Spin Probe Analysis of Microtubules Structure and Formation

Microtubules (MTs) control cell replication, material transport and motion in eukaryotic cells, but MT role in several pathologies is still unknown. These functions are related to the MT physico-chemical properties and MT formation mode starting from tubulin molecules. This study describes a new method, based on the computer aided analysis of the electron paramagnetic resonance (EPR) spectra of selected spin probes to obtain structural and dynamical information on tubulins and MTs and the kinetics of MTs formation promoted by guanosine-50-triphosphate (GTP). It was found that tubulin and MTs avoid radical quenching caused by ethylene glycol tetraacetic acid (EGTA). MT formation showed different kinetics as a function of tubulin concentration. At 5 mg/mL of tubulin, MTs were formed in 8 min. These results are also useful for getting information on MT–drug interactions

Nitroxide Radicals Interacting with Polyamine-Phenolic Ligands and Their Metal Complexes

We have analyzed the interactions of a series of nitroxide radicals with ligands containing a phenol moiety coupled with different polyarnine fragments, and their CuII and ZnII complexes, by EPR experiments. Nitroxide radicals are good mimics of the properties of pollutant molecules, e.g., organic compounds having different polarities and charges, surfactants, and polyaromatics. Charged, polar, and low-polarity sites in the ligands and complexes are all available for interactions; in particular, the ability to entrap the radicals in a vice-like structure, by the formation of phenyl sandwich complexes, determines the type of selective interaction that occurs when a chain or planar aromatic structure is present. For closely packed complexes, ZnII has a cooperative effect in the interaction with the radical; it allows both the entrance of low-polarity radicals at the low-polarity sites of the complex and the interactions of polar or charged radicals at the polar sites of the complexes. Conversely, CuII coordinates to the amino groups of the ligands and blocks the entry of radicals, which, thus, impedes their insertion into the structure of the complexes. The opposite behavior, with respect to that of CuII and ZnII, was found for the complexes that have more open structures. The complexes that have cyclic ligands exhibit a poor ability to interact with radicals, but, for the ligands in the absence of the coordinating metals, the interactions are significantly enhanced as a result of the entrance of the radicals into the cyclic structures. In all cases, stronger interactions occur when both hydrophilic and hydrophobic sites are involved. The neutral-surfactant probe forms aggregates mainly with the cooperative effect of the copper complexes. UV/Vis measurements confirmed the results obtained by EPR and indicate that weak interactions occur between the radicals and the complexes or ligands; they also confirmed the formation of 1:1 adducts

Bronchiectasis in children with recurrent pneumonia: an immunopathological damage associated with secondary ciliary dysmotility.

The aim of this study is to assess ciliary motion patterns in children with bronchiectasis unrelated to cystic fibrosis or primary ciliary dyskinesia. In 51 children with recurrent pneumonia, high resolution computed tomography (HRCT) was carried out to detect and score bronchiectasis. Moreover, ciliary ultrastructure, beat frequency and motion pattern were evaluated and compared to those observed in 30 healthy children. Bronchiectasis at HRCT was found in 31/51 children. Ciliary dysmotility was found in 20/31 children with bronchiectasis (64.5%). Overall, ciliary dysmotility was found in 39/51 patients (76.5%). Ciliary dysmotility showed a significant correlation with the HRCT score (p=0.02). Absent motion in some fields was found in 44/51 patients (86.3%) and this also showed significant correlation with the HRCT score (p=0.005). The specificity and sensitivity of ciliary dysmotility as an indicator of bronchiectasis was 74.3% and 83.3% respectively. The positive predictive value was 93.5%, and negative predictive value was 50%. Ciliary dysmotility, in children with recurrent airways infections, correlates with the presence and severity of bronchiectasis. Whether ciliary dysmotility is a cause or a consequence of anatomical lesion is a matter of speculation. Very likely there is an amplification and self-maintaining mechanism between the two events which may lead to more serious disease