Natural Killer Cell Functional Activity After 4-1BB Costimulation Shadi sadat Navabi, 1 Mehrnoosh Doroudchi, 1 Ahmad Hosseini Tashnizi, 2 and Mojtaba Habibagahi 1,3 Abstract—Reports show enhancement of CD8 T cells’ activity through CD137 (4-1BB) signal; how- ever, not all data proved similar effect in natural killer (NK) cells. Here, the impact of 4-1BB signal on NK cells’ function was assessed during short term cultures. To that end, cytokine-activated NK cells were cocultured with adenovirally transduced MCF-7 stimulator cells expressing 4-1BB ligand. Cellular cytotoxicity, cytokine production, and expression of cytotoxicity related genes were assessed after overnight cultures. Sharp decrease of CD56 + and CD56 bright NK cells was demonstrated. 4-1BB neither enhanced cellular degranulation nor improved IFN-γ production although it promoted granzyme B, perforin, and FasL gene expression. 4-1BB signal stimulated higher proportions of CD56 bright population to degranulate and express CD107a; however, it could not recover killing activity against K562 targets. Our data could not show major promotion in activity of all NK subpopulations. Due to great heterogeneity of NK cells, more investigation is needed to draw a comprehensive conclusion. KEY WORDS: CD137 (4-1BB); CD137 ligand (4-1BBL); costimulation; natural killer cell. INTRODUCTION Natural killer (NK) cells are known as CD3 - TCR - large granular cytotoxic lymphocytes with major immune reactivity against tumors and virus-infected cells without prior activation. Unique features of NK cells have inspired scientists to exploit NK cells for cancer immunotherapy. However, NK cells are not a homogeneous population and several subtypes can be recognized among these cells which might not react similarly against their targets [1]. Unlike T and B lymphocytes, receptors of NK cells do not undergo genetic rearrangement. Instead, innate immune system contains diverse arrays of NK cells with differential expression of a wide variety of receptors and functions [2– 4]. Simply, based on the expression levels of neural cell adhesion molecule (NCAM, CD56), NK cells can be subdivided as CD56 bright (high expression) and CD56 dim (low expression) which can reflect their differences in function as well [1]. In the current models of NK cell activation, NK cells govern cell activation/inhibition by keeping precise balance between signals transduced from activating and inhibitory receptors [5]. Recent evidence also show adaptive-like immune reactivities of NK cells, similar to Ag specific and memory formation of T cells, which should be incorporated in the activation scenario of NK cells. In fact, in addition to the traditional ideas of existence of “missing self receptors” and the “pathogen recognition receptors,” NK cells express arrays of other surface molecules which can participate in the process of cell activation. It has been demonstrated that NKG2D transduces activation signals to NK cells similar to CD28 to activate p85 subunit of phosphatidylinositol 3-kinase (PI3K) [6]. In this regard, it is being increasingly appreci- ated that activation of NK cells, like T cells, can take advantage of several costimulatory ligand receptor interac- tions [3, 7]. In case of T cells, multiple costimulatory interactions during Ag stimulation can shape the ultimate T cell function [8–10]. Among them, signal transduction by CD137 (4-1BB), a member of tumor necrosis factor receptor (TNFR) superfamily, costimulates T cells inde- pendent of CD28 for proliferation, and enhances T cell 1 Department of Immunology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran 2 Institute for Cancer Research, Shiraz University of Medical Sciences, Shiraz, Iran 3 To whom correspondence should be addressed at Department of Immu- nology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran. E-mail: agahim@sums.ac.ir 0360-3997/14/0000-0001/0 # 2014 Springer Science+Business Media New York Inflammation ( # 2014) DOI: 10.1007/s10753-014-0082-0