Hepatitis C virus infection in spouses: sexual transmission or common exposure to the same risk factors?

OBJECTIVES: The aim of this study was to evaluate whether the transmission of hepatitis C virus (HCV) between spouses occurs through sexual contact or through other types of exposure. METHODS: We consecutively enrolled 311 chronic HCV carriers and their spouses. The spouses underwent HCV blood testing. Exposure to parenteral risk factors was compared between couples of which both partners were HCV positive and couples with one positive partner. In couples with both partners positive, qualitative detection of serum HCV RNA and genotyping were performed. RESULTS: The prevalence among spouses was 10.3% (32/311). The mean age was higher for HCV-positive spouses (57.7 vs 49.6 yr for HCV-negative spouses; p < 0.01). The prevalence among spouses increased with the duration of marriage, whereas no difference was found in relation to the clinical status of the index case. The 32 HCV-positive spouses reported parenteral exposure (blood transfusion, drug use, and use of multiple-use glass syringes inside or outside the family) more often than the 279 HCV-negative spouses (84.4% vs 26.2%; odds ratio [OR], adjusted for age by multiple logistic regression analysis, 12.4; 95% CI = 4.5-34.0). The percentage of couples sharing glass syringes was significantly higher among those with both partners infected (65.6% vs 12.9%; OR = 12.9; 95% CI = 5.4-31.4). Qualitative serum HCV RNA was determined in 22 couples with both partners infected; in 13 of them, both partners were HCV RNA positive, whereas in the remaining nine, only one partner was positive. In eight of the 13 couples with both partners HCV RNA positive, the same genotype was found for both partners. CONCLUSIONS: The findings that the same genotype was detected for both partners in relatively few couples, and that a history of parenteral exposure was an independent predictor of HCV positivity, suggest that the risk of sexual transmission is low. The sharing of glass syringes may have played an important role in transmission between spouses.

Usefulness of human papilloma virus testing in the screening of cervical cancer precursor lesions: a retrospective study in 314 cases.

OBJECTIVE: To assess the usefulness of human papilloma virus (HPV) typing for predicting pre-malignant and malignant cervical lesions. STUDY DESIGN: 314 women, who underwent colposcopy, biopsies and high and low-risk HPV typing after a confirmed abnormal routine Pap test were studied. HPV-DNAs were typed by using PCR technique. RESULTS: We found a significant increasing rate of high-risk-HPV by the increasing severity of histology, ranging from 40% in negative cases to 86.9% in those with CIN3 lesions. The positive predictive value of high-risk-HPV ranged from 13.3% in patients with atypical squamous cells of undetermined significance (ASCUS) to 29.4% in those with HSIL. By contrast, negative predictive value was 96% in patients with ASCUS, 97.2% in low-grade squamous intraepithelial lesions (LSIL), and 71.4% in high-grade squamous intraepithelial lesions (HSIL). Sensitivity and specificity for detecting CIN2 or CIN3 was 86.0% and 41.3%, respectively. CONCLUSIONS: The high negative predictive value of high-risk HPV testing suggests that HPV negativity could be used for predicting the absence of important cervical lesions, and therefore avoiding unnecessary colposcopy in ASCUS and LSIL cases.

Molecular characterisation of HCV genotype 4 isolates circulating in Italy.

The characteristics of genotype 4 subtype variability of HCV isolates circulating in Italy were studied. The viral isolates were identified from 736 HCV-RNA positive sera originated from seroepidemiological studies undertaken in 4 different regions of North, South Italy and Sardinia. 24 out of 28 genotype 4 isolates (86%) were classified by phylogenetic analysis of E1 genome region (915-1128) as belonging to subtype 4d (Neighbour Joining Method). Three isolates classified as subtype 4a were detected in haemophilic patients, possibly related to infections from blood products. One isolate classified as a new subtype derived from an Eritrean patient subjected to haemodialysis. Very high genome homogeneity (mean 4.3%) was shown by genetic comparisons (DNA dist programs Phylip Package) for all the 4d isolates relative to the studies performed in Veneto, Calabria and Sardinia and originated from subjects from the general population and outpatients (19 subtype 4d isolates out of 24). In the 3 studies different prevalence rates of HCV genotype 4 (3.1%, 1. 3%, 14% respectively) were found. In contrast a considerable degree of heterogeneity, both intragroup and with the other groups (mean 8. 2% and 8.7%, respectively) was observed among subtype 4d isolates identified in the patients of a haemodialysis centre in Apulia region. In conclusion the subtype 4d of genotype 4 was highly prevalent and endemic in Italy. An elevated level of viral heterogeneity was observed in one study carried out in a region of Southern Italy. This can be related to a longer period of past endemicity of this genotype and to a high level of exposure to reinfections in particular categories of patients such as haemodialysis patients.

TTV infection in patients with acute hepatitis of defined aetiology and in acute non-A-E hepatitis.

BACKGROUND/AIMS: Recently, the presence of a novel nonenveloped single-stranded DNA virus (TTV) has been associated with either acute or chronic hepatitis of unknown aetiology, suggesting a possible aetiological role. The aim of this study was to evaluate the prevalence, the significance and the clinical impact of TTV infection in patients with acute viral hepatitis of defined aetiology and in patients with non-A-E acute hepatitis. METHODS: TTV-DNA was tested by hemi-nested PCR in serum samples collected from 121 patients during and after acute hepatitis (103 with acute viral hepatitis of defined aetiology and 18 with acute non-A-E hepatitis) and in 30 healthy controls. RESULTS: Overall, the rate of TTV infection was 12.6% (13/103) in patients with acute hepatitis of defined aetiology, 16.6% (3/18) in patients with non-A-E acute hepatitis and 6.6% (2/30) in the healthy control group, (p=n.s). TTV-DNA was detected in the following proportions: hepatitis B, 13.2% (7/53); hepatitis C, 16.6% (4/24); hepatitis A, 4.7% (1/21); hepatitis E 20% (1/5). Moreover, acute hepatitis with and without TTV infection/coinfection were comparable in terms of both liver biochemistry and chronicity rate. The results of TTV re-testing after serial dilutions of six TTV-DNA positive serum samples during and after the peak of liver transaminases failed to demonstrate a correlation between liver damage and viral titre. CONCLUSIONS: The prevalence of TTV infection appeared to be comparable in patients with non-A-E hepatitis, in acute hepatitis of defined aetiology and in the control group. Hence, an aetiological role of TTV for acute hepatitis of unknown aetiology seems questionable. Moreover, TTV infection does not modify the natural history of acute hepatitis of defined aetiology.

Interferon alfa-2b alone or in combination with ketoprofen as treatment for interferon-naive chronic hepatitis C patients.

BACKGROUND: Non-steroidal anti-inflammatory drugs may amplify the anti-viral effect of alpha-interferon in vitro but in vivo data are still controversial. AIM: : To test the hypothesis that ketoprofen may increase the rate of response to alpha-interferon of chronic hepatitis C patients. METHODS: Fifty patients with chronic hepatitis C who had never received alpha-interferon were randomly assigned to receive 3-8 MU of alpha2b-interferon, three times weekly for 6 months, alone or in association with ketoprofen at a dose of 200 mg/day five times weekly. The virological response to treatment (undetectable HCV RNA in serum) was evaluated after 3 months and at the end of treatment, and 6 and 12 months after therapy withdrawal. RESULTS: One patient under combination therapy stopped the ketoprofen for persisting epigastric pain. Complete response under treatment was observed in 15 out of 24 (62.5%) patients receiving alpha2b-interferon alone and in 14 out of 26 (53.8%) patients under combination therapy (P=N.S.). One year after the end of treatment, a sustained response was seen in 4 out of 24 (16.2%) patients treated with alpha2b-interferon and in 5 out of 26 (19.2%) patients having received the combination (P=N.S.). CONCLUSION: Administration of ketoprofen does not increase either the primary or the sustained response to alpha2b-interferon therapy of interferon-naive chronic hepatitis C patients.

Cytokines and the progression of liver damage in experimental bile duct ligation.

1. Cytokines are soluble factors whose action has been documented in physiological and pathological conditions. Some may be involved in the pathogenesis of cholestasis, whether of acute or chronic origin. 2. The aim of the present study was to evaluate the influence of epidermal growth factor (EGF), transforming growth factor (TGF)-beta 1, interleukin (IL)-6 and tumour necrosis factor (TNF) on cholestasis. Findings from Sprague-Dawley rats submitted to bile duct ligation for 1-28 days were compared with those from controls, which underwent laparotomy but not bile duct ligation. 3. Biochemical and morphological findings confirmed that the experimental procedure was successful. At the end of each follow-up period, the hepatic levels of the cytokines were determined and compared with liver histology findings. 4. The four cytokines studied showed different patterns of activation: hepatic levels of EGF, higher in the experimental than the control group, were comparable with the proliferative picture. The TGF-beta 1 pattern was correlated with data of periportal, perivenular and perineoductular fibrosis, confirming that this cytokine has a role in mediating the synthesis of matrix proteins. A fluctuating, phasic pattern was found for TNF in the experimental group, with high values on day 0, a decrease on the first and second postoperative days and then two peaks on days 8 and 14. Finally, immediately after surgical manipulation, high levels of IL-6 were found in the experimental group, followed by a decrease in levels until zero values were obtained. 5. This suggests that the obstructive condition produces several cytokine responses, each of which contributes to determine the cholestatic condition.

High prevalence of HCV-RNA in the saliva cell fraction of patients with chronic hepatitis C but no evidence of HCV transmission among sexual partners.

The aims of this study were to evaluate the prevalence of HCV-RNA in different fractions of saliva taken from patients with chronic hepatitis C, to establish whether virologic parameters or disease severity exert any influence on the detectability of HCV-RNA in saliva, and to evaluate the prevalence of HCV infection in partners of HCV-infected subjects with respect to the presence of HCV-RNA in saliva. Sera samples and different fractions of saliva (whole saliva, surnatant, and cell fraction) from 48 subjects (45 with chronic hepatitis C and three healthy anti-HCV+ carriers) were examined for HCV-RNA by RT nested PCR and DEIA hybridization. HCV-RNA-positive sera were also tested for genotype and viral titer (bDNA2 method). Twenty-seven stable sexual partners (25 females and 2 males) were screened for anti-HCV antibodies at least twice over a minimum of 12 months. HCV-RNA was detected in the sera of 39/45 patients and of 22/39 viremic patients. In all of the latter, the presence of HCV-RNA was restricted to the cell fraction. Viral titer was significantly higher in patients with HCV-RNA in saliva than in those without (12.3 x 10(6) versus 4.6 x 10(6) eq/ml, P < 0.01). HCV-RNA positivity was unrelated to genotype, duration of disease, Hepatitis Activity Index scores or transaminase levels. Anti-HCV was positive in one of 13 sexual partners of patients with HCV-RNA in saliva and in 1/14 of those without (P = NS). In conclusion, HCV-RNA is detectable in the cell fraction of saliva in a high proportion of highly viremic patients with chronic hepatitis C, but its presence does not seem to be associated with an increased risk of HCV transmission among sexual partners.

Tumour necrosis factor alpha and cellular proliferation in primary biliary cirrhosis.

AIMS: 1) To evaluate serum levels and tissue expression of Tumour necrosis factor alpha in primary biliary cirrhosis: 2) to correlate serum tumour necrosis factor alpha levels and cellular proliferation with the severity and prognosis of liver disease. METHODS: Twenty-nine primary biliary cirrhosis patients (6 stage I, 8 II, 8 III, and 7 IV) entered the study. Serum tumour necrosis factor alpha was measured by EIA (Innogenetics, Antwerp, Belgium). Tissue tumour necrosis factor alpha and Ki-67 were tested by indirect immunoperoxidase staining on liver sections. RESULTS: Serum tumour necrosis factor alpha increased with the severity of histological stage (from 10.8 +/- 11 pg/ml in stage II to 17.1 +/- 10 in stage III and 22.8 +/- 8.7 in stage IV, p < 0.036). A positive correlation was also found between tumour necrosis factor alpha serum levels and the Mayo score (p < 0.05). A weak and sporadic expression of tumour necrosis factor alpha was observed in the inflammatory infiltrate around the bile ducts. Tissue Ki-67 (expressed as the labelling index in the hepatocellular nuclei) was evaluated in all stages of the disease (1.09 +/- 0.6% in stage I, 1.14 +/- 0.6% in stage II, 2.11 +/- 1.9% in stage III, and 2.67 +/- 2.8% in stage IV; the labelling index was significantly lower in early stages (I/II) than in late stages (III/IV), p < 0.05. A strong correlation between Ki-67 and the Mayo score was observed (p < 0.0005). CONCLUSIONS: 1) tumour necrosis factor alpha production seems related to the severity and the prognosis of primary biliary cirrhosis; 2) liver mononuclear cells in the inflammatory infiltrate do not seem to be the major site of tumour necrosis factor alpha release; 3) cellular proliferation is correlated with the severity of liver disease.

[Recurrent aphthous stomatitis: which role for viruses, food, and dental materials?]. / Stomatite aftosa ricorrente: quale ruolo per i virus, gli alimenti ed i materiali odontoiatrici?

Recurrent aphthous stomatitis is a frequently occurring disorder which may be a clinical feature of systemic disease. For many other patients, it is a tedious problem often having no known cause. The aim of this study was to verify if immune responses to common foods and/or viruses are involved in the etiopathogenesis of recurrent aphthous stomatitis. Sixteen patients with this disorder were studied by measurement of immunoglobulin classes (IgG, IgA, IgM), blood lymphocyte subpopulations, blood circulating immune complexes, and complement fractions (C3 and C4). Intradermal skin tests for common food and inhalant allergens were performed in all cases. In 5 patients with positive skin tests, serum specific IgE were tested for the same allergens by radioallergosorbent test. Skin patch tests for dental material were performed in all cases. Oral mucosal biopsies and/or cytology samples were taken in 10 cases for histopathological evaluation and in situ hybridization for Papillomavirus, Cytomegalovirus, Herpes simplex virus I and II, Epstein Barr virus. In 13 patients, lymphocyte subpopulations were altered, with a reduced CD4/CD8 ratio. No other alterations of serum immunological parameters were observed. Skin patch tests for dental material were negative in all cases, while skin tests for food allergens were positive in 5 cases (not confirmed by radioallergosorbent test or food challenge tests). Virus antigen and DNA were not found in mucosal specimens, although one patient was positive for Epstein Barr virus DNA by in situ hybridization. An alteration of the CD4/CD8 ratio was demonstrated in most of the patients with recurrent aphthous stomatitis, although immune responses to food and/or dental material and/or common viruses did not seem to be involved in the etiopathogenesis of this disorder.

HGV/GBV-C in liver tissue and in sera from patients with chronic hepatitis C.

Forty-eight persons (M = 45, F = 3; age range = 20-53, mean = 32.2) affected with chronic hepatitis C were tested for HGV/GBV-C RNA and HCV-RNA by nested PCR and DEIA in serum and in liver specimens to evaluate the prevalence and the impact of HGV/GBV-C coinfection in patients with chronic HCV-related hepatitis. Sera were also assayed for antibodies to HGV/GBV-C E2 protein. Serum HGV/GBV-RNA could be detected in nine (19%) patients, and anti-E2 antibodies in 22 (46%) patients. The presence of HGV/GBV-C RNA or anti-E2 antibodies was mutually exclusive. The cumulative prevalence of HGV/GBV-C infection was 65% (31/48); the majority of these patients (26/31, 84%) were intravenous drug users (IVDUs). In eight of nine patients viraemic for HGV/GBV-C, RNA positivity could be revealed even in liver specimens; these eight patients were also positive for HCV-RNA both in serum and the liver and did not exhibit any specific association with HCV genotype. HGV/GBV-C RNA negative strand RT-PCR testing was negative in all of the eight liver specimens, providing little support to the hypothesis that liver represents the primary site of HGV/GBV-C replication. Moreover, patients with HGV/GBV-C and HCV coinfection were comparable to those with HCV infection alone in terms of biochemistry and liver histology.

HGV/GBV-C infection in patients with acute hepatitis of different etiology and in patients with chronic hepatitis C.

To investigate the prevalence of hepatitis G virus (HGV/GBV-C) in patients with liver disease and to confirm its hypothesized ability to cause liver damage, we studied 130 subjects; 61 had chronic hepatitis C virus infection and 69 had acute hepatitis of either defined etiology (n = 57) or of unknown origin (n = 12). Positivity for HGV/GBV-C RNA was detected in 10 of the 61 subjects with chronic hepatitis C (16.3%) and in 11 of the 57 subjects with acute hepatitis of defined etiology (19%), whereas we failed to detect HGV/ GBV-C viremia in subjects with hepatitis of nonestablished etiology. Patients exhibiting positivity for HGV/GBV-C RNA were found to be comparable to those exhibiting negativity for HGV/GBV-C RNA in terms of both liver function tests and Knodell's score (in liver biopsies); the affect of HGV/GBV-C infection on the biohumoral and histological activity in patients with chronic hepatitis C therefore appears to be minimal or absent. Similar clinical features were observed in patients with acute hepatitis of known etiology whether they were positive or negative for HGV/GBV-C RNA. However, long-term clinical studies are still required to clarify the actual impact of HGV/GBV-C co-infection. In our geographic, i.e., a region or north-east Italy, HGV/GBV-C infection appears to be strictly related to intravenous drug use, and this agent does not seem to be responsible for acute hepatitis of unknown etiology; other etiological agents are probably involved.

A DNA hybridization method for typing hepatitis C virus genotype 2c.

A high prevalence of hepatitis C virus (HCV) genotype 2c (22%) was detected in sera from 459 italian patients by core-region amplification and hybridization with specific probes by DNA enzyme immunoassay. Amplified fragments failed to hybridize with 1a, 1b, 2a, 2b and 3a subtype-specific and 4, 5, 6 type-specific oligonucleotides in 105 patients. Hybridization of these samples with type 2 probe, which recognized all the subtypes sequences, showed evidence for genotype 2 distinct from 2a and 2b. Fourteen out of these 105 isolates were cloned and sequenced. The results were consistent with genotyping assay. Nucleotide sequences were partially related to types 2a, 2b, 2d, 2e and 2f (87.0-93.5% of identity). The average nucleotide identity was highest for genotype 2c (95.87%). On the basis of sequence analysis, subtype 2c specific probe was derived. Hybridization efficiency with the newly designed probe was very high and more than 95% (100/105) of type 2 cases were classified as 2c. Evidence of different outcome of therapy inside the same HCV major type account for the need of accurate subtyping. In this study, amplification of the core region followed by hybridization with highly specific probes enabled distinction between HCV subtypes.

Comparison of competitive and non-competitive reverse transcription-polymerase chain reaction (RT-PCR) for the quantification of hepatitis C virus (HCV) RNA.

A competitive reverse transcription (RT)-nested polymerase chain reaction (PCR) assay for HCV RNA was compared with the Roche Amplicor HCV Monitor assay, based on non-competitive, single step RT-PCR. A total of 83 serum samples were tested in parallel by both assays. All samples could be quantified by competitive RT-PCR (cPCR), whereas seven were negative by the non-competitive assay (ncPCR). The HCV RNA titre of the discordant samples assessed by cPCR was significantly lower than that of the remaining 76 (P < 0.001). Absolute HCV RNA titres were higher by cPCR than by ncPCR (P < 0.001), even if the results of the two methods were statistically correlated (P < 0.001). HCV RNA titre tested by cPCR was not significantly different between samples infected with genotype 1 or 2. However, values obtained by ncPCR were higher in samples with genotype 1 (P < 0.001). Furthermore, all seven discordant samples were infected with genotype 2. When both methods were used to measure serial dilutions of standard HCV RNA, we observed a bias to lower measurements with the ncPCR kit. This study shows a good correlation between the results of two PCR-based methods for the quantification HCV RNA. However, the degree of sensitivity and the absolute HCV RNA titre measured may vary according to the assay used.

Prognostic value of proliferating cell nuclear antigen (PCNA) expression in unresectable hepatocellular carcinoma.

Recent studies indicate that evaluation of cell proliferation in mallignant tumors may have prognostic value, but limited data are available on its expression in hepatocellular carcinoma. We therefore evaluated the prognostic tool of PCNA antigen expression on paraffin-embedded sections of 54 patients with hepatocellular carcinoma and concomitant cirrhosis. According to the number of positive cells, the PCNA expression ranged between 0.8 and 47%, and was divided into two groups (Group A, PCNA less than or equal to 5.5%; Group B, PCNA >5.5%). The two groups of patients were numerically well balanced. Median survival was 16 months for group A and 12 months for group B. According to the Cox multivariate analysis, PCNA expression (P=0.002),TNM stage (P=0.009) and ECOG performance status (P<0.001) significantly correlated with survival. In conclusion, PCNA antigen expression was found to be a significant prognostic faeature in unresectable hepatocellular carcinoma and may be a useful tool for future trials.

Altered lipid peroxidation/glutathione ratio in experimental extrahepatic cholestasis.

1. Lipid peroxidation can occur in the presence of a cellular antioxidant-oxidant imbalance, but the role of lipid peroxides in cholestasis is not well understood. 2. This study was undertaken in order to: (i) evaluate the behaviour of a product of lipid peroxidation (thiobarbituric acid-reactive species), and of an important antioxidant tripeptide, reduced glutathione, in the course of experimental extrahepatic cholestasis; and (ii) ascertain whether there was a link between this aspect and the alterations in liver morphology. 3. Forty-five male Sprague-Dawley rats (250-300 g) were double bile duct ligated and followed from 1 to 28 days. At the end of each experimental period, blood and liver samples were collected for thiobarbituric acid-reactive species and glutathione determinations. 4. Bile duct ligated rats showed a marked increase in liver weight which was related to cholestasis duration and to some anatomical alterations such as bile duct proliferation and dilation and liver fibrosis (periportal, perivenular, perineoductular and parenchymal). 5. An increase in serum lipid peroxidation was also observed but this was not linked to hepatic thiobarbituric acid-reactive species. Erythrocyte and hepatic glutathione decreased in relation to cholestasis duration. Serum lipid peroxides and erythrocyte glutathione were correlated with liver cell necrosis. 6. In conclusion, experimental extrahepatic cholestasis determines bile duct proliferation and fibrosis, the degree of which is directly related to the duration of cholestasis itself and to liver cell necrotic phenomena.(ABSTRACT TRUNCATED AT 250 WORDS)

[A case of embryonal rhabdomyosarcoma of the cheek in adolescence. Its cytological diagnosis by fine-needle aspiration via the gingival fornix]. / Su di un caso di rabdomiosarcoma embrionale della guancia in età giovanile. Diagnosi citologica mediante aspirazione con ago sottile attraverso il fornice gengivale.

The authors present a case of embryonal rhabdomyosarcoma of the cheek in a young patient. The histological diagnosis, on the tumoral mass, was preceded by cytological researches with fine needle aspiration biopsy, carried out on the gingival fornix. The cytology, by immunocytochemical techniques, made possible the diagnosis of rhabdomyosarcoma. Therefore the authors stress the high capacity of these cytochemical and immunocytochemical methods to demonstrate, not in a invasive way, the small round cell tumors of the skull and the neck in the youngs. This early diagnosis may orient the clinical and therapeutic management of the case.