Prevalence of the C282Y, H63D, and S65C mutations of the HFE gene in 1,146 newborns from a region of Northern Spain

In Spain, 85% of patients with genetic hemochromatosis (GH) are homozygous for the C282Y mutation of the HFE gene. H63D and S65C mutations of HFE may also play some role in the disease. The aim of this study was to establish the prevalence of C282Y, H63D, and S65C mutations of the HFE gene in newborns in Catalonia, Spain. One thousand one hundred forty-six newborn screening cards were selected randomly. DNA from these cards was extracted and HFE mutations were analyzed with the LightCycler equipment (Roche Diagnostics Gmbh, Mannheim, Germany). Sufficient DNA sample was obtained to screen for the three mutations in 1,043 cases (91%). The allelic frequencies of C282Y, H63D, and S65C mutations were 0.03 (IC 95% 0.022-0.037), 0.2 (IC 95% 0.19-0.22), and 0.01 (95% confidence interval [CI] 0.006-0.015), respectively. The frequency of C282Y homozygous newborns was 0.001 (95% CI 0.0005-0.0014). The frequencies of newborns doubly heterozygous for C282Y/H63D and C282Y/S65C were 0.01 (95% CI 0.005-0.02) and 0.002 (95% CI 0.0002-0.01), respectively. The allelic frequency of C282Y mutation is similar to that observed in Southern France, in the Czech Republic and in some areas of Italy. The allelic frequency of H63D mutation in Catalonia is the highest reported to date. Nevertheless, S65C is infrequent. These data should be kept in mind when designing hemochromatosis genotypic screening programs in Catalonia.     

Transforming growth factor beta1 involvement in the conversion of fibroblasts to smooth muscle cells in the rabbit bladder serosa

In an attempt to identify the growth factors or cytokines involved in the serosal thickening that occurs in rabbit bladder subjected to partial outflow obstruction, the following growth factors – transforming growth factor 1, platelet-derived growth factor, epidermal growth factor, granulocyte colony-stimulating factor and granulocyte–monocyte colony-stimulating factor – were delivered separately onto the serosal surface of the intact bladder via osmotic minipumps. The proliferative/differentiative cellular response of the rabbit bladder wall was evaluated by bromodeoxyuridine incorporation and immunofluorescence staining with a panel of monoclonal antibodies to cytoskeletal proteins (desmin, vimentin, keratins 8 and 18 and non-muscle myosin) and to smooth muscle (-actin, myosin and SM22) proteins. Administration of the transforming growth factor, but not of the other growth factors/cytokines, was effective in inducing serosal thickening. Accumulating cells in this tissue were identified as myofibroblasts, i.e. cells showing a mixed fibroblast–smooth muscle cell differentiation profile. The phenotypic pattern of myofibroblasts changed in a time-dependent manner: 21 days after the growth factor delivery, small bundles of smooth muscle cells were found admixed with myofibroblasts, as occurs in the obstructed bladder. These ectopic muscle structures displayed a variable proliferating activity and expressed an immature smooth muscle cell phenotype. The complete cellular conversion to smooth muscle cells was not achieved if transforming growth factor 1 was delivered to fibroblasts of subcutaneous tissue. These findings suggest a tissue-specific role for this growth factor in the cellular conversion from myofibroblast to smooth muscle cells. © 1998 Chapman & Hall     

Motor activity as an unbiased variable to assess anaphylaxis in allergic rats

The release of mediators by mast cells triggers allergic symptoms involving various physiological systems and, in the most severe cases, the development of anaphylactic shock compromising mainly the nervous and cardiovascular systems. We aimed to establish variables to objectively study the anaphylactic response (AR) after an oral challenge in an allergy model. Brown Norway rats were immunized by intraperitoneal injection of ovalbumin with alum and toxin from Bordetella pertussis. Specific immunoglobulin (Ig) E antibodies were developed in immunized animals. Forty days after immunization, the rats were orally challenged with the allergen, and motor activity, body temperature and serum mast cell protease concentration were determined. The anaphylaxis induced a reduction in body temperature and a decrease in the number of animal movements, which was inversely correlated with serum mast cell protease release. In summary, motor activity is a reliable tool for assessing AR and also an unbiased method for screening new anti-allergic drugs.     

uPA deficiency exacerbates muscular dystrophy in MDX mice

Duchenne muscular dystrophy (DMD) is a fatal and incurable muscle degenerative disorder. We identify a function of the protease urokinase plasminogen activator (uPA) in mdx mice, a mouse model of DMD. The expression of uPA is induced in mdx dystrophic muscle, and the genetic loss of uPA in mdx mice exacerbated muscle dystrophy and reduced muscular function. Bone marrow (BM) transplantation experiments revealed a critical function for BM-derived uPA in mdx muscle repair via three mechanisms: (1) by promoting the infiltration of BM-derived inflammatory cells; (2) by preventing the excessive deposition of fibrin; and (3) by promoting myoblast migration. Interestingly, genetic loss of the uPA receptor in mdx mice did not exacerbate muscular dystrophy in mdx mice, suggesting that uPA exerts its effects independently of its receptor. These findings underscore the importance of uPA in muscular dystrophy.     

The human DNA repair factor XPC-HR23B distinguishes stereoisomeric benzo[a]pyrenyl-DNA lesions

Benzo[a]pyrene (B[a]P), a known environmental pollutant and tobacco smoke carcinogen, is metabolically activated to highly tumorigenic B[a]P diol epoxide derivatives that predominantly form N(2)-guanine adducts in cellular DNA. Although nucleotide excision repair (NER) is an important cellular defense mechanism, the molecular basis of recognition of these bulky lesions is poorly understood. In order to investigate the effects of DNA adduct structure on NER, three stereoisomeric and conformationally different B[a]P-N(2)-dG lesions were site specifically incorporated into identical 135-mer duplexes and their response to purified NER factors was investigated. Using a permanganate footprinting assay, the NER lesion recognition factor XPC/HR23B exhibits, in each case, remarkably different patterns of helix opening that is also markedly distinct in the case of an intra-strand crosslinked cisplatin adduct. The different extents of helix distortions, as well as differences in the overall binding of XPC/HR23B to double-stranded DNA containing either of the three stereoisomeric B[a]P-N(2)-dG lesions, are correlated with dual incisions catalyzed by a reconstituted incision system of six purified NER factors, and by the full NER apparatus in cell-free nuclear extracts.     

The evolutionary genetics of emerging plant RNA viruses

Over the years, agriculture across the world has been compromised by a succession of devastating epidemics caused by new viruses that spilled over from reservoir species or by new variants of classic viruses that acquired new virulence factors or changed their epidemiological patterns. Viral emergence is usually associated with ecological change or with agronomical practices bringing together reservoirs and crop species. The complete picture is, however, much more complex, and results from an evolutionary process in which the main players are ecological factors, viruses' genetic plasticity, and host factors required for virus replication, all mixed with a good measure of stochasticity. The present review puts emergence of plant RNA viruses into the framework of evolutionary genetics, stressing that viral emergence begins with a stochastic process that involves the transmission of a preexisting viral strain into a new host species, followed by adaptation to the new host.     

Enhancement of antibody synthesis in rats by feeding cis-9,trans-11 conjugated linoleic acid during early life

Previous studies have demonstrated that the intake of a 1% conjugated linoleic acid (CLA) diet in an 80:20 mixture of cis-9,trans-11 and trans-10,cis-12 exerts age-specific effects on the immune system: immunoglobulin enhancement and proliferative down-modulation in neonatal and adult rats, respectively. The present study evaluates the influence of the same diet on antibody synthesis of early infant Wistar rats during suckling and/or after weaning. Dietary supplementation was performed during suckling and early infancy (4 weeks), only during suckling (3 weeks), or only in early infancy (1 week). CLA content in plasma and serum immunoglobulin (Ig) G, IgM and IgA concentration were determined. Proliferation, cytokines and Ig production were evaluated on isolated splenocytes. Cis-9,trans-11- and trans-10,cis-12-CLA isomers were detected in the plasma of all CLA-supplemented animals, and the highest content was quantified in those rats supplemented over the longest period. These rats also exhibited higher concentrations of serum IgG, IgM and IgA. Moreover, splenocytes from CLA-supplemented rats showed the highest IgM and IgG synthesis and interleukin (IL)-6 production, whereas their proliferative ability was lower. In summary, in infant rats, we observed both the enhance antibody synthesis previously reported in neonates, and the reduced lymphoproliferation previously reported in adults.     

The first radiation hybrid map of a perch-like fish: the gilthead seabream (Sparus aurata L)

Among Teleosts, Perciformes are the largest order of fishes and include numerous species of commercial importance. Perciformes also comprise species of primary interest for evolutionary studies and analysis of the sex determination systems and sex chromosome plasticity. Unfortunately, genomics tools and resources for Perciformes remain to be developed. Here, we report the production of a seabream whole-genome radiation hybrid (RH) panel in which quality was ascertained by the construction of a 2-Mb-resolution RH map. The map encompasses 440 markers (288 microsatellites, 82 gene-based markers, and 70 STS) suitable for linkage analysis and comparative mapping studies. Achievement of a RH panel and a whole-genome RH map should contribute to establishing seabream as a fish model among the Perciformes and should be of importance in aquaculture for marker-assisted selection, improvement of growth performance, and disease management. Development of RH maps in a cost-effective manner for other fishes with the described methodology will offer a powerful approach in aquaculture and will provide extended capabilities for comparing vertebrate genome evolution.     

Differential expression of MHC class II molecules in highly metastatic breast cancer cells is mediated by the regulation of the CIITA transcription Implication of CIITA in tumor and metastasis development

We analyzed the differential gene expression between variants of MDA-MB-435 human breast cancer cell line that share an identical genetic background but have different metastatic ability. The major histocompatibility complex class II was found down-regulated in highly metastatic cells and correlated with MHC transactivator (CIITA) expression. Constitutive CIITA expression observed in poorly metastatic is driven by promoters III and IV of CIITA gene. Conversely, both promoters were ineffective in highly metastatic cells. The MHC class II and CIITA expression was restored in these cells upon stimulation with IFNgamma or by the treatment with a hypomethylating agent. Both treatments induced USF-1 and IRF binding complexes to promoter IV but only IFNgamma induced the binding of 435-Lung2 nuclear proteins to an ARE-1 site at the promoter III. Neither Southern blot nor bisulfite sequencing of promoter IV demonstrated strong hypermethylation of this promoter at the IFNgamma-responsive elements such as GAS, E-box or IRF-1. We suggest that partial or hemimethylation of promoter IV is sufficient to silence the CIITA expression in highly metastatic cells and that this epigenetic mechanism is responsible for the lack of MHC-II expression. Forced CIITA expression restored the MHC-II antigen expression in 435-Lung2 cells and abrogates spontaneous lung metastasis in both SCID and nude mice but also affected the tumorigenicity in nude mice. The increase in NK cell infiltration in nude mice bearing CIITA-tumors correlated with sign of tumor cell apoptosis and the increase in the number of NK cells in the spleens, suggesting that NK cells might be responsible for the observed antitumor activity.