Native chick laminin-4 containing the beta 2 chain (s-laminin) promotes motor axon growth.

After denervation of muscle, motor axons reinnervate original synaptic sites. A recombinant fragment of the synapse specific laminin beta 2 chain (s-laminin) was reported to inhibit motor axon growth. Consequently, a specific sequence (leucine-arginine-glutamate, LRE) of the laminin beta 2 chain was proposed to act as a stop signal and to mediate specific reinnervation at the neuromuscular junction (Porter, B.E., J. Weis, and J.R. Sanes. 1995. Neuron. 14:549-559). We demonstrate here that native chick laminin-4, which contains the beta 2 chain and is present in the synaptic basement membrane, does not inhibit but rather promotes motor axon growth. In native heterotrimeric laminin, the LRE sequence of the beta 2 chain is found in a triple coiled-coil region that is formed by all three subunits. We show here that the effect of LRE depends on the structural context. Whereas a recombinant randomly coiled LRE peptide indeed inhibited outgrowth by chick motoneurons, a small recombinant triple coiled-coil protein containing this sequence did not

Neural networks on chemically patterned "cultured probe" electrode arrays: network growth and activity patterns

A 'cultured probe' is a hybrid type of neural information transducer or prosthesis, for stimulation and/or recording of neural activity in the brain or the spinal cord (ventral motor region or dorsal sensory region). It consists of a micro electrode array (MEA) on a planar substrate, each electrode being covered and surrounded by a locally confined network of cultured neurons, obtained by chemical patterning of the substrate. The purpose of the cultured cells is that they act as intermediates for collateral sprouts from the in vivo system, thus allowing for an effective and selective neuron electrode interface. As the local neural network will become spontaneously active and has the capability of information processing, one may envisage future applications of these intermediary networks as 'front-end' signal processors. Two aspects of the development of this kind of cultured probe device are described. First, it is shown how substrates can be chemically modified to confine developing networks, cultured from dissociated rat cortex cells, to the surrounding of an electrode site. Secondly, the paper presents results on neuronal activity in such confined, circular networks and synchronized activity between two such interconnected networks

A parallel-plate flow chamber to study initial cell adhesion on a nanofeatured surface

Cells in the human body come across many types of information, which they respond to. Both material chemistry and topography of the surface where they adhere have an effect on cell shape, proliferation, migration, and gene expression. It is possible to create surfaces with topography at the nanometric scale to allow observation of cell-topography interactions. Previous work has shown that 100-nm-diameter pits on a 300-nm pitch can have a marked effect in reducing the adhesion of rat fibroblasts in static cultures. In the present study, a flow of cell suspension was used to investigate cell adhesion onto nanopits in dynamic conditions, by means of a parallel-plate flow chamber. A flow chamber with inner nanotopography has been designed, which allows real-time observation of the flow over the nanopits. A nanopitted pattern was successfully embossed into polymethylmethacrylate to meet the required shape of the chamber. Dynamic cell adhesion after 1 h has been quantified and compared on flat and nanopitted polymethylmethacrylate substrates. The nanopits were seen to be significantly less adhesive than the flat substrates (p<0.001), which is coherent with previous observations of static cultures

Development of a DC-DC buck boost converter using fuzzy logic control

A fuzzy controller of DC-DC Buck-boost converter is designed and presented in this project. In order to control the output voltage of the buck-boost converter, the controller is designed to change the duty cycle of the converter. The mathematical model of buckboost converter and fuzzy logic controller are derived to design simulation model. The simulation is developed on Matlab simulation program. To verity the effectiveness of the simulation model, an experimental set up is developed. The buck-boost circuit with mosfet as a switching component is developed. The fuzzy logic controller to generate duty cycle of PWM signal is programmed. The simulation and experimental results show that the output voltage of the buck-boost converter can be controlled according to the value of duty cycl

Adhesion and growth of electrically-active cortical neurons on polyethyleimine patterns microprinted on PEO-PPO-PEO triblockcopolymer-coated hydrophobic surfaces

This paper describes the adhesion and growth of dissociated cortical neurons on chemically patterned surfaces over a time period of 30 days. The presence of neurons was demonstrated by measurement of spontaneous bioelectrical activity on a micropatterned multielectrode array. Chemical patterns were prepared with a combination of neurophobic layers of polyethylenoxide-polypropylenoxide-polyethylenoxide (PEO-PPO-PEO) triblockcopolymers adsorbed onto hydrophobic surfaces and neurophilic microprinted tracks of polyethylenimine (PEI). Results showed that commercially available PEO-PPO-PEO triblockcopolymers F108 and F127 (Synperonics, ICI) significantly reduced the adhesion of neuronal tissue when adsorbed on hydrophobic Polyimide (PI) and Fluorocarbon (FC) surfaces over a time period of eight days. In general, both F108- and F127-coated PI displayed equal or better neurophobic background properties after 30 days. Viability of neuronal tissue after 30 days on PEI microprinted F108- and F127-coated PI was comparable with relatively high viability factors between 0.9 and 1 (scale from 0 to 1). Summarizing, the strategy to combine the neurophobic adsorbed triblock-copolymers F108 and F127 onto hydrophobic surfaces with neurophilic microprinted PEI resulted in relatively long-term neuronal pattern preservation with high numbers of viable neurons present after 30 days

Template assisted surface micro microstructuring of flowable dental composites and its effect on the microbial adhesion properties

Despite their various advantages, such as good esthetic properties, absence of mercury and adhesive bonding to teeth, modern dental composites still have some drawbacks, e.g., a relatively high rate of secondary caries on teeth filled with composite materials. Recent research suggests that microstructured biomaterials surfaces may reduce microbial adhesion to materials due to unfavorable physical material–microbe interactions. The objectives of this study were, therefore, to test the hypotheses that (i) different surface microstructures can be created on composites by a novel straightforward approach potentially suitable for clinical application and (ii) that these surface structures have a statistically significant effect on microbial adhesion properties.Peer ReviewedPostprint (author's final draft

Differentiation of Human Embryonic Stem Cells into Cells with Corneal Keratocyte Phenotype

Corneal transparency depends on a unique extracellular matrix secreted by stromal keratocytes, mesenchymal cells of neural crest lineage. Derivation of keratocytes from human embryonic stem (hES) cells could elucidate the keratocyte developmental pathway and open a potential for cell-based therapy for corneal blindness. This study seeks to identify conditions inducing differentiation of pluripotent hES cells to the keratocyte lineage. Neural differentiation of hES cell line WA01(H1) was induced by co-culture with mouse PA6 fibroblasts. After 6 days of co-culture, hES cells expressing cell-surface NGFR protein (CD271, p75NTR) were isolated by immunoaffinity adsorption, and cultured as a monolayer for one week. Keratocyte phenotype was induced by substratum-independent pellet culture in serum-free medium containing ascorbate. Gene expression, examined by quantitative RT-PCR, found hES cells co-cultured with PA6 cells for 6 days to upregulate expression of neural crest genes including NGFR, SNAI1, NTRK3, SOX9, and MSX1. Isolated NGFR-expressing cells were free of PA6 feeder cells. After expansion as a monolayer, mRNAs typifying adult stromal stem cells were detected, including BMI1, KIT, NES, NOTCH1, and SIX2. When these cells were cultured as substratum-free pellets keratocyte markers AQP1, B3GNT7, PTDGS, and ALDH3A1 were upregulated. mRNA for keratocan (KERA), a cornea-specific proteoglycan, was upregulated more than 10,000 fold. Culture medium from pellets contained high molecular weight keratocan modified with keratan sulfate, a unique molecular component of corneal stroma. These results show hES cells can be induced to differentiate into keratocytes in vitro. Pluripotent stem cells, therefore, may provide a renewable source of material for development of treatment of corneal stromal opacities. © 2013 Chan et al