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Apigenin, a flavone abundant in parsley and celery, is known to act on several CNS receptors, but its very poor water solubility (<0.001 mg/mL) impedes its absorption in vivo and prevents clinical use. Herein, apigenin was directly conjugated with glycine, l-phenylalanine, and l-lysine to give the corresponding carbamate derivatives, all of which were much more soluble than apigenin itself (0.017, 0.018, and 0.13 mg/mL, respectively). The Lys-apigenin carbamate 10 had a temporary sedative effect on the mice within 5 min of intraperitoneal administration (single dose of 0.4 mg/g) and could be detected in the mice brain tissues at a concentration of 0.82 μg/g of intact Lys-apigenin carbamate 10 and 0.42 ug/g of apigenin at 1.5 h. This study accomplished the delivery of apigenin across the BBB in a manner that might be applicable to other congeners, which should inform the future development of BBB-crossing flavonoids.

Apigenin trimethyl ether (5,7,4'-trimethoxyflavone, ATE), one of the key polymethoxyflavones present in black ginger (rhizome of Kaempferia parviflora) possesses various health-promoting activities. To optimize its medicinal application, the pharmacokinetics of ATE was assessed in Sprague-Dawley rats with emphases to identify the impacts from dose and repeated dosing on its major pharmacokinetic parameters. Plasma ATE levels were monitored by liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. Upon single intravenous administration (2 mg/kg), plasma levels of ATE declined through an apparent first-order process while dose-escalation to 4 and 8 mg/kg led to its non-linear disposition, which could be described by the Michaelis-Menten model. Similarly, dose-dependent oral pharmacokinetics was confirmed and when the dose was escalated from 5 to 15 and 45 mg/kg, much longer mean residence time (MRT 0→last ), higher dose-normalized maximal plasma concentration (C max /Dose) and exposure (AUC/Dose) were observed at 15 and/or 45 mg/kg. One-week daily oral administration of ATE at 15 mg/kg caused its accelerated elimination and the plasma exposure (AUC) after intravenous (2 mg/kg) and oral administration (15 mg/kg) dropped ~40 and 60%, respectively. As ATE displayed both dose- and time-dependent pharmacokinetics, caution is needed in the medicinal applications of ATE and/or black ginger., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The aim of this study was to investigate the protective effects and mechanism of isovitexin, a glycosylflavonoid isolated from rice hulls of Oryza sativa, on Lipopolysaccharide (LPS)/d-galactosamine (D-Gal)-induced acute liver injury. The mice were randomly divided into five groups: control group, LPS/D-Gal group, and LPS/D-Gal + isovitexin groups. The mice of LPS/D-Gal group were received of LPS (50 μg/kg) and D-gal (800 mg/kg) intraperitoneal. The mice of LPS/D-Gal + isovitexin groups were received isovitexin (25, 50, 100 mg/kg) 1 h before LPS/D-Gal treatment. The results showed that the severity of liver injury was attenuated by treatment of isovitexin, as confirmed by the decreased liver histopathologic changes, as well as serum AST and ALT levels. Furthermore, the levels of TNF-α in serum and liver tissues, MPO activity and MDA content were significantly inhibited by isovitexin. In addition, isovitexin significantly attenuated NF-κB phosphorylation induced by LPS/D-Gal. The expression of Nrf2 and HO-1 were significantly up-regulated by isovitexin. In conclusion, isovitexin could protect against LPS/D-Gal-induced liver injury by inhibiting inflammatory and oxidative responses. Isovitexin also had protective effects against carbon tetrachloride (CCl 4 )-induced liver injury. Isovitexin may used as a potential agent for the treatment of liver injury., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Flavones represent a class of polyphenols that are found in many plant-derived food sources. Herein, we provide evidence that the anti-inflammatory and antiproliferative effect of the flavone apigenin relies on the regulation of the gut microbiota by the NOD-like receptor family pyrin domain containing 6 (Nlrp6). When challenged by dextran sulfate sodium (DSS) in drinking water, mice were protected against colitis upon cohousing with apigenin-treated animals. In contrast, the protective effect was lost in the absence of Nlrp6. Sequencing of the 16S ribosomal RNA gene revealed a shift in the composition of the gut microbiota in apigenin-treated mice that was not observed in the absence of Nlrp6. Equally important, we find that the antiproliferative effect of apigenin was dominantly transmitted after cohousing, while being compromised in Nlrp6-deficient mice. In contrast, the symptoms of colitis were alleviated upon apigenin administration even in the absence of either caspase-1/11 or Asc. Collectively, these data indicate that apigenin modulated an inflammasome-independent mechanism by which Nlrp6 reprograms the gut microbiota for protecting mice against colitis. Our study highlights a modulation of the Nlrp6 signaling pathway by a prominent constituent of the human diet that may point toward improved ways to treat inflammatory bowel diseases.

Background: we have evaluated the antihypertensive effect of several flavonoid extracts in a rat model of arterial hypertension caused by chronic administration (6 weeks) of the nitric oxide synthesis inhibitor, L-NAME., Methods: Sprague Dawley rats received L-NAME alone or L-NAME plus flavonoid-rich vegetal extracts (Lemon, Grapefruit + Bitter Orange, and Cocoa) or purified flavonoids (Apigenin and Diosmin) for 6 weeks., Results: L-NAME treatment resulted in a marked elevation of blood pressure, and treatment with Apigenin, Lemon Extract, and Grapefruit + Bitter Orange extracts significantly reduced the elevated blood pressure of these animals. Apigenin and some of these flavonoids also ameliorated nitric oxide-dependent and -independent aortic vasodilation and elevated nitrite urinary excretion. End-organ abnormalities such as cardiac infarcts, hyaline arteriopathy and fibrinoid necrosis in coronary arteries and aorta were improved by these treatments, reducing the end-organ vascular damage., Conclusions: the flavonoids included in this study, specially apigenin, may be used as functional food ingredients with potential therapeutic benefit in arterial hypertension., Competing Interests: The authors declare no conflict of interest.

Alzheimer's disease (AD) is pathologically characterized by excessive accumulation of amyloid-beta (Aβ) within extracellular spaces of the brain. Aggregation of Aβ has been shown to trigger oxidative stress, inflammation, and neurotoxicity resulting in cognitive dysfunction. In this study, we use models of cerebral Aβ amyloidosis to investigate anti-amyloidogenic effects of scutellarin in vitro and in vivo. Our results show that scutellarin, through binding to Aβ42, efficiently inhibits oligomerization as well as fibril formation and reduces Aβ oligomer-induced neuronal toxicity in cell line SH-SY5Y. After nine months of treatment in APP/PS1 double-transgenic mice, scutellarin significantly improves behavior, reduces soluble and insoluble Aβ levels in the brain and plasma, decreases Aβ plaque associated gliosis and levels of proinflammatory cytokines TNF-α and IL-6, attenuates neuroinflammation, displays anti-amyloidogenic effects, and highlights the beneficial effects of intervention on development or progression of AD-like neuropathology., Competing Interests: The authors declare no conflict of interest.

Background: Scutellarin is the major constituent responsible for the clinical benefits of Erigeron breviscapus (Vant.) Hand.-Mazz which finds a long history of ethnopharmacological use in Traditional Chinese Medicine. Scutellarin as a pure compound is now under investigation for its protections against various tissue injuries., Purpose: This study aims to examine the effects of scutellarin on oxidative stress-induced vascular endothelial dysfunction and endothelial cell damage, and then to evaluate the therapeutic efficacy of scutellarin in preventing atherosclerosis in rats., Methods: Radical scavenging ability of scutellarin was determined in vitro. Impact of scutellarin on endothelium-dependent relaxation (EDR) of rabbit thoracic aortic rings upon 1, 1-diphenyl-2-picrylhydrazyl (DPPH) challenge was measured. Influences of scutellarin pre-treatment on the levels of reactive oxygen species (ROS), activities of antioxidant enzymes superoxide dismutase (SOD), glutathione peroxidase and catalase, and the expression of SOD1 and NADPH oxidase 4 (Nox4) in human umbilical vein endothelial cells (HUVECs) injured by H 2 O 2 were examined. Anti-atherosclerotic effect of scutellarin was evaluated in rats fed with high fat diet (HFD)., Results: Scutellarin showed potent antioxidant activity in vitro. Pretreatment of scutellarin retained the EDR of rabbit thoracic aortic rings damaged by DPPH. In H 2 O 2 injured-HUVECs the deleterious alterations in ROS levels and antioxidant enzymes activity were reversed by scutellarin and the mRNA and protein expression of SOD1 and Nox4 were restored also. Oral administration of scutellarin dose-dependently ameliorated hyperlipidemia in HFD-fed rats and alleviated oxidative stress in rat serum, mimicking the effects of reference drug atorvastatin., Conclusion: Scutellarin protects against oxidative stress-induced vascular endothelial dysfunction and endothelial cell damage in vitro and prevents atherosclerosis in vivo through antioxidation. The results rationalize further investigation into the clinical use of scutellarin in cardiovascular diseases., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

Liver cancer is a leading cause of cancer death worldwide, and novel chemotherapeutic drugs to suppress liver cancer are urgently required. Isovitexin (IV), a glycosylflavonoid, is extracted from rice hulls of Oryza sativa, and has various biological activities. However, the anti-tumor effect of IV against liver cancer has not yet been demonstrated in vitro or in vivo. In the present study, we showed that IV significantly suppressed the growth of liver cancer cells. Mechanistic studies indicated that IV induced apoptosis by the mitochondrial apoptotic pathway, as evidenced by the increase of Bax, cleaved Caspase-3, poly (ADP-ribose) polymerase (PARP), and cytoplasm Cyto-c released from mitochondria. In addition, IV resulted in autophagy in liver cancer cells, supported by the enhancement of LC3II, autophagy-related protein (Atg) 3, Atg5 and Beclin1. Suppressing autophagy using bafilomycin A1 (BFA) or siRNA Atg-5 reduced apoptotic cells in IV-treated cells, demonstrating that autophagy induction regulated apoptosis. Moreover, IV was found to cause endoplasmic reticulum (ER) stress in liver cancer cells, along with the promotion of ER stress-related molecules, including inositol-requiring enzyme 1α (IRE1α), X-box-binding protein-1s (XBP-1s), C/EBP homologous protein (CHOP) and glucose-regulated protein (GRP)-78. Of note, inhibition of ER stress by use of its inhibitor, tauroursodeoxycholate (TUDCA), significantly reversed IV-induced apoptosis and autophagy. In vivo, IV treatment showed significant tumor growth inhibition compared to the non-treated group. IV could therefore be a strong candidate for liver cancer prevention., (Copyright © 2018. Published by Elsevier Inc.)

A simple and rapid liquid chromatography-tandem mass spectrometry method was firstly developed for simultaneous quantification of neomangiferin, mangiferin, regaloside A, regaloside I, timosaponin BII, anemarsaponin E and timosaponin AIII in rat plasma after oral administration of Baihe Zhimu decoction, which plays an important role for the treatment of depression. The plasma samples were pretreated by a one-step direct protein precipitation with methanol. Separation of the seven components and scutellarin (IS) from endogenous components with high selectivity and sensitivity (LLOQ, 0.1-1.0ng/mL) was achieved within 10min using Poroshell 120 EC-C 18 column (150mm×3.0mm, 2.7μm). A gradient mobile phase consisting of acetonitrile and water (containing 5mM ammonium acetate) was applied at a flow rate of 0.4mL/min. Detection and measurement were performed on an AB Sciex QTRAP ® 5500 mass spectrometer in multiple reactions monitoring mode. The intra- and inter-day precisions were all within 15% and the accuracies were in the range of -10.4% to 14.5%. The recovery ranged from 90.8 to 113.8%. The validated method was successfully applied to pharmacokinetic study of the seven components in normal and chronic unpredicted mild stress-induced depression model rats., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Chronic cerebral hypoperfusion is considered as a pivotal factor of cognitive impairment that occurs in cerebrovascular diseases. This study investigated the ameliorating effect of scutellarin (SCT) on spatial cognitive impairment and β-amyloid (Aβ) formation in rats with chronic cerebral hypoperfusion induced by permanent bilateral common carotid artery occlusion (pBCAO). SCT is a flavonoid in medicinal herb of Erigeron breviscapus (vant.) Hand. Mazz. known to have neuroprotective, antioxidative and anti-inflammatory effects. However, the beneficial effect and pivotal mechanism of SCT on cognitive impairment are still unclear. SCT was treated orally with two doses (10 or 30 mg/kg) for 4 weeks. Results of Morris water maze test performed on the ninth week after pBCAO revealed that SCT (30 mg/kg)-treated rats had significantly shortened escape latencies in acquisition training trials, significantly prolonged swimming time at the platform and its surrounding zone, significant increase in memory score, significant reduction in the number of target heading, and significant reduction in the time required for the first target heading during the retention trial compared to rats in the sham-control group. SCT significantly inhibited the production of Aβ(1-40) and Aβ(1–42) in brain tissues. However, SCT significantly upregulated the expression levels of amyloid precursor protein and β-site APP-converting enzyme-1 in the hippocampus. In addition, SCT significantly inhibited the activation of Iba1-expressing microglia in brain tissues. The results suggest that SCT can exert ameliorating effect on spatial cognitive impairment caused by chronic cerebral hypoperfusion through suppressing Aβ formation and microglial activation in brain tissues. Therefore, SCT can be used as a beneficial drug for vascular dementia and Alzheimer's disease.

We aimed to construct a novel nano-assembly carrying apigenin (APG), a hydrophobic drug, and to evaluate its in vitro targeting ability for A549 cells overexpressing CD44 receptors. The apigenin-loaded sodium hyaluronate nano-assemblies (APG/SH-NAs) were assembled by multiple non-covalent interactions between sodium hyaluronate (SH) and APG. The prepared APG/SH-NAs exhibited a small average size and narrow particle size distribution. In addition, satisfactory encapsulation efficiency and drug loading were obtained. The drug release curves indicated that APG/SH-NAs achieved a sustainable drug-release effect due to the presence of hydrophilic materials. The in vitro cytotoxicity of APG/SH-NAs against A549 cells and HepG2 cells was evaluated, and the results indicated that the prepared APG/SH-NA showed higher cytotoxicity compared to apigenin suspensions. When CD44 receptors on the surface of A549 cells were blocked by the addition of excess SH, the cytotoxicity of APG/SH-NA was significantly reduced. However, similar phenomena were not observed in HepG2 cells with relatively low CD44 receptor expression. The resulting APG/SH-NAs could efficiently facilitate the internalization of APG into A549 cells, which might be due to their high affinity for CD44 receptors. Moreover, the apoptotic rate of APG/SH-NAs through receptor-mediated endocytosis mechanism was higher than that of the other groups in A549 cells. Thus, such nano-assemblies were considered to be an effective transport system with excellent affinity for CD44 receptors to allow the SH-mediated targeted delivery of APG., (Copyright © 2017. Published by Elsevier Ltd.)

Background: Apigenin is a flavonoid compound, widely distributed in natural plants. Various studies have suggested that apigenin has inhibitory effects towards several drug transporters, such as the organic anion transporting (OAT) polypeptides, 1B1 and 1B3 (OATP1B1 and OATP1B3). However, the mechanism by which apigenin interacts with OAT1 has not been well studied., Methods: MDCK cells stably-expressing OAT1 were used to examine the inhibitory effects of apigenin on OAT1. UPLC-MS/MS was used to evaluate the in vitro and in vivo effects of apigenin on the uptake of acyclovir by OAT1. Cytotoxicity was determined by the cell viability, MTT assays., Results: Apigenin effectively inhibited the activity of OAT1 in a dose-dependent manner with an IC50 value of 0.737μM. Pre-incubation of cells with apigenin caused a time-dependent inhibition (TDI) of OAT1. Additionally, we examined the interactions between apigenin and acyclovir or adefovir. Data showed that apigenin (1μM) significantly blocked the uptake of acyclovir by OAT1 in vitro with an inhibition rate of 55%. In vivo, apigenin could increase the concentration of acyclovir in plasma when co-administered with acyclovir. Importantly, the MTT assays showed that, at a dose of 50μM, apigenin significantly reduced the cytotoxicity of adefovir and substantially increased cell viability from 50.6% to 112.62%., Conclusion: Our results demonstrate that apigenin regulates OAT1, and can cause TDI or herb-drug interaction (HDI) when used in combination with acyclovir or adefovir. Therefore, apigenin could be used as a nephroprotective agent when used in combination with the substrates of OAT1., (Copyright © 2017 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.)

The purpose of this study was to prepare, optimize, and characterize a cationic lipid nanoparticle (CLN) system containing multicomponent drugs using a molecular dynamics model as a novel method of evaluating formulations. Puerarin (PUE) and scutellarin (SCU) were used as model drugs. CLNs were successfully prepared using melt-emulsion ultrasonication and low temperature-solidification technique. The properties of CLNs such as morphology, particle size, zeta potential, entrapment efficiency (EE), drug loading (DL), and drug release behavior were investigated. The CLNs were evaluated by corneal permeation, preocular retention time, and pharmacokinetics in the aqueous humor. Additionally, a molecular dynamics model was used to evaluate the formulation. Electron microscopy results showed that the nanoparticles were approximately spherical in shape. The EE (%) and DL (%) values of PUE and SCU in the optimal formulation were 56.60±3.73, 72.31±1.96 and 1.68±0.17, 2.44±1.14, respectively. The pharmacokinetic study in the aqueous humor showed that compared with the PUE and SCU solution, the area under the concentration-time curve (AUC) value of PUE was enhanced by 2.33-fold for PUE-SCU CLNs ( p <0.01), and the SCU AUC was enhanced by 2.32-fold ( p <0.01). In the molecular dynamics model, PUE and SCU passed through the POPC bilayer, with an obvious difference in the free energy well depth. It was found that the maximum free energy required for PUE and SCU transmembrane movement was ~15 and 88 kJ·mol -1 , respectively. These findings indicated that compared with SCU, PUE easily passed through the membrane. The diffusion coefficient for PUE and SCU were 4.1×10 -3 ±0.0027 and 1.0×10 -3 ±0.0006 e -5 cm 2 ·s -1 , respectively. Data from the molecular dynamics model were consistent with the experimental data. All data indicated that CLNs have a great potential for ocular administration and can be used as an ocular delivery system for multicomponent drugs. Moreover, the molecular dynamics model can also be used as a novel method for evaluating formulations., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

The purpose of this study was to develop different injectable nanosized drug delivery systems (NDDSs) i.e. liposome, lipid nanocapsule (LNC) and polymeric nanocapsule (PNC) encapsulating apigenin (AG) and compare their characteristics to identify the nanovector(s) that can deliver the largest quantity of AG while being biocompatible. Two liposomes with different surface characteristics (cationic and anionic), a LNC and a PNC were prepared. A novel tocopherol modified poly(ethylene glycol)-b-polyphosphate block-copolymer was used for the first time for the PNC preparation. The NDDSs were compared by their physicochemical characteristics, AG release, storage stability, stability in serum, complement consumption and toxicity against a human macrovascular endothelial cell line (EAhy926). The diameter and surface charge of the NDDSs were comparable with previously reported injectable nanocarriers. The NDDSs showed good encapsulation efficiency and drug loading. Moreover, the NDDSs were stable during storage and in fetal bovine serum for extended periods, showed low complement consumption and were non-toxic to EAhy926 cells up to high concentrations. Therefore, they can be considered as potential injectable nanocarriers of AG. Due to less pronounced burst effect and extended release characteristics, the nanocapsules could be favorable approaches for achieving prolonged pharmacological activity of AG using injectable NDDS., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Apigenin is a natural flavonoid compound widely distributed in a variety of vegetables, medicinal plants and health foods. This study aimed to examine the protective effect of apigenin against d-galactosamine (D-GalN)/lipopolysaccharide (LPS)-induced mouse liver injury and to investigate the potential biochemical mechanisms. The results showed that after oral administration of apigenin 100-200 mg/kg for 7 days, the levels of serum alanine aminotransferase and aspartate aminotransferase were decreased, and the severity of liver injury was alleviated. Importantly, apigenin pretreatment increased the levels of hepatic nuclear factor erythroid 2-related factor 2 (Nrf-2) and peroxisome proliferator-activated receptor γ (PPARγ) protein expressions as well as superoxide dismutase, catalase, glutathione S-transferase and glutathione reductase activities, decreased the levels of hepatic nuclear factor-κB (NF-κB) protein expression and tumor necrosis factor-α. These findings demonstrated that apigenin could prevent the D-GalN/LPS-induced liver injury in mice, and its mechanisms might be associated with the increments of Nrf-2-mediated antioxidative enzymes and modulation of PPARγ/NF-κB-mediated inflammation., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Aim: The capability of reducing fibrotic and inflammatory responses in lung tissues represents a gold standard for evaluating the efficacy of therapeutic interventions for treating idiopathic pulmonary fibrosis (IPF). A wide variety of therapeutic strategies have been employed in clinic to treat PF, but limited success has been obtained. Apigenin (4, 5, 7-trihydroxyflavone) is a member of flavonoid family that exerts anti-inflammatory and anti-fibrosis effects. In this study, we explore the potential therapeutic effect of apigenin in lung fibrosis., Materials and Methods: Apigenin was employed to treat IPF in a bleomycin-induced PF rat model. Apigenin was loaded onto a biodegradable polymer carrier (nanoparticle, NP) to improve its bio-solubility and bio-availability. The properties (e.g. size, apigenin loading and release profile) of the apigenin loaded polymer carrier were well-characterized. In vitro study was performed to assess the impact of apigenin on pulmonary cell viability, growth, as well as inflammatory and pro-fibrosis responses in pulmonary cells. The impact of apigenin on the production of inflammatory cytokines (e.g. TGF-β, TNF-α) and pro-fibrosis factors in bronchoalveolar lavage fluid and pulmonary cells from lung tissues was also investigated., Results: Our results showed, apigenin has anti-fibrosis effect by inhibition fibrosis related cytokines expression. And compared with apigenin in soluble form, the strategic release of apigenin is more effective in inhibiting pulmonary fibrosis and inflammation., Conclusion: Our finding suggested that apigenin loaded on polymeric carrier might be an effective treatment for pulmonary fibrosis patients.

Objectives: Scutellarin (SCU) is a traditional Chinese medicine used for the treatment of ischaemic cerebrovascular disease, but its clinic applications have been limited due to its poor water solubility, poor bioavailability and short half-life. In comparison with the conventional oral and intravenous administration, nasal administration may help targeting the drug more directly to brain. Thus, we proposed to employ a novel SCU-loaded HP-β-CD/chitosan nanoparticles (CD/CS-SCU-NPs) to deliver SCU to brain through the nasal route., Methods: CD/CS-SCU-NPs were prepared by an ionic cross-linking method. The NPs formulation was tested in vivo in C57BL mice. The concentrations of SCU in brain and plasma after intranasal and oral administration of the CD/CS-SCU-NPs and after intranasal administration of SCU solution (SCU-SL) were determined and brain targeting parameters were calculated., Key Findings: Compared to the intranasal administration of SCU-SL, intranasal and oral administration of the CD/CS-SCU-NPs increased accumulation of SCU in brain, indicating that CD/CS-SCU-NPs have obvious brain targeting advantage, although the advantage is more evident after intranasal administration., Conclusions: Findings from in-vivo study indicated that much higher SCU brain exposure was observed after intranasal administration of the CD/CS-SCU-NPs. Administration of CD/CS-SCU-NPs through the nasal route would have potential to treat ischemic cerebrovascular disease., (© 2017 Royal Pharmaceutical Society.)

The apigenin-phospholipid phytosome (APLC) was developed to improve the aqueous solubility, dissolution, in vivo bioavailability, and antioxidant activity of apigenin. The APLC synthesis was guided by a full factorial design strategy, incorporating specific formulation and process variables to deliver an optimized product. The design-optimized formulation was assayed for aqueous solubility, in vitro dissolution, pharmacokinetics, and antioxidant activity. The pharmacological evaluation was carried out by assessing its effects on carbon tetrachloride-induced elevation of liver function marker enzymes in a rat model. The antioxidant activity was assessed by studying its effects on the liver antioxidant marker enzymes. The developed model was validated using the design-optimized levels of formulation and process variables. The physical-chemical characterization confirmed the formation of phytosomes. The optimized formulation demonstrated over 36-fold higher aqueous solubility of apigenin, compared to that of pure apigenin. The formulation also exhibited a significantly higher rate and extent of apigenin release in dissolution studies. The pharmacokinetic analysis revealed a significant enhancement in the oral bioavailability of apigenin from the prepared formulation, compared to pure apigenin. The liver function tests indicated that the prepared phytosome showed a significantly improved restoration of all carbon tetrachloride-elevated rat liver function marker enzymes. The prepared formulation also exhibited antioxidant potential by significantly increasing the levels of glutathione, superoxide dismutase, catalase, and decreasing the levels of lipid peroxidase. The study shows that phospholipid-based phytosome is a promising and viable strategy for improving the delivery of apigenin and similar phytoconstituents with low aqueous solubility., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: Although apigenin possesses diverse pharmacological activities, its utilization as a bioactive substance is limited by poor oral bioavailability. The aim of this study was to improve the bioavailability of apigenin by co-administration of friedelin., Methods: To achieve this, the intestinal permeability of apigenin in the absence or presence of friedelin was investigated in both Caco-2 cells and single-pass rat intestinal perfusion models., Results: The apparent permeability coefficients (P app ) of apigenin in the presence of friedelin were substantially increased by 1.63- and 1.60-fold in Caco-2 cells and single-pass rat intestinal perfusion models, respectively. Such increases in the P app indicated that friedelin could significantly enhance the absorption of apigenin into the body. The increased bioavailability of apigenin in rats following the oral administration of apigenin 50mg/kg body weight with friedelin 50mg/kg body weight was further confirmed by increases in the peak concentration of apigenin (C max ), elimination half-life (T 1/2 ) and area under the plasma concentration-time curve (AUC)., Conclusions: Friedelin suppressed ATPase activity of P-glycoprotein (P-gp) indicated that the improved bioavailability of apigenin may be ascribed to P-gp inhibition by the co-administered friedelin., (Copyright © 2017 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.)

Considerable attention has been paid to the introduction of novel naturally occurring plant-derived radiosensitizer compounds in order to augment the radiation efficacy and improve the treatment outcome of different tumors. This study was therefore undertaken to evaluate the antitumor, antiangiogeneic, and synergistic radiosensitizing effects of apigenin, a dietary flavonoid, and/or cryptotanshinone, a terpenoid isolated from the roots of Salvia miltiorrhiza, against the growth of solid Ehrlich carcinoma in female mice. Apigenin (50 mg/kg body weight) and/or cryptotanshinone (40 mg/kg body weight) was intraperitoneally (i.p.) injected into non-irradiated or γ-irradiated (6.5 Gy whole-body γ-irradiation) solid Ehrlich carcinoma-bearing mice for 30 consecutive days. Investigations included molecular targets involved in proliferation, inflammation, angiogenesis, and tumor invasiveness. Treatment with apigenin and/or cryptotanshinone significantly suppressed the growth of solid Ehrlich carcinoma tumors and demonstrated a synergistic radiosensitizing efficacy together with γ-irradiation. These effects were achieved through downregulating the expression of angiogenic and lymphangiogenic regulators, including signal transducer and activator of transcription 3, vascular endothelial growth factor C, and tumor necrosis factor alpha, suppressing matrix metalloproteinase-2 and -9 activities, which play a key role in tumor invasion and metastasis, and enhancing apoptosis via inducing cleaved caspase-3 and granzyme B levels. Histological findings of solid Ehrlich carcinoma tumors verified the recorded data. In conclusion, a synergistic radiosensitizing efficacy for apigenin and cryptotanshinone was demonstrated against Ehrlich carcinoma in the current in vivo murine model, representing therefore a potential therapeutic strategy for increasing the radiation response of solid tumors.

Objective: Ficus deltoidea Jack (mistletoe fig) is an ornamental plant found in various parts of the world and used as traditional herbal medicine in some countries. This study investigated the potential use of F deltoidea leaf extract to mitigate osteoarthritis (OA) in ovariectomized (estrogen-deficient postmenopausal model) rats and the mechanisms involved. Diclofenac was used for comparison., Methods: Sprague-Dawley female rats (12 weeks old) were divided randomly into five groups (n = 6): healthy; nontreated OA; OA + diclofenac (5 mg/kg); OA + extract (200 mg/kg); and OA + extract (400 mg/kg). Two weeks after bilaterally ovariectomy, OA was induced by intra-articular injection of monosodium iodoacetate into the right knee joints. After 28 days of treatment, the rats were evaluated for knee OA via physical (radiological and histological observations), biochemical, enzyme-linked immunosorbent assay, and gene expression analysis, for inflammation and cartilage degradation biomarkers., Results: The osteoarthritic rats treated with the extract, and diclofenac showed significant reduction of cartilage erosion (via radiological, macroscopic, and histological images) compared with untreated osteoarthritic rats. The elevated serum interleukin-1β, prostaglandin E2, and C-telopeptide type II collagen levels in osteoarthritic rats were significantly reduced by F deltoidea leaf extract comparable to diclofenac. The extract significantly down-regulated the interleukin-1β, prostaglandin E2 receptor, and matrix metalloproteinase-1 mRNA expressions in the osteoarthritic cartilages, similar to diclofenac., Conclusions: F deltoidea leaf extract mitigated postmenopausal osteoarthritic joint destruction by inhibiting inflammation and cartilage degradation enzymes, at an effective extract dose equivalent to about 60 mg/kg for humans. The main bioactive compounds are probably the antioxidative flavonoids vitexin and isovitexin.

Scutellaria altissima L. is a common traditional Chinese medicine used to treat inflammation in some countries. Scutellarin, an active major flavone glycoside isolated from the traditional Chinese medicine Scutellaria altissima L., has been shown to offer various beneficial biochemical effects on cerebrovascular diseases and inflammation. However, the antiproliferative effects of Scutellarin in prostate cancer and the underlying mechanism are not fully elucidated. In the present study, we aimed to ascertain whether Scutellarin inhibits cancer cell growth and to further explore the molecular mechanism. Scutellarin enhanced the sensitivity of prostate cancer cells to cisplatin. MTT assays revealed that cell viability was significantly decreased in the prostate cancer cells treated with Scutellarin. Flow cytometric analysis indicated that Scutellarin suppressed cell proliferation by promoting G2/M arrest and inducing apoptosis. We employed western blotting to delineate the underlying mechanisms involved in the G2/M arrest and apoptosis. Comet assay and γH2AX immunocytochemistry were used to detect levels of DNA damage in PC3 cells exposed to Scutellarin and/or cisplatin. Our data revealed that Scutellarin significantly induced prostate cancer cell apoptosis by activating the caspase cascade. An increase in the Bax/Bcl-2 ratio, depolarization of mitochondrial membrane potential and cell cycle arrest at G2/M phase were accompanied by the apoptosis induction. Additionally, Scutellarin altered the protein expression of cell cycle and apoptosis regulatory genes by downregulating Cdc2, cyclin B1 and Bcl-2 and upregulating caspase-3, caspase-9 and Bax in prostate cancer cells. Furthermore, Scutellarin sensitized PC3 cells to cisplastin treatment in a dose-dependent manner. Taken together, our data confirmed the cytotoxicity of Scutellarin against prostate cancer PC3 cells and provide new findings in regards to Scutellarin sensitizing prostate cancer cells to chemotherapy. Our findings suggest that Scutellarin has potential to be used as a novel antineoplastic therapeutic candidate for prostate cancer patients.

Drug resistance in chemotherapy is a serious obstacle for the successful treatment of cancer. Drug resistance is caused by various factors, including the overexpression of P‑glycoprotein (P‑gp, MDR1). The development of new, useful compounds that overcome drug resistance is urgent. Apigenin, a dietary flavonoid, has been reported as an anticancer drug in vivo and in vitro. In the present study, we investigated whether apigenin is able to reverse drug resistance using adriamycin‑resistant breast cancer cells (MCF‑7/ADR). In our experiments, apigenin significantly decreased cell growth and colony formation in MCF‑7/ADR cells and parental MCF‑7 cells. This growth inhibition was related to the accumulation of cells in the sub‑G0/G1 apoptotic population and an increase in the number of apoptotic cells. Apigenin reduced the mRNA expression of multidrug resistance 1 (MDR1) and multidrug resistance‑associated proteins (MRPs) in MCF‑7/ADR cells. Apigenin also downregulated the expression of P‑gp. Apigenin reversed drug efflux from MCF‑7/ADR cells, resulting in rhodamine 123 (Rho123) accumulation. Inhibition of drug resistance by apigenin is related to the suppression of the signal transducer and activator of transcription 3 (STAT3) signaling pathway. Apigenin decreased STAT3 activation (p‑STAT3) and its nuclear translocation and inhibited the secretion of VEGF and MMP‑9, which are STAT3 target genes. A STAT3 inhibitor, JAK inhibitor I and an HIF‑1α inhibitor decreased cell growth in MCF‑7 and MCF‑7/ADR cells. Taken together, these results demonstrate that apigenin can overcome drug resistance.

The aim of this study was to develop a novel mix micelles system composing of two biocompatible copolymers of Soluplus ® and Pluronic F127 to improve the solubility, oral bioavailability of insoluble drug apigenin (AP) as model drug. The AP-loaded mixed micelles (AP-M) were prepared by ethanol thin-film hydration method. The formed optimal formulation of AP-M were provided with small size (178.5 nm) and spherical shape at ratio of 4:1 (Soluplus ® :Pluronic F127), as well as increasing solubility of to 5.61 mg/mL in water which was about 3442-fold compared to that of free AP. The entrapment efficiency and drug loading of AP-M were 95.72 and 5.32%, respectively, and a sustained release of AP-M was obtained as in vitro release study indicated. Transcellular transport study showed that the cell uptake of AP was increased in Caco-2 cell transport models. The oral bioavailability of AP-M was 4.03-fold of free AP in SD rats, indicating the mixed micelles of Soluplus ® and Pluronic F127 is an industrially feasible drug delivery system to promote insoluble drug oral absorption in the gastrointestinal tract.

Background: Respiratory diseases are mainly derived from acute and chronic inflammation of the alveoli and bronchi. The pathophysiological mechanisms of pulmonary inflammation mainly arise from oxidative damage that could ultimately lead to acute lung injury. Apigenin (Api) is a natural polyphenol with prominent antioxidant and anti-inflammatory properties in the lung. Inhalable formulations that consist of nanoparticles (NPs) have several advantages over other administration routes, and therefore, this study investigated the application of apigenin-loaded bovine serum albumin nanoparticles (BSA-Api-NPs) for pulmonary delivery., Methods: Dry powder formulations of BSA-Api-NPs were prepared by spray drying and characterized by laser diffraction particle sizing, scanning electron microscopy, differential scanning calorimetry, and powder X-ray diffraction. The influence of dispersibility enhancers (lactose monohydrate and l-leucine) on the in vitro aerosol deposition using a next-generation impactor was investigated in comparison to excipient-free formulation. The dissolution of Api was determined in simulated lung fluid by using the Franz cell apparatus. The antioxidant activity was determined by 2,2-diphenyl-1-picrylhydrazyl (DPPH ⋅ ) free radical scavenging assay., Results: The encapsulation efficiency and the drug loading were measured to be 82.61% ± 4.56% and 7.51% ± 0.415%. The optimized spray drying conditions were suitable to produce particles with low residual moisture content. The spray-dried BSA-Api-NPs possessed good aerodynamic properties due to small and wrinkled particles with low mass median aerodynamic diameter, high emitted dose, and fine particle fraction. The aerodynamic properties were enhanced by leucine and decreased by lactose, however, the dissolution was reversely affected. The DPPH ⋅ assay confirmed that the antioxidant activity of encapsulated Api was preserved., Conclusion: This study provides evidence to support that albumin nanoparticles are suitable carriers of Api and the use of traditional or novel excipients should be taken into consideration. The developed BSA-Api-NPs are a novel delivery system against lung injury with potential antioxidant activity.

To develop an alternative treatment for lung cancer, a combination of two potent chemotherapeutic agents - liposomal apigenin and tyroservatide - was developed. The therapeutic potential of this combination was investigated using A549 cells. Apigenin and tocopherol derivative-containing D-alpha-tocopheryl polyethylene glycol 1000 succinate (TPGS) liposomes might improve the delivery of apigenin to tumor cells, both in vitro and in vivo. Importantly, compared to either agent alone, the combination of apigenin TPGS liposomes and tyroservatide exhibited superior cytotoxicity, induced stronger G2 arrest, and suppressed A549 cancer cell invasion at a lower dose. The proapoptotic synergistic effects were also observed in A549 cells using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling, flow cytometry, and Western blot analysis. More importantly, in vivo results showed that the combination of apigenin TPGS liposomes and tyroservatide exhibited tumor-growth inhibitory effects in A549 cell-bearing mice. In conclusion, our study showed that this combination therapy could serve as a promising synergistic therapeutic approach to improve outcomes in patients with lung cancer., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

The aim of the present research was to study the protective effects and underlying mechanisms of apigenin on d-galactose-induced aging mice. Firstly, apigenin exhibited a potent antioxidant activity in vitro. Secondly, d-galactose was administered by subcutaneous injection once daily for 8 weeks to establish an aging mouse model to investigate the protective effect of apigenin. We found that apigenin supplementation significantly ameliorated aging-related changes such as behavioral impairment, decreased organic index, histopathological injury, increased senescence-associated β-galactosidase (SAβ-gal) activity and advanced glycation end product (AGE) level. Further data showed that apigenin facilitated Nrf2 nuclear translocation both in aging mice and normal young mice, and the Nrf2 expression of normal young mice was higher than that of natural senile mice. In addition, the expressions of Nrf2 downstream gene targets, including HO-1 and NQO1, were also promoted by apigenin administration. Moreover, apigenin also decreased the MDA level and elevated SOD and CAT activities. In conclusion, focusing on the Nrf2 pathway is a suitable strategy to delay the aging process, and apigenin may exert an anti-senescent effect process via activating the Nrf2 pathway.

Background: The potential application of Ficus deltoidea and vitexin for the management of symptomatologies associated with diabetes mellitus (DM) has gained much attention. However, less firm evidence comes from data to augment our understanding of the role of F. deltoidea and vitexin in protecting pancreatic β-cells. The aim of this study was to assess histological and oxidative stress changes in the pancreas of streptozotocin (STZ)-induced diabetic rats following F. deltoidea extract and vitexin treatment., Methods: F. deltoidea and vitexin was administrated orally to six-weeks STZ-induced diabetic rats over 8 weeks period. The glucose and insulin tolerances were assessed by intraperitoneal glucose (2 g/kg) tolerance test (IPGTT) and intraperitoneal insulin (0.65 U/kg) tolerance test (IPITT), respectively. Subsequently, insulin resistance was assessed by homeostasis assessment model of insulin resistance (HOMA-IR), quantitative insulin sensitivity check index (QUICKI) and the insulin/triglyceride-derived McAuley index. The histological changes in the pancreas were then observed by hematoxylin-eosin (H&E) staining. Further, the pattern of fatty acid composition and infrared (IR) spectra of the serum and pancreas were monitored by gas chromatography (GC) method and Fourier Transform Infrared (FT-IR) spectroscopy., Results: F. deltoidea and vitexin increased pancreatic antioxidant enzymes and promoted islet regeneration. However, a significant increase in insulin secretion was observed only in rats treated with F. deltoidea. More importantly, reduction of fasting blood glucose is consistent with reduced FT-IR peaks at 1200-1000 cm -1 ., Conclusions: These results accentuate that F. deltoidea and vitexin could be a potential agent to attenuate pancreatic oxidative damage and advocate their therapeutic potential for treating DM.

Angiogenesis plays important roles in progression of hepatocellular carcinoma. The antiangiogenic mechanisms of vitexicarpine are not fully defined. Therefore, we conducted the following study to evaluate the antiangiogenic mechanism and antitumor activity of vitexicarpine in vivo model of hepatocellular carcinoma through modulation of vascular endothelial growth factor signaling pathway. Hepatocellular carcinoma was induced in Sprague Dawley rats by thioacetamide. Hepatocellular carcinoma was assessed by measuring serum alpha-fetoprotein and investigating liver sections stained with hematoxylin/eosin. Hepatocellular carcinoma rats were injected with vitexicarpine (150 mg/kg) for 2 weeks. Hepatic vascular endothelial growth factor was measured by enzyme-linked immunosorbent assay. Protein and expression of hepatic phospho-Ser473-AKT (p-AKT) and phospho-Tyr419-Src (p-Src) were determined. The apoptotic pathway was evaluated by assessment of protein expression of caspase-3. Vitexicarpine increased rats' survival time and decreased serum alpha-fetoprotein as well as it ameliorated fibrosis and massive hepatic tissue breakdown. It attenuated hepatocellular carcinoma-induced protein and gene expression of vascular endothelial growth factor, p-AKT, p-Src, and caspase-3. In conclusion, this study suggests that vitexicarpine possesses both antiangiogenic and antitumor activities through inhibition of vascular endothelial growth factor, p-AKT/AKT, and p-Src with subsequent inhibition of apoptotic pathway.

Pancreatic beta cells are very sensitive to oxidative stress, which is one of the major causes of cell damages in diabetes. Growing interest has focused on the development of effective therapeutics to protect pancreatic cells from oxidative stress and searching for potentially protective antioxidants for treating diabetes. Apigenin, a plant-derived flavonoid, was investigated to determine whether it could protect rat insulinoma cell lines (RINm5F pancreatic beta cells) against streptozotocin (STZ)-induced oxidative damages and the mechanisms implicated. Our results showed that STZ treatment could induce oxidative stress and consequent cytotoxic effects in RINm5F cells. Pretreatment with apigenin effectively decreased the intracellular reactive oxygen species (ROS) production, attenuated cellular DNA damage, diminished lipid peroxidation, relieved protein carbonylation, and restored the cell apoptosis of pancreatic beta cells stressed by STZ. Our further experiments demonstrated that the beneficial effects of apigenin were related to ameliorate the loss of antioxidant enzymes of the STZ-treated cells in the level of gene transcription, protein expression, and enzyme activity. That suggested apigenin was not only a free radical scavenger but also a regulator to antioxidant defenses of pancreatic cells. Taken all together, our findings suggested that apigenin could attenuate the STZ-induced oxidative damages in pancreatic beta cells and might serve as a novel agent for the treatment of diabetes.

The aim of this study was to evaluate the inhibiting effect of apigenin on liver cancer in vivo based on the optical molecular imaging method. Subcutaneous liver tumor models were established using respective 1 × 10 6 firefly luciferase (fLuc) and green fluorescent protein (GFP) labeled human hepatocellular carcinoma cells (HepG2-fLuc and HepG2-GFP cells) in 20 BALB/c nude mice which were randomly divided into two groups, 10 in each group. After the tumor cells were implanted 15 days, apigenin was administered through intraperitoneal injection in group B, the other ten mice as control group A. Bioluminescence imaging (BLI) and fluorescence molecular imaging (FMI) were carried out for the follow-up of subcutaneous tumor model. As time goes on, intensity and distribution of bioluminescence and fluorescence of tumours increased gradually with the growth of tumours little by little. The whole process of observation was in accordance with known activities of HCC in the human liver. The tumor volume and tumor weight were significant lower in group B than in group A (p < 0.05), Subcutaneous tumours in the apigenin treatment group B based on BLI and FMI were significantly inhibited compared to the control group A (p < 0.05). Apigenin could be expected as a new drug to treat hepatocellular carcinoma. Optical molecular imaging technology enabled the non-invasive and reliable assessment of anti-tumor drug efficacy on liver cancer., (Copyright © 2017 Elsevier Inc. All rights reserved.)

We substantiated the role of peroxisome proliferator-activated receptor-γ (PPAR-γ) activation in the protective effect of apigenin against the myocardial infarction (MI) in diabetic rats. Diabetes was induced by intraperitoneal administration of a single dose of streptozotocin (55 mg/kg). The study groups included diabetic rats receiving vehicle, apigenin (75 mg/kg/day, orally), GW9662 (1 mg/kg/day, intraperitoneally), and a combination of apigenin and GW9662 for 14 days. The MI was induced in all the study groups except the diabetic control group by subcutaneous injection of 100 mg/kg/day of isoproterenol on the two terminal days. The diabetes and isoproterenol-induced MI was evident as a reduction in the maximal positive and negative rate of developed left ventricular pressure and an increase in the left ventricular end-diastolic pressure. The activities of creatine kinase on myocardial bundle (CK-MB) and lactate dehydrogenase (LDH) were also reduced. Apigenin treatment prevented the hemodynamic perturbations, restored the left ventricular function and reinstated a balanced redox status. It protected rats against an MI by attenuating myonecrosis, edema, cell death, and oxidative stress. GW9662, a PPAR-γ antagonist reversed the myocardial protection conferred by apigenin. Further, an increase in the PPAR-γ expression in the myocardium of the rats receiving apigenin reinforces the role of PPAR-γ pathway activation in the cardioprotective effects of apigenin.

Apigenin is an important component of fruits and vegetables in human daily diets. Several cellular and animal models have been performed to demonstrate its anti-oxidant and anti-inflammatory bioactivities. However, the cardioprotective effects of apigenin in diabetic cardiomyopathy (DCM) remain unclear. In this study, we intended to explore the roles of apigenin in cardiac remodeling of DCM. Male C57BL/6 J mice were treated with streptozotocin (STZ, 50 mg/kg) for 5 consecutive days to induce DCM. The echocardiography and catheter-based measurements of hemodynamic parameters were performed to evaluate the cardiac function. Paraffin slices of harvested hearts were prepared for histological pathological analysis and TUNEL assay. Oxidative assay kits were used to detect Glutathione Peroxidase (GPx), Lipid Peroxidation Malondialdehyde (MDA), and Superoxide Dismutase (SOD). Western blot and real-time PCR were used for accessing the expressions of protein and mRNA. Diabetes mellitus exacerbated the cardiac dysfunction, fibrosis, and overaccumulation of 4-hydroxynonenal accompanying with down-regulation of Bcl2, GPx, and SOD, up-regulation of MDA, cleaved caspase3, and pro-apoptotic protein Bax, and contribution to the translocation of NF-κB. All these pathological changes could be effectively blunted by treatment of apigenin in vivo. Finally, H9c2 treated with high glucose or apigenin was used for further investigation of these effects in vitro; what is more, we also compared the effects between apigenin and Resveratrol in in vitro experiments. Our experiments have demonstrated that apigenin may be a potential drug for diabetic patients suffering from DCM.

The bioflavonoid apigenin has been shown to possess cancer-preventive and anti-cancer activities. In a drug screening, we found that apigenin can inhibit Wnt/β-catenin signaling, a pathway that participates in pivotal biological functions, which dis-regulation results in various human diseases including cancers. However, the underlying mechanism of apigenin in this pathway and its link to anti-cancer activities remain largely unknown. Here we showed that apigenin reduced the amount of total, cytoplasmic, and nuclear β-catenin, leading to the suppression in the β-catenin/TCF-mediated transcriptional activity, the expression of Wnt target genes, and cell proliferation of Wnt-stimulated P19 cells and Wnt-driven colorectal cancer cells. Western blotting and immunofluorescent staining analyses further revealed that apigenin could induce autophagy-mediated down-regulation of β-catenin in treated cells. Treatment with autophagy inhibitors wortmannin and chloroquine compromised this effect, substantiating the involvement of autophagy-lysosomal system on the degradation of β-catenin during Wnt signaling through inhibition of the AKT/mTOR signaling pathway. Our data not only pointed out a route for the inhibition of canonical Wnt signaling through the induction of autophagy-lysosomal degradation of key player β-catenin, but also suggested that apigenin or other treatments which can initiate this degradation event are potentially used for the therapy of Wnt-related diseases including cancers.

Background: Diabetic retinopathy is the most common complication in diabetic patients relates to high expression of VEGF and microaneurysms. Scutellarin (Scu) turned out to be effective against diabetes related vascular endothelial cell dysfunction. However, its clinical applications have been limited by its low bioavailability. In this study, we formulated and characterized a novel intestinal target nanoparticle carrier based on amphiphilic chitosan derivatives (Chit-DC-VB12) loaded with scutellarin to enhance its bioavailability and then evaluated its therapeutic effect in experimental diabetic retinopathy model., Results: Chit-DC-VB12 nanoparticles showed low toxicity toward the human colon adenocarcinoma (Caco-2) cells and zebra fish within concentration of 250 μg/ml, owing to good biocompatibility of chitosan. The scutellarin-loaded Chit-DC-VB12 nanoparticles (Chit-DC-VB12-Scu) were then prepared by self-assembly in aqueous solution. Scanning electron microscopy and dynamic light scattering analysis indicated that the Chit-DC-VB12-Scu nanoparticles were spherical particles in the sizes ranging from 150 to 250 nm. The Chit-DC-VB12-Scu nanoparticles exhibited high permeation in Caco-2 cell, indicated it could be beneficial to be absorbed in humans. We also found that Chit-DC-VB12 nanoparticles had a high cellular uptake. Bioavailability studies were performed in Sprague-Dawley rats, which present the area under the curve of scutellarin of Chit-DC-VB12-Scu was two to threefolds greater than that of free scutellarin alone. Further to assess the therapeutic efficacy of diabetic retinopathy, we showed Chit-DC-VB12-Scu down-regulated central retinal artery resistivity index and the expression of angiogenesis proteins (VEGF, VEGFR2, and vWF) of retinas in type II diabetic rats., Conclusions: Chit-DC-VB12 nanoparticles loaded with scutellarin have better bioavailability and cellular uptake efficiency than Scu, while Chit-DC-VB12-Scu nanoparticles alleviated the structural disorder of intraretinal neovessels in the retina induced by diabetes, and it also inhibited the retinal neovascularization via down-regulated the expression of angiogenesis proteins. In conclusion, the Chit-DC-VB12 nanoparticles enhanced scutellarin oral delivery efficacy and exhibited potential as small intestinal target promising nano-carriers for treatment of type II diabetes induced-retinopathy.

Background: Although 5- Fluorouracil (5-FU) has exhibited effectiveness against cancer, novel therapeutic strategies are needed to enhance its antitumor efficiency and modulate its cytotoxity. Apigenin, a flavonoid present in fruits and vegetables, is a potent dietary phytochemical effective in cancer chemoprevention., Aim: This study was undertaken to investigate the potential synergistic antitumor activity of apigenin and 5-FU on Solid Ehrlich carcinoma (SEC)., Methods: Eighty Swiss albino male mice were divided into four equal groups: vehicle treated control SEC, SEC+5-FU, SEC+apigenin, SEC+ 5-FU+apigenin. Beclin-1 and caspases 3, 9 and JNK activities were estimated by ELISA; mRNA expression levels of the antiapoptotic gene Mcl-1 were estimated using quantitative real-time RT-PCR, while tissue malondialdehyde (MDA), glutathione peroxidase and total antioxidant capacity were evaluated spectrophotometrically. A part of the tumor was examined for histopathological and Ki-67 immunohistochemistry analysis., Results: 5-FU and/or apigenin caused significant increase in tissue levels of Beclin-1, caspases 3, 9 and JNK activities, MDA with significant decrease in tumor volume, Mcl-1expression, tissue glutathione peroxidase and total antioxidant capacity and alleviated the histopathological changes with significant decrease of Ki-67 proliferation index compared to vehicle treated SEC control group., In Conclusion: The combination of 5-FU and apigenin had a greater effect than each of 5-FU or apigenin alone against solid Ehrlich carcinoma in mice., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Drug delivery of phytochemicals has gained interest recently due to their remarkable health effects. Apigenin, a plant flavonoid, has antioxidant, anti-inflammatory and anticancer activities but its delivery is challenging. It could be absorbed through the whole intestine, however, it has poor bioavailability due to its low aqueous solubility. In Europe, the daily intake was estimated to be as low as 3 ± 1 mg. Pellets offer several advantages such as improved bioavailability and various resultant drug release profiles can be obtained by simply mixing pellets with different coatings., Objective: The objective of our study was to develop a carrier system containing 20 mg apigenin thus enhancing intake and to offer reduction of oxidative stress which can cause inflammation in the intestine., Method: The apigenin powder was dispersed in aqueous solution of binding material and layered onto the inert cores in a fluidized bed apparatus. The layered cores were further coated with enteric polymers and the process parameters were optimized., Results: The prepared pellets met with the requirements and have good physical characteristic. 10% (w/w) Eudragit® L was suitable for enteric coating with a complete release at pH 6.8 within 1 hour. 15% (w/w) Eudragit® FS coating ensured acid resistance ability and colonic delivery. The therapeutic efficiency was confirmed with antioxidant activity measurement by using DPPH* assay., Conclusion: Enteric coated spheres allow targeted delivery into the intestine and colon thus reaching the main absorption site. Pellets were proved to be an optimal delivery system for apigenin thus providing enhanced apigenin intake., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.)

Background: Ovarian cancer is most lethal among all gynecologic malignancies. Paclitaxel (PTX) is well used chemotherapeutic regimen for cancer control; however its undesired toxicity has been a matter of concern for clinicians. Here, we used the graphene oxide coated nanotised apigenin (GO-NA) to enhance the efficacy of paclitaxel., Objective: The combined use of paclitaxel (PTX) and nanotised apigenin (NA) may reduce the PTX dose and increase the efficacy., Methods: GO and GO-Apigenin was prepared by modified Hummers method and the nanoparticles were characterized by dynamic light scattering and transmission electron microscopy. SKOV-3 cells were treated by DMSO, Group I (Control)-McCoy's 5A Medium, Group II-Paclitaxel (5nM), Group III- Nanotised Apigenin (GO-NA-10µM), Group IV- Paclitaxel (5nM) + GO-NA (10µM). Cell viability and IC-50 value were determined by MTT assay, synergism by Compusyn software, ROS by DCFH-DA assay, SOD activity by kit and MMP were examined by JC-1 and mitotracker/DAPI staining, cell cycle by flow cytometry, mRNA and protein level by Real Time-PCR and Western blot respectively Results: Results showed that GO-NA-PTX enhanced the anti-proliferative effect in synergistic manner as compare to GO-NA and PTX alone. GO-NA-PTX significantly suppressed the SOD activity, promotes the ROS accumulation, mitochondrial depolarization, DNA integrity and cell cycle arrest collectively accord the apoptosis. Results of immunocytochemistry, RT-PCR and western blot showed up-regulation of caspase-3, Bax, and down-regulation of Bcl-2., Conclusion: The combination of PTX with GO-NA produces synergistic effects in SKOV-3 cells via the modulation of pro and anti-apoptotic gene and may reduce side effects of PTX., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.)

Tibial dyschondroplasia (TD) is an important tibiotarsal bone disorder characterized by an avascular and nonmineralized growth plate; it is attributed to abnormal differentiation of chondrocytes and causes lameness. Heat-shock protein 90 (Hsp90) is a proangiogenic factor in animal tissues; however, its gene expression increases in cases of chicken TD. The objective of this study was to evaluate the efficacy of epigallocatechin-3-gallate (EGCG) and apigenin in thiram-induced TD birds; these substances were used because of their Hsp90 inhibitory activities. The histologic study of growth plates was carried out with hematoxylin and eosin staining, and the Hsp90 gene expression was examined by reverse transcription quantitative real-time PCR. Results showed that as compared to a control group, TD-affected birds displayed changes in chondrocyte differentiation, with lack of blood vessels, and an increased expression of Hsp90 was observed significantly (P < 0.05). However, on administering the EGCG and apigenin to TD-affected birds, the normal chondrocyte columnar organization was restored with vascularization and decreased Hsp90 expression activity (P < 0.05), which ultimately abrogated the lameness. Our results suggested that Hsp90 is the key factor in the development of TD, and EGCG and apigenin have a novel effect on Hsp90 inhibition properties, thus reducing the lameness and leg deformity in chicken broilers.

Angiographic vasospasm, especially in the early phases (<72h) of subarachnoid hemorrhage (SAH), is one of the major complications after an aneurysm rupture and is often the cause of delayed neurological deterioration. Scutellarin (SCU), a flavonoid extracted from the traditional Chinese herb Erigeron breviscapus, has been widely accepted as an antioxidant, but the effect of SCU on vasospasm after SAH remains elusive. Endovascular perforation was conducted to induce SAH in Sprague-Dawley rats. Then, the underlying mechanism of the anti-vasospasm effect of SCU was investigated using a modified Garcia scale, India ink angiography, cross-sectional area analysis, immunohistochemistry staining and western blot. SCU (50μM, 100mg/kg) alleviated angiographic vasospasm and improved neurological function 48h after SAH and enhanced the expression of endothelial nitric oxide synthase (eNOS) at the intima of cerebral arteries. In addition, SCU upregulated the expression of phosphorylated extracellular-regulated kinase 5 (p-Erk5) and Kruppel-like factor 2 (KLF2) at 48h after SAH. However, the effects of SCU were reversed by the Erk5 inhibitor XMD8-92. Our results indicate that SCU could attenuate vasospasm and neurological deficits via modulating the Erk5-KLF2-eNOS pathway after SAH, which may provide an experimental basis for the clinical use of SCU treatment in SAH patients., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Background/aims: Hepatic injury caused by ischemia/reperfusion (I/R) is a clinical problem associated with major liver surgery. Among other flavonoids, apigenin has shown a promising effect on I/R cases. In this study, we have investigated the effects of apigenin after liver I/R injury in rats., Materials and Methods: Forty eight rats were randomized into the following eight groups: (1) Control-sham group: rats subjected to the surgical procedure, except for liver I/R; (2) DMSO group: rats subjected to surgery, except for liver I/R given the apigenin solvent dimethyl-sulfoxide intraperitoneally; (3) C60 group; (4) C120 group; (5) C240 group: rats underwent liver ischemia for 45 min followed by reperfusion for 60 min, 120 min, and 240 min; (6) AP60 group; (7) AP120 group; (8) AP240 group: rats underwent liver ischemia for 45 min, and then given apigenin (5 mg) intraperitoneally followed by reperfusion for 60 min, 120 min, and 240 min. Reverse transcription polymerase chain reaction was performed on liver tissues to measure BCL-2/BAX expression, enzyme-linked immunosorbent assay to measure M30/M65 and ICAM-1. Immunohistochemistry was used to identify M30 biomarker in liver tissues., Statistical Analysis: Quantitative variables were tested by Kolmogorov-Smirnov test, repeated measures analysis of variance/Friedman test. Gene levels were assessed by Student's t-test/Mann-Whitney U-test., Results: BCL-2 levels were significantly higher in I/R apigenin groups than in I/R control groups. BAX levels were lower in the AP240 group than in C240 group. Prolongation of reperfusion resulted in increased activation of M30. ICAM-1 levels were lower in the AP240 group than in C240 group., Conclusions: Apigenin seems to inhibit the process of apoptosis and ameliorate the hepatic I/R injury., Competing Interests: There are no conflicts of interest.

Vicenin-1 (fenugreek glycoside) has been proven to possess potent anti-inflammatory and anti-oxidant activity. The objective of the present investigation was to determine in-vivo acute and subacute (28-days repeated dose) oral toxicity of Vicenin-1 isolated from fenugreek seed. Vicenin-1 (93%) was isolated from a hydroalcoholic extract of fenugreek seed and characterized using HPLC, TLC, 1 H NMR and 13 C NMR. Acute oral toxicity (AOT) and subacute toxicity studies of Vicenin-1 were carried out according to OECD 425 (up-and-down procedure) and OCED 407 guidelines in Swiss albino mice. In AOT, Vicenin-1 showed 10% mortality when administered at a dose of 5000 mg/kg. However, when vicenin-1 was administered for at doses of 37.5, 75, or 150 mg/kg 28-days it did not show any mortality at the administered doses. Vicenin-1 (75 mg/kg) did not show observational, behavioral, biochemical or histopathological toxic effects. There were minor alterations in body weight, hematology, and histopathology of mice administered with Vicenin-1 (150 mg/kg), but these changes were within normal laboratory ranges. The highest concentration of Venicin-1 was found in liver (3.46%) followed by lung (0.65%). In conclusion, Vicenin-1 showed median lethal dose (LD 50 ) of 4837.5 mg/kg with no-observed-adverse-effect levels (NOAEL) at 75 mg/kg and lowest adverse effect levels (LOAEL) at 150 mg/kg for both sexes of mice during AOT and sub-acute toxicity study, respectively., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Efflux transporters are extensively distributed and expressed in the intestinal epithelium and contribute to the low oral bioavailability of flavonoids and flavonoid glucuronides by pumping these compounds back into intestinal lumen. Our previous study has shown the inhibitory effect of Cremophor EL, a non-ionic surfactant, on efflux transporter multidrug resistance-associated protein (MRP) 2. In the current study, by using membranes overexpressing several common ATP-binding cassette (ABC) transporters including P-glycoprotein (P-gp), MRP1, MRP2, MRP3 and breast cancer resistance protein (BCRP), scutellarin, a poorly water-soluble flavonoid, was identified as the substrate of MPR2, MRP3 and BCRP. The effects of Cremophor EL on the transmembrane transportation of scutellarin by MRP2, BCRP, and MRP3 were investigated with inside-out Sf9 vesicles. Results showed that at nontoxic concentrations, Cremophor EL enhanced the transportation of scutellarin by MRP3 and inhibited the efflux transportation of scutellarin by MRP2 and BCRP concurrently. The relations between Cremophor EL and these transporters were explored using MDCK II-MRP2, MDCK II-BCRP, and MDCK II-MRP3 cell models. Compared with the control group, 5μg/ml Cremophor EL decreased the Papp(BL-AP) of scutellarin in MDCK II-MRP2 cell monolayers by >4 fold (from 13.57±0.76×10(-7) to 2.90±0.14×10(-7)cm/s), and the Papp(BL-AP) in MDCK II-BCRP cell monolayers decreased from 9.12±0.15×10(-7) to 6.34±0.08×10(-7)cm/s. On MDCK II-MRP3 cell monolayers, 5μg/ml Cremophor EL increased the Papp(AP-BL) of scutellarin by 3.5 fold (from 7.88±0.43×10(-7) to 2.79±1.61×10(-6)cm/s), and caused an over 5-fold increase in Papp(AP-BL)/Papp(BL-AP). These findings suggested that Cremophor EL possesses the potent ability of inhibiting MRP2 and BCRP, as well as activating MRP3 effectively. In vivo pharmacokinetic research in rats further confirmed the improvement of oral absorption of scutellarin by Cremophor EL. In summary, our present study has identified a new mechanism for increasing the oral absorption and bioavailability of poorly absorbed drugs in which Cremophor EL increased MRP3 mediated transport but reduced MRP2 and BCRP mediated efflux concurrently, thereby enhancing the entry of drugs from enterocytes into the blood circulation., (Copyright © 2016 Elsevier B.V. All rights reserved.)

We analyzed the physical properties and digestibility of apigenin-loaded emulsions as they passed through a simulated digestion model. As the emulsion passed through the simulated stages of digestion, the particle size and zeta potential of all the samples changed, except for the soybean oil-Tween 80 emulsion, in which zeta potential remained constant, through all stages, indicating that soybean oil-Tween 80 emulsions may have an effect on stability during all stages of digestion. Fluorescence microscopy was used to observe the morphology of the emulsions at each step. The in vivo pharmacokinetics revealed that apigenin-loaded soybean oil-Tween 80 emulsions had a higher oral bioavailability than did the orally administrated apigenin suspensions. These results suggest that W/O/W multiple emulsions formulated with soybean oil and tween 80 have great potential as targeted delivery systems for apigenin, and may enhance in vitro and in vivo bioavailability when they pass through the digestive tract., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The composition of Morinda citrifolia (M. citrifolia) was determined using high-performance liquid chromatography (HPLC), and the anticancer effects of M. citrifolia extract evaluated in HepG2, Huh7, and MDA-MB-231 cancer cells. M. citrifolia fruit extracts were obtained by using five different organic solvents, including hexane (Hex), methanol (MeOH), ethyl acetate (EtOAc), chloroform (CHCl3), and ethanol (EtOH). The water-EtOAc extracts from M. citrifolia fruits was found to have the highest anticancer activity. HPLC data revealed the predominance of chrysin in water-EtOAc extracts of M. citrifolia fruit. Furthermore, the combined effects of cotreatment with apigenin and chrysin on liver and breast cancer were investigated. Treatment with apigenin plus chrysin for 72-96 h reduced HepG2 and MDA-MB-231 cell viability and induced apoptosis through down-regulation of S-phase kinase-associated protein-2 (Skp2) and low-density lipoprotein receptor-related protein 6 (LRP6) expression. However, the combination treatment for 36 h synergistically decreased MDA-MB-231 cell motility but not cell viability through down-regulation of MMP2, MMP9, fibronectin, and snail in MDA-MB-231 cells. Additionally, chrysin combined with apigenin also suppressed tumor growth in human MDA-MB-231 breast cancer cells xenograft through down-regulation of ki-67 and Skp2 protein. The experimental results showed that chrysin combined with apigenin can reduce HepG2 and MDA-MB-231 proliferation and cell motility and induce apoptosis. It also offers opportunities for exploring new drug targets, and further investigations are underway in this regard.

Apigenin is a natural flavonoid compound that can inhibit hypoxia-inducible factor (HIF)-1α expression in cultured tumor cells under hypoxic conditions. Hypertension-induced cardiac hypertrophy is always accompanied by abnormal myocardial glucolipid metabolism due to an increase of HIF-1α. However, whether or not apigenin may ameliorate the cardiac hypertrophy and abnormal myocardial glucolipid metabolism remains unknown. This study aimed to examine the effects of apigenin. Rats with cardiac hypertrophy induced by renovascular hypertension were treated with apigenin 50-100 mg kg(-1) (the doses can be achieved by pharmacological or dietary supplementation for an adult person) by gavage for 4 weeks. The results showed that after treatment with apigenin, the blood pressure, heart weight, heart weight index, cardiomyocyte cross-sectional area, serum angiotensin II, and serum and myocardial free fatty acids were reduced. It is important to note that apigenin decreased the expression level of myocardial HIF-1α protein. Moreover, apigenin simultaneously increased the expression levels of myocardial peroxisome proliferator-activated receptor (PPAR) α, carnitine palmitoyltransferase (CPT)-1, and pyruvate dehydrogenase kinase (PDK)-4 proteins and decreased the expression levels of myocardial PPARγ, glycerol-3-phosphate acyltransferase genes (GPAT), and glucose transporter (GLUT)-4 proteins. These findings demonstrated that apigenin could improve hypertensive cardiac hypertrophy and abnormal myocardial glucolipid metabolism in rats, and its mechanisms might be associated with the down-regulation of myocardial HIF-1α expression and, subsequently increasing the expressions of myocardial PPARα and its target genes CPT-1 and PDK-4, and decreasing the expressions of myocardial PPARγ and its target genes GPAT and GLUT-4.

The present study was designed to elucidate the protective effects of dietary apigenin (API) enrichment in a chronic colitis model induced by DSS in mice. Inflammatory mediators and the possible role of canonical and non-canonical NLRP3 inflammasome signaling pathways in the beneficial effects of API under chronic inflammatory conditions were also explored. Six-week-old mice were randomized in four dietary groups: sham and control groups received standard diet (SD), and other two groups were fed with API at 0.1%. After 30days, all groups except sham received 3% DSS in drinking water for 5days followed by a regime of 21days of water. Our results revealed that dietary API supplementation decreased the macroscopic and microscopic damage signs of colitis; also, it was capable to down-regulate mPGES, COX-2 and iNOS enzyme colonic expressions and to decrease serum matrix metalloproteinase (MMP-3) levels. Similarly, API diet reduced IL-1β and TNF-α proinflammatory cytokine secretions in primary LPS-stimulated splenocytes. Furthermore, we demonstrated that API anti-inflammatory activity was related with an inhibition of both canonical and non-canonical NLRP3 inflammasome pathways by decreasing proinflammatory IL-1β and IL-18 cytokine levels as a consequence of regulation of cleaved caspase-1 and caspase-11 enzymes. We conclude that API supplement might provide a basis for developing a new dietary strategy for the prevention of chronic ulcerative colitis., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The increasing prevalence of inflammatory diseases and the adverse effects associated with the long-term use of current anti-inflammatory therapies prompt the identification of alternative approaches to reestablish immune balance. Apigenin, an abundant dietary flavonoid, is emerging as a potential regulator of inflammation. Here, we show that apigenin has immune-regulatory activity in vivo. Apigenin conferred survival to mice treated with a lethal dose of Lipopolysaccharide (LPS) restoring normal cardiac function and heart mitochondrial Complex I activity. Despite the adverse effects associated with high levels of splenocyte apoptosis in septic models, apigenin had no effect on reducing cell death. However, we found that apigenin decreased LPS-induced apoptosis in lungs, infiltration of inflammatory cells and chemotactic factors' accumulation, re-establishing normal lung architecture. Using NF-κB luciferase transgenic mice, we found that apigenin effectively modulated NF-κB activity in the lungs, suggesting the ability of dietary compounds to exert immune-regulatory activity in an organ-specific manner. Collectively, these findings provide novel insights into the underlying immune-regulatory mechanisms of dietary nutraceuticals in vivo.

Background: The treatment for leishmaniasis is currently based on pentavalent antimonials and amphotericin B; however, these drugs result in numerous adverse side effects. The lack of affordable therapy has necessitated the urgent development of new drugs that are efficacious, safe, and more accessible to patients. Natural products are a major source for the discovery of new and selective molecules for neglected diseases. In this paper, we evaluated the effect of apigenin on Leishmania amazonensis in vitro and in vivo and described the mechanism of action against intracellular amastigotes of L. amazonensis., Methodology/principal Finding: Apigenin reduced the infection index in a dose-dependent manner, with IC50 values of 4.3 μM and a selectivity index of 18.2. Apigenin induced ROS production in the L. amazonensis-infected macrophage, and the effects were reversed by NAC and GSH. Additionally, apigenin induced an increase in the number of macrophages autophagosomes after the infection, surrounding the parasitophorous vacuole, suggestive of the involvement of host autophagy probably due to ROS generation induced by apigenin. Furthermore, apigenin treatment was also effective in vivo, demonstrating oral bioavailability and reduced parasitic loads without altering serological toxicity markers., Conclusions/significance: In conclusion, our study suggests that apigenin exhibits leishmanicidal effects against L. amazonensis-infected macrophages. ROS production, as part of the mechanism of action, could occur through the increase in host autophagy and thereby promoting parasite death. Furthermore, our data suggest that apigenin is effective in the treatment of L. amazonensis-infected BALB/c mice by oral administration, without altering serological toxicity markers. The selective in vitro activity of apigenin, together with excellent theoretical predictions of oral availability, clear decreases in parasite load and lesion size, and no observed compromises to the overall health of the infected mice encourage us to supports further studies of apigenin as a candidate for the chemotherapeutic treatment of leishmaniasis.

Previous research in our laboratory found that the absolute bioavailability of vitexin-2''-O-rhamnoside (VR) was quite low at 4.89%. A rapid and sensitive UHPLC method using hesperidin as an internal standard was therefore developed and validated to investigate the reasons for this by determining VR in rat plasma after administering intravenously, intraportally (5 mg/kg), intraduodenally and intragastrically (40 mg/kg) to the rat model of the hepatic, gastric and intestinal first-pass effects. As only a high intestinal first-pass effect of VR was found, that is, there existed a low bioavailability of VR (2.40%), inhibitors of P-glycoprotein (P-gp) and cytochrome P450 3A (CYP3A), including verapamil, cyclosporin A and midazolam, and absorption enhancers, including bile salts and borneol, combined with VR, were instilled into duodenum to evaluate the effects on bioavailability of VR. The results demonstrated that area under the concentration-time curve (AUC) values of VR slightly increased after administration of verapamil, cyclosporin A and midazolam, indicating that CYP3A and P-gp do not play an important role in the first-pass effect in the intestine. AUC values of VR significantly increased after administering bile salts or borneol, indicating that the low bioavailability of VR was mainly related to its poor absorption in the intestine., (Copyright © 2015 John Wiley & Sons, Ltd.)