UV photoreaction pathways of acetylacetaldehyde trapped in cryogenic matrices.

The broadband UV photochemistry kinetics of acetylacetaldehyde, the hybrid form between malonaldehyde and acetylacetone (the two other most simple molecules exhibiting an intramolecular proton transfer), trapped in four cryogenic matrices, neon, nitrogen, argon, and xenon, has been followed by FTIR and UV spectroscopy. After deposition, only the two chelated forms are observed while they isomerize upon UV irradiation toward nonchelated species. From previous UV irradiation effects, we have already identified several nonchelated isomers, capable, in turn, of isomerizing and fragmenting; even fragmentation seems to be most unlikely due to cryogenic cages confinement. Based on these findings, we have attempted an approach to understand the reaction path of electronic relaxation. Indeed, we have demonstrated, in previous studies, that in the case of malonaldehyde, this electronic relaxation pathway proceeds through singlet states while it proceeds through triplet ones in the case of acetylacetone. We observed CO and CO2 formations when photochemistry is almost observed among nonchelated forms, i.e., when the parent molecule is almost totally consumed. In order to identify a triplet state transition, we have tried to observe a "heavy atom effect" by increasing the weight of the matrix gas, from Ne to Xe, and to quench the T1 state by doping the matrices with O2. It appears that, as in the case of acetylacetone, it is the nonchelated forms that fragment. It also appears that these fragmentations certainly take place in the T1 triplet state and originate in an Π* ← n transition.

Formation of coronene:water complexes: FTIR study in argon matrices and theoretical characterisation.

In this paper, we report a combined theoretical and experimental study of coronene:water interactions in low temperature argon matrices. The theoretical calculations were performed using the mixed density functional-based tight binding/force field approach. The results are discussed in the light of experimental matrix isolation FTIR spectroscopic data. We show that, in the solid phase, (C24H12)(H2O)n (n ≤ 6) σ-type complexes, i.e. with water molecules coordinated on the edge of coronene, are formed, whereas in the gas phase, π-interaction is preferred. These σ-complexes are characterised by small shifts in water absorption bands and a larger blue shift of the out-of-plane γ(CH) deformation of coronene, with the shift increasing with the number of complexed water molecules. Such σ interaction is expected to favour photochemical reaction between water and coronene at the edges of the coronene molecule, leading to the formation of oxidation products at low temperature, even in the presence of only a few water molecules and at radiation energies below the ionisation potential of coronene.

Antibiotic prescription by general practitioners for urinary tract infections in outpatients.

In July 2008, in France, guidelines for antibiotic prescriptions for urinary tract infections (UTIs) were amended. As general practitioners (GPs) treat numerous UTIs, we wanted to evaluate whether they followed these guidelines. In order to do this, we performed a prospective study. The point of call was urinalyses. Using this selection method together with criteria diagnostic for urinalysis, we confirmed that patients presented a UTI. Each GP was contacted. Prescriptions were analysed and compared to the 2008 French guidelines for UTIs. Our study included 185 urinalyses. UTIs diagnosed by GPs were as follows: acute cystitis: 72.4 %, prostatitis: 13.5 %, nephritis: 8.7 % and asymptomatic bacteriuria: 5.4 %. The principal antibiotics used were: quinolone (59.5 %), furan (17.8 %) and cotrimoxazole (6.5 %). Only 20 % of the prescriptions were compliant with the guidelines. The correct antibiotic but not the dose or the duration of prescription was selected in 8.1 % of the prescriptions. For cystitis, inappropriate prescription was associated with an extra cost of 694 , namely, 7.4 per treatment. GP prescriptions for UTIs do not follow the guidelines. Even if GPs assert that they are aware of the emergence of resistant strains, it seems that they do not take into account the objective of quinolone restriction, which was one of the backbones of these guidelines.

Analysis of three genetic markers in IgA nephropathy patients from a single region.

BACKGROUND: IgA nephropathy (IgA-N) is the most common glomerular disease. Various genetic factors have been suspected to influence the disease, but they never have been studied in the same cohort of patients. METHODS: In 125 IgA-N biopsy-proven cases, we studied by DNA techniques the allele distribution of 3 polymorphic loci: the angiotensin-converting enzyme (ACE) gene, the specific HLA-DQB1 gene and the hs1,2 enhancer of the alpha1 gene of the IgH locus. Patients were classified as progressive and non-progressive based on a creatininemia above 150 microl/ml or/and a deterioration of the clearance greater than 3 ml/min/year. We analyzed the influence of the polymorphism on the development and the progression of the disease. The control group consisted of 83 heathly subjects. RESULTS: The frequency of HLA-DQB1*0602 was decreased in IgA-N patients (3.6% vs 10.2%, Pc = 0.04, RR = 0.36), suggesting a protective effect of this allele for IgA-N. Kaplan-Meyer analysis with the Cox-proportional hazard model revealed a shorter time between diagnosis and renal failure in patients with the B allele for the al gene hs1,2 enhancer (p = 0.04). ACE polymorphism did not influence the development or the progression of the disease. CONCLUSION: Genes controlling the immune response, such as HLA DQB1 and the alpha1 transcriptional enhancer gene, may influence the development and/or the progression of IgA-N nephropathy. Patients who develop an IgA-N nephropathy have a higher risk of severe evolution if they have a profile of high IgA humoral responder.

Polymorphism of the human alpha1 immunoglobulin gene 3' enhancer hs1,2 and its relation to gene expression.

We studied the hs1,2 transcriptional enhancer identified downstream of the human alpha1 gene of the immunoglobulin H (IgH) locus, for which two different allelic configurations (a and b) were previously reported by Southern blotting. By using a polymerase chain reaction (PCR) method we amplified minisatellites within the hs1,2 core enhancer, with variable numbers of tandem repeats (VNTR) defining three 'PCR alleles' alpha1A, alpha1B and alpha1C (including one, two and three repeats, respectively). Five different alpha1 h1,2 genotypes were encountered in a population of 513 donors, representing 13.8, 34.5, 49.7, 1.3 and 0.6% for the AA, BB, AB, AC and BC genotypes, respectively. Luciferase assays showed that increasing the number of minisatellites increased the transcriptional strength of the alpha1 hs1,2 enhancer. Simultaneous determination of Southern blot alleles and VNTR alleles only showed a partial linkage between both types of polymorphism, altogether defining at least six different allelic forms of the 3'alpha1 region. In conclusion, the present study further demonstrates the genetic instability of the 3'alpha region, for which multiple alleles have been generated through inversions and internal deletions and/or duplications. This study also strengthens the hypothesis that the polymorphism at the IgH 3' regulatory region of the alpha1 gene could play a role in the outcome of diseases involving immunoglobulin secretion.

Alleles of the alpha1 immunoglobulin gene 3' enhancer control evolution of IgA nephropathy toward renal failure.

BACKGROUND: IgA nephropathy is the most common glomerular disease. Mechanisms leading to its occurrence and controlling the evolution of the disease remain largely unknown. Various genetic factors have been found, mostly implicating immunologically relevant genes (IgH, TCR, human lymphocyte antigen, and complement loci). A regulatory region recently identified downstream, the alpha1 gene of the IgH locus, was a likely candidate for the control of IgA1 production in patients. Alleles of this region, differing by size, sequence, and orientation of the alpha1 hs1,2 transcriptional enhancer, were first identified through Southern blot hybridization. METHODS: We established a polymerase chain reaction (PCR) method suitable for routine testing that amplifies minisatellites within the alpha1 hs1, 2 enhancer, with variable numbers of tandem repeats (VNTR) defining the two alleles. This assay allowed the typing of 104 patients with IgAN and 83 healthy volunteers. Results from typing of alpha1 hs1,2 alleles were compared with long-term clinical outcome in patients. Enhancer alleles were compared in a luciferase reporter gene assay. RESULTS: The alpha1 hs1,2 alleles do not constitute a predictive factor for IgA nephropathy, since similar allelic frequencies were observed in healthy individuals and in unrelated European patients. In contrast, among patients, homozygosity for the weakest enhancer allele (AA genotype) was significantly correlated with a milder form of the disease, whereas the allele B was associated with severe evolution. The minisatellite region within the alpha1 hs1,2 enhancer carried potential transcription factor-binding sites, and its duplication increased the transcriptional strength of the alpha1 hs1, 2 allele B over that of allele A. CONCLUSION: Altogether, these alleles may constitute a risk factor for the prognosis of IgA nephropathy.

Deregulated platelet-activating factor levels and acetylhydrolase activity in patients with idiopathic IgA nephropathy.

BACKGROUND: Platelet-activating factor (PAF) is a phospholipid mediator with potent inflammatory activities. PAF stimulates IgA synthesis by B cells while IgA aggregates enhance PAF production by neutrophils and mesangial cells. These results led us to investigate blood PAF levels and plasma acetylhydrolase (AHA, the PAF catabolic enzyme) activity in patients with idiopathic IgA nephropathy (IgAN). METHODS: PAF and AHA levels were investigated using the platelet aggregation assay and degradation of (3)H-labelled PAF, respectively. The genotype of AHA with regard to the G994-->T mutation in exon 9 was assessed by an allele-specific polymerase chain reaction. RESULTS: Blood PAF levels were significantly (P:=0.003, Mann-Whitney U:-test) elevated in IgAN patients (50.6+/-6.8 pg/ml, n=33) compared with healthy controls (18+/-5 pg/ml, n=18). In contrast, plasma AHA levels were significantly (P:=0.0001, Mann-Whitney U:-test) reduced in patients with IgAN (61+/-2 nmol/ml/min, n=51) compared with healthy controls (78+/-4 nmol/ml/min, n=53). G994-->T transversion in exon 9 of AHA was not found in any of the IgAN patients. CONCLUSION: Elevated circulating levels of PAF in IgAN patients might result from an insufficient AHA probably related to environmental factors rather than genetic ones. The mechanism and the precise role of the PAF/AHA deregulation in IgAN patients remain to be clarified.

CTLA4 exon 1 dimorphism in bullous and cicatricial pemphigoid.

The human cytotoxic T-lymphocyte antigen 4 (CTLA4) gene encodes proteins regulating the immune response. The polymorphism of this gene is associated with some autoimmune diseases. In this study, we analysed the distribution of the dimorphisms of exon 1 (+ 49 A/G) in bullous pemphigoid (BP) and cicatricial pemphigoid (CP), two types of autoimmune bullous skin diseases that occur in elderly people. The frequency of the exon 1 A-G genotype was marginally decreased in patients (36.4%; n = 55) compared with controls (52.8%, n = 53), but the results were not statistically significant (P = 0.09).

Optimisation of HLA-B27 testing by association of flow cytometry and DNA typing.

HLA-B27 typing contributes to the diagnosis of ankylosing spondylitis. The classical technique of microlymphocytotoxicity is costly and can give false-negative results. We have compared 304 samples using two relatively new methods - flow cytometry and PCR-SSP - and evaluated their respective uses in routine analysis. Flow cytometric HLA-B27 testing was performed using three monoclonal anti-B27 antibodies (HLA-ABC-m3, GS145.2 and FD705 clones). Cut-off values were established to differentiate HLA-B27-positive from HLA-B27-negative samples with ROC curves. Although flow cytometric analysis with a reliable monoclonal antibody (mAb) is valuable for HLA screening, none of the HLA-B27 flow cytometry protocols was sufficient on its own to ascertain the HLA phenotype in 100% of samples. Two false negatives were observed with the FD705 mAb and the use of two different monoclonal antibodies did not increase the accuracy of HLA-B27 typing. HLA-B27 typing using molecular biology is a reliable but costly technique. Therefore we suggest that DNA typing could be used as a complementary technique and applied to samples whose HLA-B27 phenotype cannot be determined by flow cytometry. The association of flow cytometry and DNA typing is, in our experience, an economical and reliable approach.

Identification of a homolog of the C alpha 3'/hs3 enhancer and of an allelic variant of the 3'IgH/hs1,2 enhancer downstream of the human immunoglobulin alpha 1 gene.

Although four regulatory elements are known downstream of the mouse IgH alpha gene, a single enhancer homologous to hs1,2 has been thus far described downstream of each human alpha gene (Chen, C. and Birshtein, B. K., J. Immunol. 1997. 159: 1310). We characterized a 10-kb region downstream of the human alpha 1 gene. Two B cell-specific regulatory elements homologous to the murine C alpha 3'/hs3 and hs1,2,3' enhancers were found, which are duplicated downstream of alpha 2. The hs1,2 element is in inverted orientation by comparison with a recently reported alpha 1 hs1,2 element: it appears as a common allelic variant carrying an internal tandem repeat insertion and its prevalence in the human population is 60%. As in the mouse, the human hs1,2 enhancer is flanked with long inverted repeats which may have promoted inversion events through homologous recombination. Although the palindromic organization of the region is maintained in human, sequence identity with rodents focuses on core enhancer elements rather than on flanking repeats. Concerted divergence of both sides of the dyad symmetry suggests that inverted repeats are not just evolutionary remnants but rather play an architectural role in the LCR function.