Codon optimization of the human papillomavirus 11 (HPV 11) L1 gene leads to increased gene expression and formation of virus-like particles in mammalian epithelial cells

The 505 amino acid L1 protein of the human papillomavirus type 11 (HPV 11) is the major capsid polypeptide that has been shown to self-assemble into virus-like particles (VLPs) in vivo and in vitro. While L1 is essential for viral infection, expression studies in mammalian cells have been hampered by different codon preference between the virus and its host. To optimize L1 gene expression in mammalian cells, we converted wild-type HPV 11 L1 (11 L1wt) codons to those more common in human genes. The modified HPV 11 L1 gene (11 L1h) generated protein levels that were at least 100-fold higher than those of wild-type HPV 11 L1, while no obvious differences were seen in the level of mRNA. HPV 11 L1 protein was detected in mammalian epithelial and fibroblast cells, by immunoblotting and indirect immunofluorescence (IIF) techniques. Unlike the situation in situ, IIF revealed the presence of L1 mainly at perinuclear sites. Virus-like particles assembled intranuclearly only to a low extent, as indicated by transmission electron microscopy. DNA vaccination using the HPV 11 L1h gene yielded a drastic increase in L1-specific antibody production in mice as compared to immunization with the wild-type gene.     

Increased Immunostaining of Fibulin-1, an Estrogen-Regulated Protein in the Stroma of Human Ovarian Epithelial Tumors

Fibulin-1, an extracellular matrix protein, is secreted by human ovarian metastatic cancer cell lines under estrogen stimulation. Fibulin-1 expression was quantified by immunohistochemistry and computer-aided image analysis in 44 human ovarian epithelial tumors and 14 normal ovaries. The fibulin-1 staining intensity in proximal stroma, close to the surface of epithelial cells and tumor cells, progressively increased from normal ovaries to serous carcinomas. In all lesions, excluding cystadenomas, fibulin-1 accumulation was higher in proximal stroma than in distant stroma. In situ hybridization demonstrated strong fibulin-1 gene expression in epithelial cells of serous ovarian carcinomas and some cysts. The weak expression of fibulin-1 RNA in some stromal cells of these tumors could not explain the strong fibulin-1 protein accumulation in tumor stroma, which was therefore mostly produced by tumor epithelial cells. In carcinomas, fibulin-1 staining was not correlated with the percentage of estrogen receptor-α (ERα)-stained nuclei but was inversely correlated with the progesterone receptor. However, in cystadenomas and borderline tumors, both fibulin-1 and ERα protein levels increased, in comparison with normal ovaries, suggesting an effect of estrogens in the early steps of tumorigenesis. This fibulin-1 overexpression, demonstrated in vivo in ovarian carcinomas, might be a useful indicator for predicting cancer risk and/or aggressiveness.     

The anti-apoptotic factor Bcl-2 can functionally substitute for the B cell survival but not for the marginal zone B cell differentiation activity of BAFF

The TNF family ligand B cell-activating factor (BAFF, BLyS, TALL-1) is an essential factor for B cell development. BAFF binds to three receptors, BAFF-R, transmembrane activator and CAML interactor (TACI), and B cell maturation antigen (BCMA), but only BAFF-R is required for successful survival and maturation of splenic B cells. To test whether the effect of BAFF is due to the up-regulation of anti-apoptotic factors, TACI-Ig-transgenic mice, in which BAFF function is inhibited, were crossed with transgenic mice expressing FLICE-inhibitory protein (FLIP) or Bcl-2 in the B cell compartment. FLIP expression did not rescue B cells, while enforced Bcl-2 expression restored peripheral B cells and the ability to mount T-dependent antibody responses. However, many B cells retained immaturity markers and failed to express normal amounts of CD21. Marginal zone B cells were not restored and the T-independent IgG3, but not IgM, response was impaired in the TACI-IgxBcl-2 mice. These results suggest that BAFF is required not only to inhibit apoptosis of maturating B cells, but also to promote differentiation events, in particular those leading to the generation of marginal zone B cells.     

Allergy to cooked white potatoes in infants and young children: A cause of severe,chronic allergic disease

BACKGROUND: Cases of allergy to cooked potato in children have been reported, some with immediate and others with late reactions. The clinical effects of chronic allergic reactions to potato and the effectiveness of diet on such reactions have not been described previously. OBJECTIVE: We sought to evaluate the importance of cooked potato as an allergenic food in individual cases of atopy in children. METHODS: Eight atopic children were selected on the basis of suspicion of allergy to cooked potatoes: all had potato-specific IgE, 2 of 8 had experienced immediate allergic reactions, and 6 of 8 had eczema that improved with a potato-elimination diet (decrease in severity scoring of atopic dermatis [SCORAD] index of >50%). The patients were evaluated by using skin prick tests with homemade cooked and noncooked potato extracts and with a commercial extract and by using IgE immunoblots from SDS-PAGE patterns of potato extract. Seven patients were challenged with cooked potato. The control group consisted of 9 age-matched atopic children, 8 of them with eczema. RESULTS: The mean SCORAD index decreased from 43.3 before to 11.5 after elimination of potato from the diet. Potato CAP values ranged from 3.71 to greater than 100 kUa/L. Potato challenge results were positive in 7 of 7 patients. Skin prick test responses were positive for cooked potato extracts in 7 of 7 patients, for noncooked extracts in 7 of 7 patients, and for the commercial extract in 8 of 8 patients compared with in 0 of 9, 1 of 9, and 1 of 9 subjects in the control group, respectively. During immunoblotting, 8 of 8 patient sera recognized one or more protein bands compared with 0 of 9 control subject sera. CONCLUSION: Allergy to cooked potatoes is a cause of severe allergic disease, with immediate reactions and eczema in some atopic infants and young children.     

Photo-oxydation des polyéther-bloc-polyamides, 2. Influence de la composition des polymères multiséquencés

Analytical and kinetical aspects of the photooxidation in the solid state of two polyether-block-polyamides 1c and 1d are described. The block copolymers studied contain different polyether sequences at different percentages. In copolymer 1d as well as in copolymers 1a and 1b , previously studied, the photooxidation concerns mainly the ether groups. High maximal concentrations of hydroperoxy groups, ? CH(OOH)? O? CH₂? , are observed (0,12 mol.kg^?1). These hydroperoxides are converted into hemi-acetals, ? CH(OH)? O? CH₂? , saturated esters and formate through photolysis or thermolysis above 60°C. The hemi-acetal groups are thermo-unstable too and decompose into alcohol and aldehyde groups. The photooxidation of copolymer 1c presents different features. At long wavelengths, low stationary concentration of hydroperoxy groups are observed (0,02 mol.kg^?1). Hydroperoxidation of the polyamide 6 sequences is revealed by the formation of imide groups. Hydroperoxidation of the poly(propylene glycol) sequences are surprisingly located on the ether methylene groups. Excitation of polyamide 6 sequences at short wavelength (254 nm) induces the formation of crotonized aldehyde groups.     

Fourteen novel OPA1 mutations in autosomal dominant optic atrophy including two de novo mutations in sporadic optic atrophy

The OPA1 gene, encoding a dynamin-related GTPase that plays a role in mitochondrial biogenesis, is implicated in most cases of autosomal dominant optic atrophy (ADOA). Sixty-nine pathogenic OPA1 mutations have been reported so far. Most of these are truncating mutations located in the GTPase domain coding region (exons 8-16) and at the 3'-end (exons 27-28). We screened 44 patients with typical ADOA using PCR-sequencing. We also tested 20 sporadic cases of bilateral optic atrophy compatible with ADOA. Of the 18 OPA1 mutations found, 14 have never been previously reported. The novel mutations include one nonsense mutation, 3 missense mutations, 6 deletions, one insertion and 3 exon-skipping mutations. Two of these are de novo mutations, which were found in 2 patients with sporadic optic atrophy. The recurrent c.2708_2711delTTAG mutation was found in 2 patients with a severe congenital presentation of the disease. These results suggest that screening for OPA1 gene mutations may be useful for patients with optic atrophy who have no affected relatives, or when the presentation of the disease is atypical as in the case of early onset optic atrophy.     

Anti-soluble liver antigen (SLA) antibodies in chronic HCV infection

Hepatitis C infection is associated with autoimmune disorders, such as the production of autoantibodies. Anti-LKM1 and anti-LC1, immunomarkers of type 2 autoimmune hepatitis, have been previously associated with a HCV infection. Anti-Soluble-Liver-Antigen autoantibodies (SLA) are specifically associated with type 1 and type 2 autoimmune hepatitis and more closely related to patients who relapse after steroid therapy. The recent molecular cloning of the soluble liver antigen provides the opportunity to develop more specific tests for the detection of antibodies against it. The aim of this work is to characterize anti-soluble-liver autoantibodies in sera from patients chronically infected by HCV. A recombinant cDNA from activated Jurkat cells coding for the full length tRNP(Ser)Sec/SLA antigen was obtained. ELISA, Western Blot and immunoprecipitation tests were developed and used to search for linear and conformational epitopes recognized by anti-SLA antibodies in sera from patients chronically infected by HCV. Anti-soluble liver antigen antibodies were found in sera from 10.4% of HCV-infected patients. The prevalence was significantly increased to 27% when anti-LKM1 was also present. Most anti-SLA reactivity was directed against conformational epitopes on the antigen. The means titers by ELISA were lower than those obtained in type 2 AIH. The result of autoantibody isotyping showed a subclass restriction to IgG1 and also IgG4. This study shows the presence of anti-SLA antibodies in approximately 10% of HCV infected patients. The prevalence of SLA autoantibodies in HCV infected patients increases when LKM1 autoantibodies are also present. The relationship between the prevalence of this characteristic autoimmune hepatitis autoantibody and the implication of an autoimmune phenomenon in the liver injury of patients chronically infected by HCV needs further investigation.