Search

Abstract: The occurrence of COVID-19 pandemic had a significant negative effect on health care systems over the last year. Health care providers were forced to focus mainly on COVID-19 patients, neglecting in many cases equally important diseases, both acute and chronic. Therefore, also screening and diagnostic strategies for HIV could have been significantly impaired.This retrospective, multicenter, observational study aimed at assessing the number and characteristics of new HIV/AIDS diagnoses during COVID-19 pandemic in Italy and compared characteristics of people living with HIV at diagnosis between pre- and post-COVID-19 era (2019 vs 2020).Our results showed a significant reduction of HIV diagnoses during pandemic. By contrast, people living with HIV during pandemic were older and were diagnosed in earlier stage of disease (considering CD4+ T cell count) compared to those who were diagnosed the year before. Moreover, there was a significant decrease of new HIV diagnoses among men who have sex with men, probably for the impact of social distancing and restriction applied by the Italian Government. Late presentation incidence, if numbers in 2020 were lower than those in 2019, is still an issue.Routinely performing HIV testing in patients with suspected SARS-CoV-2 infection is identifying and linking to care underdiagnosed people living with HIV earlier. Thus, combined tests (HIV and SARS-CoV-2) should be implemented in patients with SARS-CoV-2 symptoms overlapping HIV's ones. Lastly, our results lastly showed how urgent implementation of a national policy for HIV screening is necessary., Competing Interests: The authors report no conflicts of interest., (Copyright © 2021 the Author(s). Published by Wolters Kluwer Health, Inc.)

Objectives: HIV establishes a latent infection at different degrees within naïve (TN) or central (TCM) and effector memory (TEM) CD4 T cell. Studying patients in whom HIV production was suppressed by combined antiretroviral therapy, our main aim was to find which factors are related or can influence intracellular viral reservoir in different CD4 T-cell subsets., Methods: We enrolled 32 HIV patients successfully treated for more than 2 years, with a CD4 T-cell count more than 500 cells/μl and plasma viremia undetectable from at least 1 year. Proviral HIV-DNA, the amount of cells expressing signal-joint T-cell receptor rearrangement excision circles and telomere length were quantified by droplet digital PCR in highly purified, sorted CD4 T-cell subsets; plasma IL-7 and IL-15 were measured by ELISA., Results: HIV-DNA was significantly lower in TN cells compared with TCM or to TEM. Conversely, TN cells contained more signal-joint T-cell receptor rearrangement excision circles compared with TCM or to TEM; no appreciable changes were observed in telomere length. HIV-DNA content was significantly higher in TN and TCM cells, but not in TEM, from patients with shorter time of treatment, or in those with lower CD4 : CD8 ratio., Conclusion: Length of treatment or recovery of CD4 : CD8 ratio significantly influences viral reservoir in both TN and TCM. Measuring HIV-DNA in purified lymphocyte populations allows a better monitoring of HIV reservoir and could be useful for designing future eradication strategies.

Introduction: Scanty data exist on the phenotype and functionality of invariant natural killer T (iNKT) cells in HIV-infected (HIV+) patients., Methods: By flow cytometry, we studied iNKT cells from 54 HIV+ patients who started combined antiretroviral therapy and had undetectable viral load for more than 1 year. Twenty-five maintained a CD4/CD8 ratio less than 0.4, whereas 29 reached a ratio more than 1.1; 32 age-matched and sex-matched patients were healthy controls (CTR)., Results: Patients with low ratio had lower percentage of CD4 iNKT cells compared with patients with high ratio and higher CD8 iNKT cell percentage; double-negative iNKT cells were lower in HIV+ patients compared with CTR. Patients with low ratio had higher percentage of CD4 and double-negative iNKT cells expressing CD38 and HLA-DR compared with patients with high ratio. CD4 iNKT cells expressing PD-1 were higher in patients with CD4/CD8 ratio less than 0.4, whereas double-negative iNKT cells expressing PD-1 were lower compared with patients with ratio more than 1.1. Patients with low ratio had higher CD4 iNKT cells producing IL-17, CD8 iNKT cells producing IFN-γ, TNF-α or IFN-γ and TNF-α, and double-negative iNKT cells producing IL-17 or IL-17 and IFN-γ compared with CTR. Activated CD4 (or CD8) T cells correlated with activated CD4 (or CD8) iNKT cells, as well as the percentages of CD4 (or CD8) T cells expressing PD-1 was correlated to that of CD4 (or CD8) iNKT cells expressing PD-1., Conclusion: Low CD4/CD8 ratio despite effective combined antiretroviral therapy is associated with altered iNKT cell subsets, enhanced activation, and prominent Th1/Th17 proinflammatory profile.

Objective: Few studies have investigated the importance of different components of the inflammasome system and of innate mitochondrial sensing (IMS) pathways in HIV infection and its treatment. We analysed the expression of several components of the inflammasome and of the IMS in HIV-positive patients taking successful combination antiretroviral therapy (cART)., Methods: We enrolled 20 HIV-positive patients under cART, who achieved viral suppression since at least 10 months and 20 age and sex-matched healthy donors. By RT-PCR, using peripheral blood mononuclear cells (PBMCs), we quantified the mRNA expression of 16 genes involved in inflammasome activation and regulation (AIM2, NAIP, PYCARD, CASP1, CASP5, NLRP6, NLRP1, NLRP3, TXNIP, BCL2, NLRC4, PANX1, P2RX7, IL-18, IL-1β, SUGT1) and eight genes involved in IMS (MFN2, MFN1, cGAS, RIG-I, MAVS, NLRX1, RAB32, STING)., Results: Compared with controls, HIV-positive patients showed significantly lower mRNA levels of the mitochondrial protein NLRX1, which plays a key role in regulating apoptotic cell death; main PBMC subpopulations behave in a similar manner. No differences were observed in the expression of inflammasome components, which however showed complex correlations., Conclusion: The decreased level of NLRX1 in HIV infection could suggest that the virus is able to downregulate mechanisms linked to triggering of cell death in several immune cell types. The fact that HIV-positive patients did not show altered expression of inflammasome components, nor of most genes involved in IMS, suggests that the infection and/or the chronic immune activation does not influence the transcriptional machinery of innate mechanisms able to trigger inflammation at different levels.

Background: The activity of virus-specific T lymphocytes, among which those capable of a polyfunctional response against the viral protein gag, is crucial to control HIV infection., Objective: The objective of this study is to investigate the polyfunctionality of gag-specific T cells in different phases of HIV infection, analyzing markers related to T-helper cell 1 (Th1) and degranulation/cytotoxicity, and the production of Th1 cytokines in peripheral blood lymphocytes from patients experiencing an acute primary infection, long-term nonprogressors, patients naive for antiretroviral drugs, and patients taking HAART., Materials and Methods: Cells were stimulated with a pool of gag-derived peptides or with a superantigen (staphylococcal enterotoxin B). Using eight-color polychromatic flow cytometry, we analyzed the expression of interleukin-2, interferon-gamma, CD154, and CD107a by CD4 and CD8 T cells., Results: The main finding was that in all HIV-positive patients, about half gag-specific CD4 T cells were CD107a, that is, able to degranulate. CD4CD154 cells unable to produce Th1 cytokines were the second most represented population. Truly polyfunctional CD4 T cells were very rare and present only in a few long-term nonprogressors. Superantigen stimulation showed that CD4 T lymphocytes from all patients displayed a typical Th response, including interleukin-2 and interferon-gamma production, lacking CD107a expression., Conclusion: In all the aforementioned phases of HIV infection, the large majority of gag-specific CD4 T lymphocytes cannot be identified by the sole expression of interleukin-2 and interferon-gamma, which is early impaired. Degranulation and helper functions other than Th1 cytokine production are the predominant features of HIV-specific CD4 lymphocytes.

Objective: Many patients infected with HIV still present with an AIDS diagnosis. The aim of this study was to evaluate the virological, immunological and clinical outcomes of HAART in these patients., Design: The present study was a multi-cohort study. All patients with an AIDS diagnosis between 30 days before and 14 days after HIV diagnosis, recruited between 1997 and 2004 from eight hospital cohorts, were evaluated., Results: A total of 760 patients were included [268 (35.3%) had pneumocystis and 168 (22.1%) tuberculosis]. Six hundred and twenty-four patients (82.1%) started HAART a median of 31 days after HIV diagnosis. One hundred and fifty-three patients started a nonnucleoside transcriptase inhibitor-based regimen (20.1%), 409 a protease inhibitor-based regimen (53.8%) and 62 other regimens (8.2%). In adjusted analyses, HAART was started sooner in more recent years, in patients with lower CD4 cell count and in those with Kaposi's sarcoma, whereas it was started later in those with tuberculosis. Five hundred and five patients (89%) attained a viral load of less than 500 copies/ml. The factors associated with a better virological response were later calendar year, lower initial viral load and cytomegalovirus disease. Virological rebound was more common in those receiving nucleoside reverse transcriptase inhibitor-based regimens, in those with tuberculosis and in earlier calendar years. One hundred and twenty-five (16%) patients died; 61 had received HAART (48.6%). Mortality was more likely in those who were older, those with a higher viral load at diagnosis, those with nonsexual HIV risks and those with lower CD4 cell count and haemoglobin levels over follow-up., Conclusion: Virological suppression was achieved in most AIDS patients, though mortality remains high in these individuals.

Objective: To investigate immunological changes during CD4-guided therapy interruption in HIV(+) patients who suspended HAART., Patients: Seventeen patients aged > 18 years, who had received HAART for at least 12 months, and had a pre-interruption CD4+ cell count > 500 cells/microl, interrupted treatment. Median nadir CD4(+) cell count was 288 cells/microl. HIV plasma viral load at discontinuation was < 50 or > 50 copies/ml. Criteria for restarting treatment were: a CD4(+) T-lymphocyte count < 350 cells/microl on two separate occasions, a clinical manifestation of AIDS, and the patient's desire to resume HAART. Eleven patients were still off therapy after 12 months (group A); according to the first criterion, six patients restarted therapy within 12 months (group B)., Methods: Haematological, viro-immunological, cytofluorimetic and molecular assays were performed at baseline and every 2 months following standard methods. Statistical analysis was performed under Stata 7.0., Results: In the first 2 months of treatment interruption, a significant increase in viral load and CD8(+) lymphocyte activation occurred. Then such parameters decreased and remained stable. In all patients, a decrease in CD4(+) lymphocytes took place as well, that affected in a similar manner naive, central memory, effector memory and terminally differentiated cells. Group B always presented lower amounts of CD4(+) effector memory lymphocytes. The expression of CD127 was always higher in group A., Conclusions: The loss of CD4(+) lymphocytes upon viral rebound is equal among naive and memory subsets. Patients with higher expression of CD127, who are likely to exert a better capacity to utilize endogenous interleukin-7 by T cells, could remain off therapy for longer periods.

Background: HIV infection per se and HAART can alter mitochondrial functionality, leading to a decrease in mitochondrial DNA content., Objective: To evaluate whether treatment interruption monitored by CD4 cell count can restore mitochondrial DNA content in peripheral blood lymphocytes., Methods: Mitochondrial DNA content was measured in platelet-free CD4 and CD8 T cells by real-time polymerase chain reaction; flow cytometry was used to identify and quantify activated CD4 and CD8 T lymphocytes., Results: The 30 patients had been treated for a mean of 107 months (range, 27-197). Median CD4 cell count at discontinuation was 702 cells/microl (range, 547-798). Median observational time from HAART discontinuation was 11.3 months (range, 4-26). Discontinuation of treatment provoked significant increases in mitochondrial DNA in CD8 T cells, which started only 6 months after therapy discontinuation [5.12 copies/cell per month from 0 to 6 months (P = 0.37) and 26.96 copies/cell per month from 6 to 12 months (P < 0.0001)]., Conclusions: This study is the first showing that mitochondrial DNA content can increase in peripheral blood lymphocytes during treatment interruption, but only after at least 6 months of interruption. Consequently, interruptions of shorter periods, whether by clinician or patient decision, are unlikely to allow restoration of mitochondrial DNA and so decrease HAART-related toxicity.

Background: Little is known about CD4 cell count changes in patients with high CD4 cell counts who interrupt antiretroviral therapy, especially in those with a nadir of 250-350 x 10 cells/l., Methods: Data derived from 139 patients from seven prospective cohorts who had > 12 months highly active antiretroviral therapy (HAART), CD4 cell count nadir of > 250 x 10 cells/l and at pre-interruption of > 500 x 10 cells/l. Endpoint was time to CD4 cell count < 350 x 10 cells/l or reinitiation of treatment., Results: At interruption, the median CD4 cell count was 800 x 10 cells/l, median viral load was 1.7 log10 copies/ml. At the time of analysis, 63 (45.3%) had resumed therapy or experienced < 350 x 10 cells/l CD4 cells over a median interruption of 75 weeks. Of these, 33 (52.4%) experienced a decline to < 350 x 10 cells/l and 30 (47.6%) restarted therapy before their CD4 cell count had fallen below this level. In 43 patients with CD4 cell nadir of 251-350 x 10 cells/l, median time to therapy resumption or CD4 cell count < 350 x 10 cells/l was 61 weeks. Higher CD4 cell count nadir, longer duration of viral load suppression on therapy, and higher viral load level at interruption were independently associated with longer time to restart therapy. The risk of clinical events was 5 per 1000 person-years of follow-up., Conclusions: Patients who started therapy with CD4 cell count of 250-350 x 10 cells/l and who later interrupted therapy appear able to remain off therapy with a CD4 cell count > 350 x 10 cells/l for a substantial period of time.

Objective: To identify crucial immunological characteristics of a group of patients, defined 'CD4-exploders', who were able to fully reconstitute their immune system after receiving highly active antiretroviral therapy (HAART)., Patients: Among a population of 540 HIV-positive patients treated with HAART, six individuals were identified who experienced a nadir of less than 85 x 106 CD4+ cells/l, had major opportunistic infections (four out of six), started HAART in 1996 or 1997, and showed a rapid and relevant CD4+ lymphocyte increase (mainly due to virgin cells), in some cases regardless of virological control., Methods: Enzyme-linked immunosorbent assay for the determination of interleukin (IL)-7 plasma levels; flow cytometry to analyse surface antigens and CD127 (IL-7 receptor alpha-chain) expression; quantitative-competitive (QC) PCR for detecting cells containing T-cell receptor rearrangement excision circles (TREC); chest-computed tomography (CT) to analyse thymus volume and content., Results: In 'CD4-exploders', high levels of TREC+ lymphocytes were found among CD4+ T cells, which also contained a high percentage of naive cells. However, CT revealed a dramatic depletion of intrathymic lymphoid tissue. Plasma levels of IL-7 were significantly high. Most CD4+ and CD8+ T lymphocytes expressed CD127, whose level was similar to that of healthy donors. CD127 expression on CD8+ lymphocytes was markedly higher than in HIV-positive individuals treated with the same therapy or in patients with AIDS., Conclusion: In 'CD4-exploders', HAART-induced reconstitution of the T-cell compartment is independent from thymus volume, and is favoured by the upregulation of the IL-7/IL-7 receptor system.

Objective: To study alterations of mitochondrial membrane potential (delta psi) and the propensity to undergo apoptosis in peripheral blood lymphocytes (PBL) from subjects with acute HIV syndrome; and to evaluate possible modulations of these phenomena by antioxidants that can be used in therapy, such as N-acetyl-cysteine (NAC), nicotinamide (NAM), or L-acetyl-carnitine (LAC)., Methods: Mitochondrial function and the tendency of PBL to undergo spontaneous apoptosis were studied on freshly collected PBL from patients with symptomatic, acute HIV-1 primary infection, which were cultured for different durations in the presence of absence of NAC. NAM or LAC. By a cytofluorimetric method allowing analysis of delta psi in intact cells, we studied the function of these organelles under the different conditions. PBL apoptosis was evaluated by the classic cytofluorimetric method of propidium iodide staining, capable of revealing the typical DNA hypodiploid peak., Results: Significant delta psi alterations and tendency to undergo apoptosis were present in PBL from the subjects we studied. Indeed, when cultured even for a few hours in the absence of any stimulus, a consistent number of cells died. However, the presence of even different levels of NAC, NAM or LAC was able to rescue most of them from apoptosis. Both a fall in delta psi and apoptosis were evident in PBL collected in the earliest phases of the syndrome (before seroconversion), and changed significantly after a few days. A significant correlation was found between spontaneous apoptosis and tumour necrosis factor (TNF)-alpha or p24 plasma levels, as well as between apoptosis and the percentages of circulating CD4+ or CD8+ T cells., Conclusions: PBL from patients with acute HIV syndrome are characterized by both significant mitochondrial alterations and a dramatic tendency to undergo apoptosis. The use of NAC, NAM or LAC seems to rescue cells through a protective effect on mitochondria, a well-known target for the action of TNF-alpha and for reactive oxygen species, the production of which is strongly induced by this cytokine. Thus, our data could provide the rationale for the use of such agents in addition to antiviral drugs in primary infection.

Objective: To study the V beta T-cell repertoire in peripheral blood lymphocytes (PBL) during acute HIV syndrome by using several anti-V beta monoclonal antibodies (MAb) and to analyse its functionality by stimulating PBL with superantigens (SAg) such as Staphylococcus aureus enterotoxins., Methods: Cytofluorimetric analysis of V beta T-cell-receptor expression was performed on PBL from eight patients with symptomatic, acute HIV-1 primary infection, showing a dramatic decrease of CD4+ PBL accompanied by a marked increase in activated/memory CD8+ T cells, and on 12 age- and sex-matched healthy controls. PBL were then isolated, stimulated with different SAg, anti-CD3 MAb or phytohaemagglutinin and cultured for 3 days. PBL capability to progress through cell cycle was studied by the classic cytofluorimetric method of bromodeoxyuridine incorporation and DNA staining with propidium iodide., Results: Despite the presence of a few expansions of some V beta families among CD8+ T lymphocytes, no gross alterations in T-cell repertoire were present in patients with acute HIV syndrome. Its functionality was maintained overall, as PBL responsiveness to SAg was well preserved. Interestingly, all CD8+ T cells, although bearing different V beta T-cell receptors, expressed marked signs of activation, i.e., CD45R0, CD38 and major histocompatibility complex class II molecules, and also high amounts of CD11a and CD18., Conclusions: Our data suggest, at least in the early phases and in the acute form of the infection, that HIV is not likely to act as a SAg. However, further studies are needed to analyse other sites, such as lymph nodes, where HIV could exert other, significant effects, and to study the expression of other V beta families than those investigated here.