Sublingual immunotherapy in polysensitized patients: effect on quality of life.

BACKGROUND: Quality of life (QOL) is an important issue in allergic rhinitis and has been evaluated in a number of studies that have shown how it is impaired in untreated patients and improved by effective treatment. However, there are no data concerning QOL after sublingual immunotherapy (SLIT) in polysensitized patients. OBJECTIVE: To evaluate the effect, in real-life clinical practice, of SLIT on QOL in a population of polysensitized patients with allergic rhinitis. METHODS: We prospectively evaluated 167 consecutively enrolled polysensitized patients with allergic rhinitis. QOL was measured in all cases with the Rhinoconjunctivitis Quality of Life Questionnaire at baseline and after 1 year of SLIT (performed in approximately 70% of cases using single allergen extracts provided by the same manufacturer). RESULTS: The most frequent causes of sensitization were grass pollen, Parietaria, and house dust mites. The mean number of sensitizations per patient was 3.65. SLIT was performed with 1 extract in 123 patients (73.6%), with 2 extracts in 31 patients (18.6%), and with more than 2 extracts in 13 patients (7.8%). The mean values of all the QOL items improved significantly (P < .01 in all cases), with the following reductions noted: activities, 3.96 to 2.89; sleep, 2.07 to 1.56; general problems, 2.16 to 1.5; practical problems, 3.69 to 2.58; nasal symptoms, 3.57 to 2.50; eye symptoms, 2.92 to 1.83; and emotional aspects, 2.2 to 1.44. CONCLUSIONS: This study provides evidence that QOL can be improved in polysensitized patients treated with SLIT, and that the use of just 1 or 2 allergen extracts seems to be sufficient and effective in terms of improving QOL.

The autologous serum skin test in the follow-up of patients with chronic urticaria.

BACKGROUND: The presence of anti-FcepsilonRI and anti-IgE autoantibodies in a subset of patients with chronic urticaria suggests their aetiopathogenetic role. In clinical practice, the presence of these antibodies is usually considered when the autologous serum skin test (ASST) is positive. AIMS: To evaluate if the positive ASST follows up the activity of chronic urticaria. METHODS: Autologous serum skin test and thyroid autoantibody detection were performed in 82 patients with chronic urticaria and repeated 1 year later, when the vast majority of patients were symptom-free. Twenty patients with Hashimoto thyroiditis (HT), who had never suffered from urticaria, represented the control group. RESULTS: At the start of the study, the prevalence of positive ASST was 46.6%. The association of HT-urticaria was 29.3%. ASST was positive in 62 and 39% of patients with and without HT, respectively (P > 0.05 ns). One year later, 28 of 34 patients with a positive ASST were symptom-free, but 50% of them were positive for ASST. The ASST was positive in 86.7 and 8% of patients with and without HT, respectively (P < 0.001). In the control group, ASST was always negative. CONCLUSIONS: The co-existence of autoimmune thyroiditis with chronic urticaria seems to induce a significant difference in the persistence of a positive ASST. Consistent with previous reports, a positive ASST correlates with disease exacerbation in chronic urticaria patients without thyroiditis. In patients with thyroiditis and urticaria, positive ASST persists even after the urticaria has disappeared, thus questioning whether a positive ASST to be a surrogate marker of the functional role of anti-FcepsilonRI and anti-IgE autoantibodies.

Churg-Strauss syndrome: clinical and serological features of 19 patients from a single Italian centre.

OBJECTIVE: Churg-Strauss syndrome is a rare multisystem vasculitis of unknown aetiology. Due to the rarity of the disease, few single-centre case series have been described. The aim of this study was to evaluate a small series from a single Italian centre in order to describe the clinical features of the disease, the treatment and long-term follow-up. METHODS: Nineteen Churg-Strauss syndrome patients were selected from the medical records of all vasculitis patients attending the Immunology Unit at the Department of Internal Medicine of the University of Pisa in the decade between 1989 and 2000. Data were obtained retrospectively. RESULTS: All the patients had asthma and hypereosinophilia. As in other case series, the lungs, skin and peripheral nervous system were the most commonly involved organs. The majority of our patient received i.v. pulses of methylprednisolone followed by i.v. pulses of cyclophosphamide. The outcome and long-term follow-up were good. There were no fatalities observed in this series during the follow-up period. CONCLUSIONS: Churg-Strauss syndrome is a systemic vasculitis occurring in patients with a history of asthma and allergic rhinitis. The positive results of the treatment protocol used in this preliminary study deserve to be tested in controlled multicentre studies.

Structural evidence for a common zinc binding domain in archaeal and eukaryal transcription factor IIB proteins.

X-ray absorption spectroscopy has been used to compare the metal coordination of the N-terminal zinc binding domain of eukaryal human transcription factor (TF) IIB to the previously reported structure of archaeal Pyrococcus furiosus (Pf) TFB. Full length and N-terminal fragments for both PfTFB and human TFIIB were cloned, expressed, and purified. The [C10H] variant of PfTFB was constructed to resemble the metal binding motif of higher eukaryal TFIIB proteins by mutating the second cysteine ligand to a histidine. All five proteins bind zinc in a 1:1 ratio. Zn X-ray absorption spectroscopy of human TFIIB and [C10H]PfTFB mutant are consistent with ZnS3(N,O) ligation, and further suggest that the N/O ligand is an imidazole.

Kinetic and spectroscopic characterization of the gamma-carbonic anhydrase from the methanoarchaeon Methanosarcina thermophila.

The zinc and cobalt forms of the prototypic gamma-carbonic anhydrase from Methanosarcina thermophila were characterized by extended X-ray absorption fine structure (EXAFS) and the kinetics were investigated using steady-state spectrophotometric and (18)O exchange equilibrium assays. EXAFS results indicate that cobalt isomorphously replaces zinc and that the metals coordinate three histidines and two or three water molecules. The efficiency of either Zn-Cam or Co-Cam for CO(2) hydration (k(cat)/K(m)) was severalfold greater than HCO(3-) dehydration at physiological pH values, a result consistent with the proposed physiological function for Cam during growth on acetate. For both Zn- and Co-Cam, the steady-state parameter k(cat) for CO(2) hydration was pH-dependent with a pK(a) of 6.5-6.8, whereas k(cat)/K(m) was dependent on two ionizations with pK(a) values of 6.7-6.9 and 8.2-8.4. The (18)O exchange assay also identified two ionizable groups in the pH profile of k(cat)/K(m) with apparent pK(a) values of 6.0 and 8.1. The steady-state parameter k(cat) (CO(2) hydration) is buffer-dependent in a saturable manner at pH 8. 2, and the kinetic analysis suggested a ping-pong mechanism in which buffer is the second substrate. The calculated rate constant for intermolecular proton transfer is 3 x 10(7) M(-1) s(-1). At saturating buffer concentrations and pH 8.5, k(cat) is 2.6-fold higher in H(2)O than in D(2)O, suggesting that an intramolecular proton transfer step is at least partially rate-determining. At high pH (pH > 8), k(cat)/K(m) is not dependent on buffer and no solvent hydrogen isotope effect was observed, consistent with a zinc hydroxide mechanism. Therefore, at high pH the catalytic mechanism of Cam appears to resemble that of human CAII, despite significant structural differences in the active sites of these two unrelated enzymes.

Monitoring protein expression in whole bacterial cells with MALDI time-of-flight mass spectrometry.

We report the application of matrix-assisted laser desorption ionization (MALDI) to monitor recombinant protein expression in whole bacteria. This technique is characterized by rapid sample preparation that provides analysis of samples extracted directly from the growth media in less than 10 min. The mass spectrometric method holds several advantages over gel electrophoresis, the conventional method for examining the protein content of cells. Comparisons between the two methods of analysis are presented in terms of increased speed, efficiency, resolution, and mass accuracy. Delayed extraction time-of-flight mass spectrometry identifies posttranslational modifications and other changes in the expected structure which are not recognized by gel electrophoresis. The utility of this method is demonstrated for proteins with molecular masses ranging from 5 to 50 kDa. Low molecular mass proteins (< 10 kDa) can be efficiently analyzed without any treatment of the bacterial broth prior to MALDI sample preparation. The MALDI analysis of higher molecular weight proteins shows enhanced sensitivity when the bacterial solutions are first sonicated.

Assembly of a [2Fe-2S]2+ cluster in a molecular variant of Clostridium pasteurianum rubredoxin.

The rubredoxin from Clostridium pasteurianum contains a single iron atom bound to the polypeptide chain by cysteines 6, 9, 39, and 42. The C42A variant of this protein has been prepared by site-directed mutagenesis and heterologous expression of the gene in Escherichia coli. The mutated protein was found to contain an unexpected chromophore that has been characterized by a variety of techniques. UV-visible absorption and resonance Raman spectra were strongly reminiscent of those of [2Fe-2S] proteins. Mössbauer spectra of the oxidized chromophore isolated in oxygen-free conditions indicated low-temperature diamagnetism resulting from antiferromagnetically coupled high-spin ferric ions. Analysis of X-ray absorption fine structure spectra yielded an Fe-Fe distance of 2.68 A. Colorimetric assays of iron and inorganic sulfide showed that the two elements are present in a 1:1 ratio. Electrospray-ionization mass spectra displayed a major component at M = 6190 Da, i.e. the molecular mass of the C42A apoprotein plus two atomic masses of iron and two atomic masses of sulfur. Taken together, these data show that a mere point mutation allows the stabilization of a binuclear [2Fe-2S] cluster in a protein that normally accommodates a mononuclear Fe(Scys)4 site. Assembly of a [2Fe-2S] cluster may occur because rubredoxin assumes a similar fold around its metal center as the [2Fe-2S] Rieske protein. Alternatively, a more extensive structural rearrangement of the polypeptide chain of the C42A rubredoxin variant may be considered as well.

The N-terminal domain of TFIIB from Pyrococcus furiosus forms a zinc ribbon.

The three-dimensional structure of the N-terminal domain of an archaeal TFIIB, which has high sequence homology with eucaryal analogues, is strikingly similar to that of the C-terminal zinc ribbon of the eucaryal transcription elongation factor TFIIB.