Magnetic nanoparticles containing labeling reagents for cell surface mapping

Cell surface proteins play an important role in understanding cell-cell communication, cell signaling pathways, cell division and molecular pathogenesis in various diseases. Commonly used biotinylation regents for cell surface mapping have shown some potential drawbacks such as crossing the cell membrane, difficult recovery of biotinylated proteins from streptavidin/avidin beads, interference from endogenous biotin and nonspecific nature of streptavidin. With aim to solve these problems, we introduced sulfo-N-hydroxysuccinimidyl (NHS) ester functionalized magnetic nanoparticles containing cleavable groups to label solvent exposed primary amine groups of proteins. Silica coated iron oxide magnetic nanoparticles (Fe3O4@SiO2 MNPs) were linked to NHS ester groups via a cleavable disulfide bond. Additionally, the superparamagnetic properties of Fe3O4@SiO2 MNPs facilitate efficient separation of the labeled peptides and removal of the detergent without any extra step of purification. In the last step, the disulfide bond between the labeled peptides and MNPs was cleaved to release the labeled peptides. The disulfide linked NHS ester modified Fe3O4@SiO2 MNPs were tested using a small peptide, and a model protein (bovine serum albumin) followed by liquid chromatography-tandem mass spectrometry analysis (LC-MS/MS) of labeled peptides. In the next step, disulfide linked, NHS ester modified Fe3O4@SiO2 MNPs (150 nm) successfully labeled the solvent exposed cell surface peptides of Saccharomyces cerevisae. Electron microscopic analysis confirmed the cell surface binding of NHS ester modified Fe3O4@SiO2 MNPs. Mass spectrometric analysis revealed the presence of 30 unique proteins containing 56 peptides. Another MNPs based labeling reagent was developed to target solvent exposed carboxyl acid residues of peptides and proteins. The surface of Fe3O4@SiO2 MNPs was modified with free amine groups via a disulfide bond. Solvent exposed carboxyl groups of ACTH 4-11 and BSA were labeled by using1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) chemistry. Upon cleaving the disulfide bond, labeled peptides were analyzed by LC-MS/MS. The MNPs containing labeling reagents offers specific labeling under physiological conditions and rapid magnetic separation of labeled peptides prior to mass spectrometric analysis. The ability of large Fe3O4@SiO2 MNPs to specifically attach to cell surface makes them a potential candidate to study the surface of variety of different cell types and complex proteins surrounded by lipid bilayer

A study of minimally invasive percutaneous plate osteosynthesis for tibial plateau fractures

Background: Fractures of proximal tibia involve a major weight-bearing joint and are serious injuries, which, if not treated well, result in functional impairment. To preserve normal knee function one must strive to maintain joint congruity, preserve the normal mechanical axis, ensure joint stability and restore a full range of motion. This is a formidable task to accomplish, especially in the face of associated medical conditions of the patients.Methods: In our study, 30 cases were selected. Selection of cases were done on the basis of X-rays. Schatzker type I, II, III, IV, V & VI included in study. Criteria for acceptable reduction 1) <5 mm of articular step; 2) <5 mm of articular depression. Each case is referred to one set of tibial plateau fracture, showing distribution of tibia plateau fractures that we treated with MIPPO. Clinical follow-up examination was performed at 4, 6, 10, 12 weeks and 3, 6 months. Clinico-radiological assessment was done at 3 month and grading was done. Patients were evaluated according to grading minimum 3 months after injury.Results: Our study of 30 tibial plateau fractures confirms that the MIPPO technique is an excellent treatment modality in case of tibial plateau fractures. We observed these fractures mainly in age group of 30-40 years, which were involved in road traffic accident. Tibial plateau fractures seen in elder age group were mainly due to abnormal loading patterns on the leg. We have found oblique views very much informative especially for posterolateral or posteromedial displacement, articular depression which helps to plan the position and direction of screws to be used for fixation. Conclusions: In view of the excellent results obtained with this technique, we advocate MIPPO over conventional open reduction and internal fixation technique for tibial plateau fracture fixation

From academic research to marketing practice: Exploring the marketing science value chain

We aim to investigate the impact of marketing science articles and tools on the practice of marketing. This impact may be direct (e.g., an academic article may be adapted to solve a practical problem) or indirect (e.g., its contents may be incorporated into practitioners' tools, which then influence marketing decision making). We use the term "marketing science value chain" to describe these diffusion steps, and survey marketing managers, marketing science intermediaries (practicing marketing analysts), and marketing academics to calibrate the value chain.In our sample, we find that (1) the impact of marketing science is perceived to be largest on decisions such as the management of brands, pricing, new products, product portfolios, and customer/market selection, and (2) tools such as segmentation, survey-based choice models, marketing mix models, and pre-test market models have the largest impact on marketing decisions. Exemplary papers from 1982 to 2003 that achieved dual - academic and practice - impact are Guadagni and Little (1983) and Green and Srinivasan (1990). Overall, our results are encouraging. First, we find that the impact of marketing science has been largest on marketing decision areas that are important to practice. Second, we find moderate alignment between academic impact and practice impact. Third, we identify antecedents of practice impact among dual impact marketing science papers. Fourth, we discover more recent trends and initiatives in the period 2004-2012, such as the increased importance of big data and the rise of digital and mobile communication, using the marketing science value chain as an organizing framework

Radiological correlation with functional outcome in displaced proximal humerus fractures treated with locking plates: a prospective study of 35 cases

Background: The objective of this prospective study is to investigate the relationship between the functional outcome and the radiographic results of two-, three- and four part proximal humeral fractures treated with open reduction and fixation with PHILOS locking plate.Methods: 35 patients (30 males and 5 females) with displaced proximal humerus fracture were treated with PHILOS plating and studied prospectively. According to Neer’s classification there were 16 two part fracture (45%), 15 three part fracture (42%) and 4 four part fracture (13%). The healing of fracture, head shaft angle (HSA) and complications were evaluated radiologically with plain radiographs whereas the functional outcome was assessed by the Shoulder Pain and Disability Index (SPADI) at 6 weeks, 3 months, 6 months and 1 year after surgery.Results: In our study, patients treated with locking plates had mean forward flexion of 123°, abduction of 124°, mean external rotation of 30° and internal rotation 53° at 1 year follow-up. Maximum improvement in range of movement occurred between interval of 12 weeks to 24 weeks due to complete fracture union occurred approximately up to 12 weeks. The mean SPADI score at end of 1 year were 10.22, 10.38 and 10.20 for two, three and four part fractures respectively. Average postoperative and follow-up HSA's were 134.1±4.4 and 132.5±1.2 degrees, respectively. Varus progression was more prominent in patients with postoperative HSA <130 (p<0.001). Complications included impingement in 1 case, joint penetration by screw in 1 case, failure of fixation in 1 case and infection in 1 case.Conclusions: Fixed angle locked plate is an extremely useful implant for reconstruction and salvage of proximal humeral fractures. Precise surgical technique, stable fracture fixation and restoration of anatomical reduction are absolutely necessary for improved outcome

A Randomized Controlled Trial to Compare the Safety and Efficacy of Tadalafil and Tamsulosin in Relieving Double J Stent Related Symptoms

Objectives. To evaluate the safety and efficacy of Tadalafil and Tamsulosin in treating Double J stent related symptoms. Methods. In a prospective study, 161 patients with DJ related symptoms were randomized into 3 groups: Group A patients (54), Group B patients (53), and Group C patients (54). They were given Tadalafil, Tamsulosin, and placebo, respectively, at 1st week till removal of DJ stent at 3rd week. All patients completed Ureteral Stent Symptom Questionnaire (USSQ) at 1st week and at 3rd week. The statistical significant difference among groups was determined by the t-test, Kruskal-Wallis test and multivariate analysis were used to assess association of the variables within the three groups, and the level of significance was set at P<0.05. Results. Tadalafil and Tamsulosin were comparable in relieving urinary symptoms, general health, and work performance (OR = 0.65, 1.8, and 0.92). But Tadalafil was more effective in relieving body pain, sexual problems, and additional problems than Tamsulosin (OR = 5.95, 19.25, and 2.69) and was statistically significant as P<0.05. Conclusion. Tadalafil was as effective as Tamsulosin in relieving urinary symptom but more effective in relieving sexual symptoms and body pain

Signaling-Biased and Constitutively Active Dopamine D2 Receptor Variant

A dopamine D2 receptor mutation was recently identified in a family with a novel hyperkinetic movement disorder. Compared to the wild type D2 receptor, the novel allelic variant D2-I212F activates a Gαi1β1γ2 heterotrimer with higher potency and modestly enhanced basal activity in human embryonic kidney (HEK) 293 cells and has decreased capacity to recruit arrestin3. We now report that omitting overexpressed G protein-coupled receptor kinase-2 (GRK2) decreased the potency and efficacy of quinpirole for arrestin recruitment. The relative efficacy of quinpirole for arrestin recruitment to D2-I212F compared to D2-WT was considerably lower without overexpressed GRK2 than with added GRK2. D2-I212F exhibited higher basal activation of GαoA than Gαi1 but little or no increase in the potency of quinpirole relative to D2-WT. Other signs of D2-I212F constitutive activity for G protein-mediated signaling, in addition to basal activation of Gαi/o, were enhanced basal inhibition of forskolin-stimulated cyclic AMP accumulation that was reversed by the inverse agonists sulpiride and spiperone and a ∼4-fold increase in the apparent affinity of D2-I212F for quinpirole, determined from competition binding assays. In mouse midbrain slices, inhibition of tonic current by the inverse agonist sulpiride in dopamine neurons expressing D2-I212F was consistent with our hypothesis of enhanced constitutive activity and sensitivity to dopamine relative to D2-WT. Molecular dynamics simulations with D2 receptor models suggested that an ionic lock between the cytoplasmic ends of the third and sixth α-helices that constrains many G protein-coupled receptors in an inactive conformation spontaneously breaks in D2-I212F. Overall, these results confirm that D2-I212F is a constitutively active and signaling-biased D2 receptor mutant and also suggest that the effect of the likely pathogenic variant in a given brain region will depend on the nature of G protein and GRK expression

Development of an automated DNA purification module using a micro-fabricated pillar chip

We present a fully automated DNA purification module comprised of a micro-fabricated chip and sequential injection analysis system that is designed for use within autonomous instruments that continuously monitor the environment for the presence of biological threat agents. The chip has an elliptical flow channel containing a bed (3.5 &times; 3.5 mm) of silica-coated pillars with height, width and center-to-center spacing of 200, 15, and 30 &micro;m, respectively, which provides a relatively large surface area (ca. 3 cm2) for DNA capture in the presence of chaotropic agents. We have characterized the effect of various fluidic parameters on extraction performance, including sample input volume, capture flow rate, and elution volume. The flow-through design made the pillar chip completely reusable; carryover was eliminated by flushing lines with sodium hypochlorite and deionized water between assays. A mass balance was conducted to determine the fate of input DNA not recovered in the eluent. The device was capable of purifying and recovering Bacillus anthracis genomic DNA (input masses from 0.32 to 320 pg) from spiked environmental aerosol samples, for subsequent analysis using polymerase chain reaction-based assays.<br /