[Effect of anti-retroviral therapy on body composition changes: a literature review]. / Variazioni della composizione corporea associate alla terapia antiretrovirale: una rassegna della letteratura.

Protein-energetic malnutrition, characterized by both lean mass and fat depletion, was common in the pre-HAART era, and was associated with shortened survival and diminished quality of life. The pathogenesis of protein-energy malnutrition was multifactorial, and nutritional treatments were largely ineffective in the absence of disease stabilization. The introduction of HAART brought markedly improved outcomes, including a decrease in the incidence of malnutrition. However, other nutritional and metabolic alterations were noticed, and included changes in body shape, both lipoatrophy and lipohypertrophy, as well as changes in metabolism, notably hyperlipidemia and insulin resistance. These conditions, though sometimes occurring together, may occur independently, suggesting a complex, multifactorial cause. Several mechanisms have been hypothesized, including impairment to adipocyte differentiation and adipokine regulation, production of proinflammatory cytokines and mitochondrial toxicity. The role of the single drug class is still unclear, because both PI and NRTI have been associated with the syndrome, and the therapeutic protocols include both groups. Most of the medical therapies proposed for lipodystrophy are ineffective, and even if surgery remains an alternative, it is not associated with long lasting outcomes.

Quantitative viremia test for early prediction of a biochemical response in patients with chronic hepatitis C treated with interferon.

To evaluate the importance of the changes in viremia as an early predictor of the outcome of interferon (IFN) therapy, we assayed the levels of hepatitis C virus (HCV)-RNA in stored serum samples obtained from 34 patients with chronic hepatitis C who showed different biochemical responses to therapy. Serum samples obtained before the start of therapy and after 1 and 3 months were used, and viremia levels were determined by "branched DNA (bDNA)" technique. Viremia levels at 1 month of therapy were lower than pre-therapy levels in all 19 patients who had shown a persistent normalization of ALT during therapy (responder patients). The bDNA test was negative, i.e. the levels of viremia were below the sensitivity threshold of the method, in 12 (63.1%) patients at 1 month and in 13 (68.4%) at the 3rd month of therapy, whereas the bDNA test was negative in none of the 15 non-responder patients at the 1st month and in only 2 (13.3%) of them at the 3rd month of therapy. The bDNA test was superior to the ALT test both in predicting the non-response and the biochemical response to IFN after 1 month of therapy. The bDNA test results, instead, were not predictive of the duration of the response to IFN, either at the 1st or 3rd months of therapy. These results seem to indicate the usefulness of measuring the HCV-RNA levels at the beginning and after 1 month of IFN therapy in order to envisage or exclude a possible biochemical response early on in treatment.

[Experience at a branch center of the National HIV Protease Inhibitor Registry]. / L'esperienza di un centro periferico sul registro nazionale degli inibitori delle proteasi di HIV.

In the present study we show our experience as one of the Centers (center 006) of the Italian Index of Protease Inhibitors (PI). In this multicentric, observational study we studied HIV-positive naive patients for antiretroviral therapy with protease inhibitors. From 8/1/97 to 31/1/98 in our department 64 patients were enrolled and followed up through an electronic medical sheet in order to evaluate efficacy, tolerability, toxicity and compliance of treatment with PI. Our results show a reduction in AIDS-related mortality and a decrease in AIDS defining diseases: in particular data show an increase in CNS pathologies in comparison with other opportunistic events. Moreover, in patients non complaints to therapy, the progression of the disease was more frequent. The side effects more frequently observed during PI treatment were hypertrigliceridaemia and lipodystrophy; gynaecomastia and hyperprolactinaemia were even present, perhaps as expression of direct metabolic action of HAART therapy.

Value of the test for serum HCV-RNA in clinical practice.

The test to identify genomic RNA is the only tool currently available to directly evidence the presence of the hepatitis C virus (HCV) in infected subjects. In this review we examined the most commonly used qualitative and quantitative methods to detect HCV-RNA in serum or plasma, focusing particular attention on commercially available methods. Furthermore, we report the clinical conditions in which the viremia test is useful or even mandatory for the diagnosis of HCV infection, and comment on the usefulness of the test for monitoring patients in the course of antiviral therapy.

Biochemical response to interferon of chronic hepatitis in drug-addict patients infected with human immunodeficiency virus.

The effects of IFN treatment were retrospectively evaluated for 18 drug-addict patients with symptomatic HIV infection and chronic hepatitis C. Most of the patients were receiving concomitant treatment with zidovudine. Seven out of the 18 patients (39%) stopped IFN after less than three months, most of them for non-compliance. Among the 11 patients who completed a 6-12 month period of IFN treatment, 3 (27%) normalized and maintained normal ALT levels during therapy: for 2 of them the response was sustained after IFN discontinuation. The response to IFN therapy was neither correlated to the CD4+ levels nor to the clinical stage of the HIV infection. Instead, the response seemed to be influenced by pre-therapy ALT levels and liver histology. Tolerance to IFN treatment was good. These data show that IFN may be indicated in the therapy of chronic HCV infection for HIV-positive patients.

The action of VIRKON No Foam on the hepatitis B virus.

The aim of this work was to verify the in vitro virucidal activity of VIRKON No Foam (VIRKON NF) against the HBV DNA, notoriously one the most resistant viruses to heat and to the most commonly used disinfectants. VIRKON NF, an oxidizing agent, has a synergic effect on nucleic acids, polypeptides, glycoproteins, and structural proteins. The experiment was conducted using the serum of a patient with HBsAg+/HBeAg+/HBVDNA+ with a DNA concentration 100 pg/ml. The virucidal activity was tested using the Slot Blot method, the result being evaluated as an inhibition of the autoradiographic signal because of contact of different concentrated solutions and various contact times with the serum. Preliminary tests were carried out after contact of VIRKON NF with serum solutions at 0.1-0.5-1-4% concentrations for a 15 min contact time. VIRKON NF showed an inhibition of autoradiographic signal equal respectively to 0%, 25-50%, 50-70%, 100%. Further experiments were carried out to compare VIRKON NF virucidal activity on HBV to the activity of the most common disinfectants, such as formic aldehyde, Amuchina, glutaraledhyde and phenol. In this way, the infected serum was put into contact with both the most common disinfectant and 1-1.5-2-3-4% VIRKON NF solutions in contact times of 15, 10, 5, 2 min. Using 3% concentrations for a contact time of 10 min, VIRKON NF showed an autoradiographic inhibition signal of 90%. Regarding other disinfectants, only a 2% glutaraldehyde solution for a contact time of 15 min showed similar virucidal activity.

[Diagnosis of HCV infection by 3rd-generation ELISA in HCV-RNA positive hypertransaminasemic patients with 2nd-generation ELISA negative results]. / Diagnosi di infezione da HCV con ELISA di 3 generazione in pazienti con ipertransaminasemia cronica HCV-RNA positivi ma negativi all'ELISA di 2 generazione.

To evaluate the improved sensitivity of 3rd-generation assays for the detection of ani-HCV antibodies in diagnosing cases of HCV infection, we have re-tested by 3rd-generation ELISA test (ELISA-3) serum samples from immunocompetent patients with chronic hypertransaminasemia who were HCV-RNA positive but tested negative with 2nd-generation ELISA (ELISA-2). Out of 21 HCV-RNA positive/ELISA-2 negative samples, 3 (14.3%) were ELISA-3 positive. Among the ELISA-3 reactive samples, two were indeterminate by RIBA-3 (one was reactive with c1 00 and the other with c22), and one was negative. These results demonstrate that even in the clinical setting ELISA-3 improves the diagnosis of HCV infection. The improvement seems to be related to a better reactivity of HCV peptides rather than to the inclusion of the new determinant NS5. However, the sensitivity of the tests for the detection of anti-HCV antibodies remains to be improved.

Semiquantitative anti-HBc IgM detection in children with chronic hepatitis B: a long-term follow-up study.

Serum anti-HBc IgM titres were monitored monthly by a semiquantitative method in 14 children with HBeAg positive chronic hepatitis B followed up for 18-65 months. All patients, but one, were treated with alfa-interferon (IFN) at different times. On the whole, 12 flare-up episodes were observed and 7 patients cleared HBV-DNA and seroconverted to anti-HBe. Seroconversion occurred only in patients with pretreatment anti-HBc IgM index greater than 0.15 and serum HBV-DNA concentration below 100 pg/ml; the pretreatment alanine aminotransferase (ALT) value was not predictive of response. Combining anti-HBc IgM results and serum HBV-DNA levels observed during the pre-IFN period allowed a precise identification of patients who were likely to respond to IFN therapy. Patients who seroconverted to anti-HBe showed a progressive reduction in serum anti-HBc IgM titres within 6 months. Interestingly, one child, in whom HBV-DNA reappeared and who reconverted to HBeAg 7 months after treatment, showed no anti-HBc IgM decrease after the transient clearance of HBV-DNA and anti-HBe seroconversion. Semiquantitative anti-HBc IgM detection is a useful tool in the decision making process for children with chronic hepatitis B.

[Role of hepatitis C virus in mixed cryoglobulinemia in patients with chronic hepatitis]. / Ruolo del virus dell'epatite C nella crioglobulinemia mista in pazienti con epatite cronica.

To evaluate the role played by hepatitis C virus (HCV) in mixed cryoglobulinemia in subjects with chronic hepatitis, we studied 72 consecutive patients: 43 had HCV-related chronic hepatitis, 19 HBV-related chronic hepatitis, and 10 chronic hepatitis of other etiology. We used second generation ELISA and RIBA to test for mixed cryoglobulinemia and anti-HCV antibodies in both serum and cryoprecipitates; HCV RNA were evaluated by "nested" PCR. Serum levels of rheumatoid factor and complement were also determined. The immunoglobulins in the cryoimmunoprecipitate were characterized by immunofixation electrophoresis. Cryoglobulinemia was present in 47% of the patients with chronic hepatitis C but in none of the sera of patients with HBV-related chronic hepatitis nor in those with chronic hepatitis of non-viral etiology. Type II mixed cryoglobulinemia was observed in 45% of the cases, and type III in 55%. HCV RNA and anti-HCV antibodies were present in all the cryoimmunoprecipitates. Ninety-five percent of the cryoglobulinemic patients had serum rheumatoid factor and 80% of them had low serum levels of C4. Our data indicate that mixed cryoglobulinemia is frequently associated with HCV-related chronic hepatitis, and that HCV and anti-HCV antibodies play an essential role in the development of mixed cryoglobulinemia.

Treatment of chronic hepatitis B in children with prednisone followed by alfa-interferon: a controlled randomized study.

The efficacy and safety of sequential treatment with prednisone and interferon was evaluated in a randomized, controlled study on 43 children with biopsy proven HBsAg/HBeAg/hepatitis B virus-DNA positive, anti-delta negative, chronic hepatitis (34 chronic persistent hepatitis, 9 chronic active hepatitis). Patients received either a 1-month course of prednisone (0.6 to 0.3 mg/kg per day) followed by interferon alfa-2a (3 MU/m2, thrice weekly, for 12 months; 22 patients) or no treatment (21 patients). At the end of the study (20 months), clearance of hepatitis B virus-DNA and HBeAg seroconversion were observed in nine (41%) of the patients treated with prednisone and interferon and in two (9.5%) of the untreated controls (p = 0.020). Two of the treated patients who lost HBeAg, also cleared HBsAg. In the treated group, 13 (59%) patients had stable normal levels of alanine aminotransferase on their last examination. The baseline serum level of hepatitis B virus-DNA was an important predictor of response. In fact, HBeAg clearance was observed in 75% of patients with a baseline hepatitis B virus-DNA level lower than 100 pg/ml and in none with a level above 100 pg/ml. We suggest that combined treatment with prednisone followed by alfa-interferon may be safe and effective in inducing a stable clearance of HBeAg and, in some cases, of HBsAg in children with chronic hepatitis B and with a low level of viral replication. For children with high levels of viral replication, this regimen seems to be ineffective.

Cytomegalovirus as a co-factor of disease progression in human immunodeficiency virus type 1 infection.

Cytomegalovirus has been suggested as a co-factor of disease progression in patients with human immunodeficiency virus type 1 infection. Cytomegalovirus infection is highly prevalent among populations at risk of human immunodeficiency virus 1 infection, and has been associated with both an increased susceptibility to infection and a more rapid course of the disease towards immunodeficiency. Cytomegalovirus can have a direct immunosuppressive effect (through infection of immune cells) and can enhance the replication of human immunodeficiency virus (through the transactivation of the genic immunodeficiency virus expression, the stimulation of cytokine production, and the increase in Fc receptor expression on target cells). The role of cytomegalovirus as a co-factor of the progression towards immunodeficiency in subjects infected with the human immunodeficiency virus type 1 needs to be elucidated with more extensive clinical studies and the application of new molecular biology techniques.

Identification of a human immunodeficiency virus type 1 TAR binding protein in human hepatoblastoma HepG2 cells that trans-activates HIV-1 LTR-directed gene expression.

Recently, we have shown that the human immunodeficiency virus (HIV-1) long terminal repeat (LTR) directed chloramphenicol acetyltransferase (CAT) gene is efficiently expressed in human hepatoblastoma HepG2 cells and these cells can support productive HIV-1 replication. In this study we show that HepG2 cells contain a nuclear factor that binds to the HIV-1 trans-activating region (TAR), which we named HepG2-derived TAR binding protein (HTBP). Gel retardation assays using synthetic oligonucleotide probes carrying different mutations in the TAR region and competition DNA mobility-shift experiments using these oligonucleotides revealed the binding site encompassing between +7 and +13 nucleotides (5'-TCTGGTT-3') in the HIV-1 LTR. An in vivo CAT competition assay using -65HIV-1 LTR CAT as a reporter plasmid and various competitor plasmids containing these mutated oligonucleotides also demonstrated that HTBP can influence the HIV-1 LTR-directed CAT gene expression in HepG2 cells by interaction with a specific sequence in the TAR region.

Inhibition of HIV-1 productive infection in hepatoblastoma HepG2 cells by recombinant tumor necrosis factor-alpha.

OBJECTIVE: To evaluate the role of liver cells in and the effect of tumor necrosis factor-alpha (TNF-alpha) on HIV-1 replication. METHODS: Human hepatoblastoma HepG2 cells were infected with various strains of HIV-1 and the effect of TNF-alpha treatment, either before or after infection, was monitored by p24 antigen assays. Northern blot analysis and gel retardation assays were performed to determine the expression of CD4 and HIV-1 trans-acting region (TAR)-binding proteins in these cells, respectively. RESULTS: HepG2 cells are CD4+ and support active HIV-1 replication, producing infectious virions, as measured by both p24 production and ability to infect T-cell lines with the virus produced by HepG2 cells. In contrast to the stimulatory effect of TNF-alpha on HIV-1 replication in T-cells and monocytes, up to 200 U/ml TNF-alpha treatment, at various times, either before or after HIV-1 infection, substantially inhibited p24 antigen production in HepG2 cells without causing any remarkable cytotoxicity. Gel-retardation assay revealed enhancement of a DNA-binding protein in TNF-alpha-treated HepG2 cells that binds to a specific sequence of the HIV-1 TAR, compared with the untreated control. CONCLUSIONS: These results indicate the importance of cellular factor(s) in HIV-1 infection and suggest that cytokines in different tissues can induce opposite effects. TAR-binding protein may act as an inhibitory factor for HIV-1 replication in the HepG2 cell line.

Relation of changes in hepatitis B virus replication markers to the outcome of interferon treatment in patients with chronic hepatitis.

The relationship between the behavior of hepatitis B virus (HBV) replication markers and the response to treatment with recombinant alpha-2b interferon (IFN) was investigated in 11 patients with chronic hepatitis. At the end of 6 months of treatment, 4 patients showed a complete response to IFN: 2 more patients had seroconversion to HBeAb after 8 and 9 months of follow-up, respectively. The response to IFN was partial in the remaining patients. Pre-treatment levels of HBV DNA in patients showing complete response were lower than pre-treatment levels in patients with partial response: in addition, serum HBV DNA clearance during the treatment was associated with sustained remission more frequently than changes in the HBeAg/HBeAb system.

Levels of anti-albumin autoantibodies in acute and chronic hepatitis B virus infection.

Levels of anti-albumin autoantibodies (AAA) of the IgG class were determined by ELISA in sera of patients with acute and chronic hepatitis B virus (HBV) infection. Mean AAA levels were higher than normal in both acute and chronic hepatitis patients. AAA levels were higher than the upper normal limits in practically all patients with acute self-resolving hepatitis, and decreased to normal levels when the patients recovered. Enhanced IgG AAA levels were observed in many patients with chronic hepatitis and serological markers of HBV replication. Elevated AAA levels were not associated with either more elevated transaminase levels or more severe histological forms of chronic hepatitis. The results of this study suggest that the interaction of albumin with HBV determinants is involved in AAA elevations, probably by mediating an increase in albumin immunogenicity. Moreover, the fall in AAA levels in the recovery phase of acute hepatitis, the coexistence of elevated levels with HBeAg and HBV-DNA, and the lack of correlation between AAA levels and different evolutive forms of chronic hepatitis, seem to exclude AAA from playing a relevant role, be it protective or damaging, during HBV infection.

Subclass characterization of IgG anti-S.typhi lipopolysaccharide antibodies with commercial monoclonal antibodies.

We have evaluated several commercial monoclonal antibodies, specific for human IgG subclasses, for their reactivity in an ELISA test for the characterization of subclasses of IgG anti- Salmonella typhi lipopolysaccharide (LPS) antibodies. Four monoclonals, each specific for a single IgG subclass, were chosen for their good reactivity. In 19 typhoid and non-typhoid serum samples, the sum of the ELISA values obtained with the four subclass-specific monoclonals was highly correlated with the ELISA values obtained with a monoclonal anti-total IgG. Moreover, there was no competition among the various IgG subclasses of anti-LPS antibodies. These data indicate that the subclass distribution of IgG anti-LPS antibodies, calculated on the basis of ELISA values in the subclass-specific assays, is likely to reflect the actual distribution of the different subclasses in whole serum. Different subclass patterns of IgG anti-LPS antibodies were observed in serum samples from 11 patients with typhoid fever, with IgG1 and IgG2 being the most represented subclasses. The ELISA method described here will be useful to elucidate the factors that influence the anti-LPS subclass profile during the humoral immune response to Salmonella typhi.

An extensive search for RFLP in the human glucose-6-phosphate dehydrogenase locus has revealed a silent mutation in the coding sequence.

The genetic polymorphism of an approximately 100-kb DNA region comprising and flanking the glucose-6-phosphate dehydrogenase (G6PD) gene on human chromosome Xq28 has been analyzed in detail. By using 14 unique sequence probes and 18 restriction enzymes, we have characterized 257 restriction fragments or 370 restriction sites. On testing 12-57 individual X chromosomes, all sites but one were nonpolymorphic. However, a PstI site that maps to exon 10 of the G6PD gene, which is still monomorphic in all British and Italian subjects tested, is polymorphic in west-African people. Specifically, it is absent from 22% of Nigerian X chromosomes. By sequence analysis we have shown that the absence of this PstI site results from a G----A replacement at position 1116, corresponding to the third base of a glutamine codon; no amino acid change is produced in the protein. Thus, a polymorphic silent mutation is demonstrated in a human gene.

Performance characteristics of an enzyme-linked immunosorbent assay for the determination of immunoglobulin M anti-Salmonella typhi lipopolysaccharide antibodies.

Some characteristics of an enzyme-linked immunosorbent assay for the determination of immunoglobulin M anti-Salmonella lipopolysaccharide antibodies have been assessed. Immunoglobulin M anti-lipopolysaccharide antibodies in serum samples from most patients with typhoid fever cross-reacted with S. typhimurium lipopolysaccharide. However, absorbance values with S. typhi lipopolysaccharide were always higher than with the heterologous antigens. Serum samples from two patients with S. paratyphi B infection failed to show significant immunoglobulin M binding to S. typhi lipopolysaccharide. Serum samples from six rheumatoid arthritis patients were negative in this immunoenzymatic assay with S. typhi or S. typhimurium lipopolysaccharide as coating antigen. Finally, interference by specific immunoglobulin G in the immunoglobulin M anti-S. typhi lipopolysaccharide antibodies determination was excluded.

Sensitive ELISA for IgG and IgM anti-albumin autoantibodies not influenced by HBsAg-associated polyalbumin receptors.

An enzyme immunoassay for the detection of IgG and IgM anti-polymerized albumin autoantibodies (AAA) is described. It was found that polyalbumin receptors on HBsAg particles interfere in the detection of IgG AAA when polymerized human albumin (pHSA), but not polymerized bovine albumin (pBSA), is used as coating antigen. Polyalbumin receptors do not appear to interfere in the detection of IgM AAA, with either pHSA or pBSA as coating antigen. All normal sera showed evidence of AAA, of both IgG and IgM classes. Levels of IgG and IgM AAA in sera from most type A and type B acute hepatitis patients were above the range of normal controls. ELISA detection of AAA distinct from HBsAg reactivity can help in understanding the role of these autoantibodies in HBV infection.