Escin-sorafenib synergy up-regulates LC3-II and p62 to induce apoptosis in hepatocellular carcinoma cells.

INTRODUCTION: Hepatocellular carcinoma (HCC) is a common solid cancer and the leading cause of cancer deaths worldwide. Sorafenib is the first drug used to treat HCC but its effectiveness needs to be improved, and it is important to find ways to treat cancer that combine sorafenib with other drugs. Synergistic therapies lower effective drug doses and side effects while enhancing the anticancer effect. PURPOSE: In the present study, the therapeutic potential of sorafenib in combination with escin and its underlying mechanism in targeting liver cancer has been established. STUDY DESIGN/METHODS: The IC50 of sorafenib and escin against HepG2, PLC/PRF5 and Huh7 cell lines were determined using MTT assay. The combination index, dose reduction index, isobologram and concentrations producing synergy were evaluated using the Chou-Talaly algorithm. The sub-effective concentration of sorafenib and escin was selected to analyze cytotoxic synergistic potential. Cellular ROS, mitochondrial membrane potential, annexin V and cell cycle were evaluated using a flow-cytometer, and autophagy biomarkers were determined using western blotting. Moreover, autophagy was knocked down using ATG5 siRNA to confirm its role. A DEN-induced liver cancer rat model was developed to check the synergy of sorafenib and escin. RESULTS: Different concentrations of escin reduced the IC50 of sorafenib in HepG2, PLC/PRF5 and Huh7 cell lines. Chou-Talaly algorithm determined cytotoxic synergistic concentrations of sorafenib and escin in these cell lines. Mechanistically, this combination over-expressed p62 and LC-II, reflecting autophagy block and induced late apoptosis, further reconfirmed by ATG5 knockdown. Sorafenib and escin combination  reduced HCC serum biomarker α-feto protein (α-FP) by 1.5 folds. This combination restricted liver weight, tumor number and size, also, conserved morphological features of liver cells. The combination selectively targeted the G0 /G1 phase of cancer cells. CONCLUSION: Escin and sorafenib combination potentially up-regulates p62 to block autophagy to induce late apoptosis in liver cancer cells.

ß-escin activates ALDH and prevents cigarette smoke-induced cell death.

BACKGROUND: The tobacco use is one of the biggest public health threats worldwide. Cigarette smoke contains over 7000 chemicals among other aldehydes, regarded as priority toxicants. ß-escin (a mixture of triterpenoid saponins extracted from the Aesculus hippocastanum. L) is a potent activator of aldehyde dehydrogenase (ALDH) - an enzyme catalyzing oxidation of aldehydes to non-toxic carboxylic acids. PURPOSE: The aim of this study was to evaluate the effect of ß-escin on ALDH activity, ALDH isoforms mRNA expression and cytotoxicity in nasal epithelial cells exposed to cigarette smoke extract (CSE). METHODS: Nasal epithelial cells from healthy non-smokers were treated with ß-escin (1 µM) and exposed to 5% CSE. After 6- or 24-hours of stimulation cell viability, DNA damage, ALDH activity and mRNA expression of ALDH isoforms were examined. RESULTS: 24 h ß-escin stimulation revised CSE induced cytotoxicity and DNA damage. Cells cultured with ß-escin or exposed to CSE responded with strong increase in ALDH activity. This effect was more pronounced in cultures treated with combination of ß-escin and CSE. The strongest stimulatory effect on ALDH isoform mRNA expression was observed in cells cultured simultaneously with ß-escin and CSE: at 6 h for ALDH1A1 and ALDH3A1, and at 24 h for ALDH1A3, ALDH3A2, ALDH3B1, and ALDH18A1. Combined ß-escin and CSE treatment prevented the CSE-induced inhibition of ALDH2 expression at 24 h. CONCLUSIONS: ß-escin is an effective ALDH stimulatory and cytoprotective agent and might be useful in the prevention or supportive treatment of tobacco smoke-related diseases.

Toxicity Derived from Interaction between Natural Compounds and Cancer Therapeutic Drugs Metabolized by CYP3A4: Lessons Learned from Two Clinical Case Reports.

The use of natural compounds and, in general, the use of Complementary and Alternative Medicine (CAM), is growing steadily worldwide, both due to commercial pressure and the increasing use of self-medication and the desire to manage one's own personal health and well-being. Patients facing a cancer diagnosis are also strongly pressured to use these compounds, which are often added to standard therapeutic regimens, that should instead be based solely on diagnostic and therapeutic care pathways (DTCP) or evidence-based medicine (EBM). This study presents two clinical cases of cancer patients who presented to the pharmaceutical consultation service (PCD-Pharmacy Clinical Desk) established at the CRO Institute in Aviano, Italy. Both patients were using natural products along with prescribed chemotherapy. In the first case, a 55-year-old woman diagnosed with bilateral breast cancer with bone metastases, who was using natural compounds based on diosmin, escin (or aescin) and resveratrol in combination with ribociclib anticancer therapy, a severe ADR (neutropenia) was identified as a consequence of the drug-natural product interaction. In the second case, following a detailed medication review by the PCD, we avoided taking a therapeutic treatment (with natural compounds) that in itself could potentially render chemotherapy ineffective in a 57-year-old woman with multiple infiltrating ductal carcinoma of the left breast; the patient was planning to take a natural product containing St. John's Wort tincture and lemon balm tincture, in combination with paclitaxel and trastuzumab. In addition, we describe the corrective actions taken, thus outlining the main objectives of the activity of the PCD's pharmacy counseling service: first, to identify, report, and manage adverse drug reactions (ADRs), and second, to identify therapeutic combinations that present potential risks of toxicity or ineffectiveness of the drug therapy itself.

ß-Escin: An Updated Review of Its Analysis, Pharmacology, Pharmacokinetics, and Toxicity.

[Formula: see text]-Escin is an oleanane-type pentacyclic triterpenoid saponin extracted from the seeds of Aesculus hippocastanum (AH), which is more widely distributed. [Formula: see text]-Escin sodium has been approved by the American FDA for clinical usage. This paper is intended to summarize an updated and comprehensive review of the pharmacological activities, pharmacokinetic properties, toxicity, and analytical methods of [Formula: see text]-escin. Studies have shown that [Formula: see text]-escin has significant antitumor, antiviral, anti-inflammatory, and other activities alongside less adverse effects and higher safety than other compounds. The review shows that the pharmacological effects of [Formula: see text]-escin involve mechanisms such as ATM/[Formula: see text]H2AX, RhoA/Rock, GSK-3[Formula: see text]/[Formula: see text]-Catenin, HER2/HER3/Akt, and PI3K/Akt signaling pathways, and Cyclin A, p21[Formula: see text], survivin, Bcl-2, Mcl-1, Caspases, TGF-[Formula: see text], MMPs, and TNF-[Formula: see text] among other inflammatory factors. [Formula: see text]-Escin has significant cytotoxicity; the use of the chitosan/xanthan gum-based polyelectrolyte complexes PA1 and PC-11 to modify it not only to reduces its toxicity, but also improves its drug efficacy. Because of this, these compounds may become a new research hotspot. [Formula: see text]-Escin in vivo metabolism can be converted by the CYP1A2 enzyme in the intestinal flora to produce [Formula: see text]-escin, deacylated, deglycosylated, and 21[Formula: see text]-[Formula: see text]-crotonoyl-protoescin, and the binding rate of the plasma proteins is higher than 90%. These are mainly metabolized by the liver, kidneys, and other organs, and excreted in the form of urine and feces. The number of reports on the specific mediators of the metabolism of [Formula: see text]-escin and their mechanisms and metabolites is relatively small; furthermore, the results are vague. Therefore, a complete and in-depth exploration of the pharmacokinetic characteristics of [Formula: see text]-escin is needed to provide a more complete and effective theoretical reference for the study of its pharmacodynamic activity.

Chitosan biopolymer functionalized with graphene oxide and titanium dioxide with Escin metallic nanocomposites for anticancer potential against colon cancer.

Our study produced GO-TiO2-chitosan-escin nanocomposites (GTCEnc), characterized them using physical and biological methods, and evaluated their potential as cancer treatment candidates. Standard protocols were used to produce GTCEnc. Nanocomposites are created using XRD, FTIR, UV-Vis, and PL spectroscopy analysis. The morphology and ultrastructure of nanocomposites were investigated using SEM and TEM. Nanocomposites containing TiO2, GO, chitosan, and escin nanostructures were characterized using diffraction, microscopy, and spectroscopy; the antimicrobial activity of GTCEnc was investigated. Various methods were used to test the anticancer activity of GTCEnc against COLO 205 cell lines, including MTT, EtBr/AO, DAPI, JC-1, Annexin-V/FITC, cell cycle analysis, and activation of pro-apoptotic markers, such as caspase-3, -8, and -9. The nanocomposites were cytotoxic to COLO 205 cells, with an IC50 of 22.68 µg/mL, but not to 293T cells. In cells treated with nanomaterials, cytotoxicity, nuclear damage, apoptosis induction, and free radical production were significantly increased. Our finding suggests that GTCEnc has potent anticancer and antibacterial activity in vitro because of its unique nanocomposite properties and antibacterial and anticancer activity in vitro. Additional research is required to understand the clinical efficacy of these nanocomposites.

Escin avoids hemorrhagic transformation in ischemic stroke by protecting BBB through the AMPK/Cav-1/MMP-9 pathway.

BACKGROUND: Hemorrhagic transformation (HT) seriously affects the clinical application of recombinant tissue plasminogen activator (rt-PA). The main strategy for combating HT is to keep the blood-brain barrier (BBB) stable. Escin is the active ingredient of Aesculus hippocastanum and a natural mixture of triterpene saponins, and may play a part in mitigation of HT. PURPOSE: This study sought to investigate the effect of Escin in improving rt-PA-induced HT, explore possible mechanisms, and provide new ideas for the treatment of clinical HT. STUDY DESIGN AND METHODS: In in vivo experiments, transient middle cerebral artery occlusion (tMCAO) was undertaken in 6-week-old and 12-month-old mice, and rt-PA was administered to induce HT injury. The inhibitory effect of Escin on HT and its protective effect on neurobehavior, the BBB, and cerebrovascular endothelial cells was determined. In in vitro experiments, bEnd.3 cells were injured by oxygen-glucose deprivation/reperfusion (OGD/R) and rt-PA. The protective effect of Escin was measured by the CCK8 assay, release of lactate dehydrogenase (LDH), and expression of tight junction (TJ) proteins. In mechanistic studies, the effect of Escin on the adenosine monophosphate-activated kinase / caveolin-1 / matrix metalloprotease-9 (AMPK/Cav-1/MMP-9) pathway was investigated by employing AMPK inhibitor and Cav-1 siRNA. RESULTS: In mice suffering from ischemia, rt-PA caused HT as well as damage to the BBB and cerebrovascular endothelial cells. Escin reduced the infarct volume, cerebral hemorrhage, improved neurobehavioral deficits, and maintained BBB integrity in rt-PA-treated tMCAO mice while attenuating bEnd.3 cells damage caused by rt-PA and OGD/R injury. Under physiological and pathological conditions, Escin increased the expression of p-AMPK and Cav-1, leading to decreased expression of MMP-9, which further attenuated damage to cerebrovascular endothelial cells, and these effects were verified with AMPK inhibitor and Cav-1 siRNA. CONCLUSION: We revealed important details of how Escin protects cerebrovascular endothelial cells from HT, these effects were associated with the AMPK/Cav-1/MMP-9 pathway. This study provides experimental foundation for the development of new drugs to mitigate rt-PA-induced HT and the discovery of new clinical application for Escin.

Yeast lacking the sterol C-5 desaturase Erg3 are tolerant to the anti-inflammatory triterpenoid saponin escin.

Escin is a mixture of over 30 glycosylated triterpenoid (saponin) structures, extracted from the dried fruit of horse chestnuts. Escin is currently used as an anti-inflammatory, and has potential applications in the treatment of arthritis and cancer. Engineered yeast would enable production of specific bioactive components of escin at industrial scale, however many saponins have been shown to be toxic to yeast. Here we report that a Saccharomyces cerevisiae strain specifically lacking the sterol C-5 desaturase gene ERG3, exhibits striking enhanced tolerance to escin treatment. Transcriptome analyses, as well as pre-mixing of escin with sterols, support the hypothesis that escin interacts directly with ergosterol, but not as strongly with the altered sterols present in erg3Δ. A diverse range of saponins are of commercial interest, and this research highlights the value of screening lipidome mutants to identify appropriate hosts for engineering the industrial production of saponins.

Molecular regulation of autophagy and suppression of protein kinases by aescin, a triterpenoid saponin impedes lung cancer progression.

Lung cancer is the most common and lethal cancer worldwide, yet there are no adequate and novel medications to control this illness. Previous reports suggested the potential of protein kinases to target lung cancer by regulating autophagy. This study establishes the role of aescin, a triterpenoid saponin, in targeting protein kinases responsible for lung cancer proliferation and mobility. The experimental data revealed that aescin significantly impedes lung cancer cell proliferation by downregulating protein kinases such as AKT, mTOR, MEK, and ERK. Downregulation of AKT-mTOR may promote a string of events inducing cytotoxic autophagy-mediated apoptosis in the presence of aescin. Besides, aescin decreases mobility and invasion by downregulating HIF-1α and VEGF gene expressions. Moreover, it successfully monitors EGFR gene expression, improves lung histology, and regulates biochemical parameters in a pre-clinical DEN-induced lung cancer model. Aescin was observed to be safe and non-toxic in both in silico toxicity predictions and ex vivo erythrocyte fragility assays. Hence, this study elucidates the molecular mechanism of aescin in targeting protein kinases and suggests that it could be a safer and more viable therapeutic agent for lung cancer treatment.

Role of Escin in breast cancer therapy: potential mechanism for inducing ferroptosis and synergistic antitumor activity with cisplatin.

Breast cancer (BC) has threatened women worldwide for a long time, and novel treatments are needed. Ferroptosis is a new form of regulated cell death that is a potential therapeutic target for BC. In this study, we identified Escin, a traditional Chinese medicine, as a possible supplement for existing chemotherapy strategies. Escin inhibited BC cell growth in vitro and in vivo, and ferroptosis is probable to be the main cause for Escin-induced cell death. Mechanistically, Escin significantly downregulated the protein level of GPX4, while overexpression of GPX4 could reverse the ferroptosis triggered by Escin. Further study revealed that Escin could promote G6PD ubiquitination and degradation, thus inhibiting the expression of GPX4 and contributing to the ferroptosis. Moreover, proteasome inhibitor MG132 or G6PD overexpression could partially reverse Escin-induced ferroptosis, when G6PD knockdown aggravated that. In vivo study also supported that downregulation of G6PD exacerbated tumor growth inhibition by Escin. Finally, our data showed that cell apoptosis was dramatically elevated by Escin combined with cisplatin in BC cells. Taken together, these results suggest that Escin inhibits tumor growth in vivo and in vitro via regulating the ferroptosis mediated by G6PD/GPX4 axis. Our findings provide a promising therapeutic strategy for BC.

Escin's Multifaceted Therapeutic Profile in Treatment and Post-Treatment of Various Cancers: A Comprehensive Review.

Although modern medicine is advancing at an unprecedented rate, basic challenges in cancer treatment and drug resistance remain. Exploiting natural-product-based drugs is a strategy that has been proven over time to provide diverse and efficient approaches in patient care during treatment and post-treatment periods of various diseases, including cancer. Escin-a plant-derived triterpenoid saponin-is one example of natural products with a broad therapeutic scope. Initially, escin was proven to manifest potent anti-inflammatory and anti-oedematous effects. However, in the last two decades, other novel activities of escin relevant to cancer treatment have been reported. Recent studies demonstrated escin's efficacy in compositions with other approved drugs to accomplish synergy and increased bioavailability to broaden their apoptotic, anti-metastasis, and anti-angiogenetic effects. Here, we comprehensively discuss and present an overview of escin's chemistry and bioavailability, and highlight its biological activities against various cancer types. We conclude the review by presenting possible future directions of research involving escin for medical and pharmaceutical applications as well as for basic research.

Chemotherapy-induced phlebitis via the GBP5/NLRP3 inflammasome axis and the therapeutic effect of aescin.

BACKGROUND AND PURPOSE: Intravenous infusion of chemotherapy drugs can cause severe chemotherapy-induced phlebitis (CIP) in patients. However, the underlying mechanism of CIP development remains unclear. EXPERIMENTAL APPROACH: RNA-sequencing analysis was used to identify potential disease targets in CIP. Guanylate binding protein-5 (GBP5) genetic deletion approaches also were used to investigate the role of GBP5 in NLR family pyrin domain containing 3 (NLRP3) inflammasome activation in lipopolysaccharide (LPS) primed murine bone-marrow-derived macrophages (BMDMs) induced by vinorelbine (VIN) in vitro and in mouse models of VIN-induced CIP in vivo. The anti-CIP effect of aescin was evaluated, both in vivo and in vivo. KEY RESULTS: Here, we show that the expression of GBP5 was upregulated in human peripheral blood mononuclear cells from CIP patients. Genetic ablation of GBP5 in murine macrophages significantly alleviated VIN-induced CIP in the experimental mouse model. Mechanistically, GBP5 contributed to the inflammatory responses through activating NLRP3 inflammasome and driving the production of the inflammatory cytokine IL-1ß. Moreover, aescin, a mixture of triterpene saponins extracted from horse chestnut seed, can alleviate CIP by inhibiting the GBP5/NLRP3 axis. CONCLUSION AND IMPLICATIONS: These findings suggest that GBP5 is an important regulator of NLRP3 inflammasome in CIP mouse model. Our work further reveals that aescin may serve as a promising candidate in the clinical treatment of CIP.

Anti-obesity effect of escin: a study on high-fat diet-induced obese mice.

OBJECTIVE: Obesity is characterized by excess fat accumulation and closely associated with insulin resistance and type 2 diabetes. We aimed at exploring the potential effect and mechanism of escin for the treatment of obesity using network pharmacology, and to verify the effect of escin on obese mice. MATERIALS AND METHODS: Escin targets were predicted by DrugBank and SwissTarget database. Potential targets for the treatment of obesity were identified based on the DisGeNET database. Comparative analysis was used to investigate the overlapping genes between escin targets and obesity treatment-related targets. Using STRING database and Cytoscape to analyze interactions among overlapping genes, hub genes were identified. Gene Ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were conducted in DAVID. High-fat diet (HFD) -induced obese mice were used to observe the anti-obesity effects of escin. The body weight, relevant biochemical markers and HE staining of fat and liver tissues were determined after escin was administered for 18 weeks. RESULTS: We screened 53 overlapping genes for escin and obesity. The mechanism of intervention of escin in treating obesity may involve 10 hub targets (STAT3, MTOR, NR3C1, IKBKB, PTGS2, MMP9, PRKCA, PRKCD, AR, CYP3A4). The screening and enrichment analysis revealed that the treatment of obesity using escin primarily involved 10 GO enriched terms and 13 related pathways. In vivo, escin can reduce the body weight of obese mice induced by HFD and improve lipid metabolism through lowering triglycerides (TG), total cholesterol (TC), and density lipoprotein (LDL) levels and increasing high density lipoprotein (HDL) levels and decreasing leptin level and increasing adiponectin (ADPN) level. Escin can regulate glucose metabolism caused by obesity through decreasing fasting glucose, postprandial blood glucose and regulating the level of insulin. These obese mice induced by HFD displayed the increased insulin resistance that was associated with the increased inflammatory cytokines, including interleukin 6 (IL-6), tumor necrosis factor α (TNF-α), and monocyte chemoattractant protein-1 (MCP-1). Escin may antagonize the increase of MCP-1 and partially antagonize the low-grade inflammation caused by obesity. From the morphological changes of fat and liver tissues stained by HE stain, escin could decrease the size of adipocytes and improve liver necrosis and fatty degeneration in obese mice fed by HFD. CONCLUSIONS: The network pharmacology of escin in treating obesity may involve 10 hub targets (STAT3, MTOR, NR3C1, IKBKB, PTGS2, MMP9, PRKCA, PRKCD, AR, CYP3A4), 10 GO enriched terms and 13 related pathways. In vivo, escin can be potentially used to prevent or treat obesity through reducing the weight, improving glucose and lipid metabolism, partially antagonizing the low-grade inflammation, and improved insulin resistance.

Escin suppresses immune cell infiltration and selectively modulates Nrf2/HO-1, TNF-α/JNK, and IL-22/STAT3 signaling pathways in concanavalin A-induced autoimmune hepatitis in mice.

The current study aims to investigate the possible protective effect of escin, the active constituent of a natural mixture of triterpene saponin glycoside, against immune-mediated hepatitis driven by concanavalin A (Con A) and to elucidate its possible underlying mechanisms. Adult male mice were administered Con A (15 mg/kg, intravenously) for 8 h. In the treated groups, mice were pretreated with escin daily (10 mg/kg in CMC, orally) for 4 days before Con A intoxication. In addition, escin was administered in a group to examine its effect on normal mice. Our results showed that escin inhibited Con A-induced elevation in liver enzymes (ALT, AST, and LDH) and curbed the Con A-induced hepatocyte necrosis and apoptosis together with abrogating the death pathway, JNK. Coincidentally, escin has shown a reduction in neutrophil, CD4+ T cell, and monocyte infiltration into the liver. In addition, escin modulated the cellular oxidant status by compensating for the Con A-depleted expression of the transcription factor Nrf2 and the stress protein hemeoxygenase-1. These effects were in good agreement with the restraining effect of escin on Con A-instigated overexpression of NF-κB and the pro-inflammatory cytokines TNF-α and IL-17A. Interestingly, Con A provoked the cellular protective pathway IL-22/STAT3, which was revoked by the escin pretreatment. In conclusion, escin shows extended antioxidant, anti-inflammatory, antinecrotic, and anti-apoptotic effects against Con A-induced immune-mediated hepatitis. These effects may collectively be via suppressing immune cell infiltration into the liver and selective modulation of Nrf2/HO-1, TNF-α/NF-κB, TNF-α/JNK, and IL-22/STAT3 signaling pathways.

Integrated systematic pharmacology analysis and experimental validation to reveal the mechanism of action of Semen aesculi on inflammatory bowel diseases.

BACKGROUND AND ETHNOPHARMACOLOGICAL RELEVANCE: Semen aesculi (SA), a traditional Chinese herb, has been used in the treatment of gastrointestinal disease for thousands of years. The escin was the main components of SA. A growing number of research showed that escin has a wide range of pharmacological activities in intestinal barrier dysfunction. AIM OF THE STUDY: Inflammatory bowel diseases (IBD) are an idiopathic disease of the intestinal tract with the hallmark features of mucosal inflammation and loss of barrier function. The theory of traditional Chinese medicine (TCM) suggests that SA plays a potential role in protecting the gastrointestinal diseases. The present study aimed to explore the effects of SA on the intestinal barrier under existing inflammatory conditions and elucidate underlying mechanisms. MATERIALS AND METHODS: The bioactive components of SA and their predicted biological targets were combined to develop a compound target pathway network. It is used to predict the bioactive components, molecular targets, and molecular pathways of SA in improving IBD. The ingredients of SA were extracted by decoction either in water and ethanol and separated into four fractions (AE, EE, PEE and PCE). The effects of extractions were evaluated in the lipopolysaccharide (LPS)-induced RAW264.7 macrophages cell model, LPS-induced intestinal barrier injury model and imodium-induced constipation model. The high-performance liquid chromatography (HPLC) analysis was performed to identify the bioactive components. RESULTS: The compound-target pathway network was identified with 10 bioactive compounds, 166 IBD-related targets, and 52 IBD-related pathways. In LPS-induced RAW264.7 cells, PEE and PCE significantly decreased nitric oxide (NO) production and TNF-α level. In mice, PEE and PCE administration improved intestinal barrier damage, increased intestinal motility, reduced levels of TNF-α and diamine oxidase (DAO). Furthermore, PEE and PCE administration not only decreased expression of p-Akt, p-IκBα, nuclear p-p65, and TNF-α level, but also increased expression of the zonula occludin-1 (ZO-1) in LPS-induced intestinal barrier injury model. The escin content of AE, EE, PEE and PCE gradually increased with an increase of the bioactivity. CONCLUSIONS: Escin was the main bioactive components of SA. The effects of SA on IBD were mediated by repairing the intestinal barrier and promoting intestinal motility. The mechanism of action of SA is related to inhibiting the Akt/NF-κB signaling pathway in intestinal tissue, at least, in part. Our results provide a scientific basis for further exploring the mechanisms involved in the beneficial effects of SA in IBD.

ß-Escin reduces cancer progression in aggressive MDA-MB-231 cells by inhibiting glutamine metabolism through downregulation of c-myc oncogene.

BACKGROUND: The c-myc oncogene, which causes glutamine dependence in triple negative breast cancers (TNBC), is also the target of one of the signaling pathways affected by ß-Escin. METHODS AND RESULTS: We sought to determine how c-myc protein affects glutamine metabolism and the proteins, glutamine transporter alanine-serine-cysteine 2 (ASCT2) and glutaminase (GLS1), in ß-Escin-treated MDA-MB-231 cells using glutamine uptake and western blot analysis. Cell viability, colony formation, migration and apoptosis were also evaluated in MDA-MB-231 cells in response to ß-Escin treatment using MTS, colony forming, wound healing, and Annexin-V assay. We determined that ß-Escin decreased glutamine uptake and reduced c-myc and GLS1 protein expressions and increased the expression of ASCT2. In addition, this inhibition of glutamine metabolism decreased cell proliferation, colony formation and migration, and induced apoptosis. CONCLUSIONS: In this study, it was suggested that ß-Escin inhibits glutamine metabolism via c-myc in MDA-MB-231 cells, and it is thought that as a result of interrupting the energy supply in these cells via c-myc, it results in a decrease in the carcinogenic properties of the cells. Consequently, ß-Escin may be promising as a therapeutic agent for glutamine-dependent cancers.

Escin attenuates oxidative damage, apoptosis and lipid peroxidation in a model of cyclophosphamide-induced liver damage.

To investigate the effects of escin (ES) on acute damage induced by alkylating agent, experimental rats were injected with cyclophosphamide (CPM) to cause liver damage. The animals were divided into four groups: Control Group, CPM (200 mg/kg), ES (10 mg/kg), CPM, and ES Groups. Immunohistopathological, hepatic histopathological, and biochemical changes were analyzed. The activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), malondyaldehyde (MDA), glutathion (GSH), total oxidant status (TOS) and total antioxidant status (TAS) in serum were all determined. Serum and immunohistopathology analysis revealed that MDA, ALT, AST, LDH, TOC and OSI, caspase-3 and Bax levels had increased while GSH, TAC, Bcl- 2 and OSI levels decreased in CPM Group when compared to Control Group. These findings appear to account for the severe damage detected. In the CPM + ES treated group, positive improvements were found in biochemical parameters as well as in cell-death and tissue-related damage parameters.The results show that ES considerably protects the rat liver against CPM-induced hepatotoxicity thanks to because of its anti-oxidant and anti-apoptotic properties.

Effectiveness of early administration of a single dose of steroids and escin after loss of signal on electromyographic signal recovery during neuromonitored thyroidectomy.

BACKGROUND: The aim of this study was to evaluate the effect of a single early administration of dexamethasone and escin after loss of signal (LOS) during a neuromonitored total thyroidectomy. METHODS: A retrospective analysis of results concerning consecutive patients undergoing total thyroidectomy was performed. Patients included in the study were divided into two groups: Group 1 for which a "wait and see" strategy was used; Group 2, receiving dexamethasone and escin immediately after LOS detection. RESULTS: Overall 37 patients were included in Group 1 and 35 in Group 2. LOS recovery occurring in 29.7% of cases (n. 11) versus 65.7% (n. 23) respectively (p < 0.001). Postoperative fibrolayngoscopy for patients without LOS recovery showed normal cord function in 4 out of 26 cases (15.4%) in Group 1 and in 7 out of 12 (58.3%) in Group 2 (p < 0.001). CONCLUSIONS: The early administration of dexamethasone and escin after LOS detection may achieve greater EMG signal recovery than a "wait and see" strategy.

Pretreatment with Coenzyme Q10 Combined with Aescin Protects against Sepsis-Induced Acute Lung Injury.

Sepsis-associated acute lung injury (ALI) is a critical condition characterized by severe inflammatory response and mitochondrial dysfunction. Coenzyme Q10 (CoQ10) and aescin (AES) are well-known for their anti-inflammatory activities. However, their effects on lipopolysaccharide (LPS)-induced lung injury have not been explored yet. Here, we asked whether combined pretreatment with CoQ10 and AES synergistically prevents LPS-induced lung injury. Fifty male rats were randomized into 5 groups: (1) control; (2) LPS-treated, rats received a single i.p. injection of LPS (8 mg/kg); (3) CoQ10-pretreated, (4) AES-pretreated, or (5) combined-pretreated; animals received CoQ10 (100 mg/kg), AES (5 mg/kg), or both orally for 7 days before LPS injection. Combined CoQ10 and AES pretreatment significantly reduced lung injury markers; 52.42% reduction in serum C-reactive protein (CRP), 53.69% in alkaline phosphatase (ALKP) and 60.26% in lactate dehydrogenase (LDH) activities versus 44.58, 37.38, and 48.6% in CoQ10 and 33.81, 34.43, and 39.29% in AES-pretreated groups, respectively. Meanwhile, combination therapy significantly reduced interleukin (IL)-1ß and tumor necrosis factor (TNF)-α expressions compared to monotherapy (p < 0.05). Additionally, combination therapy prevented LPS-induced histological and mitochondrial abnormalities greater than separate drugs. Western blotting indicated that combination therapy significantly suppressed nucleotide-binding oligomerization domain (NOD)-like receptors-3 (NLRP-3) inflammasome compared to separate drugs (p < 0.05). Further, combination therapy significantly decreased the expression of signaling cascades, p38 mitogen-activated protein kinases (p38 MAPK), nuclear factor kappa B (NF-κB)-p65, and extracellular-regulated kinases 1/2 (ERK1/2) versus monotherapy (p < 0.05). Interestingly, combined pretreatment significantly downregulated high mobility group box-1 (HMGB1) by 72.93%, and toll-like receptor 4 (TLR4) by -0.93-fold versus 61.92%, -0.83-fold in CoQ10 and 38.67%, -0.70-fold in AES pretreatment, respectively. Our results showed for the first time that the enhanced anti-inflammatory effect of combined CoQ10 and AES pretreatment prevented LPS-induced ALI via suppression of NLRP-3 inflammasome through regulation of HMGB1/TLR4 signaling pathway and mitochondrial stabilization.

Escin inhibits angiogenesis by suppressing interleukin­8 and vascular endothelial growth factor production by blocking nuclear factor­κB activation in pancreatic cancer cell lines.

Pancreatic cancer (PaCa) is one of the most aggressive types of cancer. Thus, the development of new and more effective therapies is urgently required. Escin, a pentacyclic triterpenoid from the horse chestnut, has been reported to exhibit antitumor potential by reducing cell proliferation and blocking the nuclear factor­κB (NF­κB) signaling pathway in several types of cancer. Our previous study reported that NF­κB enhanced the secretion of interleukin (IL)­8 and vascular endothelial growth factor (VEGF), thereby inducing angiogenesis in PaCa cell lines. In the present study, it was examined whether escin inhibited angiogenesis by blocking NF­κB activation in PaCa. It was initially confirmed that escin, at concentrations >10 µM, significantly inhibited the proliferation of several PaCa cell lines. Next, using immunocytochemical staining, it was found that escin inhibited the nuclear translocation of NF­κB. Furthermore, ELISA confirmed that NF­κB activity in the escin­treated PaCa cells was significantly inhibited and reverse transcription­quantitative PCR showed that the mRNA expression levels of tumor necrosis factor­α­induced IL­8 and VEGF were significantly suppressed following escin treatment in the PaCa cell lines. ELISA also showed that escin decreased the secretion of IL­8 and VEGF from the PaCa cells. Furthermore, tube formation in immortalized human endothelial cells was inhibited following incubation with the supernatants from escin­treated PaCa cells. These results indicated that escin inhibited angiogenesis by reducing the secretion of IL­8 and VEGF by blocking NF­κB activity in PaCa. In conclusion, escin could be used as a novel molecular therapy for PaCa.

Attenuation of Cardiac Autonomic Neuropathy by Escin in Diabetic Rats.

Cardiac autonomic neuropathy (CAN) is a least diagnosed complication of diabetes. Inflammation and oxidative stress play a crucial role in the pathophysiology of cardiomyopathy and neuropathy. Escin has anti-inflammatory activity and antioxidant activity. Hence, the present study was designed to evaluate the effect of escin in the management of CAN. Diabetes was induced in Sprague Dawley rats with streptozotocin (STZ). Diabetic animals were randomized in different groups after 6 weeks. Animals in the diabetic control group received no treatment, while animals in other groups received escin at dose 5, 10, and 20 mg/kg for 4 weeks. One group was kept as normal control. Various parameters like basic hemodynamic parameters, heart rate variability (HRV), oxidative stress parameters, and matrix metalloproteinase 9 (MMP-9) were assessed at the end of study. Escin significantly normalized hemodynamic parameters and HRV as compared to diabetic animals. Escin significantly reduced the malondialdehyde level and significantly increased reduced glutathione, catalase and superoxide dismutase levels in diabetic animals. Escin treatment significantly reduced plasma MMP-9 level in diabetic rats. The improvement in the studied parameters was found mainly with administration of higher doses of escin (10 and 20 mg/kg). The escin treatment mitigates CAN in diabetic rats. The results of study indicate that escin can be useful option for management of CAN.