The newly discovered aer locus of Escherichia coli encodes a 506-residue protein with an N terminus that resembles the NifL aerosensor and a C terminus that resembles the flagellar signaling domain of methylaccepting chemoreceptors. Deletion mutants lacking a functional Aer protein failed to congregate around air bubbles or follow oxygen gradients in soft agar plates. Membranes with overexpressed Aer protein also contained high levels of noncovalently associated flavin adenine dinucleotide (FAD). We propose that Aer is a flavoprotein that mediates positive aerotactic responses in E. coli. Aer may use its FAD prosthetic group as a cellular redox sensor to monitor environmental oxygen levels.Aerotaxis, the movement of a cell or organism toward or away from oxygen, was first described in bacteria more than a century ago by Engelmann (8), Pfeffer (24), and Beijerinck (3), who observed accumulation of cells near air bubbles or other sources of oxygen. Despite considerable study, particularly in Escherichia coli (22,28,30), the molecular mechanism underlying this behavior has remained elusive. Does the organism detect oxygen directly, or does it instead sense some metabolic consequence of different oxygen environments, such as changes in electron transport activity (19), cellular redox potential (4), or proton motive force (20, 32)? We describe here a gene, dubbed aer for aerotaxis, that encodes a likely flavoprotein signal transducer for aerotaxis in E. coli. Studies of the Aer protein promise to provide a definitive answer to the longstanding puzzle of how cells detect oxygen gradients during aerotaxis.Sequence features of the aer locus. We initially identified the aer gene as an open reading frame (ORF506) discovered in the E. coli genome sequencing project (7). Its conceptual translation product, a 506-amino-acid Aer protein, exhibits several hallmarks of an aerosensing function (Fig. 1). Aer residues 10 to 110 are similar to parts of NifL, FixL, and related bacterial proteins that trigger regulatory responses to changes in environmental oxygen levels (5, 11). Residues 168 to 209 are predominantly hydrophobic and could serve to anchor Aer in the cytoplasmic membrane. Aer residues 259 to 506 are about 50% identical to the cytoplasmic signaling domains of methyl-accepting chemotaxis proteins (MCPs), the principal chemoreceptors of E. coli (14). These features suggested that Aer might generate chemotactic signals in response to oxygen gradients.Construction of an aer mutant. We constructed a large inframe deletion lacking codons 5 to 505 of the aer coding region by PCR amplification of chromosomal sequences flanking the aer locus in strain RP437 (23) by using primer pairs NSB19-NSB20 and NSB25-NSB22 (Fig. 1). The two PCR fragments were ligated at their common XbaI site, joining aer codon 4 to codon 506, and inserted into the pMAK705 vector, whose replication is temperature-sensitive (13), producing plasmid pSB25. RP437 carrying pSB25 was grown at 44°C to select recombinational insertions and then at 30°C for recombin...
The influence of tethering silicon microelectrode arrays on the cortical brain tissue reaction was compared with that of untethered implants placed in the same location by identical means using immunoflourescent methods and cell type specific markers over indwelling periods of 1-4 weeks. Compared with untethered, freely floating implants, tethered microelectrodes elicited significantly greater reactivity to antibodies against ED1 and GFAP over time. Regardless of implantation method or indwelling time, retrieved microelectrodes contained a layer of attached macrophages identified by positive immunoreactivity against ED1. In the tethered condition and in cases where the tissue surrounding untethered implants had the highest levels of ED1+ and GFAP+ immunoreactivity, the neuronal markers for neurofilament 160 and NeuN were reduced. Although the precise mechanisms are unclear, the present study indicates that simply tethering silicon microelectrode arrays to the skull increases the cortical brain tissue response in the recording zone immediately surrounding the microelectrode array, which signals the importance of identifying this important variable when evaluating the tissue response of different device designs, and suggests that untethered or wireless devices may elicit less of a foreign body response.
Blood contact with biomaterials triggers activation of multiple reactive mechanisms that can impair the performance of implantable medical devices and potentially cause serious adverse clinical events. This includes thrombosis and thromboembolic complications due to activation of platelets and the coagulation cascade, activation of the complement system, and inflammation. Numerous surface coatings have been developed to improve blood compatibility of biomaterials. For more than thirty years, the anticoagulant drug heparin has been employed as a covalently immobilized surface coating on a variety of medical devices. This review describes the fundamental principles of non-eluting heparin coatings, mechanisms of action, and clinical applications with focus on those technologies which have been commercialized. Because of its extensive publication history, there is emphasis on the CARMEDA BioActive Surface (CBAS Heparin Surface), a widely used commercialized technology for the covalent bonding of heparin.
Nerve outgrowth in the developing nervous system utilizes a variety of attractive and repulsive molecules found in the extracellular environment. In addition, physical cues may play an important regulatory role in determining directional outgrowth of nervous tissue. Here, by culturing nerve cells on filamentous surfaces and measuring directional growth, we tested the hypothesis that substrate curvature is sufficient to influence the directional outgrowth of nerve cells. We found that the mean direction of neurite outgrowth aligned with the direction of minimum principle curvature, and the spatial variance in outgrowth direction was directly related to the maximum principle curvature. As substrate size approached the size of an axon, adherent neurons extended processes that followed the direction of the long axis of the substrate similar to what occurs during development along pioneering axons and radial glial fibers. A simple Boltzmann model describing the interplay between adhesion and bending stiffness of the nerve process was found to be in close agreement with the data suggesting that cell stiffness and substrate curvature can act together in a manner that is sufficient to direct nerve outgrowth in the absence of contrasting molecular cues. The study highlights the potential importance of cellular level geometry as a fidelity-enhancing cue in the developing and regenerating nervous system.
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