Echinacoside attenuates glandular fibrosis in benign prostatic hyperplasia via inhibiting MKK6/MK2 signaling pathway

Background:Lower urinary tract symptoms commonly occur in the elderly population and seriously constrain the quality of life.Glandular fibrosis is an important pathobiological process in benign prostatic hyperplasia and is also a main inducing factor for benign prostatic hyperplasia-associated lower urinary tract symptoms.Cistanches species is an important herbal medicine resource and is traditionally used in ameliorating renal and prostatic defects.Methods:This study was to investigate the potential protective function of echinacoside(a bioactive compound from Cistanches)against prostatic fibrosis in mice and human benign prostatic hyperplasia epithelial-1 cell models.Results:It was found that echinacoside attenuated testosterone-induced prostatic hyperplasia and collagen deposition in mice,relieved prostate local inflammation and oxidative damage,and ameliorated prostatic epithelial-mesenchymal transition.Additionally,echinacoside inhibited the activation of the MKK6/MK2 signaling pathway both in vivo and in vitro.Conclusion:This study added new evidence for the anti-fibrotic function of echinacoside on the prostate and provided new insights for understanding its possible pharmacological mechanisms.

Effects of Echinacoside on Histio-central Levels of Active Mass in Middle Cerebral Artery Occlusion Rats

Objective To investigate the effects of echinacoside on the extracellular striatal levels of norepinephrine（NE）,dopamine（DA）,homovanillic acid（HVA）,3,4-dihydroxyphenylethanoid acid（DOPAC）,5-hydroxyindoleacetic acid（HIAA）,and 5-hydroxytryptamine（5-HT） in middle cerebral artery occlusion（MCAO rats.Methods The middle cerebral artery was occluded in male Sprague-Dawley rats.Three days later microdialysis probes were placed into the right striatum of MCAO rat brains and the brains were perfused with Ringer＇s solution at a rate of 1.5 μL/min.Cerebral microdialysates were collected every 30 minutes from awake and freely moving rats before assaying for NE,DA,HVA,DOPAC,HIAA,and 5-HT levels by reverse phase HPLC with electrochemistry.Results Three days after MCAO,the extracellular striatal levels of NE,DA,DOPAC,HIAA,HVA,and 5-HT of the MCAO rats increased significantly（at least P0.05 vs.control）.However,simultaneous treatment with echinacoside（30.0 or 15.0 mg/kg） attenuated these increases（at least P0.05 vs.non-treated model rats）.Conclusion These results imply that echinacoside may protect striatal dopa minergic neurons from the injury induced by MCAO and may help prevent and treat cerebral ischemic diseases.

Echinacoside suppresses cellular senescence of human fibroblastic cells by down-regulation of p53

Echinacoside is one of the phenylethanoids isolated from the stems of Cistanches salsa. Our previous research showed that echinacoside has anti-senescence activity. To investigate the mechanism of echinacoside＇s anti-senescence activity, the expressions of p53, p21, p16 and Rb at the mRNA and protein levels were determined. Results showed that the expression of p53 was down-regulated significantly in a dose dependent manner after treatment with echinacoside. Further experiments suggested that the down-regulation of p53 may be correlated with the upregulation of SIRT1. In addition, echinacoside may exhibit considerable higher affinity towards SIRT1 than resveratrol, according to our molecular docking simulation. In conclusion, we expect that echinacoside might be a promising candidate for regulating cell senescence.

Echinacoside Protects Against MPP+-Induced Neuronal Apoptosis via ROS/ATF3/CHOP Pathway Regulation

Echinacoside （ECH） is protective in a mouse model of Parkinson＇ s disease （PD） induced by 1-methyl-4- phenylpyridinium ion （MPP＋）. To investigate the mechanisms involved, SH-SYSY neuroblastoma ceils were treated with MPP＋ or a combination of MPP＋ and ECH, and the expression of ATF3 （activating transcription factor 3）, CHOP （C/EBP-homologous protein）, SCNA （synuclein alpha）, and GDNF （glial cell line-derived neurotrophic factor） was assessed. The results showed that ECH significantly improved cell survival by inhibiting the generation of MPP＋-induced reactive oxygen species （ROS）. In addition, ECH suppressed the ROS and MPP＋- induced expression of apoptotic genes （ATF3, CHOP, and SCNA）. ECH markedly decreased the MPP＋-induced cas- pase-3 activity in a dose-dependent manner. ATF3- knockdown also decreased the CHOP and cleaved caspase- 3 levels and inhibited the apoptosis induced by MPP＋. Interestingly, ECH partially restored the GDNF expression that was down-regulated by MPP＋. ECH also improved dopaminergic neuron survival during MPP＋ treatment and protected these neurons against the apoptosis induced by MPTP. Taken together, these data suggest that the ROS/ ATF3/CHOP pathway plays a critical role in mechanisms by which ECH protects against MPP＋-induced apoptosis in PD.

Echinacoside Alleviates Cognitive Impairment in Cerebral Ischemia Rats through α7nAChR-Induced Autophagy

Objectives:To evaluate the effect of echinacoside(ECH)on cognitive dysfunction in post cerebral stroke model rats.Methods:The post stroke cognitive impairment rat model was created by occlusion of the transient middle cerebral artery(MCAO).The rats were randomly divided into 3 groups by a random number table:the sham group(sham operation),the MCAO group(received operation for focal cerebral ischemia),and the ECH group(received operation for focal cerebral ischemia and ECH50 mg/kg per day),with 6 rats in each group,The infarct volume and spatial learning were evaluated by triphenyl tetrazolium chloride staining and Morris water maze.The expression of α7 nAChR in the hippocampus was detected by immunohistochemistry.The contents of acetylcholine(ACh),malondialdehyde(MDA),glutathione(GSH),superoxide dismutase(SOD),activities of choline acetyltransferase(ChAT),acetylcholinesterase(AChE),and catalase(CAT)were evaluated by enzyme linked immunosorbent assay.The neural apoptosis and autophagy were determined by TUNEL staining and LC3 staining,respectively.Results:ECH significantly lessened the brain infarct volume and ameliorated neurological deficit in infarct volume and water content(both P<0.01).Compared with MCAO rats,administration of ECH revealed shorter escape latency and long retention time at 7,14 and 28 days(all P<0.01),increased the α7 nAChR protein expression,ACh content,and ChAT activity,and decreased AChE activity in MCAO rats(all P<0.01).ECH significantly decreased MDA content and increased the GSH content,SOD,and CAT activities compared with MCAO rats(all P<0.05).ECH suppressed neuronal apoptosis by reducing TUNEL-positive cells and also enhanced autophagy in MCAO rats(all P<0.01).Conclusion:ECH treatment helped improve cognitive impairment by attenuating neurological damage and enhancing autophagy in MCAO rats.

Content Determination of Cistanches Herba in Zhenrongdan Mixture by HPLC

[Objectives]To establish a HPLC-UV method for the determination of Cistanches Herba in Zhenrongdan mixture.[Methods]According to the content determination method of Cistanches Herba in the 2020 edition of Chinese Pharmacopoeia,using HPLC-UV method,the content determination method of Cistanches Herba in Zhenrongdan mixture was established,and the content of echinacoside was determined.The related chromatographic conditions were explored,and the methodological investigation was carried out.[Results]The chromatographic conditions were determined as follows:octadecylsilane bonded silica gel as filler(C_(18));80%acetonitrile solution(containing 0.2%phosphoric acid)-0.2%phosphoric acid aqueous solution(15∶85)as mobile phase;detection wavelength was 330 nm.The number of theoretical plates should not be less than 3000 according to the peak of echinacoside.The methodological investigation showed that the method had better precision,accuracy and repeatability.Under the conditions of this study,echinacoside had better linear relationship in the range of 74.9-1498 ng,and the average recovery was 99.1%,RSD=0.5%(n=6).[Conclusions]A method for the determination of Cistanches Herba in Zhenrongdan mixture by HPLC-UV was established.The method is sensitive,rapid,accurate and suitable for the determination of Cistanches Herba in Zhenrongdan mixture.

Application of ~1H NMR-based metabolomics for discrimination of different parts and development of a new processing workflow for Cistanche deserticola

Cistanche deserticola(CD) is one of the two authoritative source plants of Cistanches Herba, a well-known medicinal plant. Herein,~1H NMR spectroscopy was employed to characterize the chemical profile and to distinguish the different parts, as well as to propose a new processing workflow for CD.Signal assignment was achieved by multiple one and two dimensional NMR spectroscopic techniques in combination with available databases and authentic compounds. The upper parts of the plant were distinguished from the lower parts by combining ~1H NMR spectroscopic dataset with multivariate statistical analysis. A new processing method that hyphenated steaming with freeze-drying, was demonstrated to be superior to either steaming coupled with oven-drying or direct freeze-drying via holistic ~1H NMR-based metabolomic characterization. Phenylethanoid glycosides, mainly echinacoside and acteoside, were screened out and confirmed as the chemical markers responsible for exhibiting the superiority of the new processing workflow, whereas serial primary metabolites, especially carbohydrates and tricarboxylic acid cycle metabolites, were found as the primary molecules governing the discrimination between the upper and lower parts of the plant. Collectively,~1H NMR spectroscopy was demonstrated as a versatile analytical tool to characterize the chemical profile and to guide the indepth exploitation of CD by providing comprehensive qualitative and quantitative information.

Prevention of Cistanche salsa Extract on Hepatic Fibrosis Induced by Carbon Tetrachloride in Rats

Objective To explore the antifibrotic effect of echinacoside on carbon tetrachloride(CCl4)-induced hepatic fibrosis in rats.Methods Male Wistar rats were randomly divided into normal control(n = 8),model(n = 14),and echinacoside treatment(n = 14) groups.The hepatic fibrosis model was induced by CCl4compositor.The rats were ig administered with echinacoside at a daily dose of 50 mg/kg.The anti-oxidant status,liver function parameters,and hepatic hydroxyproline content were detected by chromatometry.The serum levels of hyaluronic acid(HA),type IV collagen(CIV),type III precollagen(PIIIP),and laminin(LN) were assayed with radioimmunoassay.The hpatic injury was detected by haematoxylin-eosine staining.The deposition of collagen was observed with Masson staining.Results Echinacoside increased the superoxide dismutase activity and reduced the levels of malondialdehyde,aspartate aminotransferase,alanine aminotransferase,HA,CIV,PIIIP,and LN in serum.Echinacoside could also reduce the hydroxyproline content in liver,alleviate hepatic injury,and inhibit collagen deposition.Conclusion Echinacoside possesses antihepatic fibrosis effect.

Preliminary optimization of a Chinese herbal medicine formula based on the neuroprotective effects in a rat model of rotenone-induced Parkinson's disease

Objective： The main objective of this study was to preliminarily determine the optimum formulation of a Chinese herbal formula that may have neuroprotective effects against rotenone-induced Parkinson＇s disease （PD）. Methods： Seven recipes were made from Dihuang （DH, Rehmanniaglutinosa Libosch）, Roucongrong （RCR, Cistanche deserticola Y.C.Ma）, Niuxi （NX, Achyranthes bidentata BI.） and Shanzhuyu （SZY, Comus officinalis Sieb. et Zucc） in different proportions, according to the principles of uniform design （4 factors 7 levels）. Tyrosine hydroxylase （TH）-positive neurons in substantia nigra pars compacta （SNpc） were detected by immunohistochemistry and rotenone-exposure days necessary to induce PD symptoms were recorded. To probe one likely mechanism of the formulas, echinacoside （ECH） concentrations of all seven recipes were determined by high-performance liquid chromatography and related to number of TH-positive neu- rons. Results： The data showed that recipe 4 （DH：RCR：SZY：NX = 1：1：1：1 ） and recipe 7 （DH：RCR：SZY：NX = 7：5：3： 1） partially reversed rotenone-induced death of TH-positive neurons in the SNpc and significantly increased rotenone-e^posed days compared with model group. Pharmacologically, there was not a strong correlation between ECH concentration and TH-positive neurons. Conclusion： The investigated formulations of Chinese herbs had neuroprotective effects against PD mod- els, and the neuroprotective effects were weakly related to the proportion of key herbs. However the neu- roprotective effects of the formula may not result from a single active constituent.