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Transforming growth factor (TGF)-β regulates the function of vascular endothelial cells and may be involved in endothelial dysfunction. The canonical TGF-β pathway involves TGF-β receptor-mediated carboxy-terminal phosphorylation of Smad2; however, TGF-β signalling also activates numerous serine/threonine kinases that phosphorylate Smad2 in its linker region. The expression of phosphorylated Smad linker proteins were determined following TGF-β stimulation in the absence and presence of different serine/threonine kinase inhibitors in vascular endothelial cells.
Proteins were quantified by Western blotting using specific antibodies to individual phosphorylated Smad2 linker region residues.
TGF-β mediated the phosphorylation of all four Smad2 linker region residues of interest. Erk and Jnk specifically phosphorylate Ser245 while all mitogen-activated protein kinases phosphorylate Ser250 and Ser255. Thr220 and Ser245 are phosphorylated by phosphoinositide 3 kinase (PI3K), while Ser255 was phosphorylated by the PI3K/Akt pathway. CDK and GSK-3 were shown to phosphorylate Thr220 and Ser245. TGF-β also mediated plasminogen activator inhibitor-1 gene expression that was attenuated by p38 and CDK inhibitors.
TGF-β-mediated phosphorylation of individual serine/threonine sites in the linker region of Smad2 occurs in a highly specific manner by kinases. These phosphorylations provide an opportunity to further understand a therapeutically targeted and very specific signalling pathway in vascular endothelial cells.
The sublingual mucosa provides a promising route for the delivery of glutathione (GSH). However, challenges are encountered in developing sublingual mucoadhesive drug delivery formulations such as: prolonging drug retention, uniform drug content, desirable drug release profiles, adequate drug permeation and efficient delivery of GSH. The aim of this study was to develop a suitable mucoadhesive polymer-based sublingual film.
The mucoadhesive films were prepared by casting method. Several characterization studies including thickness, weight uniformity, surface pH, elongation, mucoadhesiveness, swelling and erosion were carried out on preliminary formulations to optimise formulations for in-vitro drug release and ex-vivo permeation studies.
The optimal mucoadhesive polymer-based films showed acceptable physical properties and good mucoadhesion, and remained attached to excised porcine sublingual mucosa for sufficient time, providing a sustained delivery of GSH through the mucosal epithelial.
The optimal mucoadhesive films may provide a promising drug delivery platform to develop commercial sublingual products of GSH as well as a wide range of protein and peptide drugs.
For anaphylaxis treatment in community settings, adrenaline (epinephrine) administration using an auto-injector in the thigh is universally recommended. Despite this, many people at risk of anaphylaxis in community settings do not carry their prescribed auto-injectors consistently and hesitate to use them when anaphylaxis occurs.The objective of this research was to study the effect of a substantial reduction in adrenaline (Epi) particle size to a few micrometres (Epi microcrystals (Epi-MC)) on enhancing adrenaline dissolution and increasing the rate and extent of sublingual absorption from a previously developed rapidly disintegrating sublingual tablet (RDST) formulation in a validated preclinical model.
The in-vivo absorption of Epi-MC 20 mg RDSTs and Epi 40 mg RDSTs was evaluated in rabbits. Epi 0.3 mg intramuscular (IM) injection in the thigh and placebo RDSTs were used as positive and negative controls, respectively.
Epimean(standard deviation) area under the plasma concentration vs time curves up to 60 min and Cmax from Epi-MC 20 mg and Epi 40 mg RDSTs did not differ significantly (P > 0.05) from Epi 0.3 mg IM injection. After adrenaline, regardless of route of administration, pharmacokinetic parameters were significantly higher (P < 0.05) than after placebo RDSTs administration (reflecting endogenous adrenaline levels).
Epi-MC RDSTs facilitated a twofold increase in Epi absorption and a 50% reduction in the sublingual dose. This novel sublingual tablet formulation is potentially useful for the first-aid treatment of anaphylaxis in community settings.
Evaluate the ability of biorelevant media to adequately predict solubility in human gastrointestinal aspirates collected in the fasted state for the sodium salt of a highly dosed, Biopharmaceutics Classification System II (BCS II) compound with weakly acidic properties (L-870,810, pKa 7.3, HA (5-(1,1-dioxothiazinan-2-yl)-N-((4-fluorophenyl)methyl)-8-hydroxy-1,6-naphthyridine-7-carboxamide)). Identify key luminal processes that dictate the behaviour of sodium salt of HA (NaA), after single-dose administrations of high (relatively to solubility limit) doses corresponding to 400 and 800 mg of HA in the fasted state.
Aspirates from stomach and upper small intestine were collected from eight healthy fasted adults, after administration of 240 ml of water. Solubilities of NaA and HA were measured in aspirated samples and biorelevant media. Dissolution experiments of NaA granules were performed in biorelevant media. Prediction of oral pharmacokinetics was evaluated in silico using Stella software.
Equilibrium solubility of NaA in fluids aspirated from the upper gastrointestinal tract is more transient than of HA. Solubility in upper gastrointestinal lumen was adequately estimated by data in biorelevant media. Supersaturation, followed by precipitation, which did not fully revert to the equilibrium solubility of HA, was observed during the dissolution of NaA granules in biorelevant media. Physiologically based pharmacokinetic modelling indicated that while intragastric processes had no significant impact on absorption kinetics, dissolution kinetics, kinetic solubility, radial transport rates and, for the 800-mg dose, precipitation kinetics in the small intestine had the greatest impact on absorption profiles.
Adequate prediction of the average plasma profile, after administration of NaA, required consideration of region-dependent dissolution rates and/or solubilisation.
An in-situ forming gel-like depot, prepared by using an appropriate polyaspartamide-polylactide graft copolymer, has been employed to release in a sustained way sulpiride.
α,β-poly(N-2-hydroxyethyl)-D,L-aspartamide-g-polylactic acid (PHEA-g-PLA) has been used as a polymer component. Its physicochemical properties make possible to dissolve it in N-methyl-2-pyrrolidone, with the obtainment of a solution able to form a gel-like depot once injected into a physiological medium. Cell compatibility of PHEA-g-PLA depot has been investigated, using murine dermal fibroblasts as cell model. 3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt assay and fluorescence microscopy have been employed to evaluate cell viability and morphology after contact with PHEA-g-PLA depot. Pharmacokinetic parameters of sulpiride released from depot have been determined following subcutaneous administration to rabbits and compared with corresponding parameters following administration of free sulpiride solution.
It has been demonstrated that the system does not affect significantly the viability of fibroblasts and is able to sustain the release of sulpiride until a week, with a burst effect dependent on the initial weight ratio polymer/drug.
In-vivo release profiles and pharmacokinetic parameters suggest that PHEA-g-PLA depot could have interesting clinical applications for a once a week administration of poorly soluble drugs to humans or animals.
Glycosylation or the modification of a cellular component with a carbohydrate moiety has been demonstrated in all three domains of life as a basic post-translational process important in a range of biological processes. This review will focus on the latest studies attempting to exploit bacterial N-linked protein glycosylation for glycobiotechnological applications including glycoconjugate vaccine and humanised glycoprotein production. The challenges that remain for these approaches to reach full biotechnological maturity will be discussed.
Oligosaccharyltransferase-dependent N-linked glycosylation can be exploited to make glycoconjugate vaccines against bacterial pathogens. Few technical limitations remain, but it is likely that the technologies developed will soon be considered a cost-effective and flexible alternative to current chemical-based methods of vaccine production. Some highlights from current glycoconjugate vaccines developed using this in-vivo production system include a vaccine against Shigella dysenteriae O1 that has passed phase 1 clinical trials, a vaccine against the tier 1 pathogen Francisella tularensis that has shown efficacy in mice and a vaccine against Staphylococcus aureus serotypes 5 and 8. Generation of humanised glycoproteins within bacteria was considered impossible due to the distinct nature of glycan modification in eukaryotes and prokaryotes. We describe the method used to overcome this conundrum to allow engineering of a eukaryotic pentasaccharide core sugar modification within Escherichia coli. This core was assembled by combining the function of the initiating transferase WecA, several Alg genes from Saccharomyces cerevisiae and the oligosaccharyltransferase function of the Campylobacter jejuni PglB. Further exploitation of a cytoplasmic N-linked glycosylation system found in Actinobacillus pleuropneumoniae where the central enzyme is known as N-linking glycosyltransferase has overcome some of the limitations demonstrated by the oligosaccharyltransferase-dependent system.
Characterisation of the first bacterial N-linked glycosylation system in the human enteropathogen Campylobacter jejuni has led to substantial biotechnological applications. Alternative methods for glycoconjugate vaccine production have been developed using this N-linked system. Vaccines against both Gram-negative and Gram-positive organisms have been developed, and efficacy testing has thus far demonstrated that the vaccines are safe and that robust immune responses are being detected. These are likely to complement and reduce the cost of current technologies thus opening new avenues for glycoconjugate vaccines. These new markets could potentially include glycoconjugate vaccines tailored specifically for animal vaccination, which has until today thought to be non-viable due to the cost of current in-vitro chemical conjugation methods. Utilisation of N-linked glycosylation to generate humanised glycoproteins is also close to becoming reality. This 'bottom up' assembly mechanism removes the heterogeneity seen in current humanised products. The majority of developments reported in this review exploit a single N-linked glycosylation system from Campylobacter jejuni; however, alternative N-linked glycosylation systems have been discovered which should help to overcome current technical limitations and perhaps more systems remain to be discovered. The likelihood is that further glycosylation systems exist and are waiting to be exploited.
A number of species belonging to herbal genus Aconitum are well-known and popular for their medicinal benefits in Indian, Vietnamese, Korean, Japanese, Tibetan and Chinese systems of medicine. It is a valuable drug as well as an unpredictable toxic material. It is therefore imperative to understand and control the toxic potential of herbs from this genus. In this review, the ethnomedicinal, phytochemistry, pharmacology, structure activity relationship and toxicology studies of Aconitum were presented to add to knowledge for their safe application.
A total of about 76 of all aconite species growing in China and surrounding far-east and Asian countries are used for various medical purposes. The main ingredients of aconite species are alkaloids, flavonoids, free fatty acids and polysaccharides. The tuberous roots of genus Aconitum are commonly applied for various diseases such as rheumatic fever, painful joints and some endocrinal disorders. It stimulates the tip of sensory nerve fibres. These tubers of Aconitum are used in the herbal medicines only after processing. There remain high toxicological risks of the improper medicinal applications of Aconitum. The cardio and neurotoxicities of this herb are potentially lethal. Many analytical methods have been reported for quantitatively and qualitatively characterization of Aconitum.
Aconitum is a plant of great importance both in traditional medicine in general and in TCM in particular. Much attention should be put on Aconitum because of its narrow therapeutic range. However, Aconitum's toxicity can be reduced using different techniques and then benefit from its pharmacological activities. New methods, approaches and techniques should be developed for chemical and toxicological analysis to improve its quality and safety.
Although the exact underlying mechanisms are still unknown, Plantago lanceolata L. (PL) water extracts are frequently used to stimulate wound healing and to drain abscesses. Therefore, in this experimental study the effect of PL water extract on skin wound healing was studied in Sprague-Dawley rats.
Two excisional and one incisional skin wounds were performed on the back of each rat. Wounds were treated for three consecutive days with two different concentrations of the aqueous extract of PL. Rats were sacrificed 7, 14, and 21 days after surgery. Samples of wounds were processed for macroscopic (excisions – wound contraction measurement), biomechanical (incisions – wound tensile strength (TS) measurement) and histological examination (excisions).
It was shown that open wounds treated with PL extract contained myofibroblasts and demonstrated significantly higher contraction rates. Furthermore, significantly increased wound TSs were recorded in treated rats as a consequence of increased organization of extracellular matrix proteins, such as the collagen type 1.
We demonstrated that PL aqueous extract improves skin wound healing in rats. However, further research need to be performed to find optimal therapeutic concentration, and exact underlying mechanism prior obtained results may be introduced into the clinical practice.
Litchi chinensis has been traditionally used in folk medicine to treat several ailments. In this study, we investigated the chemical composition, antioxidant and antinociceptive activity of L. chinensis leaves.
The antioxidant capacity of the extract, fraction and compounds was evaluated using the 1,1-diphenyl-picrylhydrazyl (DPPH) and 2,2′-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assays, and the liposome model with peroxyl radicals generated by 2,2′-azobis (2-amidinopropane) dihydrochloride radical. The pharmacological models of acute nociception used in mice were: writhing test with acetic acid (AA), hotplate (HP), glutamate (GLU), capsaicin (CP) and formalin (FM) tests.
The main compounds isolated were procyanidin A2 (PA2), procyanidin B2 (PB2) and (–)-epicatechin. The biochemical features of the crude extracts and their ethyl acetate fraction (EtOAcFR) presented high antioxidant activity, and the antioxidant activity of PA2 and PB2 was remarkably high, with DPPH and ABTS. The crude methanol extract (MeOHEXTR), EtOAcFR and PB2 were effective in reducing nociception in FM and HP models. MeOHEXTR and EtOAcFR treatments also reduced pain induced by GLU and AA. In the CP model, only EtOAcFR and PB2 were effective.
The results demonstrate the antinociceptive and antioxidant of MeOHEXTR, EtOAcFR and PB2.
The aim of this study was to extend our knowledge about the mechanism involved in the gastroprotective effect of P1G10, a proteolytic fraction rich in cysteine proteinases from Vasconcellea cundinamarcensis (syn. Carica candamarcensis) latex, which demonstrated gastric healing and protection activities in rats.
Wistar rats were submitted to gastric lesions by indomethacin and treated with P1G10 (10 mg/kg). Free thiol groups and prostaglandin E2 content were measured in gastric mucosal and gastrin levels in blood samples. To evaluate the participation of nitric oxide (NO) or proteolytic activity of P1G10 on its gastroprotective effect, animals were treated with an inhibitor of NO production (L-NAME) or the fraction inhibited by iodoacetamide, respectively. Gastric secretion study (acidity and pepsin activity) was also performed.
P1G10 (10 mg/kg) inhibited the occurrence of gastric lesions by indomethacin, restored the free thiol groups content on gastric mucosa and increased moderately prostaglandin E2 levels (34%). Furthermore, the treatment decreased the gastrin levels (95%), suggesting a possible modulation of secretory activity. This effect was accordant with attenuation of gastric acidity (42%) and pepsin activity (69%) seen in animals subjected to pyloric ligation. The inhibition of NO production or the proteolytic activity of P1G10 does not affect the gastroprotective effect.
These results can explain the gastroprotective activity of P1G10 and serve a basis for further studies of this active principle.
The aims of this study were to identify the active ingredients from Portulaca oleracea L. (PO) that could provide synergism with antibiotics against methicillin-resistant Staphylococcus aureus (MRSA) and their possible mechanisms of resistance inhibition.
High-speed counter-current chromatography (HSCCC) coupled with gas chromatography-mass spectrometry and a panel of laboratory MRSA strains were used for checkerboard and efflux inhibitory assays.
Linoleic and oleic acids were identified from HSCCC fraction 18 of PO with synergistic antibacterial activity when combined with erythromycin against RN4220/pUL5054. Ethidium bromide efflux inhibitory studies revealed that linoleic and oleic acids may interfere the activity of MsrA pump. By comparing among a panel of linoleic and oleic acids analogues, unsaturated fatty acids in salt form with cis configuration and an increase in number of double bonds were found to further increase the antibacterial activity when used alone or in combination with antibiotics.
This study reported for the first time that two active ingredients, namely linoleic and oleic acids, were identified from PO with synergistic antibacterial activity when combined with erythromycin against MRSA RN4220/pUL5054 and possibly act by inhibiting the efflux pumps of the bacteria cells.
Doxorubicin (Dox) is a frontline chemotherapeutic against osteosarcoma (OS) that is plagued by side effects, particularly in the heart. The specific objective of this article is to investigate whether low-dose Dox treatment had pro-autophagic effects in cardiomyocytes as well as osteosarcoma cells.
This study characterises apoptotic (Bax) and autophagic (Beclin-1) biomarker levels in human OS and cardiomyocyte cell lines as well as in various tissues when mice are exposed to low (1 mg/kg, thrice weekly) and high (3 mg/kg thrice weekly) dose Dox for a month.
There was a decrease in Bax and increase in Beclin-1 in cardiac tissue in the high-dose group. Dox decreased Beclin-1 in the skin and liver, with no clear indication in the stomach, small intestine and testis. At low Dox doses of 10 and 100 nm in cardiomyocytes and OS cells, there is a pro-apoptotic effect, with a quicker response in the 100-nm condition, and a slower but steady increase of a pro-apoptotic response at the lower 10-nm dose. However, electron microscopy images revealed changes to human OS cells that resembled autophagy. Human prostate, breast and colorectal cells treated with 10-nm Dox showed ∼ 40% reduction in cell viability after 24 h.
In culture, cells of both cardiomyocytes and OS revealed a predominant pro-apoptotic response at the expense of autophagy, although both seemed to be occurring in vivo.
Doxorubicin (Dox) is used clinically against various neoplasias, but suffers from serious side effects, and for the past three decades, this shortcoming has spurred research towards finding better drug delivery systems (DDSs) for this frontline drug.
A non-targeted nucleotropic Dox-loaded nanoparticle (DNP) DDS is described, which has a simple chemical design, is easy to formulate and administer, is inexpensive, non-biohazardous and may prove to be useful clinically.
The DNP formulated via vortex-assisted complex coarcevation enhanced (300-fold) cell-inhibitory activity of the drug in a panel of human cancer cells (osteosarcoma, breast, prostate and colorectal cancer) and enhanced (10-fold) efficacy against osteosarcoma (OS) in vivo. The slow-release DNPs localised to the endoplasmic reticulum disrupted the mitochondria and entered the nucleus. Prominent cytosolic vacuolisation, budding off of portions of the cytoplasm, both suggestive of autophagy, were observed. Mice that were administered with DNPs intratumorally had the smallest tumours at the end of the study, with more necrotic hotspots.
This promising nucleotropic DDS enhances the cell delivery and activity of Dox against a variety of human cancer cell lines and in OS tumours in mice.
We tested if modulation of cytosolic K+ levels by ouabain, an inhibitor of Na+/K+-ATPase, exerts cytoprotection against distinct stressful stimuli in human leukemic cells.
The cytosolic K+, Na+ or Ca2+ levels and the cytotoxicity were evaluated by flow cytometry.
Various cytotoxic chemicals and ultraviolet irradiation induced cell death and increased intracellular concentrations of K+, Na+ or Ca2+. Ouabain reduced the cytotoxicity and the elevation of cytosolic levels of K+ but not those of Na+ or Ca2+.
Our data thus suggest that elevated cytosolic K+ levels are associated with the cytotoxicity in response to distinct stressful stimuli and that ouabain exerts cytoprotection most probably by regulating intracellular K+ levels.
This study was to explore the antifibrotic effect and the possible mechanism of total polysaccharides of Yupingfeng (YPF-P) on bleomycin (BLM)-induced pulmonary fibrosis in rats.
Pulmonary fibrosis was induced in Sprague-Dawley rats by BLM (5 mg/kg), killed 14 and 28 days after BLM administration by abdominal aorta exsanguination and removed the lungs. Lung coefficient was counted at the same time. Besides, H&E and Masson's trichrome staining for histopathological changes of lung tissues were observed. Additionally, western blotting and immunohistochemical staining techniques were used to detect expression of transforming growth factor-β1 (TGF-β1), type I collagen (Col-I) and α-smooth muscle actin (α-SMA). Finally, the levels of Col-I and hydroxyproline (HYP) in lung tissues were also utilized.
YPF-P alleviated the increase of lung coefficient induced by BLM instillation in pulmonary fibrosis rat, pathologic changes and collagen distribution were obviously ameliorated, while the increase of α-SMA-positive cells and TGF-β1 expression was prevented after YPF-P treatment. Moreover, the contents of HYP and Col-I were decreased in YPF-P group.
YPF-P had antifibrotic effect in experiment, which may reduce the synthesis and promote the deposition of Col-I via suppressing the increase of TGF-β1-mediated activation of myofibroblasts.
The aim of this study was to prepare microcapsules for the oral delivery of realgar nanoparticles (RN) that are also capable of improving its stability.
RN and RN-based microcapsules (RNM) were prepared using ball milling and solvent evaporation techniques, respectively. Properties such as particle size, ζ-potential (ZP), morphology and X-ray diffractometer (XRD) were investigated. In addition, drug release, bioavailability and antitumour studies were also performed.
The nanoparticles appeared round or elliptical in shape with a mean size of 85.4 ± 3.5 nm and a ZP of −34.3 ± 1.7 mV. The obtained RNM appeared spherical and not aggregated with a relatively narrow size distribution. XRD analysis revealed that ball milling technique did not change the crystallinity of the realgar powder. RN and RNM exhibited considerable higher release of As2S2, bioavailability and antitumour efficacies compared with crude realgar. Furthermore, RNM could protect RN directly exposed to the air and light, and therefore increased the stability of the RN.
The developed RNM demonstrated a greater potential as a delivery system for realgar.
The aim of this study was to investigate the in-vitro permeation enhancement effects of the gel and whole-leaf materials of Aloe vera, Aloe marlothii and Aloe ferox using ketoprofen as a marker compound.
The permeation studies were conducted across excised female abdominal skin in Franz diffusion cells, and the delivery of ketoprofen into the stratum corneum-epidermis and epidermis-dermis layers of the skin was investigated using a tape-stripping technique.
A. vera gel showed the highest permeation-enhancing effect on ketoprofen (enhancement ratio or ER = 2.551) when compared with the control group, followed by A. marlothii gel (ER = 1.590) and A. ferox whole-leaf material (ER = 1.520). Non-linear curve fitting calculations indicated that the drug permeation-enhancing effect of A. vera gel can be attributed to an increased partitioning of the drug into the skin, while A. ferox whole leaf modified the diffusion characteristics of the skin for ketoprofen. The tape stripping results indicated that A. marlothii whole leaf delivered the highest concentration of the ketoprofen into the different skin layers.
Of the selected aloe species investigated, A. vera gel material showed the highest potential as transdermal drug penetration enhancer across human skin.
The efficiency of the Astree e-tongue and Taste Sensing system TS5000Z for the evaluation of the taste masking effect of hot melt extruded formulations was investigated in this study.
Hot melt extrusion (HME) processing was optimized using Randcastle single screw extruder (USA) to manufacture extrudates with desirable characteristics. Cationic model drug propranolol HCl (PRP) was processed with the anionic polymers – Eudragit L100 (L100) and Eudragit L100-55 (Acryl-EZE). Solid state of the drug in polymer matrices was evaluated by scanning electron microscopy (SEM), differential scanning calorimetry, particle size analysis, Fourier transform infrared (FTIR) and Nuclear magnetic resonance (NMR) analysis. In-vitro taste masking efficiency of the two polymers was performed by using two different e-tongues (Astree e-tongue and TS5000Z). The results obtained from both e-tongues were further compared and contrast to find out the sensor outputs in all formulations.
Solid state analysis of the extruded formulations revealed the presence of amorphous PRP. Both e-tongues were able to detect the taste masking variations of the extrudates and were in good agreement with the in-vivo results obtained from a panel of six healthy human volunteers (R2 > 0.84). However, each e-tongue sensor demonstrated different sensitivity, suggesting a careful consideration of the experimental findings during melt extrusion, is necessary for the development of taste-masked formulations. Furthermore, FTIR spectroscopy and NMR studies revealed possible drug polymer intermolecular interactions as the mechanism of successful taste masking.
HME can effectively be used to manufacture taste-masked extruded formulations, while both e-tongues demonstrated satisfactory taste analysis for the development of taste-masked formulations.
Phosphodiesterase (PDE)-5 inhibitors are useful as vasodilators for the treatment of pulmonary arterial hypertension. We aimed to study curcumin analogues for PDE5 inhibitory activity and vasorelaxation of rat pulmonary arteries.
Three natural curcuminoids (1–3) and six synthetic analogues (4–9) were tested for PDE5 and PDE6 inhibitory activities using enzymatic radioassay. Their vasorelaxation was measured using freshly isolated segments of rat pulmonary artery and aorta.
Curcuminoids (1–3) mildly inhibited PDE5 (half maximal inhibitory concentration (IC50) = 18 µm): the metamethoxyl of curcumin was important for PDE5 inhibition. But hydroxyl rearrangements, removing both methoxyls and one ketomethylene, yielded the potent 7 and 9 (IC50 = 4 µm) (compared with sildenafil, IC50 = 0.03 µm). Only 1, 3 and 4 were PDE5 selective over PDE6. Triazole-carboxylic addition provided water-solubility while preserving potency. All analogues possessed concentration-dependent vasorelaxant activity on pulmonary arteries (40% of maximal effective concentration (EC40) = 29–90 µm, maximum response = 60–90% at 300 µm), while compounds (1–8) were weakly acting in aorta (maximum response <40%). Only demethoxycurcumin (2) and analogues 5, 8, 9 had endothelium-dependent actions. Sildenafil was highly potent (EC40 = 0.04 µm) and highly endothelium dependent in pulmonary artery but weak on intact aorta (EC40 = 1.8 µm). Activity profiles suggest actions through additional cell pathways for promoting vasorelaxation.
Curcumin analogues are potential leads for developing efficacious and selective PDE5 inhibitors and other pathologies of pulmonary hypertension.
The aim of this work was to evaluate a continuous, small-scale extrusion process with a particular focus on powder and liquid-feeding systems, because it is likely that uniformity issues are related to small-scale production.
The study is divided into three parts. The first part investigates the uniformity and accuracy of the powder and the liquid feeders. In the second part, a solid polymer and low amounts of liquid plasticizer were combined in hot-melt extrusion. The third part deals with wet extrusion-spheronization using water as the granulation liquid.
The powder and the liquid feed rate were identified as crucial parameters in small-scale extrusion. With respect to powder feeding, the cohesiveness of the powder and electrostatic charging are the limitations, while liquid feeding is challenging based on particularly low feed rates. The hot-melt extrusion was performed using a powder feed rate of 2 g/min. When small quantities of plasticizer were applied to the hot melt extrusions (from 2.5% to 15% w/w), homogenous plasticizer distribution was found. In wet extrusion, larger quantities of water were used and the extrudates were investigated with respect to their spheronization behaviour. Spherical pellets were obtained at certain water contents.
These findings demonstrated that the extruder is a useful tool to screen formulations and perform feasibility studies on a small scale in the early stages of product development.
This study was undertaken to study the effects of melatonin on metabolic and reproductive aspects of polycystic ovary syndrome (PCOS) in rats.
PCOS was induced by daily subcutaneous administration of testosterone (20 mg/kg) to 21-day-old female rats for 35 days. Rats were given metformin (500 mg/kg), melatonin (1 mg/kg) or melatonin (2 mg/kg) along with testosterone. One group served as vehicle control. On the 36th day, the animals were euthanised, and anthropometrical, biochemical (glucose, insulin, lipids, testosterone, C reactive protein (CRP)), oral glucose tolerance test, and histopathological evaluation of ovaries, uterus and intraabdominal fat (IAF), were carried out. Daily colpocytological examination was carried out from 14th day of study until termination.
Both the doses of melatonin significantly reduced body weight, body mass index, IAF, insulin and CRP. A favourable lipid profile, normal glucose tolerance and a decrease in the percentage of estrus smears were observed. Histopathological examination of ovary, uterus and IAF revealed a decrease in the number of cystic follicles, decrease in neoplastic endometrial glands, and decrease in adipocyte hypertrophy, respectively. The effects observed with melatonin were comparable to that with metformin.
The study provides evidence of the potential beneficial effects of melatonin in PCOS.
To provide an overview of a novel anti-hepatitis C agent, sofosbuvir.
Sofosbuvir is a novel nucleotide analogue inhibitor of hepatitis C virus (HCV) NS5B polymerase that has recently been approved by the Federal Drug Administration for the treatment of HCV genotypes 1, 2, 3 or 4 in a variety of patients. The emergence of such a novel treatment provides benefit to previously untreated genotypes and patient populations, with little chance of promoting significant viral resistance. Excellent sustained virologic response rates 12 weeks after the end of treatment (SVR12) were seen in phase II and III clinical trials when sofosbuvir was used with ribavirin. Even more promising are the results from phase II and III clinical trials that evaluated sofosbuvir in combination with other oral direct acting antivirals (DAAs). Data with sofosbuvir in the hepatitis C virus (HCV)/HIV coinfected and in the pre- and post-transplantation populations are still emerging. The drug was very well tolerated in clinical studies, with the most common adverse events of headache, nausea and fatigue.
Overall, sofosbuvir presents a new and effective treatment option for HCV-infected patients.
This study aimed to investigate the stimulative and pharmacological effects of ginsenoside Rh1 (hereinafter referred to as: Rh1) on differentiation and mineralization of osteoblast and its possible mechanism of action on the expression of bone morphogenetic protein 2 (BMP-2)/Runt-related gene 2 (Runx2) signalling pathways using mouse preosteoblastic MC3T3-E1 cell line as in-vitro model.
An in-vitro stimulative activity of Rh1 was assessed by analyzing alkaline phosphatase activity (ALP), type-I collagen (Coll-I) synthesis, mineralization and glutathione content. Its antioxidant activity was measured by evaluating the reactive oxygen species (ROS) production in the presence of antimycin A (AMA), one of the mitochondrial dysfunction factors. The level of BMP-2/Runx2 signal-regulated osteoblast-specific proteins such as osteocalcin (OCN), Coll-I and ALP were detected using Western blot analysis.
Rh1 was capable to stimulate cell growth, ALP activity, Coll-I synthesis, mineralization and glutathione content in the MC3T3-E1 cells. BMP-2 and Runx2 expression were also increased by Rh1 concentration dependently. Additionally, Rh1 also showed inhibitory action on the level of ROS production enhanced by AMA in MC3T3-E1 cells. Rh1 could increase the expression level of BMP-2/Runx2 signal-regulated osteogenic markers such as ALP, Coll-I and OCN.
Rh1, a protopanaxatriol type's active ingredients of Panax ginseng Meyer, possesses osteoblast differentiation, osteogenic stimulatory and anti-oxidative activity.
To test the toxicity and antitumoral activity of the compound N-oleyl-daunorubicin (oDNR) with a cholesterol-rich nanoemulsion (LDE) formulation.
LDE-oDNR was prepared by high-pressure homogenisation of lipid mixtures. B16F10 melanoma cells and NIH/3T3 fibroblasts were used for cytotoxicity tests. The maximum tolerated dose (MTD) of both commercial and LDE-oDNR was determined in mice, and melanoma-bearing mice were used for the antitumoral activity tests.
CC50 for LDE-oDNR and DNR in melanoma cells were 200 μm and 15 μm, respectively, but LDE-oDNR was less toxic against fibroblasts than DNR. MTD for LDE-oDNR was 65-fold higher than commercial DNR. In tumour-bearing mice, LDE-oDNR (7.5 μmol/kg) reduced tumour growth by 59 ± 2%, whereas the reduction by DNR was only 23 ± 2%. LDE-oDNR increased survival rates (P < 0.05), which was not achieved by DNR treatment. The number of mice with metastasis was only 30% in LDE-oDNR-treated mice, compared with 82% under DNR treatment. By flow cytometry, there were 9% viable cells in tumours of animals treated with LDE-oDNR compared with 27% in DNR-treated animals. Less haematological toxicity was observed in LDE-oDNR-treated mice.
Compared with DNR, LDE-oDNR improved tumour growth inhibition and survival rates with pronouncedly less toxicity, and thus may become a new tool for cancer treatment.
The objective of the review is to provide an overview of the nomenclature used in the solid-state continuum and relate these to the development of drug substances and drug products.
The importance of a rational approach to solid-state form selection, including integrated decision making (ensuring equal weight is given to the needs of the drug substance and the drug product), is vital for the effective development of a drug candidate. For example, how do secondary processing considerations influence the selection of drug substance solid-state form and resulting formulation, and how can drug substance solid-state form be used to optimise secondary processing? Further, the potential use of 'crystal' engineering to optimise stability, purity and optical resolutions, and the linked regulatory requirements, will be discussed.
The nomenclature used in the solid-state continuum, which contains a large number of different crystalline and non-crystalline forms, for example, amorphous systems, was reviewed. Further, the significant role of the drug substance within the solid oral dose form from a physicochemical perspective was covered.
The purpose of this study was to investigate the central activity of the two new imidazo[2,1-f]purine-2,4-dione derivatives behaved as presynaptic 5HT1A receptor agonists and postsynaptic 5HT1A, 5HT2A and D2 receptors antagonists. The compounds were examined using animal tests towards antipsychotic, antidepressant- and anxiolytic-like properties and then compared with effects evoked by an atypical antipsychotic drug ziprasidone.
D-amphetamine-induced hyperactivity test was used to determine antipsychotic-like activity of compounds 7 and 9. The forced swim test (FST) and the four-plate test were conducted to investigate antidepressant- and antianxiety-like activity, respectively, of studied agents. The investigated compounds 7, 9 and ziprasidone were administered intraperitoneally 60 min before the tests. Diazepam and imipramine were used as standard anxiolytic and antidepressant drugs, respectively.
The obtained results demonstrate that new synthesized compound 9 evokes antipsychotic-like activity alike ziprasidone and, in contrary to the antipsychotic drug, shows antidepressant- and anxiolytic-like properties in behavioural tests in mice.
The present preclinical results indicate that one of the two investigated imidazo[2,1-f]purine-2,4-dione derivatives, compound 9, with methyl group at 7 position of imidazo[2,1-f]purine-2,4-dione fragment and the ortho-OCH3 substituent in the aryl moiety, acts as an antipsychotic drug with additional antidepressant and anxiolytic properties.
In this study, the antioxidant action of ((4-tert-butylcyclohexylidene) methyl) (4-methoxystyryl) sulfide, a novel unsymmetrical divinyl sulfide, against oxidative damage induced by sodium nitroprusside (SNP) in brains of mice was investigated.
Mice received SNP (0.335 μmol/site, intracerebroventricular) 30 min after administration of sulfide (10 mg/kg, intragastrically). After 1 h, animals were sacrificed and the brains were removed to biochemistry analysis. Thiobarbituric acid reactive species (TBARS), protein carbonyl (PC) and non-protein thiol (NPSH) levels, as well as catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione S-transferase (GST) activities were determined.
SNP increased TBARS and PC levels, CAT, GPx, GR and GST activities and reduced NPSH levels. Administration of the sulfide attenuated the changes produced by SNP and increased per se GPx activity in brains of mice. Toxicological parameters revealed that this compound did not cause acute renal or hepatic damage.
In conclusion, ((4-tert-butylcyclohexylidene) methyl) (4-methoxystyryl) sulfide protected against oxidative damage caused by SNP in mouse brain. GPx activity is involved, at least in part, in the cerebral antioxidant action of this compound.
Investigate the involvement of monoaminergic and glutamatergic systems on the antinociceptive and ataxic effects of uliginosin B, which we have already demonstrated to be a promising molecular scaffold to develop new analgesic drugs.
Uliginosin B was obtained from hexane extract of aerial parts of Hypericum polyanthemum by chromatographic methods. Uliginosin B antinociceptive and motor coordination effects were evaluated in mice by using hot-plate (15 and 90 mg/kg, i.p.) and rotarod (90 mg/kg, i.p.) tests, respectively. The mechanism of action was investigated through pretreatments with prazosin 1 mg/kg intraperitoneal (α1 receptor antagonist), yohimbine 5 mg/kg intraperitoneal (α2 receptor antagonist), pCPA 300 mg/kg intraperitoneal (serotonin synthesis inhibitor) and MK-801 0.25 mg/kg intraperitoneal (N-methyl-D-aspartic acid receptor antagonist).
The antinociceptive effect of uliginosin B (15 and 90 mg/kg, i.p.) was reduced significantly by pCPA and MK-801. Prazosin and yohimbine improved the antinociceptive effect of the highest dose (90 mg/kg, i.p.) of uliginosin B only. The ataxic effect of uliginosin B (90 mg/kg, i.p.) was completely prevented by pretreatment with pCPA or MK-801, but it was unaffected by pretreatment with prazosin or yohimbine.
These data confirm the contribution of monoaminergic neurotransmission as well as provide the first evidence of glutamatergic neurotransmission contribution to the uliginosin B effects.
Tegafur (FT), a pro-drug of 5-fluorouracil (5-FU), is a racemate consisting of two enantiomers, R and S-FT. The aim of this study was to clarify interspecies variation in the enantioselective metabolism of FT.
Plasma concentrations of FT enantiomers were determined in rats, dogs and monkeys following intravenous and oral dosing of the racemate (5 mg/kg). In addition, the enzymatic conversion of FT enantiomers to 5-FU was assayed using hepatic preparations.
Metabolic clearance of R-FT was higher than that of S-FT in rats and monkeys, but S-FT was the preferential substrate for dogs. An inhibition study revealed that cytochrome P450 is primarily responsible for the enantioselective metabolism of FT in rats and dogs. In contrast, in monkeys, thymidine phosphorylase was a determinant of the enantioselectivity in FT metabolism. Although oral bioavailability was not enantioselective, in-vitro and in-vivo kinetic studies suggested that the enantioselectivity in the hepatic intrinsic clearance of FT directly influences the body clearance in all animal species examined.
The interspecies variations were observed in the enantioselective pharmacokinetics of FT, and the in-vivo enantioselectivity could be extrapolated from the in-vitro metabolic activities.
We investigated the mechanisms underlying the relaxant effect of adrenomedullin (AM) in the rat carotid artery and verified the expression of AM system components in this tissue.
The carotid artery was isolated from male Wistar rats and immunohistochemical, Western immunoblotting, real-time polymerase chain reaction and functional assays were conducted.
Protein and mRNA expression of AM, calcitonin receptor-like receptor (CRLR) and receptor activity-modifying proteins (RAMP)1, 2, 3 were detected in carotid segments from male Wistar rats. Immunohistochemical assays showed that AM and CRLR receptors are expressed in the endothelium and smooth muscle cells. Functional assays showed that AM concentration dependently relaxed carotid rings with intact endothelium. Endothelial removal reduced, but not abolished, the relaxation induced by AM. AM22–52 (selective antagonist for AM receptors) and calcitonin gene-related peptide (CGRP)8–37 (selective CGRP receptor antagonist) reduced AM-induced relaxation in endothelium-intact rings. Pre-incubation of endothelium-intact rings with N-nitro-L-arginine methyl ester, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one or Rp-8-Bromo-?-phenyl-1,N2-ethenoguanosine 3′,5′cyclic monophosphorothioate reduced AM-induced relaxation. Inhibition of cyclooxygenase-1 and protein kinase A (PKA) reduced AM-induced relaxation. The relaxation induced by AM was attenuated by the K+ channel blockers apamin and glibenclamide. AM increased nitrate levels and 6-keto-prostaglandin F1α (stable product of prostacyclin) in the rat carotid. In endothelium-denuded rings, AM22–52, glibenclamide and PKA inhibition by H89 reduced AM-induced relaxation.
The novelty of this work is that it first demonstrated functionally that AM-induced relaxation is mediated by AM and CGRP receptors located on the endothelium and AM receptors located on smooth muscle of rat carotid arteries. AM-induced relaxation involves the nitric oxide-cGMP pathway, a vasodilator prostanoid, the opening of K+ channels and the activation of PKA.
The aim of this study was to compare lung deposition and assess the bioequivalence of two formulations containing budesonide and formoterol and being delivered via Elpenhaler and Turbuhaler, respectively. A pharmacokinetic (PK) study was conducted.
An open, randomized, two-sequence, two-period, crossover, single-dose study in 100 asthmatic patients under fasting conditions was performed. Wash out period was 6 days. Equivalence in lung deposition was assessed after a single inhalation of each treatment with concomitant oral administration of activated charcoal (40 g) to prevent gastrointestinal absorption of the drugs. Several PK parameters were estimated, the area under the drug concentration in plasma versus time curve (AUC0−t) and the maximum drug concentration in plasma (Cmax) being the primary response variables. Equivalent lung deposition was concluded if the 90% confidence interval (CI) for the Elpenhaler/Turbuhaler geometric mean ratio of AUC0−t and Cmax, for both drug substances fell within the regulatory limits (0.80–1.25).
Acceptance criteria were met. Equivalent lung deposition can be concluded. No statistically significant differences between treatments in the incidence of adverse events were found.
The formulations are bioequivalent regarding both rate and extent of absorption. The treatments were also well tolerated by the participating subjects.
To scientifically validate the traditional substitution of roots of highly expensive Aconitum heterophyllum (AH) with rhizomes of Cryptocoryne spiralis (CS) in the treatment of diarrhoea.
Different fractions from root/rhizome extract of both the plant were subjected to faecal excretion rate and castor oil-induced diarrhoea models. Further, bioactive fractions from both plants, i.e. chloroform (CAH) from AH at 50 mg/kg p.o. and ethyl acetate (EACS) from CS at 100 mg/kg p.o., were examined for small intestinal transit, intestinal fluid accumulation and PGE2-induced enteropooling models in rats. Biochemical estimations and Na+ and K+ concentration in intestinal fluid were also determined along with antibacterial studies. Phytochemical standardisation of AH and CS was performed by quantifying aconitine for the former and stigmasterol for the latter using HPLC.
CAH and EACS illustrated a significant reduction in faecal output rate and demonstrated a protection of 63.068% at CAH 50 and 59.090% at EACS 100 mg/kg p.o. in castor oil-induced diarrhoea model. The fractions also persuaded promising effects in all the other models, restored alterations in biochemical parameters and showed potential antibacterial activity.
The antidiarrhoeal potential of AH and CS may be attributed to an antimotility and antisecretory type of effect.
1,8-Cineole, the main monoterpene in many essential oils, has been used as an ingredient in flavourings and medicine. 1,8-Cineole has been shown to possess pharmacological properties, including anti-oxidative, anti-inflammatory and anti-nociceptive actions. However, to date, no studies have examined the potential of 1,8-cineole to protect against cerebral ischaemic injury.
In this study, we investigated the neuroprotective effects of 1,8-cineole against cortical neuronal/glial cell injury caused by oxygen-glucose deprivation/reoxygenation (OGD/R) in an in-vitro model of ischaemia.
1,8-Cineole significantly attenuated OGD/R-induced cortical cell injury, as well as reduced n-methyl-d-aspartate (NMDA)-induced cell injury. However, it did not inhibit NMDA-induced cytosolic calcium overload. Nevertheless, 1,8-cineole significantly reduced the OGD/R- and NMDA-induced overproduction of reactive oxygen species (ROS). These results indicate that 1,8-cineole exerts neuroprotection through its anti-oxidative rather than its anti-excitotoxic, properties. The decrease in OGD/R-induced intracellular superoxide in 1,8-cineole-treated cortical cells was associated with the upregulation of superoxide dismutase activity. Moreover, 1,8-cineole showed direct ROS scavenging activity in an assay of oxygen radical absorbance capacity.
Collectively, these results suggest 1,8-cineole as a potentially effective neuroprotective and anti-oxidative candidate for the treatment of patients with ischaemic stroke.
This review focuses on real-time analytics of drug dissolution and precipitation testing on a comparatively small scale.
Miniaturisation of test equipment is an important trend in pharmaceutics, and several small-scale experiments have been reported for drug dissolution and precipitation testing. Such tests typically employ analytics in real-time. Fibre optic ultraviolet (UV) analytics has become a well-established method in this field. Novel imaging techniques are emerging that use visible or UV light; also promising is Fourier transform infrared imaging based on attenuated total reflection. More information than just a rate constant is obtained from these methods. The early phase of a dissolution process can be assessed and drug precipitation may eventually be observed. Some real-time techniques are particularly well suited to studying drug precipitation during formulation dispersion; for example, turbidity, focused beam reflectance measurement and Raman spectroscopy.
Small-scale dissolution tests equipped with real-time analytics have become important to screen drug candidates as well as to study prototype formulations in early development. Future approaches are likely to combine different analytical techniques including imaging. Miniaturisation started with mini-vessels or small vials and future assays of dissolution research will probably more often reach the level of parallel well plates and microfluidic channels.
We aimed to highlight the utility of novel dissolving microneedle (MN)-based delivery systems for enhanced transdermal protein delivery. Vaccination remains the most accepted and effective approach in offering protection from infectious diseases. In recent years, much interest has focused on the possibility of using minimally invasive MN technologies to replace conventional hypodermic vaccine injections.
The focus of this study was exploitation of dissolving MN array devices fabricated from 20% w/w poly(methyl vinyl ether/maleic acid) using a micromoulding technique, for the facilitated delivery of a model antigen, ovalbumin (OVA).
A series of in-vitro and in-vivo experiments were designed to demonstrate that MN arrays loaded with OVA penetrated the stratum corneum and delivered their payload systemically. The latter was evidenced by the activation of both humoral and cellular inflammatory responses in mice, indicated by the production of immunoglobulins (IgG, IgG1, IgG2a) and inflammatory cytokines, specifically interferon-gamma and interleukin-4. Importantly, the structural integrity of the OVA following incorporation into the MN arrays was maintained.
While enhanced manufacturing strategies are required to improve delivery efficiency and reduce waste, dissolving MN are a promising candidate for 'reduced-risk' vaccination and protein delivery strategies.
Objectives Mutations in the p53 tumor suppressor gene are one among the most common genetic abnormalities to be described in breast cancer. However, there are a few recant reports on non-viral vector-mediated p53 gene delivery in breast cancer.
Methods A new formulation of luminescent silica nanoparticles (LSNs) for gene delivery was produced by the two-step method with slight modification.
Key findings The pp53 plasmid constructs (p53-EGFP)/LSNs complexes were transfected into human breast cancer cell (MCF-7) and transfection efficiency was determined by FACS analysis. The gene expression was determined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis respectively. Further the growth inhibition through induced apoptosis with pp53-EGFP/LSNs complex were assessed by trypan blue exclusion assay and annexin V staining, respectively. Interestingly the in vivo biodistribution of plasmid DNA study revealed the occurrence was investigated by PCR and RT-PCR. The transfection efficiency of LSNs showed the highest transfection efficiency among the LSN formulation was higher than that of commercially available Lipofectin®. The LSNs-mediated transfection of the p53 gene resulted in efficient high level of wild-type p53 mRNA and protein expression levels in MCF-7 cells. Selected tissues were analyzed for any potential toxicity by histological analysis the efficient reestablishment of wild-type p53 function in breast cancer cells restored the p53 dependent apoptotic pathway.
Conclusions Taken together, our results reveal that cationic LSN-mediated p53 gene delivery may have potential application as a non-viral vector-mediated breast cancer gene therapy due to its effective induction of apoptosis and tumor growth inhibition.
The majority of research performed on cellular stress and apoptosis focuses on mitochondrial dysfunction; however, the importance of the endoplasmic reticulum dysfunction and the link to metabolic diseases has gained a substantial interest. This review focuses on the potential of terpenoids to influence endoplasmic reticulum stress and the possible role terpenoids play as the treatment of metabolic diseases.
Metabolic diseases develop as a result of a cascade of cellular pathways. In most cases, cells are able to compensate for the disruption of the cellular homeostasis although the initiation of response pathways; however, chronic stress initiates apoptotic pathways. This reviewed (1) showed the importance of phytoterpenoids to influence endoplasmic reticulum (ER) stress and homeostasis, (2) showed how regulating ER stress affect the cell survival and death, and (3) highlighted some examples of how the progression of metabolic diseases can be influenced by ER.
Due to the substantial number of terpenoids that have been identified in literature, this review gave examples of 21 terpenoids that have been documented to have an effect on the different proteins associated with ER stress, how these plant terpenoids influence ER dysfunction and metabolic diseases such as diabetes, cancer, liver, and neurological diseases and parasitic infections.
The present study was oriented towards the development of pulmonary vaccine for Hepatitis B using probiotic biomass as an adjuvant.
The antigen was spray dried in presence of heat treated, formalin treated and live probiotic biomass.
The results indicated that the biomass itself without any additional cryoprotectant is capable of protecting the structural integrity of the antigen. We were able to retain more than 80% of the antigenicity. The scanning electron microscopic images indicated that the formulation bearing live probiotic biomass have spherical size, while the formulations with heat and formalin treated biomass shows irregular shaped particles. The developed formulations were further evaluated for in-vivo immune response. Immunoglobulin G (IgG) titre results were found to be comparable with marketed (aluminium adsorbed) formulations while significantly higher secretory immunoglobulin A titre showed better mucosal immune response than marketed formulation.
Therefore, the probiotic biomass can be utilized as a potential cryoprotectant as well as a potent immunomodulator.
Naftopidil (NAF) is a chiral compound with two enantiomers (R(+)-NAF and S(−)-NAF) and is used as a racemic mixture in clinical practice. This study aims to investigate the metabolism of NAF enantiomers in pooled human liver microsomes (HLMs) and cytochrome P450 isozymes (CYPs) involved in their metabolism.
Metabolism studies were conducted in vitro using HLMs. Specific chemical inhibitors and recombinant human CYPs were used to confirm that the CYPs contributed to the metabolism of NAF enantiomers.
Three metabolites were found and characterized in the HLMs incubations from R(+)-NAF and S(−)-NAF, respectively. The major metabolic pathways of R(+)-NAF and S(−)-NAF were demethylation and hydroxylation. CYP2C9 and CYP2C19 inhibitors strongly inhibited R(+)-NAF metabolism, and CYP1A2, CYP2C8, CYP2D6 and CYP3A4/5 inhibitors moderately inhibited R(+)-NAF metabolism. CYP2C9 inhibitors strongly inhibited S(−)-NAF metabolism, and CYP2C8, CYP2C19 and CYP3A4/5 inhibitors moderately inhibited S(−)-NAF metabolism. Consistent with the results of chemical inhibitors experiments, recombinant human CYP2C9 and CYP2C19 contributed greatly to R(+)-NAF metabolism, and CYP2C9 contributed greatly to S(−)-NAF metabolism.
Both R(+)-NAF and S(−)-NAF are metabolized to three metabolites in HLMs. CYP2C9 plays the most important role in the demethylation and hydroxylation of both NAF enantiomers, CYP2C19 is another major CYP isoform that is involved in R(+)-NAF metabolism.
Patients undergoing hip or knee replacement therapy are routinely pretreated with antibiotics before surgery. It is controversial in which antibiotic is the treatment of choice for this purpose. One possibility is the cephalosporin ceftriaxone. Here, we wanted to know if effective tissue concentrations are reached.
We studied plasma and bone kinetics of ceftriaxone in orthopaedic patients (n = 22) treated with ceftriaxone (2 g) immediately prior operation. Plasma samples were withdrawn before and at three time points after ceftriaxone infusion. After bone replacement, extracts from cancellous bone or cortical bone were obtained, and ceftriaxone was quantified using column chromatography.
The plasma kinetics of ceftriaxone and distribution into bone were analysed using a population approach (ADAPT 5). The population mean of the area under the curve (AUC) was 140 mg h/l. A cancellous bone to plasma concentration ratio of 1.12 ± 1.29 was achieved 5 h after start of infusion. The half-life of uptake into the cortical bone was less (8.4 h) than into cancellous bone (12.1 h, P < 0.05).
Under these experimental conditions, concentrations of ceftriaxone in cancellous and cortical bone should be adequate to protect the patients against usual ceftriaxone-sensitive nosocomial infections and are substantially lower than the plasma concentrations.
To assess the effect of an opioid administered topically onto a standardized skin wound in patients without significant comorbidity. Findings to date are contradictory, often obtained from multimorbid patients with wounds lacking uniformity.
Forty-four patients undergoing surgery for skin grafting were randomly assigned to receive morphine (0.25, 0.75 or 1.25 mg/100 cm2 wound size) in hydroxyethylcellulose gel or placebo applied onto the excised split-thickness donor wound at the end of surgery. Pain, supplementary systemic opioids and adverse effects were assessed during the first 24 h after application. Healing was examined when the dressings were removed.
Morphine doses ranged from 0.25 to 5.4 mg (mean 1.93 ± standard deviation 1.34 mg). No differences in pain scores or use of supplementary analgesics were found between the groups. Serious adverse effects did not occur; healing was not impaired. Large intragroup variability was observed for pain scores, wound sizes and supplementary analgesia.
Patients treated with topically applied morphine gel onto standardized skin wounds did not report lower pain scores compared with placebo-treated patients. Larger groups would be required to arrive at definitive conclusions. The split-thickness skin graft model can be used for future research.
Anticancer indirubins are poorly soluble in water. Here, digestion of four self-emulsifying drug delivery systems (SEDDS) containing E804 (indirubin-3′-oxime 2,3-dihydroxypropyl ether) was compared by dynamic lipolysis and bioavailability studies. Used lipids were either medium-chain or long-chain glycerides.
SEDDS E804 were developed. In-vitro lipolysis was carried out at pH 6.5 (37°C) by adding pancreatic lipase (800 U/ml) and controlling by CaCl2 and NaOH addition. E804 content was quantified in the aqueous micellar phase and precipitate using HPLC. Oral bioavailability was determined in rats. Plasma drug content was determined by liquid chromatography (LC)–mass spectrometry.
All formulations reserved E804 in the aqueous micellar phase up to 60 min. Precipitation proceeded towards the end of lipolysis up to 45%. Lowest level of precipitation (21%) occurred with long-chain lipids (LC-SEDDS). However, lipolysis was not really discriminative between formulations as the drug mainly stayed in solution. Oral administration of formulations resulted in similar bioavailability of E804 with no significantly different area under the concentration curve. Only medium-chain self-nanoemulsifying drug delivery systems revealed shorter Tmax compared with the other formulations.
E804 had a similar performance in four lipid/surfactant systems. All formulations increased the bioavailability of E804 with no significant difference.
We investigated the safety, tolerability, pharmacokinetics and pharmacodynamics of evacetrapib.
Healthy volunteers received multiple daily doses of evacetrapib (10–600 mg) administered for up to 15 days in a placebo-controlled study.
Mean peak plasma concentrations of evacetrapib occurred at 4–6 h and terminal half-life ranged 24–44 h. Steady state was achieved at approximately 10 days; all subjects had undetectable levels of evacetrapib 3 weeks after their last dose. The trough inhibition of cholesteryl ester transfer protein (CETP) activity was 65 and 84% at 100 and 300 mg, respectively. At the highest dose (600 mg), evacetrapib significantly inhibited CETP activity (91%), increased HDL-C (87%) and apo AI (42%), and decreased LDL-C (29%) and apo B (26%) relative to placebo. For the highest dose tested, levels of evacetrapib, CETP activity, CETP mass, HDL-C and LDL-C returned to levels at or near baseline after a 2-week washout period. Evacetrapib at the highest dose tested did not produce any significant effect on 24-h ambulatory systolic or diastolic blood pressure.
Multiple doses of evacetrapib potently inhibited CETP activity, leading to substantial elevations in HDL-C and lowering of LDL-C. Evacetrapib was devoid of clinically relevant effects on blood pressure and mineralocorticoid levels.
Pigment epithelium-derived factor (PEDF) has proven anti-osteosarcoma activity. However, the mechanism(s) underpinning its ability to reduce primary bone tumour (osteosarcoma) metastasis is unknown.
Adult and fetal murine bone were immunostained for PEDF, collagen I (major protein in bone) and its processing proteins, heat shock protein 47 (HSP47, a chaperone protein for collagen I), membrane type I matrix metalloproteinase (MT1-MMP, a collagenase), and matrix metalloproteinase 2 (MMP-2, which is activated by MT1-MMP). Immunoblotting and immunocytochemistry were used to observe levels of the above biomarkers when human osteosarcoma cells were treated with PEDF.
Immunohistochemical staining in adult and fetal bone mirrors collagen I. PEDF localised to ridges of trabecular bone in tibial cortex and to megakaryocytes within bone marrow. Second, we observed that PEDF upregulates collagen I, HSP47 and MT1-MMP, while downregulating MMP-2 in osteosarcoma cells in vitro.
PEDF is a promising antagonist to osteosarcoma cell metastasis via downregulation of MMP-2, and can induce tumour cells to further adopt differentiative properties, thereby possibly reducing their aggressive growth in vitro and in vivo.
The aim of this study was to investigate the in-vitro cytotoxic activity of new platinum(II) complexes on the human HeLa (ER−), MCF-7 (ER+) and MDA-MB 231 (ER−) cell lines. Furthermore, we investigated plasmid DNA interactions and inhibition of BamHI and HindIII restriction enzyme activity of the complex 1–4,7.
Platinum(II) complexes were synthesised from precursor complexes of [PtL2Cl2] and [PtL2I2]. Their cytotoxic activity was tested by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Their plasmid DNA interactions and restriction enzyme activities were also investigated using the agarose gel electrophoresis method.
The growth inhibitory effect results showed that the cytotoxicity of complex 2 was found to be the most active complex among the synthesised complexes.
The MTT results showed that complex 2 was found to be cytotoxic equal to cisplatin and higher than carboplatin against the MCF-7 and MDA-MB-231 cell lines. Furthermore, the estrogen or progesterone co-treatment slightly increased the cytotoxicity of complex 2, the cisplatin and carboplatin compared with the complex 2 tested alone in 50 μm concentration. According to plasmid DNA interaction and the restriction studies, complexes 1–4,7 modified the tertiary structure of pBR322 plasmid DNA, and complexes 2–4 prevented enzyme digestion at high concentrations.
Extracts of the tubers of Harpagophytum procumbens (devil's claw, DC) inhibit different proinflammatory mediators important in the pathophysiology of osteoarthritis. Many plant-derived preparations interfere with cytochrome P450 liver enzymes, which influence their different biological activities. Therefore, the present study was designed to investigate the influence of an external metabolic activation of a DC extract on the cytotoxicity and the release of proinflammatory cytokines.
A screening experiment with a panel of 12 inflammatory cytokines identified three as suitable for the study: tumour necrosis factor-α (TNF-α), interleukin (IL) IL-6 and IL-8. They were determined using enzyme-linked immunosorbent assays in lipopolysaccharide (LPS)-stimulated monocytic THP-1 cells, which were treated with rat liver S9 mix metabolically activated DC extract (DCm). For the cytotoxity experiments, a WST-1 assay was used.
DC dose-dependently suppressed the release of TNF-α, IL-6 and IL-8 in LPS-stimulated monocytic THP-1 cells at non-cytotoxic concentrations (50–250 μg/ml). The metabolic activation of the DC extract by S9 mix did not alternate its cytotoxicity and did not diminish its inhibitory effect. This effect was improved in the case of TNF-α inhibition as reflected by their EC50 values of 116 ± 8.2 μg/ml and 49 ± 3.5 μg/ml for DC and DCm (P < 0.01).
Cytokines inhibitory activity of DC was not affected after its external metabolic activation. However, the amount of harpagoside and caffeic acid derivates was decreased. Other components of the extract might have contributed to its anti-inflammatory effect.
Emerging evidence indicates that Wnt/β-catenin pathway is linked to the fibrosis of different organs including liver fibrosis. β-Catenin promotes hepatic stellate cells (HSCs) activation, a key event in the development of liver fibrosis, and has emerged as a novel mediator of fibrosis. Curcumin, a natural active ingredient derived from turmeric, possesses an inhibitory effect on liver fibrosis. This study is aimed to examine whether curcumin affects β-catenin expression/activity in HSCs and explores the underlying mechanisms.
The researchers used Western blot, real-time PCR, transfection assay and electrophoretic mobility shift assay and employed cultured HSCs and rat model of liver injury.
Results showed that curcumin could reduce β-catenin protein level in HSCs in vitro and in vivo. Both β-catenin transactivation activity and DNA-binding activity were suppressed by curcumin. Moreover, nuclear β-catenin protein level was decreased by curcumin treatment. Further experiments suggested that delta-like homologue 1 contributed to curcumin inhibition of β-catenin transactivation activity in cultured HSCs.
Curcumin affects β-catenin pathway in HSCs and might suggest a possible new explanation for the effects of curcumin on HSC activation and liver fibrosis.
To investigate the influence of co-administrated Da-Chaihu-Tang (DCT; a traditional Chinese formulation) on the pharmacokinetics of nifedipine, as well as the safe optimal dosing interval to avoid the adverse interactions.
A single dose of DCT was administered with nifedipine simultaneously, 2 h before, 30 min before or 30 min after nifedipine administration. Pharmacokinetics of nifedipine with or without DCT were compared. The influences of DCT on nifedipine intestinal mucosal and hepatic metabolism were studied by using rat in-vitro everted jejunal sac model and hepatic microsomes.
A simultaneous co-administration of DCT significantly increased the area under concentration-time curve from time zero to infinity (AUC0-inf) of nifedipine. In-vitro mechanism investigations revealed that DCT inhibited both the intestinal and the hepatic metabolism of nifedipine. Further study on the optimal dosing interval for nifedipine and DCT revealed that administration of DCT 30 min before or after nifedipine did not significantly change the AUC of nifedipine.
The bioavailability of nifedipine is significantly increased by a simultaneous oral co-administration of DCT. This increase is caused by the inhibitory effect of DCT on both the intestinal mucosal and the hepatic metabolism of nifedipine. The dose interval between DCT and nifedipine needs to be set for over 30 min to avoid such drug–drug interactions.
The phenolic components and biological activity of nine Senecio species growing in Turkey were investigated.
Senecio species were extracted with methanol. The content of total phenols was determined using Folin–Ciocalteu method, while individual phenolic acids and flavonoids were detected using HPLC analysis. Also, to determine the antioxidant capacity, phosphomolybdenum assay and 1,1-diphenyl picrylhydrazyl (DPPH) radical-scavenging activity assay were used. Antimicrobial activity of extract was determined using agar diffusion and broth microdilution method.
The total phenolic contents of the extracts were found to be highest in Senecio cilicius and Senecio mollis extracts (117.45 and 113.40 mg equivalent to gallic acid/g, respectively). S. salsuginea showed the strongest free radical-scavenging activity with IC50 (the concentration providing 50% inhibition) = 26.23 μg/ml and S. mollis showed the highest antioxidant capacity in the phosphomolybdenum method (434.48 mg equivalent to ascorbic acid/g). The extracts exerted promising antibacterial activity against most of the test bacteria (minimal inhibitory concentration = 6.25–12.5 mg/ml), but no activity was observed against Candida albicans.
The results demonstrated that nine Senecio species possess high antioxidant and antimicrobial activity in accordance with the high amount of phenolic contents in the extracts that might be natural agencies used in many areas such as food, pharmacy and alternative medicine.
Inflammation is widely acknowledged to increase morbidity and mortality in myocardial infarction (MI), and the ideal therapeutic methods should be aimed at the inflammation reaction triggers. The aim was to evaluate the protective effect of saponins extracted from Panax japonicus (SPJ) on MI, and based on these results investigate the possible involvement mechanism of the nuclear factor-kappa B (NF-κB), sirtuin1 (SIRT1) and mitogen-activated protein kinases (MAPKs) signalling pathways.
Sprague-Dawley rats were randomly divided into sham, MI, MI + SPJ 50 and SPJ 100 mg/kg groups. After administration for 3 days, MI rats were created by ligaturing coronary artery, and then underwent the same administration for 7 days as before. Cardiac function and the expressions of pro-apoptosis protein Bax, anti-apoptosis protein Bcl-2, NF-κB, SIRT1, MAPKs signal pathway-related proteins and inflammatory factor, such as tumour necrosis factor alpha (TNF-α) and monocyte chemoattractant protein 1 (MCP-1), were assessed.
SPJ might significantly improve cardiac function, decrease the serum MCP-1 and TNF-α levels, ameliorate the increased Bax protein expression and decrease Bcl-2 protein expression, and suppress the protein expressions of NF-κBp65 subunit, extracellular signal-regulated kinase 1/2 (ERK1/2) and p38 MAPK, but have no effect on c-Jun NH2-terminal kinase, and increase the expression of SIRT1. Histopathological observations provided supportive evidence for aforementioned results, and with the dose of SPJ increasing, the aforesaid improvement became more and more strong.
The studies demonstrated that SPJ exerted beneficially cardioprotective effects on MI rats, mainly inhibiting NF-κB, ERK1/2 and p38 MAPK activation, but increased the expression of SIRT1, alleviating MI injury and cardiac cell death.


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Home Improvement News: Invest in paint now to safeguard your home
By Paint Quality Institute
Now that Labor Day has come and gone, most of the country is entering the stretch run of the exterior painting season. If your house is in need of a new paint job, there's little time to waste: "Paint-friendly" weather is fast disappearing.
"Doing exterior painting in the right weather conditions is extremely important," said Debbie Zimmer, paint and color expert at the Paint Quality Institute. "Fresh paint forms a better protective film when it's applied in moderate weather like that commonly seen in early autumn. If your house needs to be painted this year, you need to paint it now."
To appreciate the urgency, it's important to understand how paint "cures." While latex paint dries to the touch very quickly, it actually takes many hours for it to form the most protective paint film. If the temperature is very hot when the paint is applied — or more likely as winter approaches, if nighttime temperatures get very cold — that can adversely affect film formation, and even lead to early flaking and peeling of the paint.
This point is vividly demonstrated on test panels at the Paint Quality Institute. More than 40 years of outdoor testing there shows that exterior paint applied in moderate conditions can last many years longer than paints applied in very hot, very cold or even very windy weather.
Those extra years of service can offer homeowners a big return on their investment in an exterior paint job. As an example, let's say a new paint job costs $3,000. If it lasts only five years, the annualized cost is $600; but if it holds up for 10 years, the annualized cost is only $300 — half as much!
As the days get shorter and winter approaches, painting in moderate weather is one of the most important factors in getting a long-lasting exterior paint job. Other things that can affect the longevity of a coating aren't so time-sensitive, according to Zimmer.
"To get the most durable paint job, it's always very important to properly prepare exterior surfaces and to use a top-quality paint," she said.
Good exterior surface preparation requires that siding and trim be clean and free of dust, dirt and mildew, so that the new paint can adhere properly. Of course, peeling, flaking or otherwise unstable paint should be completely removed, and any bare wood primed before painting.
As for the type of paint to use, studies show that high-quality, 100 percent acrylic latex paints are the most durable coatings for most home exteriors, including wood, masonry, vinyl and aluminum siding. These paints adhere well, they're tough, and they're very flexible, so they tend to expand or contract with the home exterior when temperatures rise or fall dramatically.
Which brings us back to the weather: As the song says, "For everything, there is a season." When it comes to exterior painting, that season is now.
From the Oct. 26-Nov. 1, 2011, issue

subset: commoncrawl


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NAICC: Number one key to resistance management: Teamwork
James Todd Resistance Management. There has probably not been a meeting in agriculture this winter that has not had at least one speaker on this topic. But, as I have sat through these meetings this winter, the key to resistance management has been overlooked in many instances: Teamwork. That is one of those old worn out phrases, I know, but it is essential when you think about managing resistant weeds, insects and diseases. Everyone in the game must know the game plan and know their role to be successful.
For crop consultants, we must never forget the basics of resistance management include rotating chemistry, using multiple modes of action, only treating at economic threshold and using appropriate refuges for Bt crops. Farmers must understand and be willing to implement these strategies as well, which may not mean using the cheaper alternative, but using those that will ensure the longevity of the chemistries.
In the case of resistant weeds, farmers may need to go back to using cultivators and hooded-sprayers as means of controlling weeds. Retailers and manufacturers understand these principles as well and play a significant role in making sure products are available to growers when the time comes to make an application. At times, the availability of products, due to inventory or production limitations, can be a major obstacle to implementing effective resistance management.
There are even more players in the game than just the ones mentioned above. Equipment dealers and custom harvesters must ensure equipment is clean of plant debris and weed seed when moving from one region to the next. This helps to ensure that resistant weed seed is not moved from one region to another. It has been well documented that resistant weeds are being moved in this manner. Farmers and consultants play a role here as well by ensuring their equipment is kept clean and does not become a mechanism for moving resistant weeds.
As with any team, communication becomes the key to being successful. And it is important that each player in the game communicates with each and understands the resistance management plan that is in place. This plan becomes important in making sure products are available at local retailers and retailers understand which products will be used in the upcoming year.
Furthermore, it is important in the game of resistance management to be on the offensive. In other words, waiting for resistance to occur and then developing a resistance management plan is the wrong approach to take. Farmers, consultants, manufacturers and retailers must understand how important it is to rotate chemistries and not to "over use" products. Manufacturers can take the lead here by educating growers, consultants and retailers on the mode of action found in their respective products.
I am confident that if we all work together, resistance management can be succeed and we can ensure the longevity of new products to come.
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"Cranes 101: A Basic Introduction to Cranes" is the February topic for the Tennessee Wildlife Resources Agency's Nature at Noontime program. The program will be held from noon-1 p.m. on Thursday, Feb. 7.
Melinda Welton will be the program presenter. She is a research associate with Gulf Coast Bird Observatory and an ornithological consultant under the name of Birdworks Consulting. She has been actively involved in several TWRA projects concerning songbirds and for the last two years, she has also served as co-chairwoman of the Tennessee Sandhill Crane Festival.
Standing over four feet tall with a wingspan of more than six feet, the sandhill crane is one of the largest birds found in Tennessee and the most abundant crane in the world. Over the last 60 years, the sandhill has made a comeback after being nearly wiped out in the 1800s due to breeding habitat loss and over hunting. Virtually every individual in the smaller eastern population of sandhill cranes will pass over or spend the winter at the confluence of the Tennessee and Hiwassee rivers in southeastern Tennessee. Thousands of these birds can be viewed each winter at TWRA's Hiwassee Refuge.
Hosted by the Information and Education Division, Nature at Noontime is held the first Thursday of each month. TWRA Nature at Noontime presentations are about natural resource related topics and last about 30-45 minutes, allowing time for discussion during the allotted lunch hour.
Contact Don King 615 781-6502 or by e-mail: firstname.lastname@example.com for more information.
October 30, 2014
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When the first College Football Playoff rankings were released earlier this week, Alabama football coach Nick Saban got it right on the button when he shrugged, "I don't even care, to be honest with you." "To me, none of it matters, What does it matter?" he laughed at the Tide's No. 6 ranking. "I mean, it only matters where you end up at the end. So what matters to us is how ... (click for more)


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Social Sharing Buttons:All Consumer Marketing Articles - Page 9
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Io e te nella valle delle mille luci
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Marco Brusco Artist
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Corrado Lippi Artist
Molto bella e romantica.
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Selected artists invited to exhibit in 10 galleries around the world will be announced on 15 November during Celeste Prize awards night at Assab-One, Milan. Every year partner galleries and Celeste provide exhibition opportunities for artists who enter the prize.


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Sticking With It
Sticking With It
Now that you've gotten over the first few days, it's time to make smoking a thing of the past. Sticking with it can be a challenge, but it can be done.
Plan rewards! You will save money by being smoke-free. Make a list of things you'd like to buy with your new-found fortune! See a movie. Buy a CD you've been wanting. Or buy another treat!
Wait out the urges. Remember that the urge to smoke usually lasts only three to five minutes. Try and wait it out. Take deep breaths, light a candle instead of a cigarette, or go into another location to change up the scenery and break up the desire.
Find new things to do. Leave the old habit of smoking behind and try some new habits! Take up swimming, jogging, playing tennis, or even walking your dog. How about crossword puzzles or needlework?
Watch your weight. Are you worried about gaining weight when you quit smoking? Make exercise a part of your quitting plan. Exercising can distract you from smoking and lowers the stress that makes you crave a cigarette.
Forgive yourself. If you slip up and give in to temptation, don't be discouraged. It's not a lost cause. Don't beat yourself up over it, but don't let yourself fall back into smoking, either. Feel good about all the time you went without smoking, and try to learn how to make your coping skills better.
Remember the healing effects. Your body begins to heal within 20 minutes after your last cigarette. The poison gas and nicotine start to leave your body. You can breathe easier. Your sense of taste and smell improve. Your lungs begin to repair themselves.
After you've quit, you've added healthy, full days to each year of your life. You've greatly lowered your risk of death from lung cancer and other disease including:
- Heart disease,
- Stroke,
- Chronic bronchitis,
- Emphysema,
- and Cancer.
Stay upbeat. As you go through the first days and weeks without smoking, keep a positive outlook. Quitting is a learning process – take it one day at a time.
National Cancer Institute. "Clearing the Air: Quit Smoking Today." PDF brochure downloaded from www.smokefree.gov, accessed 9/14/07.
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