Activated Notch Supports Development of Cytokine Producing NK Cells Which Are Hyporesponsive and Fail to Acquire NK Cell Effector Functions

Natural Killer (NK) cells are powerful effectors of cytotoxicity against "stressed" cells. They also produce cytokines and chemokines to activate the adaptive immune response. Understanding NK cell development and maturation may have implications for cancer therapy and for immunity against infections. We hypothesized that Notch signaling, critical for hematopoesis, would be involved in NK cell development. The role of constitutively activated Notch1 (ICN) on NK cell maturation was studied using human umbilical cord blood (UCB) progenitors cultured on a murine embryonic liver stroma cell line (EL08-1D2) and human cytokines. UCB CD34⁺/ICN⁺ sorted cells resulted in a population of CD7⁺ early lymphoid precursors and subsequent NK lineage commitment independent of stroma or IL-15. Early expression of L-selectin on ICN⁺ precursors suggested their homing competence. These precursors further committed to the NK lineage, and were capable of producing cytokines and chemokines such as interkeukin (IL)-13, granulocyte macrophage-colony stimulating factor (GM-CSF), tumor necrosis factor-alpha (TNF-α), yet poorly acquired NK inhibitory receptors and cytotoxic effector function. In the presence of stroma, ICN⁺ precursors also gave rise to a population of early T lineage committed cells characterized by expression of cytoplasmic CD3 γ, ε{lunate}, and δ chains, RAG1/2, and production of IL-2, suggesting bona fide Th1 commitment. Importantly, signals from EL08-1D2 stroma were required for this development process. In conclusion, sustained Notch signaling can replace stroma in differentiation of a common CD7⁺ lymphoid precursor from UCB CD34⁺ progenitors and induce NK cell commitment. However, these NK cells are immature in their cytokine production profile, are hyporesponsive, and poorly acquire NK cell receptors involved in self-tolerance and effector function.