI3K) were tested by Western blot. The reactive oxygen species (ROS) level was assessed with a DCFH-DA probe. Results: The results indicated that Sen dose-dependently increased cell viability and reduced the number of apoptotic cells. The ratio of P-PI3K/PI3K and P-Akt/Akt increased in a dose-dependent manner under the treatment of Sen, suggesting that Sen might activate the PI3K/Akt signaling pathway. Moreover, Sen upregulates the ratio of Bcl-2/Bax. Further study revealed that Sen can play an antioxidant role in enhancing HO-1, promoting Nrf2 nuclear translocation and reducing ROS accumulation to reduce oxidative stress. Conclusion: Sen is effective in inhibiting apoptosis and oxidative stress in Aß1-42-induced PC12 cells, which likely contribute to the development of novel therapies for AD.

Polygala tenuifolia: a source for anti-Alzheimer's disease drugs.

Context: Alzheimer's disease (AD) is a chronic neurodegenerative disease that originates from central nervous system lesions or recessions. Current estimates suggest that this disease affects over 35 million people worldwide. However, lacking effective drugs is the biggest handicap in treating AD. In traditional Chinese medicine (TCM), Polygala tenuifolia Willd. (Polygalaceae) is generally used to treat insomnia, memory dysfunction and neurasthenia.Objective: This review article explores the role of P. tenuifolia and its active components in anti-Alzheimer's disease.Methods: Literature for the last ten years was obtained through a search on PubMed, SciFinder, CNKI, Google Scholar, Web of Science, Science Direct and China Knowledge Resource Integrated with the following keywords: Polygala tenuifolia, polygalasaponin XXXII (PGS 32), tenuifolin, polygalacic acid, senegenin, tenuigenin, Alzheimer's disease.Results: Polygala tenuifolia and its active components have multiplex neuroprotective potential associated with AD, such as anti-Aß aggregation, anti-Tau protein, anti-inflammation, antioxidant, anti-neuronal apoptosis, enhancing central cholinergic system and promote neuronal proliferation.Conclusions: Polygala tenuifolia and its active components exhibit multiple neuroprotective effects. Hence, P. tenuifolia is a potential drug against Alzheimer's disease, especially in terms of prevention.

Content determination and quality analysis of five main components in Polygala tenuifolia from different habitats / 中草药

Objective: The contents of five main components in Polygala tenuifolia from different habitats were determined, which provided certain data support and theoretical basis for the quality evaluation system of P. tenuifolia. Methods: The contents of polygalaxanthone III, 3,6’-disinapoyl sucrose, polygalacic acid, senegenin and tenuifolin of roots from different wild samples were determined by HPLC. Then SPSS 20.0 and SIMCA 11.5 were used for difference analysis, correlation analysis, hierarchical cluster analysis (HCA), and principal component analysis (PCA). Results: The content of the five main components in wild P. tenuifolia samples from different habitats was significantly different. The 20 samples were placed into two clusters (I, II) by HCA and PCA. Cluster I comprised three samples with higher content of 3,6’-disinapoyl sucrose, polygalaxanthone III, senegenin and tenuifolin from Weinan, Xianyang in Shannxi Province, and Xinjiang in Shanxi Province, whereas cluster II contained the other 17 samples. Conclusion: The results showed that the main components of P. enuifolia from Weinan, Xianyang in Shannxi Province and Xinjiang in Shanxi Province were significantly higher than other origins, and which provided a reference for the quality control, selection of excellent germplasm and cultivation bases of P. tenuifolia.

Effects of different drying methods on active constituents of root bark and root of Polygala tenuifolia / 中草药

Objective To investigate the effects of different drying methods on composition and content of five active constituents in root bark and root of Polgala tenuifoliaroot. Methods The contents of polygalaxanthone III, 3,6’-disinapoyl sucrose, polygalacic acid, senegenin, and tenuifolin in root bark and root from different drying samples were determined by HPLC. Then the data analysis was performed by ANOVA and TOPSIS methods. Results There is a difference in the order of drying methods for bark and root of P. tenuifoliaroot. For P. tenuifoliaroot root bark, the order of different drying methods was microwave drying > 60 ℃ hot-air drying > 50 ℃ hot-air drying > 70 ℃ hot-air drying > freeze-drying > 40 ℃ hot-air drying > shade drying > sun drying; For P. tenuifolia root, the different drying methods were sorted by microwave drying > 60 ℃ hot-air drying > shade drying > sun drying > 50 ℃ hot-air drying > 40 ℃ hot-air drying > 70 ℃ hot-air drying > freeze-drying. Conclusion Combined with the production practice, this study suggests that microwave drying and hot-air drying at 60 ℃ are suitable drying methods for P. tenuifoliaroot bark and root, providing a basis for the determination of drying methods for the origin processing of P. tenuifolia.

Effect and mechanism of senegenin on proliferation and differentiation of neural stem cells / 国际中医中药杂志

Objective To study the effect and mechanism of senegenin on proliferation and differentiation of neural stem cells (NSCs). Methods The primary cultured NSCs were divided into the high-dose, medium-dose, low-dose group and normal control group (NC). The complete medium containing 10, 20 and 40 μmol/L senegenin was added to senegenin low-, middle-, and high- dose groups, and the NC group was routinely cultured. After 4 days of culture, CCK8 assay was used to detect cell viability, and microscopy was performed and the number of neurospheres was counted. Western blot was used to detect the expression of Nestin, TUJ1, GSK-3β, and p-GSK-3β (Ser9), and immunofluorescence staining was used to visualized Nestin and TUJ1. Results Compared with the control group, the number of NSCs neurospheres (32.78 ± 6.30, 40.93 ± 8.34, 45.37 ± 7.96 vs. 26.48 ± 5.19) and the proliferation (127.50% ± 9.31%, 138.13% ± 6.88%, 151.25% ± 9.38% vs. 100.00% ± 5.63%) in the low-, middle- and high-doses of senegenin group significantly increased (P<0.05 or P<0.01).The expression of TUJ1(2.21 ± 0.14,3.10 ± 0.16,3.30 ± 0.15 vs.1.00 ± 0.00)in the low-,middle- and high-doses of senegenin group significantly increased (P<0.05); and the expression of Nestin (0.36 ± 0.04,0.53 ± 0.05,0.46 ± 0.05 vs.1.00 ± 0.00)significantly decreased(P<0.05).The ration of p-GSK-3β(Ser9)/GSK-3β(2.31 ± 0.17,3.41 ± 0.11,3.59 ± 0.16 vs.1.00 ± 0.00)in the low-,middle-and high-doses of senegenin group significantly increased(P<0.01).The cell number of Nestin+(50.29 ± 3.18,45.28 ± 6.23,38.72 ± 5.31 vs. 75.27 ± 6.03) in the low-, middle- and high-doses of senegenin group significantly decreased (P<0.05 or P<0.01), and the cell number of TUJ1+(32.23 ± 4.36,38.23 ± 6.01,46.23 ± 4.36 vs.20.31 ± 5.23)significantly increased (P<0.01). Conclusions The senegenin may promote the proliferation and differentiation of NSCs through the activation of Wnt pathway.

Senegenin exerts anti-depression effect in mice induced by chronic un-predictable mild stress via inhibition of NF-κB regulating NLRP3 signal pathway.

Depressive disorder is a kind of affective disturbance disease. Emerging evidence has suggested that inflammation may contribute to the pathologic process of depressive disorder. Senegenin (SEN), a major bioactive constituent in Polygala tenuifolia Willd, has much bioactivity including anti-inflammatory and neuroprotection effects. However, the mechanism of its anti-depressant effect in mice remains unknown. This study aimed to explore the anti-depressant effects of SEN on behavioral changes and inflammatory responses in mice induced by chronic un-predictable mild stress (CUMS). SEN treatment remarkably ameliorated CUMS-induced behavioral abnormalities, such as improving locomotor activity, decreasing immobility time in Tail suspension test (TST) and Forced swimming test (FST), and increasing sucrose intake in Sucrose preference test (SPT). Additionally, SEN improve protein levels of Brain-derived neurotrophic factor (BDNF) and Neurotrophin-3 (NT-3) expression. In response to stress, p65 was activated to promote production of pro-IL-1ß, and then cleaved to mature IL-1ß by NOD-like receptor protein 3 (NLRP3) inflammasome pathway in hippocampus of CUMS mice. After SEN treatment, protein activation related to NLRP3 inflammasome pathway was down-regulated, which inhibited IL-1ß secretion. These results demonstrate that SEN plays an important role in treatment CUMS-induced depression in mice, possibly via suppression of pathway activation associated with NLRP3 inflammasome.

Quality research and analysis of Polygala tenuifolia with different commodity grade / 中草药

Objective To establish the main component analysis method of commodity grade Polygala tenuifolia, and to explore the correlation between the grade character of P. tenuifolia and the main chemistry. Methods P. tenuifolia of four markets were determined by high performance liquid chromatography (HPLC) to analyze the different level of medicine and explore the correlation between commodity grade and composition for P. tenuifolia. Results The main components determination of P. tenuifolia in different market grades were not completely consistent with the grade of commodity. Conclusion There are certain limitations of the commodity grading standards for P. tenuifolia in the medicinal materials market. It is necessary to establish a new grade quality standard for accurately evaluating the quality of P. tenuifolia. This study also provides some reference for the comprehensive quantitative evaluation of P. tenuifolia.

Senegenin inhibits neuronal apoptosis after spinal cord contusion injury.

Senegenin has been shown to inhibit neuronal apoptosis, thereby exerting a neuroprotective effect. In the present study, we established a rat model of spinal cord contusion injury using the modified Allen's method. Three hours after injury, senegenin (30 mg/g) was injected into the tail vein for 3 consecutive days. Senegenin reduced the size of syringomyelic cavities, and it substantially reduced the number of apoptotic cells in the spinal cord. At the site of injury, Bax and Caspase-3 mRNA and protein levels were decreased by senegenin, while Bcl-2 mRNA and protein levels were increased. Nerve fiber density was increased in the spinal cord proximal to the brain, and hindlimb motor function and electrophysiological properties of rat hindlimb were improved. Taken together, our results suggest that senegenin exerts a neuroprotective effect by suppressing neuronal apoptosis at the site of spinal cord injury.

Senegenin Ameliorate Acute Lung Injury Through Reduction of Oxidative Stress and Inhibition of Inflammation in Cecal Ligation and Puncture-Induced Sepsis Rats.

The purpose of this study was to assess the protective effect of senegenin on acute lung injury (ALI) in rats induced by sepsis. Rat ALI model was reproduced by cecal ligation and puncture (CLP). All rats were randomly divided into five groups: group 1 (control), group 2 (CLP), group 3 (CLP + senegenin 15 mg/kg), group 4 (CLP + senegenin 30 mg/kg), and group 5 (CLP + senegenin 60 mg/kg). CLP + senegenin groups received senegenin by gavage daily for consecutive 5 days, respectively, while the mice in control and CLP groups were given an equivalent volume of saline. We detected the lung wet/dry weight ratios and the histopathology of the lung. The levels of lung tissue myeloperoxidase (MPO), malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione (GSH) were determined. Meanwhile, the nuclear factor-kappa B (NF-κB) activation, tumor necrosis factor-alpha (TNF-α), and interleukin-1ß (IL-1ß) levels were studied. The results demonstrated that senegenin treatment significantly attenuated CLP-induced lung injury, including reduction of lung wet/dry weight ratio, protein leak, infiltration of leukocytes, and MPO activity. In addition, senegenin markedly decreased MDA content and increased SOD activity and GSH level. Serum levels of TNF-α and IL-1ß were also decreased by senegenin administration. Furthermore, senegenin administration inhibited the nuclear translocation of NF-κB in the lungs. These findings indicate that senegenin exerts protective effects on CLP-induced septic rats. Senegenin may be a potential therapeutic agent against sepsis.

Effects of Senegenin against hypoxia/reoxygenation-induced injury in PC12 cells.

OBJECTIVE: To investigate the effect and the potential mechanism of Senegenin (Sen) against injury induced by hypoxia/reoxygenation (H/R) in highly differentiated PC12 cells. METHODS: The cultured PC12 cells were treated with H/R in the presence or absence of Sen (60 µmol/L). Four groups were included in the experiment: control group, H/R group, H/R+Sen group and Sen group. Cell viability of each group and the level of lactate dehydrogenase (LDH) in culture medium were detected for the pharmacological effect of Sen. Hoechst 33258 staining and annexin V/propidium iodide double staining were used to analyze the apoptosis rate. Moreover, mitochondrial membrane potential (△Ψm), reactive oxygen species (ROS) and intracellular free calcium ([Ca(2+)]i) were measured by fluorescent staining and flow cytometry. Cleaved caspase-3 and activity of NADPH oxidase (NOX) were determined by colorimetric protease assay and enzyme linked immunosorbent assay, respectively. RESULTS: Sen significantly elevated cell viability (P<0.05), decreased the leakage of LDH (P<0.05) and apoptosis rate (P<0.05) in H/R-injured PC12 cells. Sen maintained the value of △Ψm (P<0.05) and suppressed the activity of caspase-3 (P<0.05). Moreover, Sen reduced ROS accumulation P<0.05) and [Ca(2+)]i increment (P<0.05) by inhibiting the activity of NOX (P<0.05). CONCLUSION: Sen may exert cytoprotection against H/R injury by decreasing the levels of intracellular ROS and [Ca(2+)]i, thereby suppressing the mitochondrial pathway of cellular apoptosis.

Effects of Senegenin against hypoxia/reoxygenation-induced injury in PC12 cells / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the effect and the potential mechanism of Senegenin (Sen) against injury induced by hypoxia/reoxygenation (H/R) in highly differentiated PC12 cells.</p><p><b>METHODS</b>The cultured PC12 cells were treated with H/R in the presence or absence of Sen (60 μmol/L). Four groups were included in the experiment: control group, H/R group, H/R+Sen group and Sen group. Cell viability of each group and the level of lactate dehydrogenase (LDH) in culture medium were detected for the pharmacological effect of Sen. Hoechst 33258 staining and annexin V/propidium iodide double staining were used to analyze the apoptosis rate. Moreover, mitochondrial membrane potential (△Ψm), reactive oxygen species (ROS) and intracellular free calcium ([Ca(2+)]i) were measured by fluorescent staining and flow cytometry. Cleaved caspase-3 and activity of NADPH oxidase (NOX) were determined by colorimetric protease assay and enzyme linked immunosorbent assay, respectively.</p><p><b>RESULTS</b>Sen significantly elevated cell viability (P<0.05), decreased the leakage of LDH (P<0.05) and apoptosis rate (P<0.05) in H/R-injured PC12 cells. Sen maintained the value of △Ψm (P<0.05) and suppressed the activity of caspase-3 (P<0.05). Moreover, Sen reduced ROS accumulation P<0.05) and [Ca(2+)]i increment (P<0.05) by inhibiting the activity of NOX (P<0.05).</p><p><b>CONCLUSION</b>Sen may exert cytoprotection against H/R injury by decreasing the levels of intracellular ROS and [Ca(2+)]i, thereby suppressing the mitochondrial pathway of cellular apoptosis.</p>

Senegenin Inhibits Hypoxia/Reoxygenation-Induced Neuronal Apoptosis by Upregulating RhoGDIα.

Neuronal apoptosis is an important event in hypoxia/reoxygenation (H/R)-induced neuronal injury. Senegenin (Sen), the predominant and most active component in Radix Polygalae root extracts, displays anti-apoptotic and anti-oxidative properties. Sen protects against H/R-induced neuronal apoptosis of highly differentiated PC12 cells and primary cortical neurons. Sen has also been investigated as a source of potential therapeutic targets. In this study, a proteomic approach was used to identify Sen-regulated proteins in PC12 cells. We found that Sen protected against H/R-induced neuronal apoptosis by upregulating RhoGDIα protein expression. The regulatory functions of RhoGDIα were investigated by knocking down RhoGDIα expression in PC12 cells using small interfering RNA (siRNA), followed by quantification of apoptosis and then altering the expression levels of apoptosis-related proteins. Our data show that after silencing RhoGDIα, the neuroprotective effects of Sen on H/R-induced PC12 cell apoptosis were absent. Furthermore, RhoGDIα silencing alleviated the Sen-mediated inhibition of the JNK pathway. Therefore, these findings indicated that Sen attenuates H/R-induced neuronal apoptosis by upregulating RhoGDIα expression and inhibiting the JNK pathway. In addition to the mechanism underlying neuroprotective effects of Sen, RhoGDIα was identified as a putative target of Sen based on a primary rat cortical neuron model of H/R-induced injury.

Protective effect of senegenin on splenectomy-induced postoperative cognitive dysfunction in elderly rats.

Postoperative cognitive dysfunction (POCD) is common in elderly patients. Senegenin, an active component of extracts from Polygala tenuifolia root, a traditional Chinese medicine, has neuroprotective and neuroregenerative effects. However, the mechanism underlying the effects of senegenin against postoperative cognitive impairment in elderly individuals has yet to be elucidated. The aim of this study was to investigate the protective effects of senegenin on the cognitive functions of elderly rats with splenectomy-induced POCD. Results from a Morris water maze test suggested that splenectomy induced a transient cognitive deficiency in the elderly rats; however, when the rats were treated with senegenin, the cognitive impairment was notably attenuated. Further experiments showed that senegenin significantly inhibited the mRNA and protein expression of several key pro-inflammatory cytokines, specifically, tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), IL-6 and IL-8, in the hippocampal tissues of elderly rats following splenectomy. In order to investigate the molecular mechanism involved, the expression and activity of the Toll-like receptor 4 (TLR4) signaling pathway was assessed. On day 1 postoperatively, it was observed that senegenin markedly suppressed the mRNA and protein expression of TLR4, myeloid differentiation factor 88 (MyD88) and TIR domain-containing adaptor-inducing interferon-ß (TRIF). Furthermore, the phosphorylation levels of nuclear factor-κB (NF-κB) p65 and inhibitor of NF-κB (IκBα) were also decreased following senegenin treatment on the first day subsequent to surgery. These results suggest that senegenin suppressed splenectomy-induced transient cognitive impairment in elderly rats, possibly by downregulating two signaling pathways involved in inflammation, TLR4/MyD88/NF-κB and TLR4/TRIF/NF-κB, to further inhibit the expression of key pro-inflammatory cytokines, specifically, TNF-α, IL-1ß, IL-6 and IL-8, and ultimately the neuroinflammation in the hippocampal tissues. In conclusion, the present study revealed that senegenin exhibited neuroprotective effects against splenectomy-induced transient cognitive impairment in elderly rats, which indicated that senegenin may be a promising agent for the treatment of POCD.

Simultaneous analysis of polygala acid, senegenin and 3,6'-disinapoylsucrose in rat plasma by liquid chromatography-tandem mass spectrometry: application to a pharmacokinetic study after oral administration.

A rapid and specific LC-MS/MS method has been developed for the simultaneous analysis of polygala acid, senegenin and 3,6'-disinapoylsucrose (DSS) in rat plasma. The method was applied to the pharmacokinetics studies of polygala acid, senegenin and DSS. The analysis was carried out on an Agilent Eclipse plus C18 reversed-phase column (100 × 4.6 mm, 3.5 µm) by gradient elution with methanol and ammonia (0.01%, v/v). The flow rate was 0.4 mL/min. All analytes including internal standard (IS) were monitored by selected reaction monitoring with an electrospray ionization source. Linear responses were obtained for polygala acid and DSS ranging from 2.5 to 2000 ng/mL, and senegenin ranging from 5 to 2000 ng/mL. The intra- and inter-day precisions (relative standard deviation) were <11.34 and 8.99%. The extraction recovery ranged from 70.89 ± 4.60 to 88.49 ± 3.26%, and that for the IS was 77.23 ± 3.68%. Stability studies showed that polygala acid, senegenin and DSS are stable during the preparation and analytical process. The validated method was successfully used to determine the concentration-time profiles of polygala acid, senegenin and DSS.

Senegenin Protects against Ischemia-reperfusion Inj ury by Ameliorating Endoplasmic Reticulum Stress in Mice / 华中科技大学学报(医学版)

Objective To investigate the protective role of senegenin(SEN)in the mouse ischemia-reperfusion injury and its mechanism.Methods Mice were divided at random into control group(the ligature was placed but not ligated for 225 min),is-chemia-reperfusion model group(the anterior descending coronary artery remained ligated for 45 min and then reperfused for 180 min)and SEN treatment group(30 mg/kg SEN was used 10 min before reperfusion).The myocardial infarct size was meas-ured by using Evans blue-TTC staining.Fluorescence assay was employed to detect the activity of Caspase-1 2 and Caspase-3 to assess the myocardial apoptosis.Western blot was used to detect the expression of two endoplasmic reticulum stress (ERS) markers,GRP78 and CHOP,in infarcted myocardia.Results Compared with the control group,the myocardial infarct size was significantly increased,the activities of Caspase-12 and Caspase-3 were increased by 4.71 fold and 3.37 fold,respectively,and the expression levels of ERS markers GRP78 and CHOP were conspicuously elevated in the ischemia-reperfusion model group.Pretreatment with SEN(30 mg/kg)could significantly reduce the myocardial infarct size,the activities of Caspase-12 and Caspase-3 and the expression levels of GRP78 and CHOP when compared with those in the ischemia-reperfusion model group and there was significant difference between the two groups(P<0.05 or P<0.01).Conclusion SEN can protect against the myocardial ischemia-reperfusion inj ury by ameliorating ERS-mediated myocardial apoptosis in mice.

Preliminary mechanism of senegenin against H/R-induced apoptosis of primary cortical neurons / 中国病理生理杂志

AIM:To explore the preliminary mechanism of senegenin ( Sen) on inhibiting hypoxia/reoxygenation ( H/R)-induced apoptosis of primary cortical neurons .METHODS:The cultured cortical neurons were randomly divided in-to normal group (control group), model group (H/R group), Sen+H/R group and Sen group.Flow cytometry was used to evaluate the effect of Sen on H/R-induced cell apoptosis .The protein levels of JNK , p-JNK, c-Jun, p-c-Jun, Bcl-2 and Bax were assessed by Western blotting .RESULTS:The apoptotic rate in H/R group was obviously higher than that in control group (P<0.05), while the apoptotic rate in Sen +H/R group was obviously lower than that in H/R group (P<0.05), suggesting that the model of apoptosis was established successfully .The results of Western blotting showed that Sen increased the expression of JNK and c-Jun, inhibited the phosphorylation of JNK and c-Jun (P<0.05), increased the protein level of Bcl-2 and inhibited the protein level of Bax in H/R treated primary cortical neurons (P<0.05).CONCLUSION:Sen has a protective effect against H/R-induced neuronal apoptosis by increasing the expression of JNK and c-Jun, inhibiting the phosphorylation of JNK and c-Jun, increasing the protein level of Bcl-2 and decreasing the protein level of Bax .