Human natural killer (NK) cells play a critical role in the control of viral infection, particularly herpesviral
infections such as Epstein-Barr virus (EBV). Their primary function of killing of virally infected target cells is
mediated by the contact-dependent lysis through directed secretion of perforin and granzymes. Despite the
demonstrated importance of their development and function, our understanding of how NK cells interact with
other cells within tissue is limited by a lack of satisfying models for studying human immunity live and in situ.
Significant challenges arise when trying to visualize cells within tissue, however it is highly relevant to human
disease to find ways to better define how particularly innate immune cells migrate and exert their cytotoxic
function. Our previous research has used 2D cell culture systems to define the importance of cell migration in
human NK cell development. In this application, we propose the adoption of previously methods of secondary
lymphoid tissue (tonsil) histoculture as a model to study human NK cell migration and function within an in vivolike
tissue microenvironment using fast, low-toxicity imaging. We will combine our expertise in human
developmental immunology with our technical skills in cutting-edge microscopy and image analysis to visualize
human NK cells within autologous tissue and measure migration in 3 dimensions. We will additionally use
multi-parametric imaging mass cytometry to localize cells within tissue (Aim 1). Further, we will extend our
model to include infection of the tonsils with EBV, which will allow us to measure differences in cell migration
and function between infected and uninfected tissue. By integrating infection of tissue histoculture with human
EBV we will enable the study of human NK cells killing physiologically relevant targets within tissue (Aim 2).
The generation of this novel model will advance future studies of human immunity and enable unprecedented
insight into the behavior of immune cells within their tissue microenvironment.