Denosumab for the treatment of HIV-associated osteoporosis with fractures in a premenopausal woman.

The prevalence of osteoporosis is about three times greater in people living with HIV than in the general population. Bisphosphonates are the only class of antiresorptive drugs which have proved to be safe and effective in HIV patients. However, bisphosphonates are not recommended in women of childbearing age due to an increased rate of associated neonatal complications. To the best of our knowledge no reports on the use of denosumab in HIV-infected individuals have been published so far. We describe a 38 year-old woman with HIV, osteoporosis and vertebral fractures treated with denosumab, a monoclonal antibody targeting RANKL. After four years of treatment, bone mineral density improved, no new fractures occurred, and neither HIV reactivation nor opportunistic infections were observed. We show that denosumab could be a safe and effective approach for osteoporosis in patients with HIV and could be considered in women of childbearing age.

The Management of Geriatric and Frail HIV Patients. A 2017 Update from the Italian Guidelines for the Use of Antiretroviral Agents and the Diagnostic Clinical Management of HIV-1 Infected Persons.

OBJECTIVE: This article deals with the attempt to join HIV and geriatric care management in the 2017 edition of the Italian guidelines for the use of antiretrovirals and the diagnostic-clinical management of HIV-1 infected persons. METHODS: The outlined recommendations are based on evidence from randomized clinical trials and observational studies published in peer-reviewed journals and/or presented at international scientific conferences in recent years. The principles of starting antiretroviral therapy in elderly patients and the viro-immunological goals are the same as in the general HIV population. However, there are some specificities to consider, related to the host as well as the therapy itself. HIV care in elderly patients must shift from a combined AntiRetroviral Therapy specific approach to a more comprehensive management, and from a disease-based model (list of co-morbidities) to a multi-morbidity and frailty standpoint. The implementation of a geriatric approach, based on the Comprehensive Geriatric Assessment, is essential and consists of a broader evaluation of health status. This multidimensional and multidisciplinary evaluation is focused on the development of a tailored intervention plan. Polypharmacy is a frequent condition in the older population and an independent risk factor for negative health-related outcomes. This can be overcome with a multidisciplinary and cooperative approach involving HIV specialists, geriatricians and primary care physicians. CONCLUSION: The inclusion of geriatric care becomes necessary due to the novel needs of an evolving patient population. It is important to underline that the HIV specialist will continue to lead multidimensional interventions and optimize quality of care for HIV-positive people.

Dolutegravir (DTG)-containing regimens after receiving raltegravir (RAL) or elvitegravir (EVG): Durability and virological response in a large Italian HIV drug resistance network (ARCA).

BACKGROUND: Dolutegravir (DTG) is a next-generation HIV integrase inhibitor (INI) with an increased genetic barrier to resistance with respect to raltegravir (RAL) or elvitegravir (EVG). Few data are available on the durability of DTG-containing regimens. OBJECTIVES: We aimed at investigating the duration of the DTG-containing regimen, the occurrence of an HIV-1 RNA blip, and factors associated with DTG virological response. STUDY DESIGN: From the Antiviral Response Cohort Analysis database, we selected 89 HIV-1-positive four-class-experienced subjects who started DTG after receiving RAL or EVG. Factors associated with durability and virological response were analysed by logistic regression. RESULTS: After a median duration of 18.8 [0.4-76.2] months, 79/89 (88.8%) subjects were still on DTG. All subjects remaining on DTG at the end of follow-up had undetectable HIV-1 RNA, compared to 5/10 subjects who discontinued DTG. DTG discontinuation was less frequent in patients who had experienced ≥10 regimens (HR 0.11, p = 0.040). The probability of having an HIV-1 RNA positive value at the last follow-up significantly increased in patients with non-B HIV-1 subtype (HR 5.77, p < .001) and significantly decreased in patients with CD4 nadir >200/µL (HR 0.29, p = 0.038), with more than 10 previous regimens (HR 0.27, p = 0.040), and who harbored virus with IN mutations (HR 0.12, p = 0.023) at DTG start. CONCLUSIONS: After previous exposure to first-generation INIs, treatment with DTG showed long durability and did not show virological rebound after virological suppression. Subjects infected with a non-B HIV-1 subtype had a greater risk of having detectable HIV-1 RNA at the last observation.

Genotypic testing on HIV-1 DNA as a tool to assess HIV-1 co-receptor usage in clinical practice: results from the DIVA study group.

PURPOSE: We have developed a sequencing assay for determining the usage of the genotypic HIV-1 co-receptor using peripheral blood mononuclear cell (PBMC) DNA in virologically suppressed HIV-1 infected patients. Our specific aims were to (1) evaluate the efficiency of V3 sequences in B versus non-B subtypes, (2) compare the efficiency of V3 sequences and tropism prediction using whole blood and PBMCs for DNA extraction, (3) compare the efficiency of V3 sequences and tropism prediction using a single versus a triplicate round of amplification. RESULTS: The overall rate of successful V3 sequences ranged from 100 % in samples with >3,000 copies HIV-1 DNA/10(6) PBMCs to 60 % in samples with <100 copies total HIV-1 DNA /10(6) PBMCs. Analysis of 143 paired PBMCs and whole-blood samples showed successful V3 sequences rates of 77.6 % for PBMCs and 83.9 % for whole blood. These rates are in agreement with the tropism prediction obtained using the geno2pheno co-receptor algorithm, namely, 92.1 % with a false-positive rate (FPR) of 10 or 20 % and of 96.5 % with an FPR of 5.75 %. The agreement between tropism prediction values using single versus triplicate amplification was 98.2 % (56/57) of patients using an FPR of 20 % and 92.9 % (53/57) using an FPR of 10 or 5.75 %. For 63.0 % (36/57) of patients, the FPR obtained via the single amplification procedure was superimposable to all three FPRs obtained by triplicate amplification. CONCLUSIONS: Our results show the feasibility and consistency of genotypic testing on HIV-1 DNA tropism, supporting its possible use for selecting patients with suppressed plasma HIV-1 RNA as candidates for CCR5-antagonist treatment. The high agreement between tropism prediction by single and triple amplification does not support the use of triplicate amplification in clinical practice.

Factors associated with virological success with raltegravir-containing regimens and prevalence of raltegravir-resistance-associated mutations at failure in the ARCA database.

Raltegravir (RAL) is the only licensed human immunodeficiency virus (HIV) integrase inhibitor. The factors associated with the virological response to RAL-containing regimens and the prevalence of integrase mutations associated with RAL failure deserve further investigation. From the Antiretroviral Resistance Cohort Analysis database, we selected triple-class-experienced subjects failing their current treatment with complete treatment history available. Selection criteria included HIV-RNA, CD4 count and HIV genotype within 3 months of RAL initiation. Factors associated with 24-week response were analysed; genotypic sensitivity scores (GSS) and weighted-GSS were evaluated. Virological response was achieved in 74.3% of 105 subjects. Mutations associated with RAL failure were detected in 12/24 subjects with an integrase genotype, with the prevalence of Q148H + G140S. Each extra unit of GSS (p 0.05, OR 2.62; 95% CI 1.00-6.87). was found to be a associated with response. Weighted-GSS had borderline statistical significance (p 0.063, OR 2.04; 95% CI 0.96-4.33) When stratifying for different cut-offs (<1 as reference, 1-1.49, ≥1.5), a borderline significant increase in the probability of response appeared for GSS ≥1.5 (p 0.053, OR 4.00; 95% CI 0.98-16.25). GSS ≥1 showed the highest sensitivity, 82.6%. Receiver operating characteristic curves depicted the widest area under the curve (0.663, p 0.054) of GSS ≥1. Unresponsiveness to RAL-containing regimens among triple-class-experienced subjects was low. The activity of the background regimen was strongly associated with response. Although few integrase genotypes were available at failure, half of these were without integrase resistance mutations. The substantial rate of RAL failure in the absence of known RAL-resistance mutations may be associated with adherence issues and this issue warrants further analysis in longer observations.

A new approach to safely type for HLA the HIV infected people eligible to abacavir therapy: saliva or buccal swab as reliable DNA sources.

BACKGROUND: Increasing the safety in Immunogenetics Labs, in the era of antiretroviral pharmacogenomics, represents an imperative goal. To this purpose, we tested saliva and buccal cells as biological sources of DNA, alternative to peripheral blood, for HLA-B*57:01 genomic typing of HIV positive patients eligible to treatment with abacavir. METHODS: Blood, saliva and buccal cells of 20 voluntary donors and 20 HIV positive patients were collected. DNA was extracted with a manual commercial kit and an automated platform. Quality and quantity of DNA was evaluated with different procedures. The suitability and reliability of DNAs for HLA-B*57:01 genotyping was checked at low and high resolution level, using PCR-SSP (sequence specific primers PCR), revPCR-SSO (reverse sequence specific oligonucleotides PCR), bead array and SBT (sequence based typing) techniques. RESULTS: DNA concentrations were qualitatively very good and quantitatively comparable in all the specimens tested with an inferior yield for cotton swabs. Comparing the results of HLA typing with different methodologies, the 100% of reproducibility was achieved. CONCLUSIONS: The viral load of buccal epithelial cells or saliva is extremely low. Here we demonstrated that the DNA from these alternative sources is appropriate for HLA-B*57:01 typing. We strongly recommend the use of this procedure to increase the safety in the lab when dealing with infectious samples.

Neuropsychological evaluation and follow up in jcv- and non-jcv-related leukoencephalopathies in HIV infection.

The introduction of highly active antiretroviral therapy does not seem to have altered the incidence of progressive multifocal leukoencephalopathy (PML) in HIV infection. Moreover, the occurrence of a HIV-related leukoencephalopathy, called not determined leukoencephalopaties (NDLE), has been reported. As neuropsychological impairment remains highly prevalent in HIV infection, the aim of this study is to describe the neuropsychological profile of PML and NDLE patients, analyzing the time-related changes. Clinical and neuropsychological data from 32 patients (17 PML, 15 NDLE) were compared with two control groups: (1) asymptomatic HIV+ patients without magnetic resonance imaging evidence of leukoencephalopathy; (2) age-/gender-/education-matched healthy subjects. Patients with rapidly worsening PML were significantly impaired on all neuropsychological tests, while PML with more benign course and NDLE groups showed a dysexecutive pattern of impairment. Asymptomatic HIV+ subjects showed mild and isolated cognitive deficits, without functional impact. Cognitive impairment should therefore be considered a key feature from HIV infection diagnosis.

HIV-related acute inflammatory leukoencephalopathy of undetermined origin: review of the literature.

HIV-related acute inflammatory leukoencephalopathy of undetermined origin (AIL) has been anecdotally described in literature as being responsible for cognitive and motor deficits. We carried out a review of all the cases of AIL published in literature. Articles were selected according to 2 criteria: acute onset of symptoms; undetermined aetiology and non-fulfilment of multiple sclerosis diagnostic criteria. They were then analyzed in terms of clinical, biological and instrumental features, therapy, diagnostic classification and prognosis. Although rare (21 patients out of about 4,000 publications), AIL is of particular interest, as the comprehension of its mechanisms could give some insight into the direct and immune-mediated actions of HIV within the brain. All the reported patients share several clinical, histopathological, radiological and CSF features, leading to hypothesize a similar aetiopathogenetic mechanism. Conversely, we observed a high heterogeneity of treatment and diagnostic classification, which could have conditioned the broad prognostic variability. The absence of a defined aetiology leads to consider these forms as a particular subgroup of not determined leucoencephalopathies (NDLE), with both MRI and histological pattern dominated by inflammation as distinctive feature.

Prevalence of transmitted HIV-1 drug resistance in HIV-1-infected patients in Italy: evolution over 12 years and predictors.

OBJECTIVES: Transmitted HIV-1 drug resistance (TDR) can reduce the efficacy of first-line antiretroviral therapy. PATIENTS AND METHODS: A retrospective analysis was performed to assess the prevalence and correlates of TDR in Italy over time. TDR was defined as the presence of at least one of the mutations present in the surveillance drug resistance mutation (SDRM) list. RESULTS: Among 1690 antiretroviral therapy-naive patients, the most frequent HIV subtypes were B (78.8%), CRF02_AG (5.6%) and C (3.6%). Overall, TDR was 15%. TDR was 17.3% in subtype B and 7.0% in non-B carriers (P < 0.001). TDR showed a slight, although not significant, decline (from 16.3% in 1996-2001 to 13.4% in 2006-07, P = 0.15); TDR declined for nucleoside reverse transcriptase inhibitors (from 13.1% to 8.2%, P = 0.003) but remained stable for protease inhibitors (from 3.7% to 2.5%, P = 0.12) and non-nucleoside reverse transcriptase inhibitors (from 3.7% to 5.8%). TDR to any drug was stable in B subtype and showed a decline trend in non-B. In multivariable analysis, F1 subtype or any non-B subtype, compared with B subtype, and higher HIV RNA were independent predictors of reduced odds of TDR. CONCLUSIONS: Prevalence of TDR to nucleoside reverse transcriptase inhibitors seems to have declined in Italy over time. Increased prevalence of non-B subtypes partially justifies this phenomenon.

Hyperbilirubinemia during atazanavir treatment in 2,404 patients in the Italian atazanavir expanded access program and MASTER Cohorts.

BACKGROUND: Although the mechanism of atazanavir (ATV)-related hyperbilirubinemia is well identified, its prevalence, risk factors, and association with transaminase flares have rarely been assessed in a large population from the "real life" setting. METHODS: Prospectively collected data on 2,404 patients from the Italian MASTER Cohort and the Italian ATV expanded access program database were examined. Uni- and multivariable Cox proportional hazards regression models were conducted to identify risk factors for grade >or= III hyperbilirubinemia during the administration of ATV. The risk of increased levels of serum alanine aminotransferase (ALT) was compared between patients with or without grade >or= III hyperbilirubinemia in a Cox regression analysis stratified by hepatitis C virus (HCV) serostatus. RESULTS: Grade III and IV hyperbilirubinemia were observed in 1,072 (44.6%) and 174 (7.2%) of the patients, respectively. Higher CD4+ T-cell counts, abnormal bilirubinemia at baseline, and ritonavir co-administration were associated with a higher risk of developing grade >or= III hyperbilirubinemia. In contrast, female gender, clinical class C, and non-nucleoside reverse transcriptase co-administration appeared to be protective. Higher bilirubinemia at baseline and the use of ritonavir were associated with a higher risk of grade IV hyperbilirubinemia. The occurrence of grade >or= III hyperbilirubinemia was not associated with severe hepatotoxicity (hazard ratio 1.00, 95% confidence interval 0.64-1.57; p = 0.997). CONCLUSIONS: Hyperbilirubinemia is a common side effect of an ATV pharmacotherapeutic regimen. However, grade IV increase in bilirubin was rarely found. In most cases, ATV hyperbilirubinemia appeared to be an innocent phenomenon as far as the risk of a subsequent increase in liver enzyme level is concerned.

Thymidine kinase and uridine-cytidine kinase from Entamoeba histolytica: cloning, characterization, and search for specific inhibitors.

Entamoeba histolytica is an intestinal parasite and the causative agent of amoebiasis, which is a significant source of morbidity and mortality in developing countries. Although anti-amoebic drugs such as metronidazole, emetine, chloroquine and nitazoxanide are generally effective, there is always potential for development of drug resistance. In order to find novel targets to control E. histolytica proliferation we cloned, expressed and purified thymidine kinase (Eh-TK) and uridine-cytidine kinase (Eh-UCK) from E. histolytica. Eh-TK phosphorylates thymidine with a Km of 0.27 microm, whereas Eh-UCK phosphorylates uridine and cytidine with Km of 0.74 and 0.22 mM, respectively. For both enzymes, ATP acts as specific phosphate donor. In order to find alternative treatments of E. histolytica infection we tested numerous nucleoside analogues and related compounds as inhibitors and/or substrates of Eh-TK and Eh-UCK, and active compounds against E. histolytica in cell culture. Our results indicate that inhibitors or alternative substrates of the enzymes, although partially reducing protozoan proliferation, are reversible and not likely to become drugs against E. histolytica infections.

Comparison of glomerular filtration rate estimates vs. 24-h creatinine clearance in HIV-positive patients.

BACKGROUND: Guidelines for kidney function monitoring and antiretroviral drug dosing are available and respectively refer to glomerular filtration rate and creatinine clearance (CrCl). OBJECTIVE: The aim of the study was to compare kidney function estimates vs. measured 24-h CrCl in HIV-infected subjects. METHODS: A cross-sectional design was used, with comparison of Cockcroft-Gault (CG), original and simplified modification of diet in renal disease (MDRD) equations vs. measured 24-h CrCl. Subjects were HIV-infected, 18-70 years old, without pre-existing kidney disease. RESULTS: Results are presented as mean (+/-standard deviation), unless otherwise stated. The study population consisted of 90 patients, of whom 71% were male, with a mean age of 45 years (+/-6.5 years). At the time of evaluation, the mean body mass index was 23 (+/-3.3); mean serum creatinine was 0.91 mg/dL (+/-0.2 mg/dL); and mean blood urea nitrogen (BUN) was 34.7 mg/dL (+/-10.6 mg/dL). Differences between paired methods were all significant (P<0.00001), except between CG and simplified MDRD (P=0.21; Pearson r=0.81). In univariate analysis, male gender, CD4 nadir, hepatitis B virus coinfection, BUN and current CD4 cell count showed a significant positive correlation (P<0.2) with the difference between measured 24-h CrCl and either CG or simplified MDRD estimates. In multivariate analysis, only BUN showed a significant positive correlation (P<0.05). CONCLUSIONS: Estimates were lower than the measurements of 24-h CrCl. Original MDRD estimates were lower than those with other equations. CG and simplified MDRD estimates showed a satisfactory correlation.

Two different patterns of mutations are involved in the genotypic resistance score for atazanavir boosted versus unboosted by ritonavir in multiple failing patients.

OBJECTIVES: The protease inhibitor atazanavir (ATV) can be used either boosted by ritonavir (ATV300/r) or unboosted (ATV400). To date, however, genotypic resistance scores (GRSs) have been developed only for boosted-ATV. We have determined GRS associated with virologic response (VR) for both ATV300/r and ATV400 in highly pre-treated HIV-1 infected patients. PATIENTS AND METHODS: We analyzed the results of genotypic tests available 0-3 months before the initiation of an ATV-containing regimen in 159 patients with HIV-RNA >or= 500 copies/ml (ATV300/r group: 74; ATV400 group: 85) who were enrolled in the CARe study through an Early Access Program. The impact of baseline protease mutations on VR (>or= 1 log(10)copies/ml HIV-RNA decrease at 12-24 weeks) was analyzed using Fisher's exact test. Mutated protease amino acid positions (MPP) with p < 0.20 were retained for further analysis. The GRSs were determined by a step-by-step analysis using the chi(2) test for trend. RESULTS: The GRSs for ATV300/r and ATV400 revealed differing sets of mutations. For ATV300/r, 12 MPPs (10C/I/V + 32I + 34Q + 46I/L + 53L + 54A/M/V + 82A/F/I/T + 84V + 90M - 15E/G/L/V - 69K/M/N/Q/R/T/Y - 72M/ T/V; p = 1.38 x 10(-9)) were the most strongly associated with VR (VR: 100%, 78.3%, 83.3%, 75% and 0% of patients with a score of -2/-1, 0, 1, 2, and >or= 3, respectively); the last three MPPs (I15/H69/I72) were associated with a better VR. For ATV400, nine MPPs (16E + 20I/M/R/T/V + 32I + 33F/I/V + 53L/Y + 64L/M/ V + 71I/T/V + 85V + 93L/M; p = 9.42 x 10(-8)) were most strongly associated with VR (VR: 83.3%, 66.7%, 5.9%, 0% of patients with 0, 1/2, 3, and >or= 4 MPP, respectively). Differences between GRSs for ATV300/r and ATV400 may be due to different ATV drug levels (boosted vs unboosted), favoring different pathways of escape from antiviral pressure. CONCLUSIONS: Both GRSs were independent predictors of response in a multivariable logistic regression model. Nevertheless, cross-validation of these GRSs on different patient databases is required before their implementation in clinical practice.

A comparison of three diagnostic techniques for malaria: a rapid diagnostic test (NOW Malaria), PCR and microscopy.

Malaria is a common, life-threatening infection in endemic tropical areas and one that presents a diagnostic challenge to laboratories in most non-endemic countries. A rapid and accurate diagnosis is a prerequisite for effective treatment, especially for the potentially fatal cases of Plasmodium falciparum infection. In the present, multi-centre study, the performances of a rapid diagnostic test (NOW) Malaria) and several, commercial, PCR-based assays (AMS61, AMS42, AMS43, AMS4 and AMS45) were compared against the results of microscopical examination of bloodsmears (the current 'gold standard'). The subjects were either non-European immigrants (N=135) or international travellers (N=171). There was good concordance between the results of all the detection methods, with kappa values of >0.8. Although the NOW Malaria rapid test was both sensitive (100%) and specific (100%) in detecting P. falciparum infections, it was less specific (93.1%) and sensitive (90.7%) in identifying the other Plasmodium species. The results from the AMS61 assay, designed to detect any malarial infection, generally parallelled those of the microscopy (kappa = 0.89), giving a specificity of 98.2% and a sensitivity of 91.0%. Although the use of species-specific molecular primers to identify pure infections with P. falciparum and P. vivax gave results that were in good agreement with those of the microscopy, the subjects who had apparently pure infections with P. ovale or P. malariae were always found PCR-negative. Compared with the standard microscopy, both the NOW Malaria test and the PCR-based assays were therefore poor at identifying mixed infections. The NOW Malaria test and the PCR-based assays clearly need to be improved, particularly for the correct identification of infections with Plasmodium spp. other than P. falciparum, including mixed infections. For now, expert microscopy must remain the mainstay of the laboratory diagnosis of malaria.

Control of human cytomegalovirus infection in patients infected with human immunodeficiency virus by high levels of specific CD8+ T-cells.

A new technique was used to simultaneously determine human cytomegalovirus (HCMV)-specific CD4(+) and CD8(+) T-cells in highly active anti-retroviral therapy (HAART)-naive and HAART-treated patients infected with human immunodeficiency virus (HIV). HIV-infected patients with HCMV infection, but without HCMV disease, showed low numbers of HCMV-specific CD4(+) cells and high numbers of CD8(+) T-cells, both before and during HAART. HIV-infected patients with HCMV disease had no HCMV-specific CD4(+) T-cells and extremely low levels of CD8(+) T-cells. Resolution of disease during HAART was associated with rescue of specific CD4(+) T-cells and a large increase in the specific CD8(+) T-cell count. Thus, HAART does not completely restore the normal immune function. In HIV-infected patients, sustained control of HCMV infection requires high frequencies of specific CD8(+) T-cells.

Key questions in antiretroviral therapy: Italian Consensus Workshop (2005).

A panel of leading Italian specialists in infectious diseases, virologists and immunologists met in Rome in 2005 to review critical data and discuss recommendations for each of the key questions in antiretroviral therapy today: When to start treatment? How to start? When to switch? What to switch to? Whether to stop or not to stop treatment, and how? The method of a nominal group meeting was used and recommendations were graded for their strength and quality using a system based on the one adopted by the Infectious Diseases Society of America. Main conclusions are summarized and critically discussed in this consensus statement, as well as some of the most recent data supporting these recommendations are provided.

Diagnosis of visceral leishmaniasis: the sensitivities and specificities of traditional methods and a nested PCR assay.

In the present study, 67 patients suspected to be cases of visceral leishmaniasis (VL) were each checked for leishmanial infection by the microscopical evaluation of various biological specimens, in-vitro culture, serology and an assay based on nested PCR. Most (35) of the subjects were immunocompetent (IC) but 32 were immunodeficient (ID) as the result of HIV infection (18 cases), treatment to prevent transplanted organs being rejected (six) or haematological malignancies (eight). Forty-one (61.2%) of the subjects (19 IC subjects, 12 HIV-positive patients, four transplant patients and six patients with malignancies) were considered true cases of VL. For the IC subjects, only the production and microscopical examination of leucocytoconcentrates and cultures of Buffy coats gave sensitivities of <80%, the results of the other methods showing higher sensitivities and almost perfect agreement with the 'gold-standard' diagnoses. For the ID subjects, however, only the serological tests and the PCR gave reasonable sensitivities (of >80%). For the initial diagnosis of leishmaniasis in ID patients, IFAT and western blots may be useful, as, among the present ID patients, they gave sensitivities (of 80.9% and 88.2%, respectively) that were almost as high as that for the PCR, and specificities of 100%. In the diagnosis of VL in either IC or ID patients, the assay based on a nested PCR appeared to be particularly reliable, with sensitivities of 88.9% and 95.2%, respectively, and a specificity of 100% in both groups of patients. The testing of bone-marrow aspirates by PCR revealed very few VL cases who were not found positive when samples of their peripheral blood were checked in the same assay. For both IC and ID subjects therefore, the use of the PCR-based method to test samples of peripheral blood (which can be collected much more easily than bone-marrow aspirates and with much less pain for the subject) is recommended.

Markers of local immunity in cervico-vaginal secretions of HIV infected women: implications for HIV shedding.

OBJECTIVES: To link local proinflammatory cytokines with HIV related nucleic acids in cervico-vaginal secretions and the factors associated with them. METHODS: An observational study on 60 HIV positive women attending the department of obstetrics and gynaecology, University of Pavia, Italy. HIV-1 RNA in plasma, proviral HIV-1-DNA, cell associated and cell free HIV-1 RNA in cervico-vaginal secretions were evaluated by competitive polymerase chain reaction (c-PCR) and reverse transcriptase PCR (cRT-PCR). IL-1beta, IL-6, and TNF-alpha were measured by ELISA in cervico-vaginal lavages. Multiple regression analysis on ordinal categorical variables was used to test for the simultaneous associations of clinical and microbiological variables on quartiles of cytokine concentrations in lavage samples. RESULTS: Proviral HIV-1 DNA, cell associated and cell free HIV-1 RNA were detected in 76.7% (46/60), 70% (42/60), and 71.7% (43/60) of the patients, respectively. IL-1beta concentration was directly correlated with proviral HIV-DNA (Spearman rho = 0.35, p = 0.01) and cell associated HIV-RNA levels (Spearman rho = 0.263, p = 0.05). IL-1beta concentration (153.9 pg/ml) was higher (p<0.05) among women with cytological squamous intraepithelial lesion (SIL) than negative controls (73.4 pg/ml). In women with vaginal infection both IL-1beta (41.7 pg/ml) and IL-6 (10.2 pg/ml) were lower (p<0.05) in comparison to negative controls (144.9 pg/ml and 23.7 pg/ml, respectively). Women receiving stable antiretroviral therapy had significantly lower TNF-alpha (34.4 pg/ml versus 44.4 pg/ml, p = 0.04) and higher IL-6 (24.0 pg/ml versus 1.4 pg/ml, p = 0.004) levels in lavage samples compared to untreated women. The associations between the presence of SIL, antiretroviral treatment, vaginal infection and cytokine concentrations in cervico-vaginal secretions were confirmed in multiple regression analysis. CONCLUSIONS: Local immune activation may modulate HIV-1 shedding in cervico-vaginal secretion with possible influence on vaginal physiology and host defence. Pharmacological agents lowering HIV-1 replication cause a shift to a pattern of cytokine production which seems less favourable to the transmission of the disease.

Assessment of atherosclerosis using carotid ultrasonography in a cohort of HIV-positive patients treated with protease inhibitors.

OBJECTIVE: Lipid disorders associated with the use of protease inhibitors (PI) may be a risk factor for premature atherosclerosis development. The aim of this study is to evaluate the extent of carotid intima media thickness (IMT) among HIV-positive patients treated with PI containing regimens compared to PI-naïve and HIV-negative subjects. METHODS: We analysed plasma lipid levels and carotid IMT in 28 HIV-positive patients treated with protease inhibitors (PIs) for a mean of 28.7 months (range 18-43) and in two control groups constituted, respectively, by 15 HIV-positive naïve patients and 16 HIV-negative subjects, that were matched for age, risk factors for HIV infection, cigarette smoke use and CD4+ cell count. RESULTS: PI-treated patients had higher triglyceride, HDL and apo B levels than controls. Carotid IMT was significantly increased in PI-treated patients compared to naïve or HIV-negative subjects. A correlation between cholesterol HDL, triglyceride and ApoB levels and IMT was observed among the entire cohort. CONCLUSIONS: Plasma lipid alterations were associated with an increased IMT and intima media thickening was more pronounced in PI-treated patients than in the two control groups. Periodical evaluation of blood lipid profile and, if required, the use of lipid-lowering agents is advisable. Moreover, physicians should address concurrent risk factor for atherosclerosis that can be modified, including smoking, hypertension, obesity and sedentary life-style.

Declining levels of rescued lymphoproliferative response to human cytomegalovirus (HCMV) in AIDS patients with or without HCMV disease following long-term HAART.

OBJECTIVE: To investigate the lymphoproliferative response (LPR) to human cytomegalovirus (HCMV) in two groups of AIDS patients undergoing long-term highly active antiretroviral therapy (HAART): group 1 ( n = 22) with nadir CD4(+) cell count <50/microl and no HCMV disease; group 2 ( n = 16) with <50/microl CD4(+) T-cell count and HCMV disease. All patients had previously undergone antiretroviral monotherapy or dual therapy before initiating HAART. STUDY DESIGN AND METHODS: The two groups of patients were tested prospectively for CD4(+) T cell count, HIV RNA load, HCMV viremia, and LPR to HCMV at baseline, and then after 3 and 4 years of HAART. A control group of 13 recently diagnosed treatment-naive AIDS patients with CD4(+) T-cell counts <100/microl was also investigated. RESULTS: No LPR to HCMV was found in any of the treatment-naive patients nor in any patient of the two groups examined at baseline, when HCMV viremia was 13.6% in the patient group without disease and 87.5% in the group with disease ( p <.0001). After 3 years of HAART, the frequency of patients who recovered an LPR to HCMV was not significantly different (81.8% in the group without HCMV disease, and 68.7% in the group with HCMV disease), whereas, compared with baseline, the HIV load decreased and the CD4(+) T-cell count increased significantly and to a comparable extent in the two groups of patients. In addition, the frequency of patients with HCMV viremia, although reduced, became comparable in both groups. After 4 years of HAART, the frequency of responders to HCMV without and with HCMV disease dropped to comparable levels (50.0 vs. 56.3%, respectively) in association with high median CD4(+) T-cell counts and low median HIV RNA plasma levels. In parallel, the frequency of patients with HCMV viremia did not change significantly. In addition, after between 3 and 4 years of HAART, although the frequency of stable responders and nonresponders remained unchanged (50%) in both groups, most of the remaining patients showed declining levels of responsiveness to HCMV. Although some patients from both groups were found to have CD4(+) T-cell counts >150/microl in the absence of LPR to HCMV, thus suggesting dissociation of specific and nonspecific immune reconstitution, a significant correlation was found between CD4(+) T-cell count and LPR to HCMV (r = 0.44; p <.001). From a clinical standpoint, anti-HCMV therapy could be safely discontinued in 8 patients with HCMV retinitis showing CD4(+) T-cell counts >150/microl, recovery of HCMV LPR, and no HCMV viremia. CONCLUSIONS: Declining levels of the previously recovered LPR to HCMV are often observed after long-term HAART. However, because the role of LPR in the evolution of HCMV infection and disease during HAART remains to be defined, the clinical impact of the declining LPR to HCMV must still be clarified in long-term prospective studies.