The ontogeny and regulation of human natural killer cells

Natural Killer (NK) cells are members of the innate lymphoid cell (ILC) family and take part in the detection and eradication of virus-infected and transformed cells. In this thesis, together with my colleagues I have investigated how NK cells and other ILCs develop and function during human fetal development, how NK cells are functionally regulated (educated) via the activating receptor KIR2DS1, and how NK cells in our body are affected during the early phase of an acute viral infection.

Little is known about the ontogeny of NK cells and other ILCs in fetal development. The characterization of ILCs has been hampered by their overlapping surface phenotypes. In contrast, ILC transcription factor expression is more specific, and by combining multicolor flow cytometry analysis of transcription factors and surface markers expressed by different fetal ILC subsets, we were able to study and model their development and differentiation. All ILC subsets were detected as early as gestational week six, and their distribution varied depending on both tissue and gestational age. Moreover, putative precursors of NK cells were identified as cells that sequentially lost CD34 and acquired CD122, Eomes, CD94/NKG2A, T-bet, and CD16. In addition putative CD34+ progenitors of RORγt+ ILCs were identified.

In the second trimester of fetal development, analysis of mature fetal NK cell subsets revealed that stage 4 and stage 5 NK cells differed in frequency in fetal organs, and the highest NK cell frequency was found in the fetal liver and lung. The vast majority of fetal NK cells were NKG2A+, and fetal lung NK cells also frequently expressed killercell immunoglobulin-like receptors (KIR). Interestingly, while NKG2A educated fetal NK cells similar to adult NK cells, KIR expression on fetal NK cells was linked to hyporesponsiveness, thus contrasting education of NK cells after birth. Nevertheless, fetal NK cells were highly responsive to cytokines, as well as to antibody-coated target cells, suggesting they may take part in fetal immune responses against in utero infections, while remaining tolerant to maternal cells crossing the placenta.

While it is established that NK cells in adults are educated via inhibitory KIRs, it is not known how activating KIRs such as KIR2DS1 affects NK cells. By combining antibodies against four inhibitory KIRs, NKG2A, and KIR2DS1, we were able to interrogate the regulation of NK cells by KIR2DS1. We found that KIR2DS1 singlepositive NK cells exist in vivo, and that the presence of the ligand for KIR2DS1, HLAC2, resulted in hyporesponsiveness of such NK cells, thus ensuring self-tolerance. Our findings represent the first identification of NK cell education via activating KIR.

The human NK cell response to viral infections is not well understood. To this end we employed the live attenuated yellow fever vaccine 17D as an in vivo model of an acute viral infection. Our results show that the vaccine primed NK cells, and that less differentiated CD57- NK cells dominated the response, which peaked at day 6-10 post vaccination. Moreover, KIR expression on NK cells did not affect their response to the vaccine, indicating that NK cells expressing self-KIR and non-self KIR contributed equally to the NK cell response to the vaccination.

List of scientific papers

I. High-resolution mapping of transcription factor expression reveals distinct developmental pathways of human fetal innate lymphoid cells. Martin A. Ivarsson, Nicole Marquardt, Liyen Loh, Jeff E. Mold, Magnus Westgren, Erik Sundström, Elisabet Åkesson, Åke Seiger, Jenny Mjösberg, Niklas K. Björkström, Douglas F. Nixon, and Jakob Michaëlsson. [Manuscript]

II. Differentiation and functional regulation of human fetal NK cells. Martin A. Ivarsson, Liyen Loh, Nicole Marquardt, Eliisa Kekäläinen, Lena Berglin, Niklas K. Björkström, Magnus Westgren, Douglas F. Nixon, and Jakob Michaëlsson. Journal of Clinical Investigation. 2013;123(9):3889–3901.
https://doi.org/10.1172/JCI68989.

III. Education of human natural killer cells by activating killer cell immunoglobulin-like receptors. Cyril Fauriat, Martin A. Ivarsson, Hans-Gustaf Ljunggren, Karl-Johan Malmberg, and Jakob Michaëlsson. Blood. 2010 115: 1166-1174.
https://doi.org/10.1182/blood-2009-09-245746.

IV. The human NK cell response to yellow fever virus is primarily governed by NK cell differentiation independently of NK cell education. Nicole Marquardt*, Martin A. Ivarsson*, Kim Blom, Veronica D. Gonzalez, Monika Braun, Karolin Falconer, Rasmus Gustafsson, Anna Fogdell-Hahn, Johan K. Sandberg, and Jakob Michaëlsson. [Manuscript]