Growth and behavior of chondrocytes on nano engineered surfaces and construction of micropatterned co-culture platforms using layer-by-layer platforms using layer -by -layer assembly lift-off method

Several approaches such as self-assembled monolayers and layer-by-layer assembled multilayer films are being used as tools to study the interactions of cells with biomaterials in vitro. In this study, the layer-by-layer assembly approach was used to create monolayer, bilayer, trilayer, five, ten and twenty-bilayer beds of eleven different biomaterials. The various biomaterials used were poly(styrene-sulfonate), fibronectin, poly-L-lysine, poly-D-lysine, laminin, bovine serum albumin, chondroitin sulfate, poly(ethyleneimine), polyethylene glycol amine, collagen and poly(dimethyldiallyl-ammonium chloride) with unmodified tissue-culture polystyrene as standard control. Three different cell lines—primary bovine articular chondrocytes, and two secondary cell lines, human chondrosarcoma cells and canine chondrocytes were used in these studies. Chondrocyte morphology and attachment, viability, proliferation, and functionality were determined using bright field microscopy, the Live/Dead viability assay, MTT assay, and immunocytochemistry, respectively. Atomic force microscopy of the nanofilms indicated an increase in surface roughness with increasing number of layers. The most important observations from the studies on primary bovine articular chondrocytes were that these cells exhibited increasing viability and cell metabolic activity with increasing number of bilayers. The increase in viability was more pronounced than the increase in cell metabolic activity. Also, bovine chondrocytes on bilayers of poly(dimethyldiallyl-ammonium chloride, poly-L-lysine, poly(styrene-sulfonate), and bovine serum albumin were substantially bigger in size and well-attached when compared to the cells grown on monolayer and trilayers. Lactate dehydrogenase assay performed on chondrosarcoma cells grown on 5- and 10-bilayer multilayer beds indicated that the 10-bilayer beds had reduced cytotoxicity compared to the 5-bilayer beds. MTT assay performed on canine chondrocytes grown on 5-, 10-, and 20-bilayer nanofilm beds revealed increasing cell metabolic activity for BSA with increasing bilayers. Micropatterned multilayer beds having poly-L-lysine, poly-D-lysine, laminin poly(dimethyldiallyl-ammonium chloride) and poly(ethyleneimine) as the terminating layers were fabricated using the Layer-by-layer Lift-off (LbL-LO) method that combines photolithography and LbL self-assembly. Most importantly, micropatterned co-culture platforms consisting of anti-CD 44 rat monoclonal and anti-rat osteopontin (MPIIIB101) antibodies were constructed using the LbL-LO method for the first time. These co-culture platforms have several applications especially for studies of stem and progenitor cells. Co-culture platforms exhibiting spatiotempora-based differentiation can be built with LbL-LO for the differentiation of stem cells into the desired cell lineage

A Proficient Method For High Eminence And Cohesive Relevant Phrase Mining

A sentence is an integral unit of semantic nature, context and significance. Visualizing sentences for each topic is an important way to investigate and interpret unstructured corporate texts in subject modeling. Usually the term mining method is double: mining phrases and modeling theme. Current methods also suffer from order-sensitive and improper segmentation problems for phrase mining, which often lead to phrases of low content. The limitations of sentences, which may undermine continuity, are not entirely taken into account by standard topic models for topic modeling. In addition, current methods are frequently subject to domain terminology loss as the effect of topical domain dissemination is disregarded. We suggest an effective approach for high-quality and coherent topical sentence mining in this article. A high-quality sentence must meet the requirements for frequency, phrasing, integrity and suitability. In order to increase the both phrase consistency and topical cohesion, we combine the quality assured phrase mining process, a novel subject models that incorporate phrasing restriction, and a novel text clustering method into an iterative system. Effective algorithm designs to perform these methods effectively are often defined

Diagnosis of Diabetic Retinopathy

ABSTRACT Diabetic retinopathy is a disease, caused by alternation in the retinal blood vessels. It is a strong sign of early blindness and if it is not treated may tend to complete blindness and the vision lost once cannot be restored once again. In this paper different image processing techniques are used to differentiate between the normal and the diseased image. The attempt is made to see where the problem actually lies so that proper diagnosis of patient can be done. Pre processing of an image, optic disk detection, Blood vessels extraction, Exudates detection are some of the methods that are applied here. Other algorithms are designed to obtain the desired result. A large number of populations are affected by this disease around the world

Induced pluripotent stem cell modelling of HLHS underlines the contribution of dysfunctional NOTCH signalling to impaired cardiogenesis

Hypoplastic left heart syndrome (HLHS) is among the most severe forms of congenital heart disease. Although the consensus view is that reduced flow through the left heart during development is a key factor in the development of the condition, the molecular mechanisms leading to hypoplasia of left heart structures are unknown. We have generated induced pluripotent stem cells (iPSC) from five HLHS patients and two unaffected controls, differentiated these to cardiomyocytes and identified reproducible in vitro cellular and functional correlates of the HLHS phenotype. Our data indicate that HLHS-iPSC have a reduced ability to give rise to mesodermal, cardiac progenitors and mature cardiomyocytes and an enhanced ability to differentiate to smooth muscle cells. HLHS-iPSC-derived cardiomyocytes are characterised by a lower beating rate, disorganised sarcomeres and sarcoplasmic reticulum and a blunted response to isoprenaline. Whole exome sequencing of HLHS fibroblasts identified deleterious variants in NOTCH receptors and other genes involved in the NOTCH signalling pathway. Our data indicate that the expression of NOTCH receptors was significantly downregulated in HLHS-iPSC-derived cardiomyocytes alongside NOTCH target genes confirming downregulation of NOTCH signalling activity. Activation of NOTCH signalling via addition of Jagged peptide ligand during the differentiation of HLHS-iPSC restored their cardiomyocyte differentiation capacity and beating rate and suppressed the smooth muscle cell formation. Together, our data provide firm evidence for involvement of NOTCH signalling in HLHS pathogenesis, reveal novel genetic insights important for HLHS pathology and shed new insights into the role of this pathway during human cardiac developmen

Chondrocyte Behavior on Micropatterns Fabricated Using Layer-by-Layer Lift-Off: Morphological Analysis

Cell patterning has emerged as an elegant tool in developing cellular arrays, bioreactors, biosensors, and lab-on-chip devices and for use in engineering neotissue for repair or regeneration. In this study, micropatterned surfaces were created using the layer-by-layer lift-off (LbL-LO) method for analyzing canine chondrocytes response to patterned substrates. Five materials were chosen based on our previous studies. These included: poly(dimethyldiallylammonium chloride) (PDDA), poly(ethyleneimine) (PEI), poly(styrene sulfonate) (PSS), collagen, and chondroitin sulfate (CS). The substrates were patterned with these five different materials, in five and ten bilayers, resulting in the following multilayer nanofilm architectures: (PSS/PDDA)5, (PSS/PDDA)10; (CS/PEI)4/CS, (CS/PEI)9/CS; (PSS/PEI)5, (PSS/PEI)10; (PSS/Collagen)5, (PSS/Collagen)10; (PSS/PEI)4/PSS, (PSS/PEI)9/PSS. Cell characterization studies were used to assess the viability, longevity, and cellular response to the configured patterned multilayer architectures. The cumulative cell characterization data suggests that cell viability, longevity, and functionality were enhanced on micropatterned PEI, PSS, collagen, and CS multilayer nanofilms suggesting their possible use in biomedical applications

Green Synthesis and Characterization of Palladium Nanoparticles Using Origanum vulgare L. Extract and Their Catalytic Activity

The synthesis of Palladium (Pd) nanoparticles by green methods has attracted remarkable attention in recent years because of its superiority above chemical approaches, owing to its low cost and ecological compatibility. In this present work, we describe a facile and environmentally friendly synthesis of Pd nanoparticles (Pd NPs) using an aqueous extract of aerial parts of Origanum vulgare L. (OV) as a bioreductant. This plant is available in many parts of the world as well as in Saudi Arabia and is known to be a rich source of phenolic components, a feature we fruitfully utilized in the synthesis of Pd NPs, using various concentrations of plant extracts. Moreover, the OV extract phytomolecules are not only accountable for the reduction and progression of nanoparticles, but they also act as stabilizing agents, which was confirmed by several characterization methods. The as-synthesized Pd nanoparticles (Pd NPs) were analyzed using ultraviolet-visible spectroscopy (UV-Vis), Fourier-transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), transmission electron microscopy (TEM), energy-dispersive X-ray spectroscopy (EDX), and thermal gravimetric analysis (TGA). Further, FT-IR study has proven that the OV not merely represents a bioreductant but also functionalizes the nanoparticles. Furthermore, the green synthesized metallic Pd NPs were successfully applied as catalysts for selective oxidation of alcohols