[The effectiveness and acceptance of a medical device for the treatment of aphthous stomatitis. Clinical observation in pediatric age]. / Efficacia e accettabilità di un dispositivo medico per le afte buccali. Osservazione clinica in età pediatrica.

BACKGROUND: To evaluate the efficacy of a new bioadhesive patch, Aloe vera hydrogel, for the treatment of aphthous stomatitis. METHODS: An open, not controlled study was performed in 31 pediatric out-patients, aged 6-14 years, affected by mouth ulcers were enrolled consecutively in the 3 Gps Depts+ of San Marino Republic. For each case, data on case history and clinical profile, patterns of the lesion, presence of spontaneous or provoked pain were collected at baseline, and a bioadhesive patch ("Alovex patch") was administered on the basis of a daily regimen of < or = 3 patches for 4 days. Data on modification of the above-mentioned parameters, with patients and physicians opinion on the therapeutical efficacy, were collected during a control visit (4 days later). Moreover, by means of a daily diary, patients recorded information on the course of the symptoms during the 4 days and were also asked to compare the current treatment with other previous therapies. RESULTS: At the control visit 77% of the patients have shown a marked resolution of spontaneous pain, while in the other patients, pain was significantly decreased to a "mild" or "moderate" level. No one child declared to suffer from severe pain. Also provoked pain resulted to be significantly decreased after treatment Global efficacy was judged positively, being the therapeutical effect in more than 80% of cases "evident or of absolute improvement" both by physicians and patients opinion. A positive improvement of symptomatology started within the 2nd day of treatment in 74% of the patients. The compliance (adhesivity, acceptability and palatability) of the formulation was judged largely favourable in more than 90% of the patients. CONCLUSIONS: The results of this study underline the good efficacy and compliance of the patch for the treatment of the aphtous stomatitis; also the limit of topical available therapies, linked to the "contact time", to develop their therapeutical action, seems not to be evinced on the basis of this study, so the application of this patch seems to be more easy and beneficial.

Electron-transfer properties of Pseudomonas aeruginosa [Lys44, Glu64]azurin.

In the hydrophobic patch of azurin from Pseudomonas aeruginosa, an electric dipole was created by changing Met44 into Lys and Met64 into Glu. The effect of this dipole on the electron-transfer properties of azurin was investigated. From a spectroscopic characterization (NMR, EPR and ultraviolet-visible) it was found that both the copper site and the overall structure of the [Lys44, Glu64]azurin were not disturbed by the two mutations. A small perturbation of the active site at high pH, similar to that observed for [Lys44]azurin, occurs in the double mutant. At neutral pH the electron-self-exchange rate constant of the double mutant shows a decrease of three orders of magnitude compared with the wild-type value. The possible reasons for this decrease are discussed. Electron transfer with the proposed physiological redox partners cytochrome c551 and nitrite reductase have been investigated and the data analyzed in the Marcus framework. From this analysis it is confirmed that the hydrophobic patch of azurin is the interaction site with both partners, and that cytochrome c551 uses its hydrophobic patch and nitrite reductase a negatively charged surface area for the electron transfer.

Expression and characterization of Pseudomonas aeruginosa cytochrome c-551 and two site-directed mutants: role of tryptophan 56 in the modulation of redox properties.

The gene coding for Pseudomonas aeruginosa cytochrome c-551 was expressed in Pseudomonas putida under aerobic conditions, using two different expression vectors; the more efficient proved to be pNM185, induced by m-toluate. Mature holo-(cytochrome c-551) was produced in high yield by this expression system, and was purified to homogeneity. Comparison of the recombinant wild-type protein with that purified from Ps. aeruginosa showed no differences in structural and functional properties. Trp56, an internal residue in cytochrome c-551, is located at hydrogen-bonding distance from haem propionate-17, together with Arg47. Ionization of propionate-17 was related to the observed pH-dependence of redox potential. The role of Trp56 in determining the redox properties of Ps. aeruginosa cytochrome c-551 was assessed by site-directed mutagenesis, by substitution with Tyr (W56Y) and Phe (W56F). The W56Y mutant is similar to the wild-type cytochrome. On the other hand, the W56F mutant, although similar to the wild-type protein in spectral properties and electron donation to azurin, is characterized by a weakening of the Fe-Met61 bond, as shown in the oxidized protein by the loss of the 695 nm band approx. 2 pH units below the wild-type. Moreover, in W56F, the midpoint potential and its pH-dependence are both different from the wild-type. These results are consistent with the hypothesis that hydrogen-bonding to haem propionate-17 is important in modulation of the redox properties of Ps. aeruginosa cytochrome c-551.

Isolation and characterization of the d1 domain of Pseudomonas aeruginosa nitrite reductase.

Proteolitic digestion of nitrite reductase from Pseudomonas aeruginosa allows to obtain and purify a domain containing only the d1 heme and constituted by two noncovalently bound peptides. This d1 domain catayzes oxygen consumption, and binds carbon monoxide with a kinetic constant slightly higher than the parental dimeric holoenzyme. The capacity to oxidize the physiological substrate, cytochrome c551, is lost, even when the proteolytic c heme domain is added to this reaction mixture. This finding suggests that the two domains do not have a significant affinity for each other, and are kept together only by being part of the same polypeptide.