Mariantonietta Di Stefano

Induction of human immunodeficiency virus type 1 replication in human glial cells after proinflammatory cytokines stimulation: Effect of IFNγ, IL1β, and TNFα on differentiation and chemokine production in glial cells

Although evidence for human immunodeficiency virus 1 (HIV‐1) presence in the central nervous system (CNS) of infected patients is well established, the intensity of viral replication within the brain is not usually known. In vitro, human embryonic microglial cells internalized HIV‐1 through a CD4‐dependent pathway but were not permissive to viral replication. We observed that HIV replication was induced when CNS cell cultures were stimulated for 14 days by a combination of proinflammatory cytokines including IFNγ, IL1β, and TNFα. After long‐term cytokine stimulation, morphologically differentiated glial cells appeared, in which HIV‐1 tat antigen was detected after infection. Thus, variations in the stage of maturation/activation of CNS cells under inflammatory conditions probably play a major role in facilitating massive production of HIV‐1. We then studied the effect of prolonged cytokine stimulation on the secretion of inflammatory mediators by glial cells. An early increased secretion of prostaglandin F2α and chemokines (RANTES>>MIP‐1α>>MIP‐1β) was observed, due to both microglia and astrocytes. In contrast to persistent PGF2α production, an extinction of RANTES and MIP‐1β but not of MIP‐1α secretion occurred during the 14 days of stimulation and was inversely correlated with the ability of glial cells to replicate HIV‐1. The study of the secretory factors produced in response to a persistent inflammation could provide a better understanding of the modulation of HIV replication in glial cells. GLIA 23:304–315, 1998.

Analysis of ENV V3 sequences from HIV-1-infected brain indicates restrained virus expression throughout the disease.

The isolation of human immunodeficiency virus type 1 (HIV‐1) from the cerebrospinal fluid (CSF) of asymptomatic virus carriers suggests that the viral infection spreading to the brain occurs early during infection. The aim of the present study was to investigate whether HIV‐1 infection of the brain parenchyma also occurs during the early phase of infection. We also wished to compare the degree of replication of the virus in the brain at different clinical stages associated with HIV‐1 infection. With the use of polymerase chain reaction (PCR), the viral genomes present in seven of eight brain specimens obtained from two asymptomatic HIV‐1 carriers and six AIDS patients were amplified. Thereafter, the number of viral copies present in each brain specimen was quantified, the third variable region (V3) of the gp120 glycoprotein was sequenced and these results compared with the histopathological findings in the tissue.

Measurement of viral sequences in cerebrospinal fluid of Aids patients with cerebral white-matter lesions using polymerase chain reaction

Objectives:To optimize the use of polymerase chain reaction (PCR) on cerebrospinal fluid (CSF) for the evaluation of central nervous system (CNS) white-matter lesions that along with clinical findings and magnetic resonance imaging (MRI) can allow a definite diagnosis to be made; also to evaluate treatment with zidovudine plus foscarnet. Design and methods:Fifteen AIDS patients with uncertain CNS white-matter lesions were identified. HIV-1 RNA, cytomegalovirus (CMV) and JC virus (JCV) DNA were measured in a total of 29 CSF samples. The results were correlated with clinical and MRI findings and treatment with zidovudine plus foscarnet was evaluated. Results:Four and five out of 15 patients with CMV DNA ≥ 1 : 625 and JCV DNA ≥ 103 copies/µl detected in the CSF were diagnosed with CMV and progressive multifocal leukoencephalopathy (PML), respectively. Six patients who were CMV/JCV-negative with the highest levels of HIV RNA (median, 6.87 log10 copies/ml) in CSF were considered as having HIV-1 encephalitis. Neurological symptoms were non-supportive for diagnosis as was MRI in 11 out of 15 patients. Nine patients completed a 21-day course of zidovudine plus foscarnet. HIV RNA decreased irrespective of neurological diagnosis. All three HIV-1 encephalitis patients and two out of three patients with CMV leukoencephalopathy improved. In these two latter patients, relief of clinical symptoms coincided with decreased CMV DNA. JCV DNA remained unchanged and all three PML patients deteriorated. Conclusions:Measurement of CSF viral sequences supports the diagnosis of CNS white-matter lesions in AIDS patients. While effective therapy for PML remains elusive, treatment including zidovudine plus foscarnet may be a promising option for HIV-1 and CMV-related manifestations.

Preliminary Data From the Study of Coagulative Profile of HIV Infected Individuals Suggest a Role For Point Mutations in the Gene in Protein S Deficiency in Individuals Undergoing Highly Antiretroviral Therapy

Background: HIV infection is a known prothrombotic condition but factors involved are still controversial. A role for antiretrovirals, especially protease inhibitors, was advocated. Objectives: The study aimed to analyze the levels of anticoagulant proteins in virally suppressed HIV-infected subjects treated with different anti-retroviral regimens. Materials and Methods: Forty-four patients were included in the study. C and PS, D-Dimers and Fibrinogen levels were determined as well as APC-resistance. PROS1 gene was sequenced in a group of patient. Results: Twelve of the 44 subjects (27%) showed reduced levels of PS, while lower levels of PC were found only in 2 patients (4,5%). No difference in the mean values of PC and PS was found stratifying the study population by antiretroviral regimen administrated (p>0.05). Three patients had higher levels of D-Dimer concentrations and in two of these patients, an association between higher D-Dimer values and lower levels of PS was observed; but however no correlation was found by statistical analysis. PROS1 gene analysis was performed in 26 of the 44 HIV-1 patients and the subjects with low levels of PS had mutation in the fifteen exon of PROS 1 gene. While among individuals with normal levels, this mutation was observed only in 8/18 (44%) of the cases (p=0,0072). Conclusion: The majority of patients with low PS levels also had mutations in the fifteen exon of PROS 1 gene. Genetic determinants, deserving further investigations, rather than antiretrovirals might cause PS deficiency in HIV-1 positive patients.