Zwitterionic pH-responsive hyaluronic acid polymer micelles for delivery of doxorubicin.

As zwitterionic polymers show great promise in drug delivery, hyaluronic acid (HA) was deacetylated and grafted with dodecylamine to prepare a pH-sensitive zwitterionic polymer dHAD used as a carrier for antitumor drugs. The polymer was negatively charged at pH 7.4 and became positive at pH 6.2. In vitro delivery of DOX against MCF-7 cells showed that the blank micelle dHAD had low cytotoxicity and the dHAD-DOX micelles could greatly prohibit the growth of the MCF-7 cells. In addition, the dHAD-DOX micelles had higher cellular uptake, indicating that the micelles were rapidly internalized into the cells via CD44 receptor-mediated endocytosis. The in vivo delivery of DOX to tumor-bearing mice confirmed that the dHAD-DOX micelles greatly inhibited the tumor growth and significantly reduced systemic toxicity of DOX. These results demonstrated that biocompatible pH-responsive zwitterionic dHAD micelles are promising carriers for the delivery of DOX.

7-O-geranylquercetin-induced autophagy contributes to apoptosis via ROS generation in human non-small cell lung cancer cells.

AIMS: To investigate the antitumor effects of 7-O-geranylquercetin (GQ), a novel O-alkylated derivative of quercetin, against non-small cell lung cancer (NSCLC) cell lines A549 and NCI-H1975 and the corresponding mechanisms. MAIN METHODS: Cell viability was assessed using MTT assay. The expression of proteins involved in apoptosis and autophagy was measured using western blotting. Besides, apoptosis was determined with DAPI staining, Annexin V-PI staining and transmission electron microscopy (TEM) assay, and autophagy was observed with TEM assay. Cell cycle and reactive oxygen species (ROS) level were detected using flow cytometry. KEY FINDINGS: GQ inhibited viability of A549 and NCI-H1975 cells in a dose- and time-dependent manner without apparent cytotoxicity to normal human lung fibroblast cells. GQ down-regulated the expression of apoptosis-related proteins pro-caspase 3 and Bcl-2, and up-regulated the expression of cleaved-PARP and Bax in A549 and NCI-H1975 cells. Meanwhile, GQ-induced cell apoptosis could be attenuated by caspase inhibitor Z-VAD-FMK. Besides, GQ induced autophagosome formation in A549 and NCI-H1975 cells, promoted the expression of autophagy-related proteins LC3-II and Beclin 1, and suppressed the expression of p62. Autophagy inhibition with chloroquine or Beclin 1 siRNA could effectively inhibit GQ-induced apoptosis. Furthermore, GQ treatment increased the generation of ROS, and ROS inhibitor N-acetylcysteine could reverse GQ-induced autophagy and apoptosis. Taken together, GQ could induce apoptosis and autophagy via ROS generation in A549 and NCI-H1975 cells, and GQ-induced autophagy contributed to apoptosis. SIGNIFICANCE: Our findings highlight that GQ is a promising anticancer agent for the treatment of lung cancer.

Synthesis, Characterization and Cytotoxicity of Alkylated Quercetin Derivatives.

Quercetin, a ubiquitous flavonol, represents a promising leading drug for development of new chemotherapeutic agents. However, its limited cytotoxicity to cancer cells hampers its clinical use. In order to obtain novel quercetin derivatives with superior cytotoxicity, seven alkylated quercetin derivatives were synthesized. Solubility of these derivatives was determined by turbidimetry. Cytotoxicity of the high-soluble derivatives against MCF-7 cells and caco-2 cells was determined using MTT assay. Among these seven products, 7-O-butylquercetin had the highest solubility in DMEM medium and 7-O-geranylquercetin had the most potent cytotoxicity. Further study on cytotoxicity of 7-O-geranylquercetin on NCI-H446, A549, MGC-803 and SGC-7901 cell lines revealed potential antiproliferative effects. The 7-O-geranylquercetin is a broad spectrum cytotoxic agent and it may be a promising leading drug for cancer chemotherapy.

The Effects of High Fat Diet and Resveratrol on Mitochondrial Activity of Brown Adipocytes

BACKGROUND: Resveratrol (RSV) is a polyphenolic phytoalexin that has many effects on metabolic diseases such as diabetes and obesity. Given the importance of brown adipose tissue (BAT) for energy expenditure, we investigated the effects of RSV on brown adipocytes. METHODS: For the in vitro study, interscapular BAT was isolated from 7-week-old male Sprague Dawley rats. For the in vivo study, 7-week-old male Otsuka Long Evans Tokushima Fatty (OLETF) rats were divided into four groups and treated for 27 weeks with: standard diet (SD); SD+RSV (10 mg/kg body weight, daily); high fat diet (HFD); HFD+RSV. RSV was provided via oral gavage once daily during the in vivo experiments. RESULTS: RSV treatment of primary cultured brown preadipocytes promoted mitochondrial activity, along with over-expression of estrogen receptor α (ER-α). In OLETF rats, both HFD and RSV treatment increased the weight of BAT and the differentiation of BAT. However, only RSV increased the mitochondrial activity and ER-α expression of BAT in the HFD-fed group. Finally, RSV improved the insulin sensitivity of OLETF rats by increasing the mitochondrial activity of BAT, despite having no effects on white adipocytes and muscles in either diet group. CONCLUSION: RSV could improve insulin resistance, which might be associated with mitochondrial activity of brown adipocyte. Further studies evaluating the activity of RSV for both the differentiation and mitochondrial activity of BAT could be helpful in investigating the effects of RSV on metabolic parameters.

Chitosan-alginate microcapsules for oral delivery of egg yolk immunoglobulin (IgY): effects of chitosan concentration.

In our previous study, chitosan-alginate microcapsules were developed to protect egg yolk immunoglobulin (IgY) from gastric inactivation. The present study was undertaken to determine the effect of chitosan concentration (0-0.8%; w/v) on various properties of the microcapsules in order to produce the optimum chitosan-alginate microcapsules for use in the oral delivery of IgY. The properties investigated included microcapsule morphology, loading capacity for IgY (expressed as the IgY loading percentage, w/w, of microcapsules), encapsulation efficiency (EE%), in vitro gastroresistance, and IgY release. IgY loading percentage and EE% were both highest at 0.2% (w/v) chitosan, and, above this level, further increases were not observed. The stability of IgY in simulated gastric fluid (pH 1.2) was significantly improved by encapsulation in alginate microcapsules (IgY retained 43.5% of its activity) and was further improved by including chitosan at any of the chitosan concentrations assessed (IgY retained an average of 69.4% activity) although there was no difference in protection of gastric inactivation among concentrations of chitosan varying from 0.05% to 0.8% (w/v). Higher chitosan concentrations (i.e., >/=0.2%; w/v) prolonged the release of IgY from the microcapsules during simulated intestinal fluid incubation (pH 6.8). However, above the 0.2% (w/v) level, no significant differences were observed. We conclude that the optimum chitosan concentration for microencapsulation is 0.2% (w/v).

Chitosan-alginate microcapsules for oral delivery of egg yolk immunoglobulin (IgY): in vivo evaluation in a pig model of enteric colibacillosis.

In our previous study, the applicability of chitosan-alginate microcapsules for oral delivery of egg yolk immunoglobulin (IgY) was established in a simulated gastrointestinal tract environment. The objective of the present study was to evaluate the protective efficacy of microencapsulated IgY against K88+ ETEC (enterotoxigenic Escherichia coli)-induced diarrhea in 40-day-old pigs. Groups of pigs orally challenged with 10(11) cfu/mL of K88+ ETEC were fed with non-encapsulated IgY, microencapsulated IgY and aureomycin-treated feed respectively. The clinical response of each group was monitored and evaluated in terms of lethargy, inappetence, occurrence of diarrhea, fecal consistency score, weight loss and recovery rate. The results showed that treatment of infected pigs with microencapsulated IgY significantly (P<0.05) reduced the K88+ ETEC-induced diarrhea at 24 h post-infection. In contrast, the diarrhea-reducing effect of non-encapsulated IgY was delayed (only evident after 72 h) while normal saline-treated pigs (controls) continued to suffer from diarrhea and dehydration. Similarly, weight gain in microencapsulated IgY-treated pigs was better and significantly different (P<0.05) than in non-encapsulated IgY and saline-treated controls. Collectively, these results support previous in vitro observations showing that chitosan-alginate microcapsules can be an effective method of protecting IgY from gastric inactivation, enabling its use for the widespread prevention and control of enteric diseases.

Efficacy of specific egg yolk immunoglobulin (IgY) to bovine mastitis caused by Staphylococcus aureus.

The objective of this study was to estimate the efficacy of specific egg yolk immunoglobulin (IgY) to bovine mastitis caused by Staphylococcus aureus. Eighteen lactating cows with clinical mastitis and 18 lactating cows with experimental mastitis (1 quarter per cow) were randomly assigned to three treatments: IgY (20mg/ml) infusion, penicillin (100mg/ml) infusion and no infusion. Treatments for clinical mastitis and experimental mastitis were performed by a 6-day course of intramammary infusion with a dosage of 10ml at an interval of 12h. Milk samples were collected at morning milking time for testing color, clot, somatic cell counts (SCC) and bacterial count. For most of the cows treated with IgY and penicillin, the milk color and clot recovered to normal form during the therapy course. The milk SCCs and bacterial counts of treated cows decreased compared to those of untreated cows (p<0.05). The cure rates by IgY for experimental and clinical mastitis were 83.3% and 50%, respectively, and those by penicillin were 66.7% and 33.3%, respectively. These results showed the potential of specific IgY to be an alternative therapy for mastitis caused by S. aureus.

Characterization of specific egg yolk immunoglobulin (IgY) against mastitis-causing Escherichia coli.

The objective of this study was to estimate the in vitro activity of egg yolk immunoglobulin (IgY) against mastitis-causing Escherichia coli. Specific IgY was produced by hens immunized with formaldehyde killed E. coli O111 in long-standing immunization response (titer > or =6400 for 100 days) and was isolated from yolks with a purity of 86% by water dilution, salt precipitations and ultrafiltration. Enzyme-linked immunosorbent assay (ELISA) indicated the produced IgY specifically targeted E. coli O111 and five other E. coli strains which were isolated from mastitic cows. The growth inhibition activity of the specific IgY to bacteria was dose-dependent with an effective concentration of 20mg purified IgY per milliliter. The phagocytic activity of E. coli either by milk macrophages (MPhi) or by polymorphonuclear neutrophil leukocytes (PMN) in the presence of specific IgY was significantly higher than that with nonspecific IgY or without IgY (p<0.05), suggesting that it enhanced phagocytic activity. The current work suggests that this specific IgY has potential as a therapeutic treatment for mastitis in dairy cows.

Chitosan-alginate microcapsules for oral delivery of egg yolk immunoglobulin (IgY).

Chitosan-alginate microcapsules were evaluated as a method of oral delivery of IgY antibodies. Physical characteristics, encapsulation efficiency (EE%), the loading capacity for IgY (IgY loading percentage, %, w/w of microcapsules), gastro-resistance, and release characteristics of these microcapsules in vitro under varying pH were investigated. Optimum physical factors were established for preparation of homogeneous, spherical, and smooth microcapsules. IgY loading% was not significantly altered by pH of the encapsulation medium. Encapsulation efficiency was highest (73.93%) at a pH of 3.5, above which EE% decreased significantly (p < 0.05). IgY was released from microcapsules upon exposure to simulated intestinal fluid (SIF, pH 6.8), and decreasing pH increased significantly IgY release (p < 0.05). The stability of IgY in simulated gastric fluid (SGF, pH 1.2) was greatly improved by encapsulation in chitosan-alginate microcapsules, and the residual activity was not affected by pH of the encapsulation medium. Moreover, microencapsulated IgY was significantly resistant to pepsin hydrolysis. This approach may enable intact IgY to reach target microorganisms within the lower digestive tract.

3,5-Dihydr-oxy-N'-[(2-hydr-oxy-1-naph-thyl)methyl-ene]benzohydrazide.

In the title compound, C(18)H(14)N(2)O(4), the dihedral angle between the benzene ring and the naphthyl ring system is 10.1 (2)°. The mol-ecule is nearly planar, with a mean deviation from the plane of 0.141 (2) Šfor 24 non-H atoms. An intra-molecular O-H⋯N hydrogen bond forms a pseudo-6-membered ring and the mol-ecules are linked into sheets by inter-molecular N-H⋯O and O-H⋯O hydrogen bonds.