Wogonin protects glomerular podocytes by targeting Bcl-2-mediated autophagy and apoptosis in diabetic kidney disease.

Diabetic kidney disease (DKD) is one of the microvascular complications of diabetes mellitus and a major cause of end-stage renal disease with limited treatment options. Wogonin is a flavonoid derived from the root of Scutellaria baicalensis Georgi, which has shown a potent renoprotective effect. But the mechanisms of action in DKD are not fully elucidated. In this study, we investigated the effects of wogonin on glomerular podocytes in DKD using mouse podocyte clone 5 (MPC5) cells and diabetic mice model. MPC5 cells were treated with high glucose (30 mM). We showed that wogonin (4, 8, 16 µM) dose-dependently alleviated high glucose (HG)-induced MPC5 cell damage, accompanied by increased expression of WT-1, nephrin, and podocin proteins, and decreased expression of TNF-α, MCP-1, IL-1ß as well as phosphorylated p65. Furthermore, wogonin treatment significantly inhibited HG-induced apoptosis in MPC5 cells. Wogonin reversed HG-suppressed autophagy in MPC5 cells, evidenced by increased ATG7, LC3-II, and Beclin-1 protein, and decreased p62 protein. We demonstrated that wogonin directly bound to Bcl-2 in MPC5 cells. In HG-treated MPC5 cells, knockdown of Bcl-2 abolished the beneficial effects of wogonin, whereas overexpression of Bcl-2 mimicked the protective effects of wogonin. Interestingly, we found that the expression of Bcl-2 was significantly decreased in biopsy renal tissue of diabetic nephropathy patients. In vivo experiments were conducted in STZ-induced diabetic mice, which were administered wogonin (10, 20, 40 mg · kg-1 · d-1, i.g.) every other day for 12 weeks. We showed that wogonin administration significantly alleviated albuminuria, histopathological lesions, and p65 NF-κB-mediated renal inflammatory response. Wogonin administration dose-dependently inhibited podocyte apoptosis and promoted podocyte autophagy in STZ-induced diabetic mice. This study for the first time demonstrates a novel action of wogonin in mitigating glomerulopathy and podocytes injury by regulating Bcl-2-mediated crosstalk between autophagy and apoptosis. Wogonin may be a potential therapeutic drug against DKD.

Identifying the molecular targets and mechanisms of xuebijing injection for the treatment of COVID-19 via network parmacology and molecular docking.

Xuebijing Injection have been found to improve the clinical symptoms of COVID-19 and alleviate disease severity, but the mechanisms are currently unclear. This study aimed to investigate the potential molecular targets and mechanisms of the Xuebijing injection in treating COVID-19 via network pharmacology and molecular docking analysis. The main active ingredients and therapeutic targets of the Xuebijing injection, and the pathogenic targets of COVID-19 were screened using the TCMSP, UniProt, and GeneCard databases. According to the 'Drug-Ingredients-Targets-Disease' network built by STRING and Cytoscape, AKT1 was identified as the core target, and baicalein, luteolin, and quercetin were identified as the active ingredients of the Xuebijing injection in connection with AKT1. R language was used for enrichment analysis that predict the mechanisms by which the Xuebijing injection may inhibit lipopolysaccharide-mediated inflammatory response, modulate NOS activity, and regulate the TNF signal pathway by affecting the role of AKT1. Based on the results of network pharmacology, a molecular docking was performed with AKT1 and the three active ingredients, the results indicated that all three active ingredients could stably bind with AKT1. These findings identify potential molecular mechanisms by which Xuebijing Injection inhibit COVID-19 by acting on AKT1.

Fabrication of Chinese Traditional Medicines incorporated collagen biomaterials for human bone marrow mesenchymal stem cells.

Chinese Traditional Medicines (CTMs) are very popular for therapeutic applications to cure several chronic diseases. Many researchers are trying to discover the potential application and actual mechanism of CTMs in order to scientifically prove their effects for commercial use. One of the main functions of CTMs is to aid stem cell regeneration. Since, this study was focused to fabricate CTMs incorporated fish collagen film, which has good biocompatibility in mammalian cell growth and thus investigated the effect on human Mesenchymal stem cells (hMSCs) proliferation and differentiation. In this study, three types of CTMs such as Genistein, Icariin, and Naringin were used for film fabrication. Mechanical properties of collagen films were improved by the addition of CTMs, especially in Collagen-Naringin films. Solubility and In-vitro biodegradation of collagen films were enhanced by the hydrophobicity and chemical interaction of CTMs with collagen. The proliferation rate was accelerated in hMSCs cultured on CTMs incorporated collagen films in a dose- and time-dependent manner. Proliferation biomarkers such as Ki-67 and BrdU levels were higher in hMSCs cultured on CTMs incorporated collagen films. The proliferative and differentiation effect of CTMs was further confirmed by higher gene expression of Collagen I, Runx2, c-Fos, SMAD3 and TGF-ß1 in hMSCs. Overall, this study provides a new insight on novel biomaterial fabrication using CTMs and fish collagen for making a compatible platform for in-vitro stem cell culture.

Protective effects of a unique combination of nutritionally active ingredients on risk factors and gene expression associated with atherosclerosis in C57BL/6J mice fed a high fat diet.

Atherosclerosis, an inflammatory disorder of the vasculature and the underlying cause of cardiovascular disease, is responsible for one in three global deaths. Consumption of active food ingredients such as omega-3 polyunsaturated fatty acids, flavanols and phytosterols has many beneficial effects on cardiovascular disease. However, their combined actions on the risk factors for atherosclerosis remains poorly understood. We have previously shown that a formulation containing each of these active components at physiologically relevant doses modulated several monocyte/macrophage processes associated with atherosclerosis in vitro, including inhibition of cytokine-induced pro-inflammatory gene expression, chemokine-driven monocyte migration, expression of M1 phenotype markers, and promotion of cholesterol efflux. The objectives of the present study were to investigate whether the protective actions of the formulation extended in vivo and to delineate the potential underlying mechanisms. The formulation produced several favourable changes, including higher plasma levels of HDL and reduced levels of macrophages and myeloid-derived suppressor cells in the bone marrow. The mRNA expression of liver-X-receptor-α, peroxisome proliferator-activated receptor-γ and superoxide dismutase-1 was induced in the liver and that of interferon-γ and the chemokine (C-X-C motif) ligand 1 decreased, thereby suggesting the potential mechanisms for many beneficial effects. Other changes were also observed such as increased plasma levels of triglycerides and lipid peroxidation that may reflect potential activation of brown fat. This study provides new insights into the protective actions and the potential underlying mechanisms of the formulation in vivo, particularly in relation to risk factors together with changes in systemic inflammation and hepatic lipid alterations associated with atherosclerosis and metabolic syndrome, and supports further assessments in human trials.

Pre-clinical evaluation of an innovative oral nano-formulation of baicalein for modulation of radiation responses.

There is an unmet medical need for non-toxic and effective radiation countermeasures for prevention of radiation toxicity during planned exposures. We have earlier shown that intraperitoneal administration of baicalein (BCL) offers significant survival benefit in animal model. Safety, tolerability, pharmacokinetics (PK) and pharmacodynamics of baicalein has been reported in pre-clinical model systems and also in healthy human volunteers. However, clinical translation of baicalein is hindered owing to poor bioavailability due to lipophilicity. In view of this, we fabricated and characterized in-situ solid lipid nanoparticles of baicalein (SLNB) with effective drug entrapment and release kinetics. SLNB offered significant protection to murine splenic lymphocytes against 4 Gy ionizing radiation (IR) induced apoptosis. Oral administration of SLNB exhibited ~70% protection to mice against whole body irradiation (WBI 7.5 Gy) induced mortality. Oral relative bioavailability of BCL was enhanced by over ~300% after entrapment in the SLNB as compared to BCL. Oral dosing of SLNB resulted in transient increase in neutrophil abundance in peripheral blood. Interestingly, we observed that treatment of human lung cancer cells (A549) with radioprotective dose of SLNB exhibited radio-sensitization as evinced by decrease in survival and clonogenic potential. Contrary to antioxidant nature of baicalein in normal cells, SLNB treatment induced significant increase in cellular ROS levels in A549 cells probably due to higher uptake and inhibition of TrxR. Thus, a pharmaceutically acceptable SLNB exhibited improved bioavailability, better radioprotection to normal cells and sensitized cancer cells to radiation induced killing as compared to BCL suggesting its possible utility as an adjuvant during cancer radiotherapy.

Alpinetin prevents inflammatory responses in OVA-induced allergic asthma through modulating PI3K/AKT/NF-κB and HO-1 signaling pathways in mice.

Allergic asthma is the most common type of asthma which characterized by inflammatory responses of the airways. Alpinetin, a flavonoid compound derived from the ginger family of medicinal herbs, possesses various biological properties including anti-inflammatory, anti-oxidant and other medical effects. In this study, we aimed to evaluate the effects of alpinetin on OVA-induced allergic asthma, and further to examine its molecular mechanisms underlying these processes in vivo and in vitro. Mice were sensitized and challenged with OVA to build allergic asthma model in vivo. Bronchoalveolar lavage fluid (BALF) was collected for inflammatory cells analysis and lung tissues were examined for histopathological examination. The levels of IL-5, IL-13, IL-4, IgE, TNF-α, IL-6 and IL-1ß were determined by the respective ELISA kits. The PI3K/AKT/NF-κB and HO-1 signaling pathways were examined by western blot analysis. The results showed that alpinetin significantly ameliorated OVA-induced pathologic changes of lungs, such as decreasing massive inflammatory cell infiltration and mucus hypersecretion, and reduced the number of inflammatory cells in BALF. Alpinetin also decreased the OVA-induced levels of IL-4, IL-5, IL-13 and IgE. Furthermore, alpinetin inhibited OVA-induced phosphorylation of p65, IκB, PI3K and AKT, and the activity of HO-1 in vivo. More importantly, these anti-inflammatory effects and molecular mechanisms of alpinetin has also been confirmed in LPS-stimulated RAW 264.7 macrophages in vitro. In conclusion, above results indicate that alpinetin exhibites a potent anti-inflammatory activity in allergic asthma through modulating PI3K/AKT/NF-κB and HO-1 signaling pathways, which would be used as a promising therapy agent for allergic asthma.

High dose of baicalin or baicalein can reduce tight junction integrity by partly targeting the first PDZ domain of zonula occludens-1 (ZO-1).

The tight junction (TJ) is the apical-most intercellular junction complex, serving as a biological barrier of intercellular spaces between epithelial cells. The TJ's integrity is maintained by a key protein-protein interaction between C-terminal motifs of claudins (CLDs) and the postsynaptic density 95 (PSD-95)/discs large/zonula occludens 1 (ZO-1; PDZ) domains of ZO-1. Weak but direct interaction of baicalin and its aglycon, baicalein-which are pharmacologically active components of Chinese skullcap (Radix scutellariae)-with ZO-1(PDZ1) have been observed in NMR experiments. Next, we observed TJ-mitigating activity of these flavonoids against Madin-Darby canine kidney (MDCK) II cells with the downregulation of subcellular localization of CLD-2 at TJs. Meanwhile, baicalein-but not baicalin-induced a slender morphological change of MDCK cells' shape from their normal cobblestone-like shapes. Since baicalin and baicalein did not induce a localization change of occludin (OCLN), a "partial" epithelial-mesenchymal transition (EMT) induced by these flavonoids was considered. SB431542, an ALK-5 inhibitor, reversed the CLD-2 downregulation of both baicalin and baicalein, while SB431542 did not reverse the slender morphology. In contrast, the MEK/ERK inhibitor U0126 reversed the slender shape change. Thus, in addition to inhibition of the ZO-1-CLD interaction, activation of both transforming growth factor-ß (TGF-ß) and MEK/ERK signaling pathways have been suggested to be involved in TJ reduction by these flavonoids. Finally, we demonstrated that baicalin enhanced the permeability of fluorescence-labeled insulin via the paracellular pathway of the Caco-2 cell layer. We propose that baicalin, baicalein, and Radix scutellariae extract are useful as drug absorption enhancers.

Self-Assembled Lecithin/Chitosan Nanoparticles Based on Phospholipid Complex: A Feasible Strategy to Improve Entrapment Efficiency and Transdermal Delivery of Poorly Lipophilic Drug.

PURPOSE: Lecithin/chitosan nanoparticles have shown great promise in the transdermal delivery of therapeutic agents. Baicalein, a natural bioactive flavonoid, possesses multiple biological activities against dermatosis. However, its topical application is limited due to its inherently poor hydrophilicity and lipophilicity. In this study, the baicalein-phospholipid complex was prepared to enhance the lipophilicity of baicalein and then lecithin/chitosan nanoparticles loaded with the baicalein-phospholipid complex were developed to improve the transdermal retention and permeability of baicalein. METHODS: Lecithin/chitosan nanoparticles were prepared by the solvent-injection method and characterized in terms of particle size distribution, zeta potential, and morphology. The in vitro release, the ex vivo and in vivo permeation studies, and safety evaluation of lecithin/chitosan nanoparticles were performed to evaluate the effectiveness in enhancing transdermal retention and permeability of baicalein. RESULTS: The lecithin/chitosan nanoparticles obtained by the self-assembled interaction of chitosan and lecithin not only efficiently encapsulated the drug with high entrapment efficiency (84.5%) but also provided sustained release of baicalein without initial burst release. Importantly, analysis of the permeation profile ex vivo and in vivo demonstrated that lecithin/chitosan nanoparticles prolonged the retention of baicalein in the skin and efficiently penetrated the barrier of stratum corneum without displaying skin irritation. CONCLUSION: These results indicate the potential of drug-phospholipid complexes in enhancing the entrapment efficiency and self-assembled lecithin/chitosan nanoparticles based on phospholipid complexes in the design of a rational transdermal delivery platform to improve the efficiency of transdermal therapy by enhancing its percutaneous retention and penetration in the skin.

Antiatherosclerotic Activity of Eriocitrin in High-Fat-Diet-Induced Atherosclerosis Model Rats.

Atherosclerosis is a multifactorial disease that develops and progresses in the arterial wall in response to a variety of stimuli. Among various other stimuli, hyperlipidemia is an extremely important factor that is correlated with the development of atherosclerosis. Lemon and citrus fruits contain various bioactive flavonoids, such as eriocitrin, that prevent obesity and related metabolic diseases. Therefore we concentrated on eriocitrin, a potent flavonoid with numerous therapeutic properties, particularly its beneficial lipid-lowering action in rats subjected to high fat diet. The anti-atherosclerotic efficacy of eriocitrin was assessed in rats administered a diet rich in fat. Wistar rats were divided into five groups consisting of six animals in all groups. Group I served the control, Group II was fed a high-fat diet (HFD), and the third and fourth groups were fed an HFD supplemented with varying doses of eriocitrin, and the last group was administered simvastatin for the last 30 days. Body weight, organ weight, lipid and lipoprotein parameters, cardiac and inflammatory markers, and histological examination were evaluated in animals induced with an HFD. Eriocitrin displayed a significant anti-atherosclerotic action by lowering the body weight, organ weight, reduction in lipid content, cardiac and inflammatory markers, myocardial changes confirmed by histopathology, malondialdehyde and increased antioxidant enzyme activities, nitric oxide, as well as 6-keto-PGF1α and high-density lipoprotein levels in rats fed an HFD. The findings of the experiment suggest that the anti-atherosclerotic action of eriocitrin was due to its modulatory activity in lipid metabolism. Considering the overall results of the study it can be validated that a use of flavonoid eriocitrin might be beneficial in altering HFD-induced alterations in atherosclerotic rats.

Influence of Hesperidin on Systemic Immunity of Rats Following an Intensive Training and Exhausting Exercise.

Intensive training and exhausting exercise can disrupt innate and acquired immunity. The flavanone hesperidin has shown immunomodulatory properties in physiological and some pathological conditions, and positive effects on exercise-induced oxidative stress. Nevertheless, it remains uncertain whether it also prevents exhausting exercise-induced immune alterations. The aim of this study was to establish the effect of oral hesperidin supplementation on the systemic immune system in rats following an intensive training and exhausting exercise. For this purpose, female Wistar rats were randomized into an intensive training group or a sedentary group. Intensive training was induced by running in a treadmill 5 days per week (including two exhausting tests) for five weeks. Throughout the training period, 200 mg/kg of hesperidin or vehicle was administered by oral gavage three times per week. At the end, blood, thymus, spleen and macrophages were collected before, immediately after and 24 h after an additional final exhaustion test. Hesperidin supplementation enhanced natural killer cell cytotoxicity and the proportion of phagocytic monocytes, attenuated the secretion of cytokines by stimulated macrophages, prevented the leukocytosis induced by exhaustion and increased the proportion of T helper cells in the thymus, blood and spleen. These results suggest that hesperidin can prevent exhausting exercise-induced immune alterations.

Naringenin ameliorates diabetic neuropathic pain by modulation of oxidative-nitrosative stress, cytokines and MMP-9 levels.

Diabetes mellitus is a serious debilitating epidemic affecting all social strata, imposing huge health, social and economic burdens. Diabetic neuropathic pain, an important microvascular complication of diabetes mellitus, characterized by allodynia and hyperalgesia, is recognized as one of the most difficult types of pain to treat. The development of tolerance, inadequate relief and potential toxicity of classical antinociceptives warrant the investigation of newer agents to relieve this pain. Reactive oxygen/nitrogen species, cytokines and matrix metalloproteinases (MMPs) are implicated in the pathogenesis of diabetic neuropathy. The present study was designed to explore the effect of naringenin, a citrus flavonoid, on streptozotocin induced diabetic neuropathic pain in Wistar rats. After 8 weeks of diabetes induction, rats developed neuropathy which was evident from marked hyperalgesia and allodynia associated with enhanced oxidative-nitrosative stress, release of inflammatory mediators (TNF-α, TGF-1ß), MMP-9 activation and decreased motor nerve conduction velocity. Treatment with naringenin (25, 50, 100 mg kg-1) for 4 weeks starting from the 5th week of streptozotocin injection significantly attenuated behavioral, biochemical and molecular changes, along with alterations in motor nerve conduction velocity in a dose-dependent manner. Moreover, diabetic rats treated with insulin-naringenin combination produced a more pronounced effect as compared to individual drugs. The major finding of the study is that insulin alone corrected the hyperglycemia and partially reversed the pain response in diabetic rats. However, combination with naringenin not only attenuated the diabetic condition but also reversed neuropathic pain through modulation of oxidative-nitrosative stress, inflammatory cytokine release and MMP inhibition in the diabetic rats. Modulation of MMP-9 by a natural flavonoid like naringenin seems to be a novel approach to target diabetic neuropathic pain.

Effect of a Combination of Citrus Flavones and Flavanones and Olive Polyphenols for the Reduction of Cardiovascular Disease Risk: An Exploratory Randomized, Double-Blind, Placebo-Controlled Study in Healthy Subjects.

A single-center, randomized, double-blind controlled trial was conducted to assess the efficacy of a food supplement based on a combination of grapefruit, bitter orange, and olive extracts administered for eight weeks (n = 51) versus placebo (n = 45) on reduction of cardiovascular risk in healthy volunteers. Study variables included flow-mediated vasodilation (FMD), blood pressure (BP), lipid profile, thrombotic status, oxidative stress biomarkers, inflammation-related biomarkers, anthropometric variables, quality of life, and physical activity. The per-protocol data set was analyzed. In the active product group, there were statistically significant within-group differences at eight weeks as compared with baseline in FMD, systolic and diastolic BP, total cholesterol, LDL-C, LDL-oxidase, oxidized/reduced glutathione ratio, protein carbonyl, and IL-6. Significant between-group differences in these variables were also found. Significant changes in anthropometric variables and quality of life were not observed in the study groups. Changes in the level of physical activity were not recorded. Treatment with the active product was well tolerated. All these findings, taken together, support a beneficial effect of supplementation with a mixture of grapefruit, bitter orange fruits, and olive leaf extracts on underlying mechanisms that may interact each other to decrease the cardiovascular risk in healthy people.

Metabolic profiling of mice plasma, bile, urine and feces after oral administration of two licorice flavonones.

ETHNOPHARMACOLOGICAL RELEVANCE: Licorice is an ancient food and medicinal plant. Liquiritigenin and liquiritin, two kinds of major flavonoes in licorice, are effective substances used as antioxidant, anti-inflammatory and tumor-suppressive food, cosmetics or medicines. However, their in vivo metabolites have not been fully explored. AIM OF STUDY: To clarify the metabolism of liquiritigenin and liquiritin in mice. MATERIALS AND METHODS: In this study, we developed a liquid chromatography coupled with quadrupole/time-of-flight mass spectrometry approach to determine the metabolites in mice plasma, bile, urine and feces after oral administration of liquiritigenin or liquiritin. The structures of those metabolites were tentatively identified according to their fragment pathways, accurate masses, characteristic product ions, metabolism laws or reference standard matching. RESULTS: A total of 26 and 24 metabolites of liquiritigenin or liquiritin were respectively identified. The products related with apigenin, luteolin or quercetin were the major metabolites of liquiritigenin or liquiritin in mice. Seven main metabolic pathways including (de)hydrogenation, (de)hydroxylation, (de)glycosylation, (de)methoxylation, acetylation, glucuronidation and sulfation were summarized to tentatively explain their biotransformation. CONCLUSION: This study not only can provide the evidence for in vivo metabolites and pharmacokinetic mechanism of liquiritigenin and liquiritin, but also may lay the foundation for further development and utilization of liquiritigenin, liquiritin and then licorice.

Eriocitrin Improves Adiposity and Related Metabolic Disorders in High-Fat Diet-Induced Obese Mice.

Eriocitrin (EC) is an abundant flavonoid in lemons, which is known as a strong antioxidant agent. This study investigated the biological and molecular mechanisms underlying the anti-obesity effect of EC in high-fat diet (HFD)-fed obese mice. C57BL/6N mice were fed an HFD (40 kcal% fat) with or without 0.005% (w/w) EC for 16 weeks. Dietary EC improved adiposity by increasing adipocyte fatty acid (FA) oxidation, energy expenditure, and mRNA expression of thermogenesis-related genes in brown adipose tissue (BAT) and skeletal muscle, whereas it also decreased lipogenesis-related gene expression in white adipose tissue. In addition to adiposity, EC prevented hepatic steatosis by diminishing lipogenesis while enhancing FA oxidation in the liver and fecal lipid excretion, which was linked to attenuation of hyperlipidemia. Moreover, EC improved insulin sensitivity by decreasing hepatic gluconeogenesis and proinflammatory responses. These findings indicate that EC may protect against diet-induced adiposity and related metabolic disorders by controlling thermogenesis of BAT and skeletal muscle, FA oxidation, lipogenesis, fecal lipid excretion, glucose utilization, and gluconeogenesis.

Tricarboxylic acid cycle dehydrogenases inhibition by naringenin: experimental and molecular modelling evidence.

The study of polyphenols' effects on health has been gaining attention lately. In addition to reacting with important enzymes, altering the cell metabolism, these substances can present either positive or negative metabolic alterations depending on their consumption levels. Naringenin, a citrus flavonoid, already presents diverse metabolic effects. The objective of this work was to evaluate the effect of maternal naringenin supplementation during pregnancy on the tricarboxylic acid cycle activity in offspring's cerebellum. Adult female Wistar rats were divided into two groups: (1) vehicle (1 ml/kg by oral administration (p.o.)) or (2) naringenin (50 mg/kg p.o.). The offspring were euthanised at 7th day of life, and the cerebellum was dissected to analyse citrate synthase, isocitrate dehydrogenase (IDH), α-ketoglutarate dehydrogenase (α-KGDH) and malate dehydrogenase (MDH) activities. Molecular docking used SwissDock web server and FORECASTER Suite, and the proposed binding pose image was created on UCSF Chimera. Data were analysed by Student's t test. Naringenin supplementation during pregnancy significantly inhibited IDH, α-KGDH and MDH activities in offspring's cerebellum. A similar reduction was observed in vitro, using purified α-KGDH and MDH, subjected to pre-incubation with naringenin. Docking simulations demonstrated that naringenin possibly interacts with dehydrogenases in the substrate and cofactor binding sites, inhibiting their function. Naringenin administration during pregnancy may affect cerebellar development and must be evaluated with caution by pregnant women and their physicians.

Protective and anticancer effects of orange peel extract and naringin in doxorubicin treated esophageal cancer stem cell xenograft tumor mouse model.

BACKGROUND: chemotherapy drugs are the common therapy for cancer cells with side effects. Recent studies reported that natural products may contribute to decreasing the side effects of chemotherapy drugs. Here, we aimed to investigate the effects of orange peel extract (OPE) and its main compound; naringin (NR) to protect the side effects of doxorubicin (Dox) in esophageal cancer stem cells (CSCs) derived tumors in vivo. METHODS: for this purpose, Esophageal cancer cell (YM1) derived spheres were treated in vitro with OPE, NR, Dox, Dox in combination with OPE or NR. The cell viability was assessed by XTT and the apoptosis was measured using Annexin/7-AAD and the cell cycle was also quantified by using PI staining method. The pluripotency related genes expression was carried out using qRT-PCR The protective effects of OPE and NR were evaluated by body weight evaluation and oxidative stress factors: malondialdehyde (MDA), total antioxidant capacity (TAC) and superoxide dismutase (SOD) measurement in xenograft mice tumor model injected with Dox. RESULTS: ESCC CSCs overexpress SOX2 and OCT4 pluripotency genes. OPE or NR can protect the cellular toxicity of Dox in vitro mainly by decreasing cellular apoptosis of ESCC CSCs however S-phase cell cycle arrest has not been affected significantly. In vivo experiments revealed that the use of Dox simultaneously with OPE or NR not only can reduce the tumor size but also the body weight of the treated nude mice were maintained in comparison to Dox alone. In contrast to Dox alone, Dox in combination with OPE or NR showed less systemic toxicity and decreased oxidative stress fraction circulation, however, OPE seemed as more protective. CONCLUSION: The results suggest that these natural compounds can be used as adjuvant therapy to lower systemic toxicity of chemotherapeutic agents like DOX in ESCC cancer stem cells treatment.

Safety and pharmacokinetics of naringenin: A randomized, controlled, single-ascending-dose clinical trial.

AIMS: To evaluate the safety and pharmacokinetics of naringenin in healthy adults consuming whole-orange (Citrus sinensis) extract. METHODS AND METHODS: In a single-ascending-dose randomized crossover trial, 18 adults ingested doses of 150 mg (NAR150), 300 mg (NAR300), 600 mg (NAR600) and 900 mg (NAR900) naringenin or placebo. Each dose or placebo was followed by a wash-out period of at least 1 week. Blood safety markers were evaluated pre-dose and 24 hours post-dose. Adverse events (AEs) were recorded. Serum naringenin concentrations were measured before and over 24 hours following ingestion of placebo, NAR150 and NAR600. Four- and 24-hour serum measurements were obtained after placebo, NAR300 and NAR900 ingestion. Data were analysed using a mixed-effects linear model. RESULTS: There were no relevant AEs or changes in blood safety markers following ingestion of any of the naringenin doses. The pharmacokinetic variables were: maximal concentration: 15.76 ± 7.88 µM (NAR150) and 48.45 ± 7.88 µM (NAR600); time to peak: 3.17 ± 0.74 hours (NAR150) and 2.41 ± 0.74 hours (NAR600); area under the 24-hour concentration-time curve: 67.61 ± 24.36 µM × h (NAR150) and 199.05 ± 24.36 µM × h (NAR600); and apparent oral clearance: 10.21 ± 2.34 L/h (NAR150) and 13.70 ± 2.34 L/h (NAR600). Naringenin half-life was 3.0 hours (NAR150) and 2.65 hours (NAR600). After NAR300 ingestion, serum concentrations were 10.67 ± 5.74 µM (4 hours) and 0.35 ± 0.30 µM (24 hours). After NAR900 ingestion, serum concentrations were 43.11 ± 5.26 µM (4 hours) and 0.24 ± 0.30 µM (24 hours). CONCLUSIONS: Ingestion of 150 to 900 mg doses of naringenin is safe in healthy adults, and serum concentrations are proportional to the dose administered. Since naringenin (8 µM) is effective in primary human adipocytes, ingestion of 300 mg naringenin twice/d will likely elicit a physiological effect.

Naringenin Increases Insulin Sensitivity and Metabolic Rate: A Case Study.

Our studies in primary human adipocytes show that naringenin, a citrus flavonoid, increases oxygen consumption rate and gene expression of uncoupling protein 1 (UCP1), glucose transporter type 4, and carnitine palmitoyltransferase 1ß (CPT1ß). We investigated the safety of naringenin, its effects on metabolic rate, and blood glucose and insulin responses in a single female subject with diabetes. The subject ingested 150 mg naringenin from an extract of whole oranges standardized to 28% naringenin three times/day for 8 weeks, and maintained her usual food intake. Body weight, resting metabolic rate, respiratory quotient, and blood chemistry panel including glucose, insulin, and safety markers were measured at baseline and after 8 weeks. Adverse events were evaluated every 2 weeks. We also examined the involvement of peroxisome proliferator-activated receptor α (PPARα), peroxisome proliferator-activated receptor γ (PPARγ), protein kinase A (PKA), and protein kinase G (PKG) in the response of human adipocytes to naringenin treatment. Compared to baseline, the body weight decreased by 2.3 kg. The metabolic rate peaked at 3.5% above baseline at 1 h, but there was no change in the respiratory quotient. Compared to baseline, insulin decreased by 18%, but the change in glucose was not clinically significant. Other blood safety markers were within their reference ranges, and there were no adverse events. UCP1 and CPT1ß mRNA expression was reduced by inhibitors of PPARα and PPARγ, but there was no effect of PKA or PKG inhibition. We conclude that naringenin supplementation is safe in humans, reduces body weight and insulin resistance, and increases metabolic rate by PPARα and PPARγ activation. The effects of naringenin on energy expenditure and insulin sensitivity warrant investigation in a randomized controlled clinical trial.

Anti-inflammatory effects of eriocitrin against the dextran sulfate sodium-induced experimental colitis in murine model.

Inflammatory bowel disease (IBD) is a continual ailment condition which engrosses the entire alimentary canal. The IBD can be primarily distinguished into two forms, ulcerative colitis, and Crohn's disease. The major symptoms of IBD include pustules or abscesses, severe abdominal pain, diarrhea, fistula, and stenosis, which may directly affect the patient's quality of life. A variety of mediators can stimulate the circumstances of IBD, some examples include infections by microbes such as bacteria, perturbation of the immune system and the surrounding environment of the intestines. Severe colitis was stimulated in the experimental animals through administering 4% dextran sulfate sodium (DSS) which is mixed in water ad libitum for 6 days. Eriocitrin (30 mg/kg) was then administered to the experimental animals followed by the induction of severe colitis to evaluate the therapeutic prospective of eriocitrin against the colon inflammation stimulated by DSS. In this study, eriocitrin (30 mg/kg) demonstrated significant (P < .05) attenuation activity against the DSS-stimulated severe colitis in experimental animals. Eriocitrin counteracted all of the clinical deleterious effects induced by DSS, such as body-weight loss, colon shortening, histopathological injury, accretion of infiltrated inflammatory cells at the inflamed region and the secretion of inflammatory cytokines. The results clearly showed that eriocitrin effectively attenuated DSS-induced acute colitis in experimental animals.

Naringin and Naringenin Relax Rat Tracheal Smooth by Regulating BKCa Activation.

Naringin and its aglycone, naringenin, occur naturally in our regular diet and traditional Chinese medicines. This study aimed to detect an effective therapeutic approach for cough variant asthma (CVA) through evaluating the relaxant effect of these two bioactive herbal monomers as antitussive and antiasthmatic on rat tracheal smooth muscle. The relaxant effect was determined by measuring muscular tension with a mechanical recording system in rat tracheal rings. Cytosolic Ca2+ concentration was measured using a confocal imaging system in primary cultured tracheal smooth muscle cells. In rat tracheal rings, addition of both naringin and naringenin could concentration dependently relax carbachol (CCh)-evoked tonic contraction. This epithelium-independent relaxation could be suppressed by BaCl2, tetraethylammonium, and iberiotoxin (IbTX), but not by glibenclamide. After stimulating primary cultured tracheal smooth muscle cells by CCh or high KCl, the intracellular Ca2+ increase could be inhibited by both naringin and naringenin, respectively. This reaction was also suppressed by IbTX. These results demonstrate that both naringin and naringenin can relax tracheal smooth muscle through opening big conductance Ca2+-activated K+ channel, which mediates plasma membrane hyperpolarization and reduces Ca2+ influx. Our data indicate a potentially effective therapeutic approach of naringin and naringenin for CVA.