Silvia Morini

Peptide-derivatized SB105-A10 dendrimer inhibits the infectivity of R5 and X4 HIV-1 strains in primary PBMCs and cervicovaginal histocultures.

Peptide dendrimers are a class of molecules that exhibit a large array of biological effects including antiviral activity. In this report, we analyzed the antiviral activity of the peptide-derivatized SB105-A10 dendrimer, which is a tetra-branched dendrimer synthetized on a lysine core, in activated peripheral blood mononuclear cells (PBMCs) that were challenged with reference and wild-type human immunodeficiency virus type 1 (HIV-1) strains. SB105-A10 inhibited infections by HIV-1 X4 and R5 strains, interfering with the early phases of the viral replication cycle. SB105-A10 targets heparan sulfate proteoglycans (HSPGs) and, importantly, the surface plasmon resonance (SPR) assay revealed that SB105-A10 strongly binds gp41 and gp120, most likely preventing HIV-1 attachment/entry through multiple mechanisms. Interestingly, the antiviral activity of SB105-A10 was also detectable in an organ-like structure of human cervicovaginal tissue, in which SB105-A10 inhibited the HIV-1ada R5 strain infection without altering the tissue viability. These results demonstrated the strong antiviral activity of SB105-A10 and suggest a potential microbicide use of this dendrimer to prevent the heterosexual transmission of HIV-1.

Effects of human immunodeficiency virus on the erythrocyte and megakaryocyte lineages

Anaemia and thrombocytopenia are haematological disorders that can be detected in many human immunodeficiency virus (HIV)-positive patients during the development of HIV infection. The progressive decline of erythrocytes and platelets plays an important role both in HIV disease progression and in the clinical and therapeutic management of HIV-positive patients. HIV-dependent impairment of the megakaryocyte and erythrocyte lineages is multifactorial and particularly affects survival, proliferation and differentiation of bone marrow (BM) CD34+ haematopoietic progenitor cells, the activity of BM stromal cells and the regulation of cytokine networks. In this review, we analyse the major HIV-related mechanisms that are involved in the genesis and development of the anaemia and thrombocytopenia observed in HIV positive patients.

HIV-1 and recombinant gp120 affect the survival and differentiation of human vessel wall-derived mesenchymal stem cells

BackgroundHIV infection elicits the onset of a progressive immunodeficiency and also damages several other organs and tissues such as the CNS, kidney, heart, blood vessels, adipose tissue and bone. In particular, HIV infection has been related to an increased incidence of cardiovascular diseases and derangement in the structure of blood vessels in the absence of classical risk factors. The recent characterization of multipotent mesenchymal cells in the vascular wall, involved in regulating cellular homeostasis, suggests that these cells may be considered a target of HIV pathogenesis. This paper investigated the interaction between HIV-1 and vascular wall resident human mesenchymal stem cells (MSCs).ResultsMSCs were challenged with classical R5 and X4 HIV-1 laboratory strains demonstrating that these strains are able to enter and integrate their retro-transcribed proviral DNA in the host cell genome. Subsequent experiments indicated that HIV-1 strains and recombinant gp120 elicited a reliable increase in apoptosis in sub-confluent MSCs. Since vascular wall MSCs are multipotent cells that may be differentiated towards several cell lineages, we challenged HIV-1 strains and gp120 on MSCs differentiated to adipogenesis and endotheliogenesis. Our experiments showed that the adipogenesis is increased especially by upregulated PPARγ activity whereas the endothelial differentiation induced by VEGF treatment was impaired with a downregulation of endothelial markers such as vWF, Flt-1 and KDR expression. These viral effects in MSC survival and adipogenic or endothelial differentiation were tackled by CD4 blockade suggesting an important role of CD4/gp120 interaction in this context.ConclusionsThe HIV-related derangement of MSC survival and differentiation may suggest a direct role of HIV infection and gp120 in impaired vessel homeostasis and in genesis of vessel damage observed in HIV-infected patients.

HIV-related mechanisms in atherosclerosis and cardiovascular diseases.

HIV-infected patients have a significantly higher risk of developing cardiovascular events during the progression of HIV disease. Atherosclerosis, myocardial infarction, cerebrovascular injury, pulmonary hypertension and thrombosis are consistently described in both combined antiretroviral therapy (cART)-treated and naive HIV-positive patients as major clinical complications. Recent studies indicate that the pathogenesis of cardiovascular lesions in HIV-positive patients is related to direct and indirect effects of HIV infection on vessel structures, independently of traditional risk factors. HIV infection strongly interferes with the biology of several cellular targets such as macrophage and endothelial cells. Moreover, HIV induces a profound derangement of lipid metabolism and inflammatory cytokine networks that are directly involved in atherogenesis and progressive impairment of the cardiovascular system. In this review, we discuss these major HIV-related mechanisms able to promote atherosclerosis and cardiovascular diseases in HIV-positive patients.

Analysis of the effects of HIV‐1 Tat on the survival and differentiation of vessel wall‐derived mesenchymal stem cells

HIV infection is an independent risk factor for atherosclerosis development and cardiovascular damage. As vessel wall mesenchymal stem cells (MSCs) are involved in the regulation of vessel structure homeostasis, we investigated the role of Tat, a key factor in HIV replication and pathogenesis, in MSC survival and differentiation. The survival of subconfluent MSCs was impaired when Tat was added at high concentrations (200–1,000 ng/ml), whereas lower Tat concentrations (1–100 ng/ml) did not promote apoptosis. Tat enhanced the differentiation of MSC toward adipogenesis by the transcription and activity upregulation of PPARγ. This Tat‐related modulation of adipogenesis was tackled by treatment with antagonists of Tat‐specific receptors such as SU5416 and RGD Fc. In contrast, Tat inhibited the differentiation of MSCs to endothelial cells by downregulating the expression of VEGF‐induced endothelial markers such as Flt‐1, KDR, and vWF. The treatment of MSCs with Tat‐derived peptides corresponding to the cysteine‐rich, basic, and RGD domains indicated that these Tat regions are involved in the inhibition of endothelial marker expression. The Tat‐related impairment of MSC survival and differentiation might play an important role in vessel damage and formation of the atherosclerotic lesions observed in HIV‐infected patients. J. Cell. Biochem. 113: 1132–1141, 2012.

Prevalence of R5 strains in multi-treated HIV subjects and impact of new regimens including maraviroc in a selected group of patients with CCR5-tropic HIV-1 infection

OBJECTIVES Maraviroc currently represents an important antiretroviral drug for multi-experienced and viremic HIV patients. This study focused on two main points: (1) determining the prevalence of R5 and X4 HIV strains in antiretroviral-experienced patients using two main tests currently in use to determine viral tropism, and (2) the follow-up to 3 years of a limited number of patients who started a new antiretroviral protocol including maraviroc. METHODS A group of 56 HIV patients, previously multi-treated, were first analyzed by genotyping assay and Trofile™ to establish their eligibility for maraviroc treatment. In addition, 25 subjects selected to follow a new therapeutic protocol including a CCR5 antagonist were monitored by HIV RNA viral load and CD4+ cell count. RESULTS The determination of viral tropism showed a large percentage of patients with an R5 profile (72% by genotyping assay and 74% by Trofile). The follow-up of most (21 out 25) patients who started the new antiretroviral protocol showed an undetectable viral load throughout the observation period, accompanied by a major improvement in CD4 cell count (cells/mm(3)) (baseline: median CD4 cell count 365, interquartile range (IQR) 204-511; 12 months: median value 501, IQR 349-677, p=0.042; 24 months: median value 503, IQR 386-678, p=0.026; 36 months: median value 601, IQR 517-717, p=0.001). Among the four non-responder subjects, two showed a lack of drug compliance and two switched from R5 to X4. CONCLUSION Although our patient cohort was small, the results showed a high prevalence of R5 viral strains in multi-experienced patients. As well as showing the advantages of genotyping, which can be performed in plasma samples with low viral load replication, the follow-up of HIV patients selected for an alternative drug protocol, including a CCR5 antagonist, showed a persistent undetectable viral replication and a good recovery of CD4 cell count in most treated HIV patients.

IFI16 Expression Is Related to Selected Transcription Factors during B-Cell Differentiation

The interferon-inducible DNA sensor IFI16 is involved in the modulation of cellular survival, proliferation, and differentiation. In the hematopoietic system, IFI16 is consistently expressed in the CD34+ stem cells and in peripheral blood lymphocytes; however, little is known regarding its regulation during maturation of B- and T-cells. We explored the role of IFI16 in normal B-cell subsets by analysing its expression and relationship with the major transcription factors involved in germinal center (GC) development and plasma-cell (PC) maturation. IFI16 mRNA was differentially expressed in B-cell subsets with significant decrease in IFI16 mRNA in GC and PCs with respect to naïve and memory subsets. IFI16 mRNA expression is inversely correlated with a few master regulators of B-cell differentiation such as BCL6, XBP1, POU2AF1, and BLIMP1. In contrast, IFI16 expression positively correlated with STAT3, REL, SPIB, RELA, RELB, IRF4, STAT5B, and STAT5A. ARACNE algorithm indicated a direct regulation of IFI16 by BCL6, STAT5B, and RELB, whereas the relationship between IFI16 and the other factors is modulated by intermediate factors. In addition, analysis of the CD40 signaling pathway showed that IFI16 gene expression directly correlated with NF-κB activation, indicating that IFI16 could be considered an upstream modulator of NF-κB in human B-cells.

HIV-1 coreceptor usage in paired plasma RNA and proviral DNA from patients with acute and chronic infection never treated with antiretroviral therapy

Although an independent evolution of viral quasispecies in different body sites might determine a differential compartmentalization of viral variants, the aim of this paper was to establish whether sequences from peripheral blood mononuclear cells (PBMCs) and plasma provide different or complementary information on HIV tropism in patients with acute or chronic infection. Tropism was predicted using genotypic testing combined with geno2pheno (coreceptor) analysis at a 10% false positive rate in paired RNA and DNA samples from 75 antiretroviral‐naïve patients (divided on the basis of avidity index into patients with a recent or long‐lasting infection). A high prevalence of R5 HIV strains (97%) was observed in both compartments (plasma and PBMCs) in patients infected recently. By contrast, patients with a long‐lasting infection showed a quite different situation in the two compartments, revealing more (46%) X4/DM in PBMCs than patients infected recently (3%) (P = 0.008). As‐ a knowledge of viral strains in different biological compartments might be crucial to establish a therapeutic protocol, it could be extremely useful to detect not only viral strains in plasma, but also viruses hidden or archived in different cell compartments to better understand disease evolution and treatment efficacy in patients infected with HIV. J. Med. Virol. 87:315–322, 2015.

Effects of Antiretroviral Molecules on Survival and Gene Expression of An Osteoblast-like Cell Line

BACKGROUND The advent of combined antiretroviral therapy effectively undermined the evolution of HIV disease. Nevertheless, clinical observations indicated a clear association between therapy and the impairment of bone mineral density. OBJECTIVE We selected some antiretroviral compounds used in clinical practice, to study their impact on bone health and their possible implication in the onset of bone disease. METHOD Scalar concentrations of several antiretroviral drugs (used in single and in combination) were tested on an osteoblast-like cell line, HOBIT cells, to analyse cell survival and gene expression of selected bone markers. RESULTS None of the tested concentrations of Tenofovir, Emtricitabine, Nevirapine, Maraviroc or Raltegravir induced any significant apoptosis activation at our experimental conditions. Only some protease inhibitors and Efavirenz, at high concentration, determined a significant activation of programmed cell death. In parallel experiments, protease inhibitors used in combination with Tenofovir and Emtricitabine, increased apoptosis. Furthermore, we performed a study of mRNA expression of specific genes involved in osteoblast biology and in bone synthesis and observed that some protease inhibitors induced a selective decrease of some osteogenic markers. CONCLUSION All the protease inhibitors included in this study trigger apoptosis at the highest concentration analysed, suggesting great caution in HIV-patients co-infected with HBV or HCV, where elevated plasma concentrations of drugs could be reached as a consequence of liver failure. Lastly, an increased apoptosis rate and an impairment of osteogenic markers were recorded only in the presence of Nelfinavir, suggesting a role of protease inhibitors in the alteration of osteoblast biology.

Meningitis Caused by Toscana Virus Is Associated with Strong Antiviral Response in the CNS and Altered Frequency of Blood Antigen-Presenting Cells

Toscana virus (TOSV) is a Phlebotomus-transmitted RNA virus and a frequent cause of human meningitis and meningoencephalitis in Southern Europe during the summer season. While evidence for TOSV-related central nervous system (CNS) cases is increasing, little is known about the host defenses against TOSV. We evaluated innate immune response to TOSV by analyzing frequency and activation of blood antigen-presenting cells (APCs) and cytokine levels in plasma and cerebrospinal fluid (CSF) from patients with TOSV neuroinvasive infection and controls. An altered frequency of different blood APC subsets was observed in TOSV-infected patients, with signs of monocytic deactivation. Nevertheless, a proper or even increased responsiveness of toll-like receptor 3 and 7/8 was observed in blood APCs of these patients as compared to healthy controls. Systemic levels of cytokines remained low in TOSV-infected patients, while levels of anti-inflammatory and antiviral mediators were significantly higher in CSF from TOSV-infected patients as compared to patients with other infectious and noninfectious neurological diseases. Thus, the early host response to TOSV appears effective for viral clearance, by proper response to TLR3 and TLR7/8 agonists in peripheral blood and by a strong and selective antiviral and anti-inflammatory response in the CNS.