Borderline ovarian tumors: laparoscopic treatment.
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Adenocarcinoma↗
Biomedical subjects
F Maggi
Publications and source records attributed to F Maggi.
Divergent evolution of hepatitis C virus in liver and peripheral blood mononuclear cells of infected patients.
In infected individuals, hepatitis C virus (HCV) exists as a variably complex population of related genetic variants known as quasispecies. The quasispecies of HCV were studied previously in 10 chronically infected patients by single-strand conformation polymorphism analysis of a segment of the envelope gene E2/NS1 containing the hypervariable region 1 and it was found that certain variants (LC variants) were present both in the liver and in peripheral blood mononuclear cells (PBMC), others (L variants) were present in the liver but not in the PBMC, and still others (C variants) showed the opposite distribution. The sequence data obtained from nine such patients are reported, indicating that, within individual subjects, L and C variants are distinct phylogenetically. Results are described on the growth of HCV in stimulated healthy donor PBMC cultures supporting the concept that genetic divergence might stem, at least in part, from virus adaptation to growth in different cell types. This information may help to understand how HCV persists and produces disease in infected patients, especially with regard to extrahepatic pathology.
Cells, Cultured↗
Detection of feline immunodeficiency proviral sequences in lymphoid tissues and the central nervous system by in situ gene amplification.
The availability of sensitive methods for detecting and localising the feline immunodeficiency virus (FIV) may help shed light on its role in generating tissue damage observed during infection. As immunohistochemical and in situ hybridisation techniques might not be sufficiently sensitive for this type of study, we adapted to FIV PCR-in situ hybridisation (PCR-ISH) that combine the extreme sensitivity of PCR with the precise localisation provided by ISH. The steps important for the success of PCR-ISH, such as sample preparation, permeabilisation, amplification profile, type of labels, and hybridisation conditions were optimised using paraformaldehyde-fixed and formalin-fixed paraffin-embedded sections of cells infected in vitro with FIV. As controls for amplification, the feline tumor necrosis factor-alpha gene (TNF-alpha) and the non-related EBNA-1 gene of the human Epstein-Barr virus were used. Once the method proved sufficiently sensitive and specific with these cells, the PCR-ISH assay was applied to paraffin sections of the lymph nodes, spleen and central nervous system of a 2-year FIV infected cat that, at the time of challenge, harboured low copy numbers of proviral genomes. Comparison of the results of PCR-ISH, competitive PCR and immunohistochemical analysis are described.
Animals↗
Autologous and heterologous neutralization analyses of primary feline immunodeficiency virus isolates.
Feline immunodeficiency virus (FIV) provides a model system with which the significance of neutralizing antibody (NA) in immunosuppressive lentivirus infections may be studied. To date, no detailed analysis of the neutralization properties of primary FIV isolates has been reported. In this study, we have conducted the first comprehensive study of the sensitivity to autologous and heterologous neutralization in a lymphoid cell-based assay of 15 primary FIV isolates and, for comparison, of one tissue culture-adapted strain. Primary isolates in general proved highly NA resistant, although there was considerable individual variation. Variation was also observed in the capacity of immune sera to neutralize heterologous FIV isolates. The ability of sera to neutralize isolates or for isolates to be neutralized by sera did not correlate with epidemiological and genetic relatedness or with the quasispecies complexity of the isolates. From the study of specific-pathogen-free cats experimentally infected with viral isolates associated with NA of different breadths, it appears that the development of FIV vaccines cannot rely on the existence of viral strains inherently capable of inducing especially broad NA responses.
Animals↗
[Analysis of a microdeletion in the AZF gene region in azoospermic patients: from research to molecular biology laboratory routine].
Microdeletions in AZF (Azoospermia Factor) region of the Y Chromosome have been recently described in azoospermic or oligozoospermic men. A Y chromosome molecular analysis was performed in peripheral blood DNA obtained from 16 azoospermic patients. Microdeletions within the interval 6 of the long arm of chromosome Y were analysed using the PCR (Polimerase Chain Reaction) technique. The presence of the DAZ (SY255) sequence was tested in 6 patients, while the remaining 10 patients were analysed for 6 different loci mapped to AZFb and AZFc regions. Microdeletion was detected in one azoospermic patient in the YRRM2 sequence. This result supports the finding of microdeletions not involving the DAZ sequence and suggests that routine investigation of azoospermic patients for microdeletions spanning the interval 6 region of y chromosome is also required.
DNA↗
Comparative evaluation of five rapid methods for identifying subtype 1b and 2c hepatitis C virus isolates.
A panel of 61 HCV isolates belonging to five different subtypes were used to evaluate five methods for rapid typing of HCV RNA: an in-house type-specific polymerase chain reaction based on the core region (type-specific PCR), a commercial amplification of the core region followed by hybridisation to probe coated wells (DEIA), a commercial amplification of the 5'-UTR region followed by hybridisation to probes immobilised on strips (LiPA), an in-house restriction fragment polymorphism analysis of the 5'UTR (RFLP), and a commercial serological method using synthetic peptides from the NS4 region (serotyping). The correct viral type was identified in 90% of cases by DEIA, in 82% of cases by type-specific PCR, in 80% of cases by LiPA and RFLP, and in 67% of cases by serotyping. Correct identification of the virus subtype was much less frequent and was beyond the performance characteristics of some assays. Major problems were found in the identification of isolates belonging to type 2. This was probably at least partly due to the fact that all type 2 isolates in the viral panel were of subtype 2c, which has been considered rare until recently.
Enzyme-Linked Immunosorbent Assay↗
Differences in hepatitis C virus quasispecies composition between liver, peripheral blood mononuclear cells and plasma.
Hepatitis C virus (HCV) exists in vivo as a highly variable mixture of closely related genomes (quasispecies), but the pathogenetic significance of such heterogeneity is still largely unknown. To investigate this issue, we compared the composition of HCV quasispecies found in the liver, peripheral blood mononuclear cells (PBMC) and plasma of ten patients by single-strand conformation polymorphism analysis of the E2/NS1 region and sequencing of the variants detected. We found considerable quasispecies differences between the liver and PBMC in all the patients, involving variant numbers, relative quantities and relative electrophoretic mobilities, but no apparent tissue-specific trend. Genome variants present in the liver and/or PBMC were not detected in the corresponding plasma samples, while certain HCV variants were present only in plasma. No dominant amino acids or amino acid pattern characteristic of variants present solely in the PBMC were detected in the E2/NS1 region sequenced.
Amino Acid Sequence↗
Subtype 2c of hepatitis C virus is highly prevalent in Italy and is heterogeneous in the NS5A region.
Hepatitis C virus (HCV) isolates, obtained from 50 Italian patients with community-acquired infection, that had previously been classified as subtype 2a or 2b by current rapid genotyping methods were further characterized by partial sequence analysis. All the isolates were reclassified: 45 within subtype 2c and the other 5 as subtype 1b, 3a, or 4d. Thus, subtype 2c is much more prevalent than previously recognized, with about 30% of all HCV strains detected in Italy being subtype 2c. In contrast, isolates of subtypes 2a and 2b appear to be infrequent, if not absent. Further studies showed that subtype 2c isolates are heterogeneous in the NS5A region, in that they may or may not contain a 57-nucleotide (nt) segment spanning from nt 7533 to nt 7589 of the viral genome. Partial nucleotide sequencing of the NS5B region of four 2c subtypes excluded the possibility that the isolates possessing or not possessing the 57-nt segment in the NS5A region may have resulted from recombination phenomena.
Adult↗
First-trimester chromosome diagnosis by lavage of the uterine cavity.
Fetal karyotyping of trophoblast cells obtained by lavage of the uterine cavity was evaluated on 86 first-trimester irrigation fluid samples. Villus fragments were observed in 72 fluid samples indicating an 83.7 per cent sampling success rate. The amount of villi in these samples ranged from 1 to 32 mg. In most cases, villus fragments showed degeneration of the external syncytiotrophoblast layer and absence of blood vessels. In the first phase of this study (15 samples), a high degree of maternal cell contamination was observed after long-term cultures. In the following phase (71 samples), this obstacle was overcome by the application of a semi-direct method. Chromosome preparations were set up after 24 h incubation of villus fragments and QFQ-banded metaphase spreads were scored for chromosome number and sex. Sixty samples showed the presence of villus fragments and the fetal karyotype was established in 40. Male and female chromosome complements were observed in 16 and 24 cases, respectively. In four cases, an abnormal fetal karyotype was diagnosed. These included trisomy of chromosomes 13, 15, and 16, and one mosaic with trisomy 12. Our results indicate that first-trimester fetal karyotyping might be feasible by a semi-direct method using chorionic villus fragments obtained at intrauterine lavage.
Cell Culture Techniques↗
Antibodies to interferon (IFN) in hepatitis C patients relapsing while continuing recombinant IFN-alpha2 therapy.
A number of trials have demonstrated that IFN-alpha is effective in chronic hepatitis C virus infection. It is known, however, that a number of chronic hepatitis C patients experience, after an initial response to IFN, disease reactivation or relapse (also called 'breakthrough') while IFN therapy is still ongoing. Since in a number of clinical conditions a significant correlation between development of antibodies to IFN and failure of therapy has been established, we addressed the possibility that the development of antibodies to IFN may take part in the relapse occurring in hepatitis C patients during recombinant IFN-alpha (rIFN-alpha) therapy. The prevalence of neutralizing (NA) and binding antibodies (BA) to rIFN-alpha2 has been evaluated in 45 patients with chronic hepatitis C treated with rIFN-alpha2a who first normalized aminotransferase (ALT) levels, and subsequently showed disease reactivation while on treatment. The presence of NA and BA was tested before therapy, during the response to IFN treatment, and at the time when ALT started to rise again to abnormal levels. The results showed that no patients had detectable antibodies to IFN before therapy and during the period of response to the therapy, while most of them (88.9%) developed NA and/or BA to IFN-alpha2 concomitantly with disease reactivation. In particular, in 29 of the 45 patients (64.4%) ALT normalized on treatment and rose to abnormal levels when NA appeared in their serum, while in 11 of the 16 (68.8%) remaining patients the relapse was associated with BA development. The frequency of seroconversion in these patients is significantly higher than that observed in the control group. These data indicate that antibodies to IFN may be responsible for breakthrough in the majority of patients showing disease reactivation while rIFN-alpha therapy is still ongoing.
Adolescent↗
Serological reactivity and viral genotypes in hepatitis C virus infection.
Patients infected with hepatitis C virus (HCV) were examined with four commercial HCV immunoblotting assays and for anti-GOR antibody to ascertain whether serological findings varied with the genotype of the infecting virus. The results indicate that patients infected with different HCV genotypes tend to show different immunoblotting profiles, mainly due to a low prevalence of antibodies to the viral region NS4 in patients infected with genotypes III and IV. Differences were more evident with second- than with third-generation assays. Patients infected with genotype IV exhibited a lower prevalence of anti-GOR antibody than patients infected with other genotypes.
Adolescent↗
Partial nucleotide sequencing of six subtype 2c hepatitis C viruses detected in Italy.
The great majority of 121 hepatitis C virus (HCV) isolates obtained from 117 Italian patients with community-acquired infection could readily be typed by genotype-specific PCR. Subtype 1b was dominant (74 isolates); subtypes 2b, 2a, and 1a followed, with 19, 14, and 8 isolates, respectively. The six isolates that remained untyped by this method were classified as subtype 2c on the basis of sequence analysis of PCR amplicors obtained from the core and NS5 genes. These findings indicate that HCV subtype 2c has a relatively high prevalence in Italy. Sequencing the core region from positions 160 to 259 is sufficient to distinguish subtype 2c from other known HCV genotypes.
Adolescent↗
First-trimester human trophoblast is class II major histocompatibility complex mRNA+/antigen.
Lack of expression of the polymorphic class I and class II MHC antigens in the cytotrophoblast is one of the major factors determining the privileged immunologic status of the placenta. In this report, we show that first-trimester human placental cytotrophoblast cells display moderate to strong expression of class II MHC (HLA-DR alpha and -DR beta) and Ii chain transcripts, apparently in absence of detectable class II antigens and Ii chain. In addition, DR alpha, DR beta, and Ii mRNAs, but not antigens, are consistently upregulated by IFN-gamma. Constitutive expression and upregulation of mRNAs are detectable in trophoblast cells kept in short term as well as prolonged (2-3 weeks) culture. These results are reminiscent of an analogous mRNA+/antigen- dissociation occurring, in the case of class I MHC gene products, in a subpopulation of first-trimester cytotrophoblast cells. Thus, analogous mechanisms prevent the expression of potentially hazardous class I and II allodeterminants at early stages of semiallogeneic pregnancy.
Blotting, Northern↗
Hepatitis C virus serological and polymerase chain reactions in human immunodeficiency virus-positive and -negative patients.
BACKGROUND: Polytransfused patients may be dually infected with hepatitis C virus (HCV) and human immunodeficiency virus (HIV). OBJECTIVES: To assess the correlation of antibodies to HCV with viral RNA in serum as determined by polymerase chain reaction (PCR) in anti HIV-positive and -negative haemophiliacs. STUDY DESIGN: Serum from 150 Patients with or without HIV infection were examined for anti-HCV by second generation enzyme-linked immunosorbent assay (ELISA) and recombinant immunoblotting assay (RIBA). A sample was also tested in a nested-reverse transcription PCR for a conserved sequence of the 5' untranslated region of HCV. PCR-positive specimens were titrated and a type-specific PCR using viral core gene sequences was used to determine distribution of HCV viral types. RESULTS: Eighty-seven percent of the patients were positive in ELISA. All the positives but 2 were either positive of indeterminate in RIBA. The frequency of indeterminate RIBA results was 33% among HIV-positive subjects and less than 1% among HIV-negative ones. PCR was positive in 68% of 73 RIBA-positive or -indeterminate individuals and negative in all HCV-seronegative individuals examined. No significant differences were observed in HCV viral type, prevalence or titers of viraemia between HIV-positive or -negative patients. CONCLUSIONS: The majority (68%) of anti-HCV-positive haemophiliacs examined in this study had HCV RNA in their sera and anti-HCV profile determined by RIBA had no apparent influence on viraemia. The presence of HIV infection in these patients had no significant impact on HCV RNA prevalence, titer or HCV type distribution.
Journal Article↗
Clinical significance of recombinant interferon-alpha 2 neutralizing antibodies in hepatitis patients.
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Antibody Specificity↗
Prevalence of hepatitis C virus genotypes in Italy.
Hepatitis C viruses (HCV) present in 110 Italian patients were characterized by genotype-specific PCRs. Among the 65 cases of community-acquired hepatitis, HCV genotype II was dominant (60%), followed by genotypes IV (15%), III (11%), and I (3%). Among the 45 hemophilia-associated cases, the distribution of the four HCV genotypes was markedly different: genotype I was the most prevalent (61%), followed by genotypes II (25%), III (4%), and IV (2%). Double infections were observed in eight patients. Two HCV remained unclassified. For the 45 community-acquired cases from which a liver biopsy was available, genotype II was associated with more severe liver damage than the other types.
Adolescent↗
Trisomy 16 confined to chorionic villi and unfavourable outcome of pregnancy.
Two cases of trisomy 16 confined to placental tissue associated with an unfavourable outcome of the pregnancy are reported. In the first case, after a diagnosis of an apparent non-mosaic trisomy 16 at chorionic villi sample (CVS), an intrauterine fetal death occurred at the 22nd week. In the second case a mosaic with trisomy 16 was found in chorionic villi and the fetus was still-born at 38 weeks. From a comparison of their cases with those of the literature, the authors conclude that a trisomy 16 confined to placental tissue has a negative effect on fetal growth and pregnancy outcome.
Adult↗
Cellular DNA content and proliferative activity evaluated by flow cytometry versus histopathological and staging classifications in human bladder tumors.
Flow cytometric analysis of cellular DNA content was performed on 78 biopsy bladder samples obtained from 61 patients with bladder tumors. All 6 normal tissue samples and 1 benign papilloma exhibited a cytometrically diploid DNA distribution, while 39 of 60 bladder carcinomas exhibited at least one aneuploid cell subpopulation. Furthermore, 13 of 39 aneuploid tumors were characterized by the presence of more than one aneuploid cell subpopulation. The results indicate a significant relationship between cytometric ploidy and morphological classification and stage: the occurrence of subpopulations with abnormal DNA content is associated with the increase in differentiation grade and stage. A significant statistical difference in the survival pattern between the diploid and aneuploid groups was observed. The percent of S cells extracted from DNA content distribution histograms indicates a statistically significant difference (p less than 0.01) between normal tissue (3.7 +/- 1.8), diploid tumor (8.4 +/- 3.9) and aneuploid tumor (14.9 +/- 6.0). Moreover the percent of S-phase cells increases with grade in only the aneuploid subgroup. Our results suggest that cytometric parameters in association with morphological and clinical criteria can contribute to a more accurate characterization of bladder tumors in prognostic terms.
Aged↗