Direct real-time measurements of superoxide release from skeletal muscles in rat limbs and human blood platelets using an implantable Cytochrome C microbiosensor.

Oxidative stress and excessive accumulation of the superoxide (O2.-) anion are at the genesis of many pathological conditions and the onset of several diseases. The real time monitoring of (O2.-) release is important to assess the extent of oxidative stress in these conditions. Herein, we present the design, fabrication and characterization of a robust (O2.-) biosensor using a simple and straightforward procedure involving deposition of a uniform layer of L-Cysteine on a gold wire electrode to which Cytochrome C (Cyt c) was conjugated. The immobilized layers, studied using conductive Atomic Force Microscopy (c-AFM) revealed a stable and uniformly distributed redox protein on the gold surface, visualized as conductivity and surface topographical plots. The biosensor enabled detection of (O2.-) at an applied potential of 0.15 V with a sensitivity of 42.4 nA/µM and a detection limit of 2.4 nM. Utility of the biosensor was demonstrated in measurements of real time (O2.-) release in activated human blood platelets and skeletal rat limb muscles following ischemia reperfusion injury (IRI), confirming the biosensor's stability and robustness for measurements in complex biological systems. The results demonstrate the ability of these biosensors to monitor real time release of (O2.-) and estimate the extent of oxidative injury in models that could easily be translated to human pathologies.

A feedback mechanism converts individual cell features into a supracellular ECM structure in Drosophila trachea.

The extracellular matrix (ECM), a structure contributed to and commonly shared by many cells in an organism, plays an active role during morphogenesis. Here, we used the Drosophila tracheal system to study the complex relationship between the ECM and epithelial cells during development. We show that there is an active feedback mechanism between the apical ECM (aECM) and the apical F-actin in tracheal cells. Furthermore, we reveal that cell-cell junctions are key players in this aECM patterning and organisation and that individual cells contribute autonomously to their aECM. Strikingly, changes in the aECM influence the levels of phosphorylated Src42A (pSrc) at cell junctions. Therefore, we propose that Src42A phosphorylation levels provide a link for the ECM environment to ensure proper cytoskeletal organisation.