The Natural Cytotoxicity Receptor NKp44 (NCR2, CD336) Is Expressed on the Majority of Porcine NK Cells Ex Vivo Without Stimulation.

Kerstin H Mair,Dante Zarlenga,Bettina Wagner,Assiatu J Crossman,Joan K Lunney,Katinka A Van Dongen,Wilhelm Gerner,Maria Stadler,Leela Noronha,Patricia Boyd,Armin Saalmüller,Susanna Babasyan

doi:10.3389/fimmu.2022.767530

Abstract

Natural killer (NK) cells have been studied extensively in humans and mice for their vital role in the vertebrate innate immune system. They are known to rapidly eliminate tumors or virus infected cells in an immune response utilizing their lytic properties. The natural cytotoxicity receptors (NCRs) NKp30 (NCR3), NKp44 (NCR2), and NKp46 (NCR1) are important mediators of NK-cell cytotoxicity. NKp44 expression was reported for NK cells in humans as well as in some non-human primates and found exclusively on activated NK cells. Previously, no information was available on NKp44 protein expression and its role in porcine lymphocytes due to the lack of species-specific monoclonal antibodies (mAbs). For this study, porcine-specific anti-NKp44 mAbs were generated and their reactivity was tested on blood and tissue derived NK cells in pigs of different age classes. Interestingly, NKp44 expression was detected ex vivo already on resting NK cells; moreover, the frequency of NKp44+ NK cells was higher than that of NKp46+ NK cells in most animals analyzed. Upon in vitro stimulation with IL-2 or IL-15, the frequency of NKp44+ NK cells, as well as the intensity of NKp44 expression at the single cell level, were increased. Since little is known about swine NK cells, the generation of a mAb (clone 54-1) against NKp44 will greatly aid in elucidating the mechanisms underlying the differentiation, functionality, and activation of porcine NK cells.

Highlights

Natural killer (NK) cells are specialized innate effector lymphocytes identified by their potent cytolytic activity against tumor or virus infected cells [1]
Based on human NK cell data, peripheral blood mononuclear cells (PBMCs) were stimulated with IL-2 and IL-15; this resulted in upregulation of staining from 3 to 9% NKp44+ PBMC
In human NK cells, the cell surface receptor NKp44 has been linked to NK-cell activation as it is only found on stimulated NK cells [7, 9, 10]

Summary

Introduction

Natural killer (NK) cells are specialized innate effector lymphocytes identified by their potent cytolytic activity against tumor or virus infected cells [1]. One important family of activating receptors are the natural cytotoxicity receptors (NCRs) comprised of three members, NKp30 (NCR1, CD337), NKp44 (NCR2, CD336) and NKp46 (NCR3, CD335), that play important roles in NK-cell recognition of aberrant or infected cells [3]. NKp44 was extensively studied on human NK cells but to date, there is no clear evidence of NKp44 expression in the mouse [4, 5]. NKp44 transcripts were found in non-human primates [6]. While five-fold higher NKp44 transcript levels were found in chimpanzees, much lower differential increases were observed upon cell activation when compared with humans. In Macaca fascicularis, NKp44 was found at the transcript level, but due to frame shifts in the sequence, only abortive transcripts were detected [6]

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