[Use of a directory of specialized services and guidance in the healthcare system: the example of the Orphanet database for rare diseases]. / Utilisation d'un annuaire des services spécialisés et orientation dans le système de soins: l'exemple d'Orphanet dans les maladies rares.

BACKGROUND: Orphanet is a database of rare diseases which includes a directory of services providing information on professional experts working either in laboratories offering diagnostic tests or in specialized outpatient clinics. The printed directory is sent to these experts, to all relevant hospital departments (public and private), healthcare authorities, and patient support groups. The directory is also available online (www.orpha.net). The aims of this study were (i) to determine how the directory is used to refer patients and send specimens, and (ii) to investigate its impact on patient referral. METHODS: Data were obtained from experts and patient support groups concerned with rare diseases, as well from non-expert health professionals and patients. Emphasis was placed on knowledge of the Orphanet database, use of the directory as a tool for referrals, opinion of users about the quality of the directory, and opinion of the referenced experts about its possible impact on their referrals. Four methods of data collection were used: (i) a postal questionnaire to all referenced experts; (ii) an on-line questionnaire posted for a few hours on the Orphanet Website that had to be completed to access the site; (iii) interviews with 25 of the referenced experts; (iv) interviews with 35 leaders of patient organizations. Data were analysed using the chi2 test and logistic regression. RESULTS: Response rates were good: 74% of laboratory experts (224/304) and 68% of clinicians (459/678) answered the questionnaire. The responders proved to be representative. Among those who responded, 85% of the laboratory experts and 80% of clinician experts used Orphanet. More than two-thirds of them used Orphanet to identify (other) laboratories to them send specimens, and (other) clinicians for patient referral. Some non-expert hospital-based clinicians had nearly the same use. Patient support groups also used the directory. Persons using the directory happened to know Orphanet in their professional environment. Conversely, patients, non-MDs healthcare professionals and professionals in private practice discovered Orphanet using search engines, often when searching information about a rare disease. Of those who had already accessed the directory, most (94%) consider that the quality of the lists was "good" or" rather good". Among the experts, 29% of laboratories and 9% of clinicians considered that Orphanet had an impact on their referrals. CONCLUSION: The Orphanet directory is used to refer patients and specimens, especially by experts and patient organizations. It appears to have more impact on referrals within the healthcare system for laboratories than for specialized outpatient clinics. The impact is strong when expertise in the field is very scarce.

Expression of the release factor eRF1 (Sup45p) gene of higher eukaryotes in yeast and mammalian tissues.

Polypeptide chain termination in eukaryotic cells is mediated in part by the release factor eRF1 (Sup45p). We have isolated and characterised cDNAs encoding this translation factor from Syrian hamster (Mesocricetus auratus) and human (Homo sapiens) Daudi cells. Comparison of the deduced amino acid sequence of these new eRF1 (Sup45p) sequences with those published for Saccharomyces cerevisiae, Arabidopsis thaliana, Xenopus laevis and human indicates a high degree of amino acid identity across a broad evolutionary range of species. Both the 5' and 3' UTRs of the mammalian eRF1 (Sup45p)-encoding cDNAs show an unusually high degree of conservation for non-coding regions. In addition, the presence of two different lengths of 3' UTR sequences in the mammalian eRF1 (Sup45p) cDNAs indicated that alternative polyadenylation sites might be used in vivo. Northern blot analysis demonstrated that eRF1 (Sup45p) transcripts of differing length, consistent with the use of alternative polyadenylation sites, were detectable in a wide range of mammalian tissues. The Xenopus, human and Syrian hamster eRF1 (Sup45p) cDNAs were shown to support the viability of a strain of S cerevisiae carrying an otherwise lethal sup45::HIS3 gene disruption indicating evolutionary conservation of function. However, the yeast strains expressing the heterogenous eRF1 (Sup45p) showed a defect in translation termination as defined by an enhancement of nonsense suppressor tRNA activity in vivo. Western blot analysis confirmed that Xenopus eRF1 (Sup45p) was primarily ribosome-associated when expressed in yeast indicating that the ribosome-binding domain of eRF1 (Sup45p) is also conserved.

The absence of human beta 2-microglobulin increases the occurrence of ankylosing enthesopathy in HLA-B27 transgenic mice.

HLA-B27 transgenic mice develop a spontaneous ankylosing enthesopathy (ANKENT). We have investigated the occurrence of ANKENT in transgenic mice carrying transgenes for human beta 2-microglobulin (M TGM), HLA-B27-heavy chain (27 TGM), or both (27M TGM). An unexpected finding was the increase in ANKENT occurrence among the HLA-B27 transgenic mice lacking the human beta 2-microglobulin transgene (27 TGM): 33% of such mice were found to develop ANKENT, whereas 19% of 27M mice were diseased. In addition, the expression of HLA-B27 molecules in individual 27 TGM was highly variable, ranging from no expression to a level similar to that observed in 27M TGM. Our results confirm that in mice the HLA-B27 transgene is a relative risk factor for ANKENT. The increase of ANKENT occurrence is HLA-B27 transgenics in the absence of human beta 2-microglobulin suggests a possible role for impaired cell surface expression of HLA-B27. The absence of human beta 2-microglobulin might entail an accumulation of unassembled HLA-B27 heavy chains. Exposure of these mice to an environmental trigger could then lead to an inappropriate immune response which might result in disease development.

Allogeneic recognition of class I molecules: anti-H-2Ld repertoire of H-2b mice includes T cells recognizing mutant class II H-2b (Abm12) molecules.

Two major histocompatibility complex (MHC) class I-reactive T cell clones derived from H-2b mice, generated against the allogeneic Ld molecule, were found to recognize the H-2b class II mutant Abm12 molecule as well. In addition, these clones also recognize the class II A(s) molecule, and display a class II-dependent reactivity to staphylococcal enterotoxin B. Neither the class I nor the class II alloreactivities of the clones were found to be dependent on other MHC molecules. Both clones express CD4+CD8- phenotypes. The CD4 molecule appears to be involved in their class II reactivity, while little or no role for CD4 could be detected in the class I reactivity. This is the first report of a class I/class II cross-reactivity being mediated by CD4+ T cells. The structural basis for this cross-reactivity is discussed.

Biochemical and functional characteristics of soluble MHC molecules derived from H-2Ld/Q10d chimeric gene.

We have constructed a chimeric class I gene in which the 5' half of the H-2Ld gene is linked to the 3' half of Q10d. The resulting H-2Ld/Q10d protein is homologous to the native H-2Ld heavy chain for the three external domains except for an Arg to His substitution at position 260. The transmembrane and intracytoplasmic domains of the H-2Ld chain are replaced by the short low hydrophobic transmembrane-like domain of the Q10d chain. Following DNA-mediated gene transfer into mouse L cells, transformants were selected for the presence of specific mRNA. Radiolabelling and immunoprecipitation analysis revealed secretion of a 48-46 kd chain weakly associated with beta 2-microglobulin. This molecule reacts with H-2Ld-specific mAb that identify determinants on the first and second domains as well as with an anti-Q10 carboxyl-terminal peptide antiserum, but is not recognized by a mAb specific for a determinant of H-2Ld third domain. The integrity of antibody reactivity of the first and second domains together with beta 2-microglobulin association suggest that our molecule may be considered a good soluble counterpart of the native membrane H-2Ld molecule with which to perform functional studies. In order to analyze the immunogenic capacities and T-cell recognition of the soluble H-2Ld molecules, T-cell lines were produced from mice of various inbred strains immunized with supernatant from H-2Ld/Q10d-transfected fibroblasts. Characterization of these T cells revealed that they expressed a CD4+CD8- phenotype, and recognized H-2Ld/Q10d products in a class II-restricted manner.