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Int. J. Mol. Sci., Volume 14, Issue 1 (January 2013) , Pages 1-2229

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Open AccessArticle
Adhesion-Induced Phase Behavior of Two-Component Membranes and Vesicles
Int. J. Mol. Sci. 2013, 14(1), 2203-2229; https://doi.org/10.3390/ijms14012203
Received: 20 December 2012 / Revised: 17 January 2013 / Accepted: 18 January 2013 / Published: 22 January 2013
Cited by 6 | Viewed by 2669 | PDF Full-text (931 KB) | HTML Full-text | XML Full-text
Abstract
The interplay of adhesion and phase separation is studied theoretically for two-component membranes that can phase separate into two fluid phases such as liquid-ordered and liquid-disordered phases. Many adhesion geometries provide two different environments for these membranes and then partition the membranes into [...] Read more.
The interplay of adhesion and phase separation is studied theoretically for two-component membranes that can phase separate into two fluid phases such as liquid-ordered and liquid-disordered phases. Many adhesion geometries provide two different environments for these membranes and then partition the membranes into two segments that differ in their composition. Examples are provided by adhering vesicles, by hole- or pore-spanning membranes, and by membranes supported by chemically patterned surfaces. Generalizing a lattice model for binary mixtures to these adhesion geometries, we show that the phase behavior of the adhering membranes depends, apart from composition and temperature, on two additional parameters, the area fraction of one membrane segment and the affinity contrast between the two segments. For the generic case of non-vanishing affinity contrast, the adhering membranes undergo two distinct phase transitions and the phase diagrams in the composition/temperature plane have a generic topology that consists of two two-phase coexistence regions separated by an intermediate one-phase region. As a consequence, phase separation and domain formation is predicted to occur separately in each of the two membrane segments but not in both segments simultaneously. Furthermore, adhesion is also predicted to suppress the phase separation process for certain regions of the phase diagrams. These generic features of the adhesion-induced phase behavior are accessible to experiment. Full article
(This article belongs to the Special Issue Self-Assembled Soft Matter Nanostructures at Interfaces)
Open AccessArticle
Effect of Repetitive Lysine-Tryptophan Motifs on the Eukaryotic Membrane
Int. J. Mol. Sci. 2013, 14(1), 2190-2202; https://doi.org/10.3390/ijms14012190
Received: 27 November 2012 / Revised: 14 January 2013 / Accepted: 15 January 2013 / Published: 22 January 2013
Cited by 12 | Viewed by 2867 | PDF Full-text (687 KB) | HTML Full-text | XML Full-text
Abstract
In a previous study, we synthesized a series of peptides containing simple sequence repeats, (KW)nNH2 (n = 2,3,4 and 5) and determined their antimicrobial and hemolytic activities, as well as their mechanism of antimicrobial action. However, (KW)5 [...] Read more.
In a previous study, we synthesized a series of peptides containing simple sequence repeats, (KW)nNH2 (n = 2,3,4 and 5) and determined their antimicrobial and hemolytic activities, as well as their mechanism of antimicrobial action. However, (KW)5 showed undesirable cytotoxicity against RBC cells. In order to identify the mechanisms behind the hemolytic and cytotoxic activities of (KW)5, we measured the ability of these peptides to induce aggregation of liposomes. In addition, their binding and permeation activities were assessed by Trp fluorescence, calcein leakage and circular dichrorism using artificial phospholipids that mimic eukaryotic liposomes, including phosphatidylcholine (PC), PC/sphingomyelin (SM) (2:1, w/w) and PC/cholesterol (CH) (2:1, w/w). Experiments confirmed that only (KW)5 induced aggregation of all liposomes; it formed much larger aggregates with PC:CH (2:1, w/w) than with PC or PC:SM (2:1, w/w). Longer peptide (KW)5, but not (KW)3 or (KW)4, strongly bound and partially inserted into PC:CH compared to PC or PC:SM (2:1, w/w). Calcein release experiments showed that (KW)5 induced calcein leakage from the eukaryotic membrane. Greater calcein leakage was induced by (KW)5 from PC:CH than from PC:SM (2:1, w/w) or PC, whereas (KW)4 did not induce calcein leakage from any of the liposomes. Circular dichroism measurements indicated that (KW)5 showed higher conformational transition compared to (KW)4 due to peptide-liposome interactions. Taken together, our results suggest that (KW)5 reasonably mediates the aggregation and permeabilization of eukaryotic membranes, which could in turn explain why (KW)5 displays efficient hemolytic activity. Full article
(This article belongs to the Special Issue Phospholipids: Molecular Sciences 2012)
Open AccessArticle
Induction of Heat Shock Protein 70 Ameliorates Ultraviolet-Induced Photokeratitis in Mice
Int. J. Mol. Sci. 2013, 14(1), 2175-2189; https://doi.org/10.3390/ijms14012175
Received: 7 November 2012 / Revised: 9 January 2013 / Accepted: 18 January 2013 / Published: 22 January 2013
Cited by 10 | Viewed by 3011 | PDF Full-text (3228 KB) | HTML Full-text | XML Full-text
Abstract
Acute ultraviolet (UV) B exposure causes photokeratitis and induces apoptosis in corneal cells. Geranylgeranylacetone (GGA) is an acyclic polyisoprenoid that induces expression of heat shock protein (HSP)70, a soluble intracellular chaperone protein expressed in various tissues, protecting cells against stress conditions. We examined [...] Read more.
Acute ultraviolet (UV) B exposure causes photokeratitis and induces apoptosis in corneal cells. Geranylgeranylacetone (GGA) is an acyclic polyisoprenoid that induces expression of heat shock protein (HSP)70, a soluble intracellular chaperone protein expressed in various tissues, protecting cells against stress conditions. We examined whether induction of HSP70 has therapeutic effects on UV-photokeratitis in mice. C57 BL/6 mice were divided into four groups, GGA-treated (500 mg/kg/mouse) and UVB-exposed (400 mJ/cm2), GGA-untreated UVB-exposed (400 mJ/cm2), GGA-treated (500 mg/kg/mouse) but not exposed and naive controls. Eyeballs were collected 24 h after irradiation, and corneas were stained with hematoxylin and eosin (H&E) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL). HSP70, reactive oxygen species (ROS) production, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and protein kinase B (Akt) expression were also evaluated. Irradiated corneal epithelium was significantly thicker in the eyes of mice treated with GGA compared with those given the vehicle alone (p < 0.01). Significantly fewer TUNEL-positive cells were observed in the eyes of GGA-treated mice than controls after irradiation (p < 0.01). Corneal HSP70 levels were significantly elevated in corneas of mice treated with GGA (p < 0.05). ROS signal was not affected by GGA. NF-κB activation was reduced but phospho-(Ser/Ther) Akt substrate expression was increased in corneas after irradiation when treated with GGA. GGA-treatment induced HSP70 expression and ameliorated UV-induced corneal damage through the reduced NF-κB activation and possibly increased Akt phosphorilation. Full article
(This article belongs to the Special Issue UV-Induced Cell Death 2012)
Open AccessReview
Innovative Therapeutic Strategies in the Treatment of Brain Metastases
Int. J. Mol. Sci. 2013, 14(1), 2135-2174; https://doi.org/10.3390/ijms14012135
Received: 9 December 2012 / Revised: 8 January 2013 / Accepted: 9 January 2013 / Published: 22 January 2013
Cited by 30 | Viewed by 3244 | PDF Full-text (310 KB) | HTML Full-text | XML Full-text
Abstract
Brain metastases (BM) are the most common intracranial tumors and their incidence is increasing. Untreated brain metastases are associated with a poor prognosis and a poor performance status. Metastasis development involves the migration of a cancer cell from the bulk tumor into the [...] Read more.
Brain metastases (BM) are the most common intracranial tumors and their incidence is increasing. Untreated brain metastases are associated with a poor prognosis and a poor performance status. Metastasis development involves the migration of a cancer cell from the bulk tumor into the surrounding tissue, extravasation from the blood into tissue elsewhere in the body, and formation of a secondary tumor. In the recent past, important results have been obtained in the management of patients affected by BM, using surgery, radiation therapy, or both. Conventional chemotherapies have generally produced disappointing results, possibly due to their limited ability to penetrate the blood–brain barrier. The advent of new technologies has led to the discovery of novel molecules and pathways that have better depicted the metastatic process. Targeted therapies such as bevacizumab, erlotinib, gefitinib, sunitinib and sorafenib, are all licensed and have demonstrated improved survival in patients with metastatic disease. In this review, we will report current data on targeted therapies. A brief review about brain metastatic process will be also presented. Full article
(This article belongs to the Special Issue Brain Metastasis)
Open AccessArticle
Low Oxygen Tension Maintains Multipotency, Whereas Normoxia Increases Differentiation of Mouse Bone Marrow Stromal Cells
Int. J. Mol. Sci. 2013, 14(1), 2119-2134; https://doi.org/10.3390/ijms14012119
Received: 21 November 2012 / Revised: 4 January 2013 / Accepted: 5 January 2013 / Published: 22 January 2013
Cited by 35 | Viewed by 3496 | PDF Full-text (857 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Optimization of mesenchymal stem cells (MSC) culture conditions is of great importance for their more successful application in regenerative medicine. O2 regulates various aspects of cellular biology and, in vivo, MSC are exposed to different O2 concentrations spanning from very [...] Read more.
Optimization of mesenchymal stem cells (MSC) culture conditions is of great importance for their more successful application in regenerative medicine. O2 regulates various aspects of cellular biology and, in vivo, MSC are exposed to different O2 concentrations spanning from very low tension in the bone marrow niche, to higher amounts in wounds. In our present work, we isolated mouse bone marrow stromal cells (BMSC) and showed that they contained a population meeting requirements for MSC definition. In order to establish the effect of low O2 on cellular properties, we examined BSMC cultured under hypoxic (3% O2) conditions. Our results demonstrate that 3% O2 augmented proliferation of BMSC, as well as the formation of colonies in the colony-forming unit assay (CFU-A), the percentage of quiescent cells, and the expression of stemness markers Rex-1 and Oct-4, thereby suggesting an increase in the stemness of culture when exposed to hypoxia. In contrast, intrinsic differentiation processes were inhibited by 3% O2. Overall yield of differentiation was dependent on the adjustment of O2 tension to the specific stage of BMSC culture. Thus, we established a strategy for efficient BMSC in vitro differentiation using an initial phase of cell propagation at 3% O2, followed by differentiation stage at 21% O2. We also demonstrated that 3% O2 affected BMSC differentiation in p53 and reactive oxygen species (ROS) independent pathways. Our findings can significantly contribute to the obtaining of high-quality MSC for effective cell therapy. Full article
(This article belongs to the Special Issue Advances in Free Radicals in Biology and Medicine)
Open AccessArticle
Metabolic Profiles of Brain Metastases
Int. J. Mol. Sci. 2013, 14(1), 2104-2118; https://doi.org/10.3390/ijms14012104
Received: 26 October 2012 / Accepted: 9 January 2013 / Published: 22 January 2013
Cited by 21 | Viewed by 3356 | PDF Full-text (2255 KB) | HTML Full-text | XML Full-text
Abstract
Metastasis to the brain is a feared complication of systemic cancer, associated with significant morbidity and poor prognosis. A better understanding of the tumor metabolism might help us meet the challenges in controlling brain metastases. The study aims to characterize the metabolic profile [...] Read more.
Metastasis to the brain is a feared complication of systemic cancer, associated with significant morbidity and poor prognosis. A better understanding of the tumor metabolism might help us meet the challenges in controlling brain metastases. The study aims to characterize the metabolic profile of brain metastases of different origin using high resolution magic angle spinning (HR-MAS) magnetic resonance spectroscopy (MRS) to correlate the metabolic profiles to clinical and pathological information. Biopsy samples of human brain metastases (n = 49) were investigated. A significant correlation between lipid signals and necrosis in brain metastases was observed (p < 0.01), irrespective of their primary origin. The principal component analysis (PCA) showed that brain metastases from malignant melanomas cluster together, while lung carcinomas were metabolically heterogeneous and overlap with other subtypes. Metastatic melanomas have higher amounts of glycerophosphocholine than other brain metastases. A significant correlation between microscopically visible lipid droplets estimated by Nile Red staining and MR visible lipid signals was observed in metastatic lung carcinomas (p = 0.01), indicating that the proton MR visible lipid signals arise from cytoplasmic lipid droplets. MRS-based metabolomic profiling is a useful tool for exploring the metabolic profiles of metastatic brain tumors. Full article
(This article belongs to the Special Issue Brain Metastasis)
Open AccessArticle
Pre-Treatment of Platinum Resistant Ovarian Cancer Cells with an MMP-9/MMP-2 Inhibitor Prior to Cisplatin Enhances Cytotoxicity as Determined by High Content Screening
Int. J. Mol. Sci. 2013, 14(1), 2085-2103; https://doi.org/10.3390/ijms14012085
Received: 7 November 2012 / Revised: 5 January 2013 / Accepted: 6 January 2013 / Published: 22 January 2013
Cited by 13 | Viewed by 4684 | PDF Full-text (535 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Platinum resistance is a major cause of treatment failure in ovarian cancer. We previously identified matrix metalloproteinase 9 (MMP-9) as a potential therapeutic target of chemoresistant disease. A2780cis (cisplatin-resistant) and A2780 (cisplatin-sensitive) ovarian carcinoma cell lines were used. The cytotoxic effect of MMP-9/MMP-2 [...] Read more.
Platinum resistance is a major cause of treatment failure in ovarian cancer. We previously identified matrix metalloproteinase 9 (MMP-9) as a potential therapeutic target of chemoresistant disease. A2780cis (cisplatin-resistant) and A2780 (cisplatin-sensitive) ovarian carcinoma cell lines were used. The cytotoxic effect of MMP-9/MMP-2 inhibitor, (2R)-2-[(4-Biphenylsulfonyl) amino]-3 phenylpropionic acid (C21H19NO4S) alone or in combination with cisplatin was determined using high content screening. Protein expression was examined using immunohistochemistry and ELISA. Co-incubation of cisplatin and an MMP-9/MMP-2 inhibitor, (2R)-2-[(4-Biphenylsulfonyl) amino]-3 phenylpropionic acid (C21H19NO4S) resulted in significantly greater cytotoxicity as compared to either treatment alone in a cisplatin resistant MMP-9 overexpressing cell line; A2780cis. In addition, pre-incubating with MMP-9i prior to cisplatin further enhances the cytotoxic effect. No significant difference was observed in MMP-9 protein in tissue but a trend towards increased MMP-9 was observed in recurrent serum. We propose that MMP-9/MMP-2i may be utilized in the treatment of recurrent/chemoresistant ovarian cancers that overexpress MMP-9 mRNA but its role in vivo remains to be evaluated. Full article
(This article belongs to the Special Issue Genes and Pathways in the Pathogenesis of Ovarian Cancer)
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Open AccessArticle
Blood microRNAs in Low or No Risk Ischemic Stroke Patients
Int. J. Mol. Sci. 2013, 14(1), 2072-2084; https://doi.org/10.3390/ijms14012072
Received: 6 November 2012 / Revised: 11 December 2012 / Accepted: 17 January 2013 / Published: 22 January 2013
Cited by 39 | Viewed by 3694 | PDF Full-text (492 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Ischemic stroke is a multi-factorial disease where some patients present themselves with little or no risk factors. Blood microRNA expression profiles are becoming useful in the diagnosis and prognosis of human diseases. We therefore investigated the blood microRNA profiles in young stroke patients [...] Read more.
Ischemic stroke is a multi-factorial disease where some patients present themselves with little or no risk factors. Blood microRNA expression profiles are becoming useful in the diagnosis and prognosis of human diseases. We therefore investigated the blood microRNA profiles in young stroke patients who presented with minimal or absence of risk factors for stroke such as type 2 diabetes, dyslipidemia and hypertension. Blood microRNA profiles from these patients varied with stroke subtypes as well as different functional outcomes (based on modified Rankin Score). These microRNAs have been shown to target genes that are involved in stroke pathogenesis. The findings from our study suggest that molecular mechanisms in stroke pathogenesis involving low or no risk ischemic stroke patients could differ substantially from those with pre-existing risk factors. Full article
(This article belongs to the Special Issue Non-Coding RNAs 2012)
Open AccessArticle
Development of Collagen/Demineralized Bone Powder Scaffolds and Periosteum-Derived Cells for Bone Tissue Engineering Application
Int. J. Mol. Sci. 2013, 14(1), 2056-2071; https://doi.org/10.3390/ijms14012056
Received: 24 October 2012 / Revised: 11 January 2013 / Accepted: 14 January 2013 / Published: 21 January 2013
Cited by 19 | Viewed by 3400 | PDF Full-text (2113 KB) | HTML Full-text | XML Full-text
Abstract
The aim of this study was to investigate physical and biological properties of collagen (COL) and demineralized bone powder (DBP) scaffolds for bone tissue engineering. DBP was prepared and divided into three groups, based on various particle sizes: 75–125 µm, 125–250 µm, and [...] Read more.
The aim of this study was to investigate physical and biological properties of collagen (COL) and demineralized bone powder (DBP) scaffolds for bone tissue engineering. DBP was prepared and divided into three groups, based on various particle sizes: 75–125 µm, 125–250 µm, and 250–500 µm. DBP was homogeneously mixed with type I collagen and three-dimensional scaffolds were constructed, applying chemical crosslinking and lyophilization. Upon culture with human periosteum-derived cells (PD cells), osteogenic differentiation of PD cells was investigated using alkaline phosphatase (ALP) activity and calcium assay kits. The physical properties of the COL/DBP scaffolds were obviously different from COL scaffolds, irrespective of the size of DBP. In addition, PD cells cultured with COL scaffolds showed significantly higher cell adhesion and proliferation than those with COL/DBP scaffolds. In contrast, COL/DBP scaffolds exhibited greater osteoinductive potential than COL scaffolds. The PD cells with COL/DBP scaffolds possessed higher ALP activity than those with COL scaffolds. PD cells cultured with COL/DBP scaffolds with 250–500 mm particle size yielded the maximum calcium deposition. In conclusion, PD cells cultured on the scaffolds could exhibit osteoinductive potential. The composite scaffold of COL/DBP with 250–500 mm particle size could be considered a potential bone tissue engineering implant. Full article
(This article belongs to the Section Materials Science)
Open AccessArticle
Corneal Stromal Cell Growth on Gelatin/Chondroitin Sulfate Scaffolds Modified at Different NHS/EDC Molar Ratios
Int. J. Mol. Sci. 2013, 14(1), 2036-2055; https://doi.org/10.3390/ijms14012036
Received: 5 November 2012 / Revised: 13 December 2012 / Accepted: 5 January 2013 / Published: 21 January 2013
Cited by 25 | Viewed by 3236 | PDF Full-text (674 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
A nanoscale modification strategy that can incorporate chondroitin sulfate (CS) into the cross-linked porous gelatin materials has previously been proposed to give superior performance for designed corneal keratocyte scaffolds. The purpose of this work was to further investigate the influence of carbodiimide chemistry [...] Read more.
A nanoscale modification strategy that can incorporate chondroitin sulfate (CS) into the cross-linked porous gelatin materials has previously been proposed to give superior performance for designed corneal keratocyte scaffolds. The purpose of this work was to further investigate the influence of carbodiimide chemistry on the characteristics and biofunctionalities of gelatin/CS scaffolds treated with varying N-hydroxysuccinimide (NHS)/1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC) molar ratios (0-1) at a constant EDC concentration of 10 mM. Results of Fourier transform infrared spectroscopy and dimethylmethylene blue assays consistently indicated that when the NHS to EDC molar ratio exceeds a critical level (i.e., 0.5), the efficiency of carbodiimide-mediated biomaterial modification is significantly reduced. With the optimum NHS/EDC molar ratio of 0.5, chemical treatment could achieve relatively high CS content in the gelatin scaffolds, thereby enhancing the water content, glucose permeation, and fibronectin adsorption. Live/Dead assays and interleukin-6 mRNA expression analyses demonstrated that all the test samples have good cytocompatibility without causing toxicity and inflammation. In the molar ratio range of NHS to EDC from 0 to 0.5, the cell adhesion ratio and proliferation activity on the chemically modified samples significantly increased, which is attributed to the increasing CS content. Additionally, the materials with highest CS content (0.143 ± 0.007 nmol/10 mg scaffold) showed the greatest stimulatory effect on the biosynthetic activity of cultivated keratocytes. These findings suggest that a positive correlation is noticed between the NHS to EDC molar ratio and the CS content in the biopolymer matrices, thereby greatly affecting the corneal stromal cell growth. Full article
(This article belongs to the Section Materials Science)
Open AccessArticle
Self-Assembly of Discrete Metal Complexes in Aqueous Solution via Block Copolypeptide Amphiphiles
Int. J. Mol. Sci. 2013, 14(1), 2022-2035; https://doi.org/10.3390/ijms14012022
Received: 11 December 2012 / Revised: 9 January 2013 / Accepted: 17 January 2013 / Published: 21 January 2013
Cited by 12 | Viewed by 3193 | PDF Full-text (2075 KB) | HTML Full-text | XML Full-text
Abstract
The integration of discrete metal complexes has been attracting significant interest due to the potential of these materials for soft metal-metal interactions and supramolecular assembly. Additionally, block copolypeptide amphiphiles have been investigated concerning their capacity for self-assembly into structures such as nanoparticles, nanosheets [...] Read more.
The integration of discrete metal complexes has been attracting significant interest due to the potential of these materials for soft metal-metal interactions and supramolecular assembly. Additionally, block copolypeptide amphiphiles have been investigated concerning their capacity for self-assembly into structures such as nanoparticles, nanosheets and nanofibers. In this study, we combined these two concepts by investigating the self-assembly of discrete metal complexes in aqueous solution using block copolypeptides. Normally, discrete metal complexes such as [Au(CN)2], when molecularly dispersed in water, cannot interact with one another. Our results demonstrated, however, that the addition of block copolypeptide amphiphiles such as K183L19 to [Au(CN)2] solutions induced one-dimensional integration of the discrete metal complex, resulting in photoluminescence originating from multinuclear complexes with metal-metal interactions. Transmission electron microscopy (TEM) showed a fibrous nanostructure with lengths and widths of approximately 100 and 20 nm, respectively, which grew to form advanced nanoarchitectures, including those resembling the weave patterns of Waraji (traditional Japanese straw sandals). This concept of combining block copolypeptide amphiphiles with discrete coordination compounds allows the design of flexible and functional supramolecular coordination systems in water. Full article
(This article belongs to the Special Issue Molecular Self-Assembly 2012)
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Open AccessArticle
Catalysis of Transesterification Reactions by a Self-Assembled Nanosystem
Int. J. Mol. Sci. 2013, 14(1), 2011-2021; https://doi.org/10.3390/ijms14012011
Received: 30 November 2012 / Accepted: 14 January 2013 / Published: 21 January 2013
Cited by 3 | Viewed by 2626 | PDF Full-text (461 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Histidine-containing peptides self-assemble on the surface of monolayer protected gold nanoparticles to form a catalytic system for transesterification reactions. Self-assembly is a prerequisite for catalysis, since the isolated peptides do not display catalytic activity by themselves. A series of catalytic peptides and substrates [...] Read more.
Histidine-containing peptides self-assemble on the surface of monolayer protected gold nanoparticles to form a catalytic system for transesterification reactions. Self-assembly is a prerequisite for catalysis, since the isolated peptides do not display catalytic activity by themselves. A series of catalytic peptides and substrates are studied in order to understand the structural parameters that are of relevance to the catalytic efficiency of the system. It is shown that the distance between the His-residue and the anionic tail does not affect the catalytic activity. On the other hand, the catalytic His-residue is sensitive to the chemical nature of the flanking amino acid residues. In particular, the presence of polar Ser-residues causes a significant increase in activity. Finally, kinetic studies of a series of substrates reveal that substrates with a hydrophobic component are very suitable for this catalytic system. Full article
(This article belongs to the Special Issue Molecular Self-Assembly 2012)
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Open AccessArticle
Quantitative and Chemical Fingerprint Analysis for the Quality Evaluation of Receptaculum Nelumbinis by RP-HPLC Coupled with Hierarchical Clustering Analysis
Int. J. Mol. Sci. 2013, 14(1), 1999-2010; https://doi.org/10.3390/ijms14011999
Received: 28 September 2012 / Revised: 10 January 2013 / Accepted: 11 January 2013 / Published: 21 January 2013
Cited by 14 | Viewed by 2853 | PDF Full-text (467 KB) | HTML Full-text | XML Full-text
Abstract
A simple and reliable method of high-performance liquid chromatography with photodiode array detection (HPLC-DAD) was developed to evaluate the quality of Receptaculum Nelumbinis (dried receptacle of Nelumbo nucifera) through establishing chromatographic fingerprint and simultaneous determination of five flavonol glycosides, including hyperoside, isoquercitrin, [...] Read more.
A simple and reliable method of high-performance liquid chromatography with photodiode array detection (HPLC-DAD) was developed to evaluate the quality of Receptaculum Nelumbinis (dried receptacle of Nelumbo nucifera) through establishing chromatographic fingerprint and simultaneous determination of five flavonol glycosides, including hyperoside, isoquercitrin, quercetin-3-O-β-d-glucuronide, isorhamnetin-3-O-β-d-galactoside and syringetin-3-O-β-d-glucoside. In quantitative analysis, the five components showed good regression (R > 0.9998) within linear ranges, and their recoveries were in the range of 98.31%–100.32%. In the chromatographic fingerprint, twelve peaks were selected as the characteristic peaks to assess the similarities of different samples collected from different origins in China according to the State Food and Drug Administration (SFDA) requirements. Furthermore, hierarchical cluster analysis (HCA) was also applied to evaluate the variation of chemical components among different sources of Receptaculum Nelumbinis in China. This study indicated that the combination of quantitative and chromatographic fingerprint analysis can be readily utilized as a quality control method for Receptaculum Nelumbinis and its related traditional Chinese medicinal preparations. Full article
Open AccessReview
The Twenty-Year Story of a Plant-Based Vaccine Against Hepatitis B: Stagnation or Promising Prospects?
Int. J. Mol. Sci. 2013, 14(1), 1978-1998; https://doi.org/10.3390/ijms14011978
Received: 17 December 2012 / Revised: 7 January 2013 / Accepted: 14 January 2013 / Published: 21 January 2013
Cited by 19 | Viewed by 4360 | PDF Full-text (591 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Hepatitis B persists as a common human disease despite effective vaccines having been employed for almost 30 years. Plants were considered as alternative sources of vaccines, to be mainly orally administered. Despite 20-year attempts, no real anti-HBV plant-based vaccine has been developed. Immunization [...] Read more.
Hepatitis B persists as a common human disease despite effective vaccines having been employed for almost 30 years. Plants were considered as alternative sources of vaccines, to be mainly orally administered. Despite 20-year attempts, no real anti-HBV plant-based vaccine has been developed. Immunization trials, based on ingestion of raw plant tissue and conjugated with injection or exclusively oral administration of lyophilized tissue, were either impractical or insufficient due to oral tolerance acquisition. Plant-produced purified HBV antigens were highly immunogenic when injected, but their yields were initially insufficient for practical purposes. However, knowledge and technology have progressed, hence new plant-derived anti-HBV vaccines can be proposed today. All HBV antigens can be efficiently produced in stable or transient expression systems. Processing of injection vaccines has been developed and needs only to be successfully completed. Purified antigens can be used for injection in an equivalent manner to the present commercial vaccines. Although oral vaccines require improvement, plant tissue, lyophilized or extracted and converted into tablets, etc., may serve as a boosting vaccine. Preliminary data indicate also that both vaccines can be combined in an effective parenteral-oral immunization procedure. A partial substitution of injection vaccines with oral formulations still offers good prospects for economically viable and efficacious anti-HBV plant-based vaccines. Full article
(This article belongs to the Special Issue Plant-Derived Pharmaceuticals by Molecular Farming 2012)
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Open AccessReview
Vitamin D and Death by Sunshine
Int. J. Mol. Sci. 2013, 14(1), 1964-1977; https://doi.org/10.3390/ijms14011964
Received: 6 December 2012 / Revised: 4 January 2013 / Accepted: 10 January 2013 / Published: 18 January 2013
Cited by 14 | Viewed by 5377 | PDF Full-text (425 KB) | HTML Full-text | XML Full-text
Abstract
Exposure to sunlight is the major cause of skin cancer. Ultraviolet radiation (UV) from the sun causes damage to DNA by direct absorption and can cause skin cell death. UV also causes production of reactive oxygen species that may interact with DNA to [...] Read more.
Exposure to sunlight is the major cause of skin cancer. Ultraviolet radiation (UV) from the sun causes damage to DNA by direct absorption and can cause skin cell death. UV also causes production of reactive oxygen species that may interact with DNA to indirectly cause oxidative DNA damage. UV increases accumulation of p53 in skin cells, which upregulates repair genes but promotes death of irreparably damaged cells. A benefit of sunlight is vitamin D, which is formed following exposure of 7-dehydrocholesterol in skin cells to UV. The relatively inert vitamin D is metabolized to various biologically active compounds, including 1,25-dihydroxyvitamin D3. Therapeutic use of vitamin D compounds has proven beneficial in several cancer types, but more recently these compounds have been shown to prevent UV-induced cell death and DNA damage in human skin cells. Here, we discuss the effects of vitamin D compounds in skin cells that have been exposed to UV. Specifically, we examine the various signaling pathways involved in the vitamin D-induced protection of skin cells from UV. Full article
(This article belongs to the Special Issue UV-Induced Cell Death 2012)
Open AccessArticle
pH Dependence of the Fluorescence Lifetime of FAD in Solution and in Cells
Int. J. Mol. Sci. 2013, 14(1), 1952-1963; https://doi.org/10.3390/ijms14011952
Received: 21 November 2012 / Revised: 27 December 2012 / Accepted: 9 January 2013 / Published: 18 January 2013
Cited by 46 | Viewed by 3729 | PDF Full-text (1030 KB) | HTML Full-text | XML Full-text
Abstract
We have studied physiological parameters in a living cell using fluorescence lifetime imaging of endogenous chromophores. In this study, pH dependence of the fluorescence lifetime of flavin adenine dinucleotide (FAD), that is a significant cofactor exhibiting autofluorescence, has been investigated in buffer solution [...] Read more.
We have studied physiological parameters in a living cell using fluorescence lifetime imaging of endogenous chromophores. In this study, pH dependence of the fluorescence lifetime of flavin adenine dinucleotide (FAD), that is a significant cofactor exhibiting autofluorescence, has been investigated in buffer solution and in cells. The fluorescence lifetime of FAD remained unchanged with pH 5 to 9 in solution. However, the fluorescence lifetime in HeLa cells was found to decrease with increasing intracellular pH, suggesting that pH in a single cell can be estimated from the fluorescence lifetime imaging of FAD without adding exogenous fluorescent probes. Full article
(This article belongs to the Section Biochemistry)
Open AccessArticle
Resveratrol Down-Regulates Myosin Light Chain Kinase, Induces Apoptosis and Inhibits Diethylnitrosamine-Induced Liver Tumorigenesis in Rats
Int. J. Mol. Sci. 2013, 14(1), 1940-1951; https://doi.org/10.3390/ijms14011940
Received: 7 November 2012 / Revised: 27 December 2012 / Accepted: 31 December 2012 / Published: 17 January 2013
Cited by 10 | Viewed by 3197 | PDF Full-text (534 KB) | HTML Full-text | XML Full-text
Abstract
Hepatocellular carcinoma (HCC) is a serious healthcare problem worldwide because of its increasing morbidity and high mortality rates. However, our understanding of the mechanism of liver tumorigenesis remains incomplete. We report the expression of myosin light chain kinase (MLCK) in the livers of [...] Read more.
Hepatocellular carcinoma (HCC) is a serious healthcare problem worldwide because of its increasing morbidity and high mortality rates. However, our understanding of the mechanism of liver tumorigenesis remains incomplete. We report the expression of myosin light chain kinase (MLCK) in the livers of rats with diethylnitrosamine (DENA)-induced HCC and investigated the correlation between MLCK and liver tumorigenesis by observing the expression of MLCK in a rat model of HCC. HCC was induced in rats by an intraperitoneal injection of DENA, and resveratrol-treated rats were orally administered resveratrol with 50 mg/kg body weight/day. The livers of rats were excised after 20 weeks and immersed in 10% formaldehyde prior to immunohistochemical and Western blot analyses for determining the level of MLCK expression. These analyses indicated that the MLCK expression was higher in the livers of HCC rats than in normal and resveratrol-treated rats. High level of MLCK expression was responsible for proliferation and anti-apoptotic effects. However, resveratrol down-regulated the expression of MLCK, which induced cell apoptosis and inhibited liver tumorigenesis in rats with DENA-induced HCC. Our results suggest that the over expression of MLCK may be related to the development of liver tumorigenesis. Full article
Open AccessArticle
Neuroprotective Effects of Ultra-Low-Molecular-Weight Heparin on Cerebral Ischemia/Reperfusion Injury in Rats: Involvement of Apoptosis, Inflammatory Reaction and Energy Metabolism
Int. J. Mol. Sci. 2013, 14(1), 1932-1939; https://doi.org/10.3390/ijms14011932
Received: 16 October 2012 / Revised: 21 December 2012 / Accepted: 26 December 2012 / Published: 17 January 2013
Cited by 13 | Viewed by 3059 | PDF Full-text (262 KB) | HTML Full-text | XML Full-text
Abstract
Previous experiments showed that ultra-low-molecular-weight heparin (ULMWH) reduced the infarct and neurologic deficit in rats followed by transient cerebral ischemia, but the mechanisms of its neuroprotective effect are unclear. This study reported the effect of ULMWH on energy metabolism, inflammatory reaction and neuronal [...] Read more.
Previous experiments showed that ultra-low-molecular-weight heparin (ULMWH) reduced the infarct and neurologic deficit in rats followed by transient cerebral ischemia, but the mechanisms of its neuroprotective effect are unclear. This study reported the effect of ULMWH on energy metabolism, inflammatory reaction and neuronal apoptosis. Male Wistar rats were subjected to middle cerebral artery occlusion (MCAO) for 2 h followed by reperfusion for 24 h. ULMWH (0.5, 1 mg/kg, i.v.) was administered after the MCAO and reperfusion. 24 h after the reperfusion, Spectrophotometric assay was used to determine the activity of ATPase and the content of lactic acid in the brain. The ICAM-1 and Caspase-3 genes were investigated by RT-PCR. Furthermore, the apoptotic percentage of cells in hippocampus was quantified by flow cytometry. Compared with the model group, ULMWH significantly decreased lactic acid content and increased ATPase activity in ischemic brain. At the same time, ULMWH inhibited the neural apoptosis and decreased the expressions of ICAM-1 and Caspase-3 mRNA in hippocampus. These findings suggest that ULMWH exhibits a neuroprotective effect against cerebral ischemia/reperfusion injury, partly through improving energy metabolism, inhibiting apoptosis and attenuating inflammatory reaction. Full article
(This article belongs to the Section Biochemistry)
Open AccessReview
Nanostructured Surfaces of Dental Implants
Int. J. Mol. Sci. 2013, 14(1), 1918-1931; https://doi.org/10.3390/ijms14011918
Received: 9 October 2012 / Revised: 21 December 2012 / Accepted: 4 January 2013 / Published: 17 January 2013
Cited by 38 | Viewed by 4517 | PDF Full-text (473 KB) | HTML Full-text | XML Full-text
Abstract
The structural and functional fusion of the surface of the dental implant with the surrounding bone (osseointegration) is crucial for the short and long term outcome of the device. In recent years, the enhancement of bone formation at the bone-implant interface has been [...] Read more.
The structural and functional fusion of the surface of the dental implant with the surrounding bone (osseointegration) is crucial for the short and long term outcome of the device. In recent years, the enhancement of bone formation at the bone-implant interface has been achieved through the modulation of osteoblasts adhesion and spreading, induced by structural modifications of the implant surface, particularly at the nanoscale level. In this context, traditional chemical and physical processes find new applications to achieve the best dental implant technology. This review provides an overview of the most common manufacture techniques and the related cells-surface interactions and modulation. A Medline and a hand search were conducted to identify studies concerning nanostructuration of implant surface and their related biological interaction. In this paper, we stressed the importance of the modifications on dental implant surfaces at the nanometric level. Nowadays, there is still little evidence of the long-term benefits of nanofeatures, as the promising results achieved in vitro and in animals have still to be confirmed in humans. However, the increasing interest in nanotechnology is undoubted and more research is going to be published in the coming years. Full article
(This article belongs to the Special Issue Bioactive Nanoparticles 2012)
Open AccessReview
Neuroprotection for Ischemic Stroke: Moving Past Shortcomings and Identifying Promising Directions
Int. J. Mol. Sci. 2013, 14(1), 1890-1917; https://doi.org/10.3390/ijms14011890
Received: 9 November 2012 / Revised: 4 January 2013 / Accepted: 10 January 2013 / Published: 17 January 2013
Cited by 18 | Viewed by 4158 | PDF Full-text (408 KB) | HTML Full-text | XML Full-text
Abstract
The translation of neuroprotective agents for ischemic stroke from bench-to-bedside has largely failed to produce improved treatments since the development of tissue plasminogen activator (tPA). One possible reason for lack of translation is the failure to acknowledge the greatest risk factor for stroke, [...] Read more.
The translation of neuroprotective agents for ischemic stroke from bench-to-bedside has largely failed to produce improved treatments since the development of tissue plasminogen activator (tPA). One possible reason for lack of translation is the failure to acknowledge the greatest risk factor for stroke, age, and other common comorbidities such as hypertension, obesity, and diabetes that are associated with stroke. In this review, we highlight both mechanisms of studying these factors and results of those that have been addressed. We also discuss the potential role of other lifestyle factors associated with an increased stroke risk such as sleep fragmentation and/or deprivation. Furthermore, many proposed therapeutic agents have targeted molecular mechanisms occurring soon after the onset of ischemia despite data indicating delayed patient presentation following ischemic stroke. Modulating inflammation has been identified as a promising therapeutic avenue consistent with preliminary success of ongoing clinical trials for anti-inflammatory compounds such as minocycline. We review the role of inflammation in stroke and in particular, the role of inflammatory cell recruitment and macrophage phenotype in the inflammatory process. Emerging evidence indicates an increasing role of neuro-immune crosstalk, which has led to increased interest in identification of peripheral biomarkers indicative of neural injury. It is our hope that identification and investigation of factors influencing stroke pathophysiology may lead to improved therapeutics. Full article
Open AccessArticle
Sericin Enhances the Bioperformance of Collagen-Based Matrices Preseeded with Human-Adipose Derived Stem Cells (hADSCs)
Int. J. Mol. Sci. 2013, 14(1), 1870-1889; https://doi.org/10.3390/ijms14011870
Received: 22 October 2012 / Revised: 23 December 2012 / Accepted: 25 December 2012 / Published: 16 January 2013
Cited by 21 | Viewed by 4124 | PDF Full-text (3223 KB) | HTML Full-text | XML Full-text
Abstract
Current clinical strategies for adipose tissue engineering (ATE), including autologous fat implants or the use of synthetic surrogates, not only are failing in the long term, but also can’t face the latest requirements regarding the aesthetic restoration of the resulted imperfections. In this [...] Read more.
Current clinical strategies for adipose tissue engineering (ATE), including autologous fat implants or the use of synthetic surrogates, not only are failing in the long term, but also can’t face the latest requirements regarding the aesthetic restoration of the resulted imperfections. In this context, modern strategies in current ATE applications are based on the implantation of 3D cell-scaffold bioconstructs, designed for prospective achievement of in situ functional de novo tissue. Thus, in this paper, we reported for the first time the evaluation of a spongious 60% collagen and 40% sericin scaffold preseeded with human adipose-derived stem cells (hADSCs) in terms of biocompatibility and adipogenic potential in vitro. We showed that the addition of the sticky protein sericin in the composition of a classical collagen sponge enhanced the adhesion and also the proliferation rate of the seeded cells, thus improving the biocompatibility of the novel scaffold. In addition, sericin stimulated PPARγ2 overexpression, triggering a subsequent upregulated expression profile of FAS, aP2 and perilipin adipogenic markers. These features, together with the already known sericin stimulatory potential on cellular collagen production, promote collagen-sericin biomatrix as a good candidate for soft tissue reconstruction and wound healing applications. Full article
(This article belongs to the Section Biochemistry)
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Open AccessArticle
Quaternized Chitosan as an Antimicrobial Agent: Antimicrobial Activity, Mechanism of Action and Biomedical Applications in Orthopedics
Int. J. Mol. Sci. 2013, 14(1), 1854-1869; https://doi.org/10.3390/ijms14011854
Received: 3 December 2012 / Revised: 8 January 2013 / Accepted: 9 January 2013 / Published: 16 January 2013
Cited by 116 | Viewed by 5486 | PDF Full-text (275 KB) | HTML Full-text | XML Full-text
Abstract
Chitosan (CS) is a linear polysaccharide with good biodegradability, biocompatibility and antimicrobial activity, which makes it potentially useful for biomedical applications, including an antimicrobial agent either alone or blended with other polymers. However, the poor solubility of CS in most solvents at neutral [...] Read more.
Chitosan (CS) is a linear polysaccharide with good biodegradability, biocompatibility and antimicrobial activity, which makes it potentially useful for biomedical applications, including an antimicrobial agent either alone or blended with other polymers. However, the poor solubility of CS in most solvents at neutral or high pH substantially limits its use. Quaternary ammonium CS, which was prepared by introducing a quaternary ammonium group on a dissociative hydroxyl group or amino group of the CS, exhibited improved water solubility and stronger antibacterial activity relative to CS over an entire range of pH values; thus, this quaternary modification increases the potential biomedical applications of CS in the field of anti-infection. This review discusses the current findings on the antimicrobial properties of quaternized CS synthesized using different methods and the mechanisms of its antimicrobial actions. The potential antimicrobial applications in the orthopedic field and perspectives regarding future studies in this field are also considered. Full article
(This article belongs to the Special Issue Antimicrobial Polymers)
Open AccessArticle
Synthesis of Bisindolylmethanes and Their Cytotoxicity Properties
Int. J. Mol. Sci. 2013, 14(1), 1843-1853; https://doi.org/10.3390/ijms14011843
Received: 19 November 2012 / Revised: 10 December 2012 / Accepted: 24 December 2012 / Published: 16 January 2013
Cited by 18 | Viewed by 2996 | PDF Full-text (221 KB) | HTML Full-text | XML Full-text
Abstract
Polymer supported dichlorophosphate (PEG-OPOCl2) is an efficient green catalyst for the electrophilic substitution reaction of indole with aromatic aldehydes, in neat condition, to afford an excellent yield of bis(indolyl) methanes with short reaction time, at room temperature. The synthesized compounds and [...] Read more.
Polymer supported dichlorophosphate (PEG-OPOCl2) is an efficient green catalyst for the electrophilic substitution reaction of indole with aromatic aldehydes, in neat condition, to afford an excellent yield of bis(indolyl) methanes with short reaction time, at room temperature. The synthesized compounds and their anti-cancer activity are evaluated. Full article
(This article belongs to the Section Materials Science)
Open AccessReview
Modulation of Cancer Traits by Tumor Suppressor microRNAs
Int. J. Mol. Sci. 2013, 14(1), 1822-1842; https://doi.org/10.3390/ijms14011822
Received: 20 November 2012 / Revised: 28 December 2012 / Accepted: 10 January 2013 / Published: 16 January 2013
Cited by 20 | Viewed by 3583 | PDF Full-text (687 KB) | HTML Full-text | XML Full-text
Abstract
MicroRNAs (miRNAs) are potent post-transcriptional regulators of gene expression. In mammalian cells, miRNAs typically suppress mRNA stability and/or translation through partial complementarity with target mRNAs. Each miRNA can regulate a wide range of mRNAs, and a single mRNA can be regulated by multiple [...] Read more.
MicroRNAs (miRNAs) are potent post-transcriptional regulators of gene expression. In mammalian cells, miRNAs typically suppress mRNA stability and/or translation through partial complementarity with target mRNAs. Each miRNA can regulate a wide range of mRNAs, and a single mRNA can be regulated by multiple miRNAs. Through these complex regulatory interactions, miRNAs participate in many cellular processes, including carcinogenesis. By altering gene expression patterns, cancer cells can develop specific phenotypes that allow them to proliferate, survive, secure oxygen and nutrients, evade immune recognition, invade other tissues and metastasize. At the same time, cancer cells acquire miRNA signature patterns distinct from those of normal cells; the differentially expressed miRNAs contribute to enabling the cancer traits. Over the past decade, several miRNAs have been identified, which functioned as oncogenic miRNAs (oncomiRs) or tumor-suppressive miRNAs (TS-miRNAs). In this review, we focus specifically on TS-miRNAs and their effects on well-established cancer traits. We also discuss the rising interest in TS-miRNAs in cancer therapy. Full article
(This article belongs to the Special Issue Non-Coding RNAs 2012)
Open AccessArticle
The Combination of Catechin and Epicatechin Gallate from Fructus Crataegi Potentiates β-Lactam Antibiotics Against Methicillin-Resistant Staphylococcus aureus (MRSA) in Vitro and in Vivo
Int. J. Mol. Sci. 2013, 14(1), 1802-1821; https://doi.org/10.3390/ijms14011802
Received: 8 October 2012 / Revised: 29 November 2012 / Accepted: 8 January 2013 / Published: 16 January 2013
Cited by 27 | Viewed by 4638 | PDF Full-text (775 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Fructus crataegi (hawthorn) is the common name of all plant species in the genus Crataegus of the Rosaceae family. In the present study, three monomers of (+)-catechin (C), (−)-epicatechin gallate (ECg) and (−)-epigallocatechin (EGC) were isolated from the hawthorn under the guide of [...] Read more.
Fructus crataegi (hawthorn) is the common name of all plant species in the genus Crataegus of the Rosaceae family. In the present study, three monomers of (+)-catechin (C), (−)-epicatechin gallate (ECg) and (−)-epigallocatechin (EGC) were isolated from the hawthorn under the guide of antibacterial sensitization activity. The bioactivity of the composite fraction in enhancing the antibacterial effect of oxacillin against methicillin-resistant Staphylococcus aureus (MRSA) was greater than that of the individual monomer of the hawthorn extract in vitro. Two-fold dilution and checkerboard methods were used to analyze antibacterial activity and screen for the combination and proportion of monomers with the best bioactivity. The result showed that C (128 mg/L) combined with ECg (16 mg/L) had the greatest effect and the combination also reduced the bacterial load in blood of septic mice challenged with a sublethal dose of MRSA, increased daunomycin accumulation within MRSA and down-regulated the mRNA expression of norA, norC and abcA, three important efflux pumps of MRSA. In summary, C and ECg enhanced the antibacterial effect of β-lactam antibiotics against MRSA in vitro and in vivo, which might be related to the increased accumulation of antibiotics within MRSA via suppression of important efflux pumps’ gene expression. Full article
(This article belongs to the Special Issue Plant-Derived Pharmaceuticals by Molecular Farming 2012)
Open AccessArticle
Modeling of Anopheles minimus Mosquito NADPH-Cytochrome P450 Oxidoreductase (CYPOR) and Mutagenesis Analysis
Int. J. Mol. Sci. 2013, 14(1), 1788-1801; https://doi.org/10.3390/ijms14011788
Received: 25 September 2012 / Revised: 19 November 2012 / Accepted: 5 January 2013 / Published: 16 January 2013
Cited by 6 | Viewed by 3832 | PDF Full-text (979 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Malaria is one of the most dangerous mosquito-borne diseases in many tropical countries, including Thailand. Studies in a deltamethrin resistant strain of Anopheles minimus mosquito, suggest cytochrome P450 enzymes contribute to the detoxification of pyrethroid insecticides. Purified A. minimus CYPOR enzyme (AnCYPOR), which [...] Read more.
Malaria is one of the most dangerous mosquito-borne diseases in many tropical countries, including Thailand. Studies in a deltamethrin resistant strain of Anopheles minimus mosquito, suggest cytochrome P450 enzymes contribute to the detoxification of pyrethroid insecticides. Purified A. minimus CYPOR enzyme (AnCYPOR), which is the redox partner of cytochrome P450s, loses flavin-adenosine di-nucleotide (FAD) and FLAVIN mono-nucleotide (FMN) cofactors that affect its enzyme activity. Replacement of leucine residues at positions 86 and 219 with phenylalanines in FMN binding domain increases FMN binding, enzyme stability, and cytochrome c reduction activity. Membrane-Bound L86F/L219F-AnCYPOR increases A. minimus P450-mediated pyrethroid metabolism in vitro. In this study, we constructed a comparative model structure of AnCYPOR using a rat CYPOR structure as a template. Overall model structure is similar to rat CYPOR, with some prominent differences. Based on primary sequence and structural analysis of rat and A. minimus CYPOR, C427R, W678A, and W678H mutations were generated together with L86F/L219F resulting in three soluble Δ55 triple mutants. The C427R triple AnCYPOR mutant retained a higher amount of FAD binding and increased cytochrome c reduction activity compared to wild-type and L86F/L219F-Δ55AnCYPOR double mutant. However W678A and W678H mutations did not increase FAD and NAD(P)H bindings. The L86F/L219F double and C427R triple membrane-bound AnCYPOR mutants supported benzyloxyresorufin O-deakylation (BROD) mediated by mosquito CYP6AA3 with a two- to three-fold increase in efficiency over wild-type AnCYPOR. The use of rat CYPOR in place of AnCYPOR most efficiently supported CYP6AA3-mediated BROD compared to all AnCYPORs. Full article
(This article belongs to the Special Issue Flavins)
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Open AccessArticle
Stable Isolation of Phycocyanin from Spirulina platensis Associated with High-Pressure Extraction Process
Int. J. Mol. Sci. 2013, 14(1), 1778-1787; https://doi.org/10.3390/ijms14011778
Received: 18 October 2012 / Revised: 17 December 2012 / Accepted: 7 January 2013 / Published: 16 January 2013
Cited by 18 | Viewed by 3990 | PDF Full-text (335 KB) | HTML Full-text | XML Full-text
Abstract
A method for stably purifying a functional dye, phycocyanin from Spirulina platensis was developed by a hexane extraction process combined with high pressure. This was necessary because this dye is known to be very unstable during normal extraction processes. The purification yield of [...] Read more.
A method for stably purifying a functional dye, phycocyanin from Spirulina platensis was developed by a hexane extraction process combined with high pressure. This was necessary because this dye is known to be very unstable during normal extraction processes. The purification yield of this method was estimated as 10.2%, whose value is 3%–5% higher than is the case from another conventional separation method using phosphate buffer. The isolated phycocyanin from this process also showed the highest purity of 0.909 based on absorbance of 2.104 at 280 nm and 1.912 at 620 nm. Two subunits of phycocyanin namely α-phycocyanin (18.4 kDa) and β-phycocyanin (21.3 kDa) were found to remain from the original mixtures after being extracted, based on SDS-PAGE analysis, clearly demonstrating that this process can stably extract phycocyanin and is not affected by extraction solvent, temperature, etc. The stability of the extracted phycocyanin was also confirmed by comparing its DPPH (α,α-diphenyl-β-picrylhydrazyl) scavenging activity, showing 83% removal of oxygen free radicals. This activity was about 15% higher than that of commercially available standard phycocyanin, which implies that the combined extraction method can yield relatively intact chromoprotein through absence of degradation. The results were achieved because the low temperature and high pressure extraction effectively disrupted the cell membrane of Spirulina platensis and degraded less the polypeptide subunits of phycocyanin (which is a temperature/pH-sensitive chromoprotein) as well as increasing the extraction yield. Full article
Open AccessArticle
Study of the UTMD-Based Delivery System to Induce Cervical Cancer Cell Apoptosis and Inhibit Proliferation with shRNA targeting Survivin
Int. J. Mol. Sci. 2013, 14(1), 1763-1777; https://doi.org/10.3390/ijms14011763
Received: 3 November 2012 / Revised: 4 January 2013 / Accepted: 6 January 2013 / Published: 16 January 2013
Cited by 12 | Viewed by 2619 | PDF Full-text (1469 KB) | HTML Full-text | XML Full-text
Abstract
Apoptosis induction by short hairpin RNA (shRNA) expression vectors could be an efficient and promising strategy for cancer gene therapy. Ultrasound-targeted microbubble destruction (UTMD) is an appealing technique. In this study, we investigated the apoptosis induction and suppression of cell proliferation in vivo [...] Read more.
Apoptosis induction by short hairpin RNA (shRNA) expression vectors could be an efficient and promising strategy for cancer gene therapy. Ultrasound-targeted microbubble destruction (UTMD) is an appealing technique. In this study, we investigated the apoptosis induction and suppression of cell proliferation in vivo transfected by the UTMD-based shRNA delivery system. Nude mice with transplanted tumors of cervical cancer were randomly arranged into three groups: control group, plasmid injection and ultrasound (P + US), P + UTMD group. Expressions of Survivin and proliferating cell nuclear antigen (PCNA), Bcl-2, Bax, Caspase-3, Ki-67, nucleostemin (NS) were investigated by immunohistochemistry. Furthermore, microvessel density (MVD) was detected by CD34 protein expressions and apoptotic index (AI) was measured by TUNEL. As compared with those in the control and P + US groups, protein expressions of PCNA, Ki-67, Bcl-2, Survivin and NS in P + UTMD groups were down-regulated markedly, while those of Bax, Caspase-3 were up-regulated significantly (p < 0.05). MVD decreased significantly, whereas AI increased remarkably (p < 0.05). We suggested that UTMD-based shRNA delivery system could induce apoptosis and inhibit proliferation significantly, without causing any apparently adverse effect, representing a new, promising technology that would be used in the future gene therapy and research. Full article
(This article belongs to the Section Biochemistry)
Open AccessArticle
Comparative Proteomic Analysis of Puccinellia tenuiflora Leaves under Na2CO3 Stress
Int. J. Mol. Sci. 2013, 14(1), 1740-1762; https://doi.org/10.3390/ijms14011740
Received: 17 October 2012 / Revised: 31 December 2012 / Accepted: 6 January 2013 / Published: 15 January 2013
Cited by 23 | Viewed by 3553 | PDF Full-text (675 KB) | HTML Full-text | XML Full-text
Abstract
Soil salt-alkalinization is a widespread environmental stress that limits crop growth and agricultural productivity. The influence of soil alkalization caused by Na2CO3 on plants is more severe than that of soil salinization. Plants have evolved some unique mechanisms to cope [...] Read more.
Soil salt-alkalinization is a widespread environmental stress that limits crop growth and agricultural productivity. The influence of soil alkalization caused by Na2CO3 on plants is more severe than that of soil salinization. Plants have evolved some unique mechanisms to cope with alkali stress; however, the plant alkaline-responsive signaling and molecular pathways are still unknown. In the present study, Na2CO3 responsive characteristics in leaves from 50-day-old seedlings of halophyte Puccinellia tenuiflora were investigated using physiological and proteomic approaches. Comparative proteomics revealed 43 differentially expressed proteins in P. tenuiflora leaves in response to Na2CO3 treatment for seven days. These proteins were mainly involved in photosynthesis, stress and defense, carbohydrate/energy metabolism, protein metabolism, signaling, membrane and transport. By integrating the changes of photosynthesis, ion contents, and stress-related enzyme activities, some unique Na2CO3 responsive mechanisms have been discovered in P. tenuiflora. This study provides new molecular information toward improving the alkali tolerance of cereals. Full article
(This article belongs to the collection Advances in Proteomic Research)
Open AccessArticle
Gamma-Aminobutyric Acid Production Using Immobilized Glutamate Decarboxylase Followed by Downstream Processing with Cation Exchange Chromatography
Int. J. Mol. Sci. 2013, 14(1), 1728-1739; https://doi.org/10.3390/ijms14011728
Received: 17 November 2012 / Revised: 1 January 2013 / Accepted: 2 January 2013 / Published: 15 January 2013
Cited by 23 | Viewed by 3077 | PDF Full-text (571 KB) | HTML Full-text | XML Full-text
Abstract
We have developed a gamma-aminobutyric acid (GABA) production technique using his-tag mediated immobilization of Escherichia coli-derived glutamate decarboxylase (GAD), an enzyme that catalyzes the conversion of glutamate to GABA. The GAD was obtained at 1.43 g/L from GAD-overexpressed E. coli fermentation and [...] Read more.
We have developed a gamma-aminobutyric acid (GABA) production technique using his-tag mediated immobilization of Escherichia coli-derived glutamate decarboxylase (GAD), an enzyme that catalyzes the conversion of glutamate to GABA. The GAD was obtained at 1.43 g/L from GAD-overexpressed E. coli fermentation and consisted of 59.7% monomer, 29.2% dimer and 2.3% tetramer with a 97.6% soluble form of the total GAD. The harvested GAD was immobilized to metal affinity gel with an immobilization yield of 92%. Based on an investigation of specific enzyme activity and reaction characteristics, glutamic acid (GA) was chosen over monosodium glutamate (MSG) as a substrate for immobilized GAD, resulting in conversion of 2.17 M GABA in a 1 L reactor within 100 min. The immobilized enzymes retained 58.1% of their initial activities after ten consecutive uses. By using cation exchange chromatography followed by enzymatic conversion, GABA was separated from the residual substrate and leached GAD. As a consequence, the glutamic acid was mostly removed with no detectable GAD, while 91.2% of GABA was yielded in the purification step. Full article
(This article belongs to the Section Biochemistry)
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