Marcello Piazza

Analysis of HIV-1 load in blood, semen and saliva : evidence for different viral compartments in a cross-sectional and longitudinal study

Objectives: To quantify the HIV‐1 load (measured as copies of viral RNA/ml using competitive reverse transcription‐polymerase chain reaction) in blood, semen and saliva and to look for relationships between the viral burden, the clinical and immunological status and antiretroviral therapy. Methods: Peripheral blood, semen and whole saliva samples were collected from 26 anti‐HIV‐1‐seropositive patients selected for a cross‐sectional study. Nine of the 26 patients provided samples of the three biological fluids for a longitudinal study. Results: HIV‐1 RNA was detected in 26 out of 26 samples of plasma, in 25 out of 26 samples of semen and in 24 out of 25 samples of saliva. The median number of HIV‐1 copies in plasma was 14 817/ml (range: 167‐254 880), in semen was 515/ml (range: 0‐196 050) and in saliva was 162/ml (range: 0‐72 080). The viral load in semen and in saliva was significantly lower than in plasma (P < 0.0001 ). The HIV‐1 RNA levels in plasma and in saliva were correlated (P < 0.05), but levels in semen were not corre‐lated with either plasma or saliva levels. The HIV‐1 copy number in plasma was significantly higher in symptomatic patients than in asymptomatic subjects (P<0.05). Plasma and saliva HIV‐1 RNA levels were higher in subjects with a CD4+ cell count < 200×106/1 than in subjects with a CD4+ cell count > 200×106/1 (P < 0.05). The HIV‐1 RNA load in either plasma, semen or saliva is not related to antiretroviral therapy. Conclusions: The absence of a correlation between plasma and semen loads suggests that semen and blood are distinct viral compartments. Viral load in semen is not related to the clinical stage of HIV infection or to the CD4+ lymphocyte count. Consequently, HIV‐1 ‐infected subjects are potentially infectious at all stages of immunodeficiency and adequate precautions must always be taken to prevent the sexual transmission of HIV.

Low frequency of severe hepatotoxicity and association with HCV coinfection in HIV-positive patients treated with HAART.

Background: Highly active antiretroviral therapy (HAART) is strongly effective in reducing morbidity and mortality in HIV‐1‐positive individuals. Its main drawback is the potential toxicity. Data on the frequency and determinants of severe hepatotoxicity in a clinical setting are still sparse. Methods: This is a prospective study of HIV‐1‐positive individuals with known HBsAg and HCV‐Ab serology. The study end point was progression to alanine aminotransferase (ALT) levels ≥200 IU/L after HAART initiation. Cumulative probability of progression to this end point was estimated by the Kaplan‐Meier method. Crude and adjusted hazard ratios (HR) were estimated by proportional hazards regression model. Results: One thousand two hundred fifty‐five patients were included. HBsAg was found in 91 (7.2%), HCV‐Ab in 578 (46.5%) patients; almost all injection drug users (451 of 482; 93.6%) were HCV‐Ab positive. Sixty‐one individuals progressed to the end point with a probability of 7.9% (95% confidence interval [CI], 5.6‐10.0) of progression at 24 months from starting. Independent factors predicting progression to the end point were baseline ALT levels (HR, 5.29; 95% CI, 3.24‐8.65; every 10 IU/L higher), HCV‐Ab positivity (HR, 4.01; 95% CI, 1.48‐10.85) or both HBsAg and HCV Ab positivity (HR, 3.85, 95% CI, 1.01‐14.61), and previous non‐HAART therapy (HR, 1.84, 95% CI, 1.04‐3.42). Patients receiving stavudine‐containing regimens had a lower risk than those receiving zidovudine‐containing regimens (HR, 0.30, 95% CI, 0.12‐0.71). Conclusions: There was a low risk of ALT ≥200 IU/L in our cohort. Hepatitis C coinfection and elevated ALT levels at HAART initiation are important predictors of progression to ALT ≥200 IU/L; stavudine‐containing regimens were associated with a lower risk compared with zidovudine‐containing regimens.

Synthetic full-length and truncated RANTES inhibit HIV-1 infection of primary macrophages.

Objective: To evaluate the role of the selective forces exerted by the host on the HIV-1 structures involved in viral entry. Design and methods: The V3 region of the env gene was analysed in cell-free HIV-1 RNA from 17 infected subjects: 11 long-term non-progressors (LTNP) and six symptomless, typical progressor patients. To evaluate the potential biological significance of one of the rare variants detected in the LTNP, it was reproduced by recombinant PCR into a HIV-1 molecular clone. Results: The intrapatient divergence of the V3-loop sequences averaged 8.62% in LTNP and 5.29% in progressors, although LTNP displayed lower divergence from the clade B consensus than progressors (16.65 and 19.76%, respectively). The analysis of non-synonymous and synonymous substitutions indicated that selective pressure was exerted in this region in both LTNP and progressors. Individual peculiarities (unique and rare V3-loop variants) emerged, however, in most sequences from LTNP, and variants bearing mutations in a domain crucial for the V3-loop structure were more prevalent in LTNP (P = 0.0012). The pNL4-3-derived mutant reproducing a V3-loop variant detected in a LTNP was efficiently expressed upon transfection, but the mutant virus was nearly completely unable to infect CD4+ cell lines, activated primary peripheral blood lymphocytes, or monocyte-derived macrophages, suggesting that a defect impaired the entry phase of the replication cycle. Conclusions: The results indicate that host factors impose selective constraints on the evolution of the HIV-1 structures involved in viral entry. In LTNP, these factors are likely to force the virus into attenuated variants.

Immunoglobulin Superantigen Protein L Induces IL-4 and IL-13 Secretion from Human Fc varepsilon RI(+) Cells Through Interaction with the kappa Light Chains of IgE.

Peptostreptococcus magnus protein L is a multidomain bacterial surface protein that correlates with virulence. It consists of up to five homologous Ig-binding domains (B1–B5) that interact with the variable domain of Ig κ L chains. Intact protein L stimulates the synthesis and the release of IL-4 and IL-13 from human basophils in vitro. A protein L fragment covering the Ig-binding domains B1–B4 also induced IL-4 and IL-13 release from basophils. There was an excellent correlation (rs = 0.82; p < 0.001) between the maximal percent IL-4 release induced by protein L and that induced by anti-IgE and between intact protein L and the B1–B4 fragment (rs = 0.90; p < 0.01). Removal of IgE bound to basophils markedly reduced the IL-4 release induced by anti-IgE, protein L, and B1–B4. Preincubation of basophils with protein L or anti-IgE caused complete cross-desensitization to subsequent challenge with the heterologous stimulus. IgE purified from myeloma patients PS and PP (λ chains) blocked anti-IgE-induced IL-4 release, but not the releasing activity of protein L. In contrast, IgE purified from myeloma patient ADZ (κ chains) blocked both anti-IgE- and protein L-induced secretion. Cyclosporin A, but not cyclosporin H, inhibited protein L-induced release of IL-4 and IL-13 from basophils. Thus, protein L acts as a bacterial Ig superantigen to induce the synthesis and release of IL-4 and IL-13 from basophils by interacting with κ L chains of the IgE isotype.

Safety and immunogenicity of hepatitis A vaccine in infants: a candidate for inclusion in the childhood vaccination programme

Forty-eight infants received a single dose (720 ELISA units = 0.5 ml) of inactivated hepatitis A vaccine at the fifth month of age with booster at the 11th month of age, together with the second and third doses of the vaccines compulsory under Italian law (diphtheria, tetanus, oral polio and hepatitis B). Overall, the seroconversion rate was 100%. The anti-HAV geometric mean titre (GMT) reached 3,021 mIU/ml in infants born to anti-HAV-negative mothers, but only 399 mIU/ml in infants born to anti-HAV-positive mothers. Hepatitis A vaccine was immunogenic, safe and well tolerated without significant side-effects. There seems to be no reason for not including it in childhood vaccination programmes particularly in low endemic HAV areas.

MASS VACCINATION AGAINST HEPATITIS B IN INFANTS IN ITALY

A field trial currently underway in an area near Naples Italy where hepatitis B is endemic is providing support for a new vaccination protocol that involves administering hepatitis B vaccine at 3 and 5 months of age with a booster at 11 months. This protocol saves 1 dose of vaccine and also allows hepatitis B vaccine to be administered at the same time as immunization against diphtheria tetanus and poliomyelitis. Measurement of anti-hepatitis B surface antigen titers in the 110 Italian infants vaccinated according to this protocol to date revealed seroconversion rates of 93.6% immediately before the 11 month booster and 96.3% 2 months after the booster injection. In addition to being effective this protocol has been highly acceptable. 99% of mothers in the area have complied with the immunization program and have welcomed the opportunity to obtain protection for their children against hepatitis B at the same time that other immunizations are administered. On the basis of these findings the Italian health authorities have adopted the new protocol for routine use in areas highly endemic for hepatitis B infection. The World Health Organization has also recommended that hepatitis B vaccine be offered at the same time as other childhood immunizations.

Myasthenia Gravis During Low-Dose IFN-α Therapy for Chronic Hepatitis C

We describe the case of a 56-year-old man who had high aminotransferase levels and anti-hepatitis C virus (HCV) antibodies. He underwent liver biopsy and biochemical screening to evaluate whether he would benefit from interferon (IFN) treatment. The patient was discharged with a diagnosis of HCV-related active chronic hepatitis, skin porphyria, and type 2 diabetes. On December 5, 1995, he began therapy with recombinant IFN-α at a dose of 3 MIU three times a week. He stopped this therapy in February 1996 because of asthenia, dyplopia, headache, and anxiety. During IFN therapy, he had normal aminotransferase levels and no detectable HCV RNA, a condition that persists to the present. Between March and May 1996, the patient was admitted several times to a neurology clinic, where myasthenia gravis was diagnosed and treatment with pyridostigmine and cyclosporine was initiated. This case and others indicate that caution should be exercised in administering IFN because low doses can be correlated with myasthenia ...

Hepatitis C virus infection in family setting

To evaluate risk factors associated with intrafamiliar transmission of hepatitis C virus (HCV), 113 hepatitis C virus index subjects with chronic HCV infection and their 267 family contacts were studied from January 1994 to October 1995. Overall, 16 family contacts (6%) were positive for anti-HCV by ELISA II generation. The prevalence was 11.3% in spouses and 2.9% in other relatives (odds ratios: 4.2; 95% CI: 1.4–12.6). Spouses who had been married to the index cases longer than 20 years had a 7.5–fold risk (95% CI: 1.0–336.3) of HCV seropositivity as compared to those married less than 20 years. In univariate analysis HCV seropositivity was associated with surgical intervention, use of glass syringes and hospitalization. The results of multivariate logistic analysis showed that any parenteral exposure (odds ratios: 3.8; 95% CI: 1.2–12.8) and sexual contact with an anti-HCV index case (odds ratios: 3.0; 95% CI: 1.0–9.4) were both independent predictors of HCV seropositivity among household contacts of HCV positive index cases. These findings indicate that sexual contact and any parenteral exposure both play an independent role in the spread of HCV infection in the family setting.

A Simple Noninvasive Score Predicts Gastroesophageal Varices in Patients With Chronic Viral Hepatitis

Background Guidelines recommend upper endoscopic screening of cirrhotic patients for gastroesophageal varices. Cirrhosis is not always distinguishable from chronic hepatitis. Goals To identify low-risk patients who can be spared upper endoscopy irrespective of a diagnosis of cirrhosis. Study We evaluated 13 nonendoscopic variables as predictors of esophagogastric varices in 254 patients with hepatitis B or hepatitis C-related chronic liver disease who underwent upper endoscopy. Results Any size varices occurred in 30.3% (77/254), and large varices in 12.2% of patients (31/254). Age >50 years [odds ratio (OR): 11.29; 95% confidence interval (CI): 2.33-54.67], platelet count <150,000/mmc (OR: 4.40; 95% CI: 1.85-10.45), albumin <3.6 g/dL (OR: 2.99; 95% CI: 1.31-6.79), and aspartate aminotransferase/alanine aminotransferase ratio >1 (OR: 2.83; 95% CI: 1.26-6.34) independently predicted varices by logistic regression. Using a score based on age >50 years, platelets <150,000/mmc, and aspartate aminotransferase/alanine aminotransferase ratio >1 (1 point/predictor), only 3.2% of patients with a score <2 had varices, all small. Conclusions Patients with chronic viral hepatitis and a score <2 need not undergo upper endoscopy, as they are unlikely to have large varices. Because about 50% of our patients had this score, 50% of upper endoscopies may be safely avoided.

Iron depletion before HCV antiviral therapy: A pilot, randomized, controlled trial

It is not known whether iron depletion before pegylated IFN or combination treatment improves sustained virological response (SVR) rate in patients with chronic hepatitis C, despite its use in clinical practice in this setting. We aimed to investigate whether blood letting improves the efficacy (SVR) and tolerability of PEG‐IFNα2b + Ribavirin in chronic hepatitis C patients. Patients with chronic hepatitis C and ferritin >100 ng/mL were randomized to: (1) repeated phlebotomies to obtain a ferritin level <50 ng/mL followed by pegylated‐Interferon α2b + ribavirin (active arm); or (2) pegylated‐Interferon α2b + ribavirin (control arm). Primary endpoint was SVR rate, secondary endpoint was frequency of clinical and laboratory grade 3–4 adverse events. Thirty‐three patients were enrolled in the study (19 in active arm, 14 in control arm). The 19 patients in the active arm underwent a median of 5 phlebotomies (range: 1–9) to achieve the targeted ferritin (<50 ng/mL). Phlebotomies significantly reduced ferritin, iron, transferrin saturation, aspartate aminotransferase, alanine aminotransferase, and hemoglobin levels. Platelet count significantly increased, whereas HCV‐RNA levels remained unchanged. After antiviral therapy overall SVR was 31.6% in active arm and 21.4% in control arm (P = 0.698). Considering only the 18 patients who were naive to antiviral therapy, SVR was 60% in active arm versus 25% in control arm (P = 0.188). Tolerability, drug dose reduction or withdrawal were similar in the two arms. In conclusion phlebotomies do not increase the overall efficacy of antiviral therapy. However, the strong trend to higher SVR in naive patients undergoing phlebotomies warrants further investigation. J. Clin. Apheresis 2009.