Novel brain-targeted nanomicelles for anti-glioma therapy mediated by the ApoE-enriched protein corona in vivo.

BACKGROUND: The interactions between nanoparticles (NPs) and plasma proteins form a protein corona around NPs after entering the biological environment, which provides new biological properties to NPs and mediates their interactions with cells and biological barriers. Given the inevitable interactions, we regard nanoparticle‒protein interactions as a tool for designing protein corona-mediated drug delivery systems. Herein, we demonstrate the successful application of protein corona-mediated brain-targeted nanomicelles in the treatment of glioma, loading them with paclitaxel (PTX), and decorating them with amyloid ß-protein (Aß)-CN peptide (PTX/Aß-CN-PMs). Aß-CN peptide, like the Aß1-42 peptide, specifically binds to the lipid-binding domain of apolipoprotein E (ApoE) in vivo to form the ApoE-enriched protein corona surrounding Aß-CN-PMs (ApoE/PTX/Aß-CN-PMs). The receptor-binding domain of the ApoE then combines with low-density lipoprotein receptor (LDLr) and LDLr-related protein 1 receptor (LRP1r) expressed in the blood-brain barrier and glioma, effectively mediating brain-targeted delivery. METHODS: PTX/Aß-CN-PMs were prepared using a film hydration method with sonication, which was simple and feasible. The specific formation of the ApoE-enriched protein corona around nanoparticles was characterized by Western blotting analysis and LC-MS/MS. The in vitro physicochemical properties and in vivo anti-glioma effects of PTX/Aß-CN-PMs were also well studied. RESULTS: The average size and zeta potential of PTX/Aß-CN-PMs and ApoE/PTX/Aß-CN-PMs were 103.1 nm, 172.3 nm, 7.23 mV, and 0.715 mV, respectively. PTX was efficiently loaded into PTX/Aß-CN-PMs, and the PTX release from rhApoE/PTX/Aß-CN-PMs exhibited a sustained-release pattern in vitro. The formation of the ApoE-enriched protein corona significantly improved the cellular uptake of Aß-CN-PMs on C6 cells and human umbilical vein endothelial cells (HUVECs) and enhanced permeability to the blood-brain tumor barrier in vitro. Meanwhile, PTX/Aß-CN-PMs with ApoE-enriched protein corona had a greater ability to inhibit cell proliferation and induce cell apoptosis than taxol. Importantly, PTX/Aß-CN-PMs exhibited better anti-glioma effects and tissue distribution profile with rapid accumulation in glioma tissues in vivo and prolonged median survival of glioma-bearing mice compared to those associated with PMs without the ApoE protein corona. CONCLUSIONS: The designed PTX/Aß-CN-PMs exhibited significantly enhanced anti-glioma efficacy. Importantly, this study provided a strategy for the rational design of a protein corona-based brain-targeted drug delivery system. More crucially, we utilized the unfavorable side of the protein corona and converted it into an advantage to achieve brain-targeted drug delivery.

Design and Synthesis of Novel Anti-Proliferative Emodin Derivatives and Studies on their Cell Cycle Arrest, Apoptosis Pathway and Migration.

Emodin is a cell arrest and apoptosis-inducing compound that is widely distributed in different plants (rhubarb, aloe), lichens and terrestrial fungi, and also isolated from marine-derived fungi and marine sponge-associated fungi. In this study, we designed and synthesized a novel series of emodin derivatives by binding emodin to an amino acid using linkers of varying lengths and composition, and evaluated their anti-proliferative activities using HepG2 cells (human hepatic carcinoma), MCF-7 cells (human breast cancer) and human normal liver L02 cells. Most of these derivatives showed moderate to potent anti-proliferative activities. Notably, compound 7a exhibited potent anti-proliferative activity against HepG2 cells with the half maximal inhibitory concentration (IC50) value of 4.95 µM, which was enhanced 8.8-fold compared to the parent compound emodin (IC50 = 43.87 µM), and it also exhibited better selective anti-proliferative activity and specificity than emodin. Moreover, further experiments demonstrated that compound 7a displayed a significant efficacy of inducing apoptosis through mitochondrial pathway via release of cytochrome c from mitochondria and subsequent activation of caspase-9 and caspase-3, inducing cell arrest at G0/G1 phase, as well as suppression of cell migration of tumor cells. The preliminary results suggested that compound 7a could be a promising lead compound for the discovery of novel anti-tumor drugs and has the potential for further investigations as an anti-cancer drug.

Inhibiting the proliferation, migration and invasion of triple negative breast cancer cells through anti-tumor human serum albumin nanoparticles loading aziditaxel as a novel taxane derivative.

Triple negative breast cancer (TNBC) is a severe breast cancer subtype with the high mortality rate, and still is lack of effective therapeutic means so far. Aziditaxel, a water-insoluble compound, is a novel taxane derivative with strong anti-tumor activity. In this study, we constructed an aziditaxel-loaded nano drug delivery system using human serum albumin as a carrier, and further investigated its anti-tumor effect on TNBC in vitro. An emulsion solvent evaporation method was employed to prepare aziditaxel-loaded human serum albumin nanoparticles (AT-NPs). Their physicochemical properties were characterized according to morphology, particle size, zeta potential, reconstitution stability and in vitro drug release. The in vitro anti-tumor effects of AT-NPs on TNBC were evaluated using a 4T1 murine triple negative mammary cancer cell lines as the TNBC model. The results showed that AT-NPs could be effectively taken up by 4T1 cells in a time-dependent manner. Cell Counting Kit-8 assay showed that the IC50 of AT-NPs was 0.17 µg/ml. Meanwhile, compared with AT, AT-NPs had a better ability to promote apoptosis and induce G2/M cycle arrest. On the other hand, AT-NPs had significantly inhibitory effects on the 4T1 cell adhesion, migration and invasion with the respective average inhibition ratios of 32.53%, 83.26% and 75.78%. Thus, our study revealed that AT-NPs had favorable antitumor activity in vitro and exhibited a good prospect for application in the field of TNBC therapy.

[The development of cell-penetrating peptides in drug delivery system].

Cell membrane serves as the natural barrier. Cell-penetrating peptides(CPPs) have been a powerful vehicle for the intracellular delivery of a large variety of cargoes cross the cell membrane. The efficiency of intracellular delivery of drugs, proteins, peptides and nucleic acid, as well as various nanoparticulate pharmaceutical carriers(e.g., liposomes, polymeric micelles and inorganic nanoparticles) has been demonstrated both in vitro and in vivo. This review focuses on the CPPs-based strategy for intracellular delivery of small molecule drugs, proteins, peptides, nucleic acid and CPP-modified nanocarriers.

Evaluation of doxorubicin-loaded pH-sensitive polymeric micelle release from tumor blood vessels and anticancer efficacy using a dorsal skin-fold window chamber model.

AIM: To evaluation the doxorubicin (DOX)-loaded pH-sensitive polymeric micelle release from tumor blood vessels into tumor interstitium using an animal vessel visibility model, the so-called dorsal skin-fold window chamber model. METHODS: DOX-loaded pH-sensitive polyHis-b-PEG micelles and DOX-loaded pH-insensitive PLLA-b-PEG micelles were prepared. The uptake of the micelles by MDA-MB-231 breast cancer cells in vitro and in vivo was examined using flow cytometry. The pharmacokinetic parameters of the micelles were determined in SD rats after intravenous injection of a DOX dose (6 mg/kg). The release of the micelles from tumor vasculature and the antitumor efficacy were evaluated in MDA-MB-231 breast cancer xenografted in nude mice using a dorsal skin-fold window chamber. RESULTS: The effective elimination half-life t1/2 of the pH-sensitive, pH-insensitive polymeric micelles and DOX-PBS in rats were 11.3 h, 9.4 h, and 2.1 h, respectively. Intravital microscopy in MDA-MB-231 breast cancer xenografted in nude mice showed that the pH-sensitive polymeric micelles rapidly extravasated from the tumor blood vessels, and DOX carried by the pH-sensitive micelles was preferentially released at the tumor site as compared to the pH-insensitive polymeric micelles. Furthermore, the pH-sensitive polymeric micelles exhibited significant greater efficacy in inhibition of tumor growth in the nude mice. CONCLUSION: When DOX is loaded into pH-sensitive polymeric micelles, the acidity in tumor interstitium causes the destabilization of the micelles and triggers drug release, resulting in high local concentrations within the tumor, thus more effectively inhibiting the tumor growth in vivo.

Improved anti-tumor efficiency against prostate cancer by docetaxel-loaded PEG-PCL micelles.

This study primarily focused on the systematic assessment of both in vitro and in vivo anti-tumor effects of docetaxel-loaded polyethylene glycol (PEG)2000-polycaprolactone (PCL)2600 micelles on hormone-refractory prostate cancer (HRPC). By using solvent evaporation method, PEG-PCL was chosen to prepare doxetaxel (DTX)-loaded mPEG-PCL micelles (DTX-PMs), with the purpose of eliminating side effects of the commercial formulation (Tween 80) and prolonging the blood circulation time. The prepared DTX-PMs had an average particle size of 25.19±2.36 nm, a zeta potential of 0.64±0.15 mV, a polydispersity index of 0.56±0.03, a drug loading of (8.72±1.05)%, and an encapsulation efficiency of (98.1±8.4)%. In vitro cytotoxicity studies indicated that DTX-PMs could effectively kill LNCap-C4-2B cells and show a dose- and time-dependent efficacy. The hemolysis test showed that DTX-PMs had less hemocytolysis than the commercial product of Duopafei®. A sustained in vitro release behavior and prolonged circulation time in blood vessels were observed in the DTX-PMs. Furthermore, when compared with Duopafei®, the DTX-PMs dramatically reduced the prostate specific antigen (PSA) level and tumor growth of prostate tumor-bearing nude mice in vivo. In conclusion, the DTX-PMs can lower systemic side effects, improve anti-tumor activity with prolonged blood circulation time, and will bring an alternative to patients with HRPC.

Preparation and characterization of polymeric nanoparticles for siRNA delivery to down-regulate the expressions of exogenous and endogenous target genes.

Gene silencing induced by RNA interference using small interfering RNA (siRNA) provides a promising therapeutic approach for cancers. However, the lack of siRNA delivery vector has limited the development of siRNA therapy. The purpose of this study was to use the novel copolymer (mPEG5k-PCL1.2k)1.4-g-PEl10k to prepare siRNA-loaded nanoparticles for siRNA delivery. The results suggested that (mPEG5k-PCL1.2k)1.4-g-PEl10k could load siRNA to form nanoparticles with particle size less than 200 nm in a narrow distribution. Moreover, a certain density of positive charge existed onto the surfaces of nanoparticles. MTT assay results demonstrated that (mPEG5k-PCL1.2k)1.4-g-PEl10k/siRNA nanoparticles showed very low cytotoxicity. The gene silencing efficiency of (mPEG5k-PCL1.2k)1.4-g-PEl10k/siRNA nanoparticles was investigated through luciferase reporter gene assays. The expression of exogenous luciferase gene was significantly downregulated at a range of N/P ratio from 50 to 125, and was maximally inhibited at the N/P ratio of 125 with 54% and 59% reduction in MCF-7 and HepG2 cells, respectively. In the 4T1-luc cell line expressing luciferase stably, the silencing of endogenous luciferase gene also has a similar overall profile with maximal 54% reduction of luciferase expression. These results suggested that (mPEG5k-PCL1.2k)1.4-g-PEI10k/SiRNA nanoparticles could serve as a kind of highly efficient siRNA delivery system for down-regulating the expression of exogenous and endogenous target genes.

[Small interfering RNA delivery mediated by mPEG-PCL-g-PEI polymer nanoparticles].

The aim of this paper is to report the synthesis of the mPEG-PCL-g-PEI copolymers as small interfering RNA (siRNA) delivery vector, and exploration of the siRNA delivery potential of mPEG-PCL-g-PEI in vitro. The diblock copolymers mPEG-PCL-OH was prepared through the ring-opening polymerization. Then, the hydroxyl terminal (-OH) of mPEG-PCL-OH was chemically converted into the carboxy (-COOH) and N-hydroxysuccinimide (NHS) in turn to prepare mPEG-PCL-NHS. The branched PEI was reacted with mPEG-PCL-NHS to synthesize the ternary copolymers mPEG-PCL-g-PEI. The structure of mPEG-PCL-g-PEI copolymers was characterized with Fourier transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR) and gel permeation chromatography (GPC). The mPEG-PCL-g-PEI/siRNA nanoparticles were prepared by complex coacervation, and the nanoparticles size and zeta potential were determined, separately. The cytotoxicities of mPEG-PCL-g-PEI/siRNA nanoparticles and PEI/siRNA nanoparticles were compared through cells MTT assays in vitro. The inhibition efficiencies of firefly luciferase gene expression by mPEG-PCL-g-PEI/ siRNA nanoparticle at various N/P ratios were investigated through cell transfection in vitro. The experimental results suggested that the ternary (mPEG5k-PCL(1.2k))1.4-g-PEI(10k) copolymers were successfully synthesized. (mPEG(5k)-PCL(1.2k))1.4-g-PEI(10k) could condense siRNA into nanoparticles (50-200 nm) with positive zeta potential. MTT assay results showed that the cytotoxicity of (mPEG(5k)-PCL(1.2k))1.4-g-PEI(10k)/siRNA nanoparticles was significantly lower than that of PEI(10k)/siRNA nanoparticles (P < 0.05). The expression of firefly luciferase gene could be significantly down-regulated at a range of N/P ratio from 50 to 150 (P < 0.01), and maximally inhibited at the N/P ratio of 125. The mPEG-PCL-g-PEI polymers could delivery siRNA into cells to inhibit the expression of target gene with very low cytotoxicity, which suggested that mPEG-PCL-g-PEI could serve as a new type of siRNA delivery vector.

[Preparation and in vitro evaluation of pH-sensitive TAT peptide conjugated micelles].

Doxorubicin loaded micelles were prepared by film-hydration method using stearyl sulfadiazine (SA-SD) which is pH sensitive, methoxy (polyethylene glycol)-2000-1, 2-dioleoyl-sn-glycero-3-phosphoethanolamine (mPEG-DOPE) and transactivator of transcription (TAT) peptide conjugated PEG-DOPE. Mean diameter of the pH-sensitive micelles was about 20 nm with a (99.1 +/- 2.1) % drug entrapment efficiency at pH 7.4. Flow cytometry studies revealed that the simple TAT micelles was taken up rapidly at the same level at pH 6.8 and pH 7.4. However, the pH-sensitive micelles entered the tumor cell less at pH 7.4 and significantly increase at pH 6.8. After 1 h incubation at pH 6.8, the amount of the pH-sensitive micelles taken up by cancer cell 4T1 was almost similar to simple TAT micelles. The confocal microscopy indicated that the pH-sensitive micelles entered the 4T1 cells at pH 6.8 more than at pH 7.4. It was indicated that the pH-sensitive micelles could shield TAT peptide at normal pH 7.4 and deshield it at pH 6.8. Hence, TAT peptides lead the drug-loaded micelles into the tumor cells and killed them selectively. The pH-sensitive micelle may provide a novel strategy for design of cancer targeting drug delivery system.

Efficient Anti-Glioma Therapy Through the Brain-Targeted RVG15-Modified Liposomes Loading Paclitaxel-Cholesterol Complex.

BACKGROUND: Glioma is the most common primary malignant brain tumor with a dreadful overall survival and high mortality. One of the most difficult challenges in clinical treatment is that most drugs hardly pass through the blood-brain barrier (BBB) and achieve efficient accumulation at tumor sites. Thus, to circumvent this hurdle, developing an effectively traversing BBB drug delivery nanovehicle is of significant clinical importance. Rabies virus glycoprotein (RVG) is a derivative peptide that can specifically bind to nicotinic acetylcholine receptor (nAChR) widely overexpressed on BBB and glioma cells for the invasion of rabies virus into the brain. Inspired by this, RVG has been demonstrated to potentiate drugs across the BBB, promote the permeability, and further enhance drug tumor-specific selectivity and penetration. METHODS: Here, we used the RVG15, rescreened from the well-known RVG29, to develop a brain-targeted liposome (RVG15-Lipo) for enhanced BBB permeability and tumor-specific delivery of paclitaxel (PTX). The paclitaxel-cholesterol complex (PTX-CHO) was prepared and then actively loaded into liposomes to acquire high entrapment efficiency (EE) and fine stability. Meanwhile, physicochemical properties, in vitro and in vivo delivery efficiency and therapeutic effect were investigated thoroughly. RESULTS: The particle size and zeta potential of PTX-CHO-RVG15-Lipo were 128.15 ± 1.63 nm and -15.55 ± 0.78 mV, respectively. Compared with free PTX, PTX-CHO-RVG15-Lipo exhibited excellent targeting efficiency and safety in HBMEC and C6 cells, and better transport efficiency across the BBB in vitro model. Furthermore, PTX-CHO-RVG15-Lipo could noticeably improve the accumulation of PTX in the brain, and then promote the chemotherapeutic drugs penetration in C6luc orthotopic glioma based on in vivo imaging assays. The in vivo antitumor results indicated that PTX-CHO-RVG15-Lipo significantly inhibited glioma growth and metabasis, therefore improved survival rate of tumor-bearing mice with little adverse effect. CONCLUSION: Our study demonstrated that the RVG15 was a promising brain-targeted specific ligands owing to the superior BBB penetration and tumor targeting ability. Based on the outstanding therapeutic effect both in vitro and in vivo, PTX-CHO-RVG15-Lipo was proved to be a potential delivery system for PTX to treat glioma in clinic.

Effects of gut microflora on pharmacokinetics of hesperidin: a study on non-antibiotic and pseudo-germ-free rats.

Hesperidin is a biologically active flavanone glycoside occurring abundantly in citrus fruits. In the present study, effects of intestinal microflora on pharmacokinetics of hesperidin were investigated using a pseudo-germ-free rat model treated with antibiotics. After administration of hesperidin to rats, hesperetin, hesperetin glucuronides, and metabolites postulated to be eriodictyol, hemoeriodictyol, and their glucuronides were detected in urine while hesperetin glucuronide was predominantly found in plasma. The plasma concentration-time profile of hesperetin was compared between non-antibiotic-exposed and pseudo-germ-free rats administered this compound. The maximal concentration (C(max)) values of hesperetin in non-antibiotic-exposed and pseudo-germ-free rats were 0.58 and 0.20 µg/ml, respectively, and area under the curve (AUC) values were 6.3 and 2.8 µg-h/ml, respectively. Thus, systemic exposure as evidenced by AUC and C(max) was significantly higher in normal compared to pseudo-germ-free rats. Fecal ß-glucosidase activities of non-antibiotic-exposed and pseudo-germ-free rats were 0.21 and 0.11 nmol/min/mg, while fecal α-rhamnosidase activities were 0.37 and 0.12 nmol/min/mg, respectively. The rate of hesperidin transformation to hesperetin was 6.9 and 2.9 nmol/min/g in fecal samples in non-antibiotic-exposed and pseudo-germ-free rats, respectively. Taken together, these results showed that pharmacokinetic differences between non-antibiotic-exposed and pseudo-germ-free rats may be attributed to differing hesperidin uptake, as well as alterations in metabolic activities of intestinal flora.

Improving antitumor outcomes for palliative intratumoral injection therapy through lecithin- chitosan nanoparticles loading paclitaxel- cholesterol complex.

BACKGROUND: Intratumoral injection is a palliative treatment that aims at further improvement in the survival and quality of life of patients with advanced or recurrent carcinomas, or cancer patients with severe comorbidities or those with a poor performance status. METHODS: In this study, a solvent-injection method was used to prepare paclitaxel-cholesterol complex-loaded lecithin-chitosan nanoparticles (PTX-CH-loaded LCS_NPs) for intratumoral injection therapy, and the physicochemical properties of NPs were well characterized. RESULTS: The particle size and zeta potential of PTX-CH-loaded LCS_NPs were 142.83±0.25 nm and 13.50±0.20 mV, respectively. Release behavior of PTX from PTX-CH-loaded LCS_NPs showed a pH-sensitive pattern. The result of cell uptake assay showed that PTX-CH-loaded LCS_NPs could effectively enter cells via the energy-dependent caveolae-mediated endocytosis and macropinocytosis in company with the Golgi apparatus. Meanwhile, PTX-CH-loaded LCS_NPs had a better ability to induce cell apoptosis than PTX solution. The in vivo antitumor results suggested that PTX-CH-loaded LCS_NPs effectively inhibited mouse mammary cancer growth and metastasis to distant organs and significantly improved the survival rate of tumor-bearing mice by intratumoral administration. CONCLUSION: In general, our study demonstrated that PTX-CH-loaded LCS_NPs used for palliative treatment by intratumoral injection showed improved safety and antitumor efficacy, which provided an alternative approach in the field of palliative chemotherapy.

Determination of macelignan in rat plasma by high-performance liquid chromatography with ultraviolet detection.

A high-performance liquid chromatographic method was developed for the determination of macelignan in rat plasma and applied to the pharmacokinetic study of macelignan in rats. Chromatographic separation was achieved on a conventional ODS column with the mobile phase of water: acetonitrile: methanol = 35:32.5:32.5 (v/v/v %). The flow rate of isocratic elution was 1 mL/min and peaks were detected at 240 nm. The limit of detection was 10 ng/mL and the limit of quantitation was 20 ng/mL. The calibration curve was linear over the concentration range of 50-5000 ng/mL. Intra-and inter-day precision for the assay over the concentration range was below 10 % and the accuracy ranged between 96.0-107% for intra-day and 98.8-114% for inter-day, respectively. The method was applied to the single dose pharmacokinetic study of macelignan in rats and the results showed that this HPLC method was adequate to support the in vivo pharmacokinetic study of macelignan.

Leucodin attenuates inflammatory response in macrophages and lipid accumulation in steatotic hepatocytes via P2x7 receptor pathway: A potential role in alcoholic liver disease.

The current study was aimed to reveal that leucodin, a sesquiterpene lactone from Artemisia capillaris could inhibit the inflammatory response in macrophages and the lipid accumulation in hepatocytes via P2x7R-NLRP3 inflammasome activation. Several types of macrophages including mouse peritoneal macrophages, mouse bone marrow-derived macrophages and human macrophages THP-1 cells were pretreated with different concentrations of leucodin for 1 h and then stimulated with LPS and ATP. LPS plus ATP initiated IL-1ß cleavage and release in mouse peritoneal macrophages and peaked at 4 h. Leucodin did not show significant toxicity within 200 µM and effectively inhibited pro-IL-1ß cleavage and release of mature-IL-1ß in macrophages. Also, P2x7R antagonist and caspase-1 inhibitor also decreased IL-1ß release and cleavage. Additionally, leucodin suppressed P2x7R, TLR4 and NLRP3 expression in LPS/ATP-stimulated macrophages. HepG2 cells were pretreated with different concentrations of leucodin for 1 h and then exposed to ethanol for 24 h. Leucodin suppressed lipid accumulation and enhanced phosphorylation of AMP-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC) in HepG2 cells exposed to ethanol. In addition, leucodin inhibited the expression of sterol regulatory element binding protein-1 (SREBP1) and ACC in ethanol-treated HepG2 cells. Leucodin possessed the capacity for inhibiting inflammatory response in macrophages and suppressing lipid accumulation in hepatocytes, suggesting a promising therapeutic potential targeting inflammation and lipid metabolism in alcoholic liver disease.

Inhibition of P2X7R-NLRP3 Inflammasome Activation by Pleurotus citrinopileatus: A Possible Protective Role in Alcoholic Hepatosteatosis.

Pleurotus citrinopileatus (golden oyster mushroom) is a widely used edible mushroom. We investigated the inhibitory effect of P. citrinopileatus aqueous extract against alcoholic steatohepatitis and its underlying mechanism. Acute and chronic ethanol-feeding murine models were established by intragastrically administering ethanol or feeding an ethanol-containing Lieber-DeCarli liquid diet to male C57BL/6 mice. In both models, P. citrinopileatus decreased serum alanine aminotransferase (ALT), aspartate transaminase (AST), triglyceride (TG), and hepatic TG levels. Hematoxylin and eosin (HE) and Oil Red O staining confirmed that P. citrinopileatus ameliorated both acute and chronic alcoholic hepatosteatosis, characterized by regulation of lipid-metabolism-related proteins, including sirtuin 1 (SIRT1), AMP-activated kinase (AMPK), and sterol regulatory element-binding protein (SREBP1). P. citrinopileatus reversed inflammatory response via modulating purinergic receptor P2X ligand-gated ion channel 7 (P2X7R)-NOD-like receptor pyrin domain 3 (NLRP3) inflammasome. P. citrinopileatus restored the expressions of those proteins to a normal level. In addition, HepG2 cells were incubated with P. citrinopileatus prior to ethanol stimulation. P. citrinopileatus reduced ethanol exposure-induced lipid deposition. Concomitantly, P. citrinopileatus increased AMPK and SIRT1 expressions, which were reduced by ethanol treatment. P. citrinopileatus ameliorated alcoholic hepatic steatosis and accompanied inflammatory response via regulating SIRT1-AMPK and P2X7R-NLRP3 inflammasome activation, highlighting a promising strategy and utility of P. citrinopileatus for alcoholic steatohepatitis as dietary health supplements.

Interaction of zalcitabine with human organic anion transporter 1.

The present study aimed to investigate the interaction of zalcitabine with human organic anion transporter 1 (hOATI) during renal excretion. Contribution of OAT1 to the renal transport of zalcitabine was examined using the transfected cell lines overexpressing the human organic anion transporter1 (CHO/hOAT1 cells). Zalcitabine exhibited the inhibition effect on the cellular uptake of [3H]-PAH in CHO/hOAT1 cells with an IC50 value of 1.23 mM. Furthermore, the cellular uptake of zalcitabine increased threefold with the enhancement of hOATI activity in CHO/hOAT1 cells, while it was significantly reduced in the presence of OAT1 inhibitors such as ketoprofen, naproxen, PAH and 6-carboxyfluorescein. Those results suggest that hOATI contributes at least in part to the cellular uptake of zalcitabine across the basolateral membrane of proximal tubular cells.

Inhibition of murine breast cancer growth and metastasis by survivin-targeted siRNA using disulfide cross-linked linear PEI.

Biodegradable disulfide-containing polyethyleneimine (PEI) derivatives showed great potential as siRNA vectors for the treatment of cancer due to the reduction-sensitive property. In this study, we developed and characterized a hyperbranched disulfide cross-linked PEI (lPEI-SS) based on linear PEI (lPEI) by ring-opening reaction of propylene sulfide. We evaluated the efficiency of lPEI-SS as a siRNA vector in vitro with luciferase reporter gene system, and investigated the anti-tumor efficacy of survivin-targeted siRNA (siRNA(sur)) on 4T1 murine breast cancer model using lPEI-SS synthesized here. Results from cytotoxicity and hemolysis assay proved that lPEI-SS showed favorable cell and blood compatibility. lPEI-SS/siRNA polyplexes prepared under the optimized condition were compact spherical particles with the average size of 229.0nm and zeta potential of 42.67mV. Cellular uptake of lPEI-SS/siRNA polyplexes was significantly improved due to the higher branching degree of lPEI-SS over the parent lPEI. lPEI-SS/siRNA(sur) exhibited great anti-proliferation effect on 4T1 cell line, which was found to be caused by the induction of apoptosis. Most importantly, results of tumor volume, tumor weight and histological observation demonstrated that lPEI-SS/siRNA(sur) polyplexes effectively inhibited the tumor growth and metastasis of 4T1 murine breast cancer model.

Lx2-32c-loaded polymeric micelles with small size for intravenous drug delivery and their inhibitory effect on tumor growth and metastasis in clinically associated 4T1 murine breast cancer.

Lx2-32c is a novel taxane derivative with a strong antitumor activity. In this study, we developed Lx2-32c-loaded polymeric micelles (Lx2-32c-PMs) with small size and investigated their antitumor efficacy against tumor growth and metastasis on 4T1 murine breast cancer cell line with Cremophor EL-based Lx2-32c solution as the control. In this study, copolymer monomethoxy polyethylene glycol2000-polylactide1300 was used to prepare Lx2-32c-PMs by film hydration method, and their physicochemical properties were characterized as well, according to morphology, particle size, zeta potential, in vitro drug release, and reconstitution stability. Under confocal laser scanning microscopy, it was observed that Lx2-32c-PMs could be effectively taken up by 4T1 cells in a time-dependent manner. Cell Counting Kit-8 assay showed that the IC50 of Lx2-32c-PMs was 0.3827 nM. Meanwhile, Lx2-32c-PMs had better ability to promote apoptosis and induce G2/M cycle block and polyploidy formation, compared with Lx2-32c solution. More importantly, in vivo animal studies showed that compared to Lx2-32c solution, Lx2-32c-PMs possessed better ability not only to effectively inhibit the tumor growth, but also to significantly suppress spontaneous and postoperative metastasis to distant organs in 4T1 orthotopic tumor-bearing mice. Consequently, Lx2-32c-PMs have significantly prolonged the survival lifetime of tumor-bearing mice. Thus, our study reveals that Lx2-32c-PMs had favorable antitumor activity and exhibited a good prospect for application in the field of antitumor therapy.

A Liquid Chromatography-Tandem Mass Spectrometry Method for Simultaneous Quantitation of 10 Bioactive Components in Rhus verniciflua Extracts.

The purpose of this study was to develop a simultaneous method to quantify 10 bioactive compounds in Rhus verniciflua extracts using high-performance liquid chromatography-tandem mass spectrometry. The chromatographic separation was performed using a C18 column under gradient elution with 0.1% formic acid and acetonitrile as the mobile phase solvents. The analytes were detected in the negative-ion mode using multiple-reaction monitoring detection with an electrospray ionization interface. The calibration curves for all the analytes showed good linearity (r(2) >0.997) over the concentration range of 1-1,000 ng/mL. The recovery values were within 89.53-110.14%, and the intra- and interday coefficients of variation were <4.86% for all the tested compounds. The developed method was successfully applied to a quality assessment of the R. verniciflua extract samples.

Role of monocarboxylic acid transporters in the cellular uptake of NSAIDs.

The present study investigated the cellular uptake mechanism of non-steroidal anti-inflammatory drugs (NSAIDs) in Caco-2 cells. Diflunisal, diclofenac, ketoprofen and naproxen exhibited a strong inhibition effect on the cellular uptake of [14C]-benzoic acid in Caco-2 cells with IC50 values of 0.05-0.44 mM. The inhibition of naproxen and ketoprofen against the membrane transport of [14C]-benzoic acid appeared to be competitive, with Ki values of 0.22 and 0.38 mM, respectively. The membrane permeability of naproxen and ketoprofen was concentration dependent, implying that the cellular uptake pathway of ketoprofen and naproxen was saturable at the higher concentration. Furthermore, the cellular accumulation of ketoprofen was significantly reduced in the presence of benzoic acid and L-lactic acid, two known substrates of monocarboxylic acid transporter 1 (MCT1). These results suggest that MCT1 contributes at least in part to the carrier-mediated transport of NSAIDs containing a carboxylic acid moiety across the apical membrane in Caco-2 cells.