Anti-West Nile virus activity of in vitro expanded human primary natural killer cells

Mingjie Zhang,Caren Chancey,Andriyan Grinev,Howard Mostowski,Sylvester Daniel,Maria Rios,Yong Huang,Andrew Dayton,Ying F Lei,Qingsheng Huang

doi:10.1186/1471-2172-11-3

Abstract

BackgroundNatural Killer (NK) cells are a crucial component of the host innate immune system with anti-viral and anti-cancer properties. However, the role of NK cells in West Nile virus (WNV) infection is controversial, with reported effects ranging from active suppression of virus to no effect at all. It was previously shown that K562-mb15-41BBL (K562D2) cells, which express IL-15 and 4-1BBL on the K562 cell surface, were able to expand and activate human primary NK cells of normal peripheral blood mononuclear cells (PBMC). The expanded NK cells were tested for their ability to inhibit WNV infection in vitro.ResultsCo-culture of PBMC with irradiated K562D2 cells expanded the NK cell number by 2-3 logs in 2-3 weeks, with more than 90% purity; upregulated NK cell surface activation receptors; downregulated inhibitory receptors; and boosted interferon gamma (IFN-γ) production by ~33 fold. The expanded NK (D2NK) cell has strong natural killing activity against both K562 and Vero cells, and killed the WNV infected Vero cells through antibody-dependent cellular cytotoxicity (ADCC). The D2NK cell culture supernatants inhibited both WNV replication and WNV induced cytopathic effect (CPE) in Vero cells when added before or after infection. Anti-IFN-γ neutralizing antibody blocked the NK supernatant-mediated anti-WNV effect, demonstrating a noncytolytic activity mediated through IFN-γ.ConclusionsCo-culture of PBMC with K562D2 stimulatory cells is an efficient technique to prepare large quantities of pure and active NK cells, and these expanded NK cells inhibited WNV infection of Vero cells through both cytolytic and noncytolytic activities, which may imply a potential role of NK cells in combating WNV infection.

Highlights

Natural Killer (NK) cells are a crucial component of the host innate immune system with anti-viral and anti-cancer properties
Co-culture with radiation killed stimulating cells in vitro significantly expanded NK cells in human peripheral blood mononuclear cells (PBMC) In co-cultures with 1 × 107 lethally radiated K562mb15-41BBL (K562D2) stimulating cells in vitro, 1 × PBMC were expanded to 1 × in 2 weeks
Expanded NK cells are cytolytic through both natural killing and antibodydependent cellular cytotoxicity (ADCC) D2NK cells expanded by co-culture with K562D2 stimulatory cells have been reported to have enhanced cytolytic function against B-lineage cell lines [12]

Summary

Introduction

Natural Killer (NK) cells are a crucial component of the host innate immune system with anti-viral and anti-cancer properties. Infection of mice with WNV transiently activates and suppresses NK cell activity [3]. WNV infection may attenuate NK cell cytotoxicity by increasing cell surface expression of MHC class I molecules [4,5,6] to overcome susceptibility to NK cell mediated lysis. Splenocytes from WNV immunized mice have poor NK cell lytic activity [7]. Mice genetically deficient in NK cells or with NK cells depleted by antiNK cell antibody demonstrate no increased morbidity or mortality for WNV infection when compared to wild type controls [2,8]. At least for WNV infection in mice, NK cells appear to be dispensable for controlling infection and disease, despite their well documented role in combating viral infection in general

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