Rapid CommunicationMycobacterium tuberculosis alters the differentiation of monocytes into macrophages in vitro
Introduction
Macrophages play an important role in granulomatous responses against Mycobacterium tuberculosis infection [1], [2]. Histological evidence in human tissue has indicated that granulomas mainly consist of infected macrophages surrounded by recruited mononuclear cells, these being mainly monocytes, lymphocytes and fibroblasts [2], [3]. Although it is difficult to study the natural history of granuloma formation in humans, studies in the zebrafish model of Mycobacterium marinum infection [4] have provided evidence for a similar structure to that of human granulomas developed as a response to M. tuberculosis infection [5], [6]. Interestingly, it has been demonstrated that alterations in granuloma organization may promote mycobacterial dissemination rather than contention [6].
The role of newly-recruited monocytes in mycobacterial control inside a granuloma is still not fully understood. Monocytes could come into contact with mycobacterial soluble products and become infected with M. tuberculosis before completing their differentiation. Innate and adaptive responses against bacilli infection may thus become compromised. It is currently unknown whether M. tuberculosis-infected monocytes are able to differentiate into macrophages.
Freshly isolated human monocytes were infected with M. tuberculosis H37Rv at low multiplicity of infection and allowed to differentiate into macrophages in the present study. It was observed that M. tuberculosis infection altered several events related to monocyte to macrophage differentiation and function, including expression of key cell markers, cytokine release, anti-mycobacterial activity and T-cell proliferation.
Section snippets
Mononuclear phagocyte differentiation
Peripheral blood mononuclear cells (PBMC) from healthy individuals containing 2.5 × 105 CD14+ cells were plated into 48-well plates (Corning Incorporated Life Science, Lowell, MA) using 1 ml of RPMI-1640 (Gibco-BRL, Grand Island, NY) plus 0.5% pooled human serum (PHS), for 4 h at 37 °C. Wells were extensively washed with pre-warmed phosphate-buffered saline (PBS, Gibco-BRL) plus 0.5% Fetal bovine serum (FBS, Gibco-BRL) to remove non-adherent cells (this was defined as differentiation time zero).
M. tuberculosis infection prevented differentiation marker up-regulation in mononuclear phagocytes
Our previous observations have shown that MDM (120 h differentiation) are more granular, adherent and larger, having increased cytoplasmic projections and increased CD36, CD86, CD68 and HLA-II expression compared to non-differentiated mononuclear phagocytes and freshly isolated cells (Castaño et al., submitted for publication). The percentage of M. tuberculosis-FDA-associated cells was thus determined before differentiation to evaluate the effect of M. tuberculosis on monocyte differentiation
Discussion
Infected macrophages attract immune cells through chemokine production [14]. Newly-recruited monocytes can be derived from the circulation or tissue reservoirs [6], [15] and may be either infected or exposed to mycobacterial or cellular products before becoming fully differentiated into macrophages; they would then not be able to properly execute some effector mechanisms. Our results showed that mononuclear phagocytes infected with live and virulent M. tuberculosis H37Rv did not completely
Conflicts of interest
The authors have neither financial nor personal conflicts of interest in the present study.
Acknowledgments
This study was financed by COLCIENCIAS (Bogotá, Colombia) Grants RC431-2004 (“Centro Colombiano de Investigación en Tuberculosis”) and 111540520270 and Programa de Sostenibilidad 2009–2010 grant from the Universidad de Antioquia. Diana Castaño was awarded a PhD scholarship by COLCIENCIAS. The authors would like to thank the healthy individuals who participated in this study.
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