Differential recruitment and activation of natural killer cell sub‐populations by Mycobacterium bovis‐infected dendritic cells

Abstract

Keywords: Cell trafficking r Dendritic cells r Infectious diseases r NK cellsAdditional supporting information may be found in the online version of this article at the publisher's web-site IntroductionTuberculosis (TB) is a chronic respiratory illness caused by mycobacterial pathogens afflicting a wide range of species. Infection in humans is most commonly as a result of Mycobacterium tuberculosis, whereas cattle are the primary host for Mycobacterium bovis. Whilst the mechanisms that govern protective immunity to mycobacteria have not been completely deciphered, IFN-γ plays an important role enabling containment of infection and Th1 polarisation [1,2].Correspondence: Dr. Jayne Hope e-mail: jayne.hope@roslin.ed.ac.uk NK cells have key roles in bridging innate and adaptive immune responses. In particular, increased numbers of activated NK cells are evident within the lungs following mycobacterial infection in mice [3,4] and have been shown in humans to lyse M. tuberculosisinfected monocytes [5]. Crucially, NK-cell-derived IFN-γ contributes to early innate resistance to bacterial infection [3] and the subsequent development of Th1-biased immune responses within draining LNs [6][7][8]. Fully functional effector responses of NK cells are however reliant upon interplay with accessory cells, particularly DCs. During this process, DCs secrete chemokines to orchestrate the recruitment of effector cells with important roles for CXCR3 and CCR5 expression by NK cells [9,10] Results NKp46+ CD2 − NK cells transcribe high levels of inflammatory and lymphoid homing chemokine receptors NK cells were identified as a small population (1-5%) of CD3 − NKp46+ lymphocytes ( Fig. 1A and B). Analysis of CD2 expression confirmed the existence of two distinct subsets of NKp46 + CD2 + and NKp46 + CD2 − NK cells (Fig. 1C). These subsets were highly purified for further analysis (Supporting Information Fig. 1).To gain an insight into the migratory characteristics of bovine NK cells, we analysed the chemokine receptor repertoire of purified NK-cell subsets by qRTPCR [24,25]. NK cells expressed transcripts for all measured chemokine receptors (Fig. 2). Despite evidence of variability in the chemokine receptor expression levels, no one receptor was transcribed at significantly different levels than the other receptors. In contrast, chemokine receptor expression levels varied significantly between CD2+ and CD2 − NK-cell subsets ( Fig. 2A − ) was reverse transcribed and subjected to qPCR analysis. Samples were tested in triplicate, quantified against a plasmid DNA standard curve and normalised to the housekeeping gene RPLPO. Each data point represents the average transcriptional value obtained from one animal, expressed as log2 of the copy number (CN) of (A) CCR1-10; (B) CXCR1-XCR1. The average transcriptional value from eight animals is represented by a solid black line. Significance between receptors was assessed using a general linear model and between NK-cell subsets using paired t-tests; *p < 0.05, **p < 0.01. (C) DCs were infected with ...

Frequency and phenotype of natural killer cells and natural killer cell subsets in bovine lymphoid compartments and blood

“…Therefore we propose that CD2 − NK cells may represent a recirculating population of lymphocytes in cattle and so may play a role in immune surveillance. Furthermore, previous studies in our laboratory have shown that the CD2 − subset of bovine NK cells are preferentially activated following in vitro co‐culture with Bacillus Calmette–Guérin‐infected dendritic cells42 and Mycobacterium bovis ‐infected dendritic cells 30. Therefore, CD2 − NK cells may migrate from sites of vaccination or infection to draining lymph nodes where they could interact with populations of antigen‐presenting cells and egress via the efferent lymphatic vessel.…”

“…This elevated expression of CD62L by CD2 − NK cells in the AL and LNs correlates with the prevalence of CD2 − NK cells within these two compartments. Bovine NK cells transcribe the LN homing chemokine receptor CCR7,30 but surface CCR7 expression could not be consistently detected by PB‐ or LN‐derived NK cells (data not shown). Using an anti‐human CCR7 antibody cross‐reactive to bovine cells, the expression of CCR7 by AL NK cells was very variable across the seven calves examined (data not shown) and expression was not detected by AL NK cells in previously published work 23.…”

“…1d) support these findings whereby the percentage of CD2 − NK cells present within the PB was low, but was significantly increased in AL and LNs, suggesting preferential recruitment of the CD2 − subset of NK cells from the tissues to the draining LNs in steady‐state conditions. In studies of human afferent lymph, CD56 bright NK cells, which are equivalent to bovine CD2 − NK cells,30 were the main subset of NK cells present 18…”

Microfluidic‐based, live‐cell analysis allows assessment of NK‐cell migration in response to crosstalk with dendritic cells

“…They are capable of killing transformed or infected cells [1,2,4,5], and/or producing cytokines/chemokines that can profoundly influence the quality and magnitude of the adaptive immune responses [6][7][8][9]. They Correspondence: Dr. Sam K. P. Kung e-mail: Sam.Kung2@med.umanitoba.ca acquire specific chemokine surface receptors during development and maturation [2,[10][11][12][13][14][15]. Chemokine receptors such as CCR7, CCR5, and CXCR3 are involved in the preferential migrations and localization of NK cells into the LNs [8,[16][17][18][19], whereas NK cells reside in blood, liver, and spleen exhibit higher CXCR1 and CX 3 CR1 expression [8,9,[20][21][22][23][24][25].…”

Immunity and Vaccination against Tuberculosis in Cattle

“…NK cells are also cytotoxic through the release of perforin and granulysin leading to killing of M. bovis-infected APCs [21][22][23]. Release of perforin is attained both in association with and without DC involvement [24]. These experiments showed that cell-cell contact is necessary to result in both NK cell activation leading to release of IFN-γ and also DC activation as measured by an increase in MHCII expression.…”