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M1 polarization of human monocyte-derived macrophages restricts pre and postintegration steps of HIV-1 replication

Cassetta, Lucaa,d; Kajaste-Rudnitski, Annab,f; Coradin, Tizianab,e; Saba, Elisaa; Della Chiara, Giuliaa,g; Barbagallo, Marialuisaa; Graziano, Francescaa,c; Alfano, Massimoa; Cassol, Edanaa,h; Vicenzi, Elisab; Poli, Guidoa,c

doi: 10.1097/QAD.0b013e328361d059
Basic Science

Objective: Functional polarization of human monocyte-derived macrophages (MDMs) into M1 cells leads to inhibition of R5 HIV-1 replication and viral DNA synthesis in comparison to control, unpolarized cells together with CD4 downregulation from the cell surface and upregulation of CCR5-binding chemokine secretion. We here investigated whether a postentry restriction of virus replication is also induced by M1 polarization of MDM.

Design: MDM were first polarized to M1 cells by 18 h stimulation with interferon-γ and tumor necrosis factor-α; the cytokines were then removed and the cells were infected with vesicular stomatitis virus G-protein pseudotyped enhanced green fluorescence protein HIV-1 (HIV-GFP) generating a single-round infection cycle.

Methods: HIV-1 expression was monitored in terms of eGFP expression by fluorescence activated cell sorter (FACS) analysis and real-time PCR analysis of total HIV-1 gag DNA, 2-long terminal repeat DNA, proviral DNA, and multiply spliced RNA transcripts. Expression of apolipopoprotein B mRNA-editing, enzyme-catalytic, polypeptide-like 3G (APOBEC3G), and APOBEC3A was tested by western blotting and FACS analysis.

Results: Inhibition of HIV-GFP expression was observed in M1-MDM along with impaired viral DNA synthesis, delayed proviral integration, and reduced proviral transcription. Although APOBEC3G levels were similar in M1 and unpolarized MDM, APOBEC 3A was selectively expressed only by M1 cells.

Conclusion: M1 polarization of in-vitro differentiated primary MDM determines a transient, but profound restriction of HIV-1 replication affecting multiple (entry and postentry) steps in the virus life cycle likely involving the upregulated expression of APOBEC3A.

aAIDS Immunopathogenesis Unit

bViral Pathogens and Biosafety Unit, Division of Immunology, Transplantation and Infectious Diseases, San Raffaele Scientific Institute

cUniversità Vita-Salute Salute San Raffaele, School of Medicine, Milan, Italy

dDepartment of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, New York, USA

eCentre for Integrative Biology, University of Trento, Mattarello, Trento

fSan Raffaele Telethon Institute for Gene Therapy, San Raffaele Scientific Institute

gDepartment of Experimental Oncology, European Institute of Oncology, Milan, Italy

hDepartment of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.

Correspondence to Professor Guido Poli, P2-P3 Laboratories, DIBIT-1, Via Olgettina n. 58, 20132 Milan, Italy. Tel: +39 02 2643 4909; fax: +39 02 2643 4905; e-mail: poli.guido@hsr.it

Received 13 January, 2012

Revised 6 March, 2013

Accepted 29 March, 2013

© 2013 Lippincott Williams & Wilkins, Inc.