Murine peripheral NK-cell populations originate from site-specific immature NK cells more than from BM-derived NK cells.

Murine NK cells can be divided by the expression of two cell surface markers, CD27 and Mac-1 (a.k.a. CD11b), into four separate subsets. These subsets suggest a linear development model: CD27(-) Mac-1(-) → CD27(+) Mac-1(-) → CD27(+) Mac-1(+) → CD27(-) Mac-1(+) . Here, we used a combination of BrdU labeling experiments and mathematical modeling to gain insights regarding NK-cell development in mouse bone marrow (BM), spleen and liver. The modeling results that best fit the experimental data show that the majority of NK cells already express CD27 upon entering the NK-cell developmental pathway. Additionally, only a small fraction of NK cells exit the BM to other sites, suggesting that peripheral NK-cell populations originate from site-specific immature NK cells more than from BM-derived mature NK cells.

Classification of human natural killer cells based on migration behavior and cytotoxic response.

Despite intense scrutiny of the molecular interactions between natural killer (NK) and target cells, few studies have been devoted to dissection of the basic functional heterogeneity in individual NK cell behavior. Using a microchip-based, time-lapse imaging approach allowing the entire contact history of each NK cell to be recorded, in the present study, we were able to quantify how the cytotoxic response varied between individual NK cells. Strikingly, approximately half of the NK cells did not kill any target cells at all, whereas a minority of NK cells was responsible for a majority of the target cell deaths. These dynamic cytotoxicity data allowed categorization of NK cells into 5 distinct classes. A small but particularly active subclass of NK cells killed several target cells in a consecutive fashion. These "serial killers" delivered their lytic hits faster and induced faster target cell death than other NK cells. Fast, necrotic target cell death was correlated with the amount of perforin released by the NK cells. Our data are consistent with a model in which a small fraction of NK cells drives tumor elimination and inflammation.

Lyn deficiency affects B-cell maturation as well as survival.

Lyn, an Src-family protein tyrosine kinase expressed in B lymphocytes, contributes to initiation of BCR signaling and is also responsible for feedback inhibition of BCR signaling. Lyn-deficient mice have a decreased number of follicular B cells and also spontaneously develop a lupus-like autoimmunity. We used flow cytometric analysis, BrdU labeling and our mathematical models of B-cell population dynamics, to analyze how Lyn deficiency impacts B-cell maturation and survival. We found that Lyn-deficient transitional 1 (T1) cells develop normally, but T2 cells develop primarily from the T1 subset in the spleen and fail to also develop directly from BM immature B cells. Lyn-deficient T2 cells either mature to the follicular B-cell type at a close to normal rate, or die in this compartment rather than access the T3 anergic subset. The ≈ 40% of WT follicular cells that were short-lived exited primarily by joining the T3 anergic subset, whereas the ≈ 15% Lyn(-/-) follicular cells that were not long lived had a high death rate and died in this compartment rather than entering the T3 subset. We hypothesize that exaggerated BCR signaling resulting from weak interactions with self-antigens is largely responsible for these alterations in Lyn-deficient B cells.

Natural killer cell inhibitory receptor expression in humans and mice: a closer look.

The Natural Killer (NK) cell population is composed of subsets of varying sizes expressing different combinations of inhibitory receptors for MHC class I molecules. Genes within the NK gene complex, including the inhibitory receptors themselves, seem to be the primary intrinsic regulators of inhibitory receptor expression, but the MHC class I background is an additional Modulating factor. In this paper, we have performed a parallel study of the inhibitory receptor repertoire in inbred mice of the C57Bl/6 background and in a cohort of 44 humans. Deviations of subset frequencies from the "product rule (PR)," i.e., differences between observed and expected frequencies of NK cells, were used to identify MHC-independent and MHC-dependent control of receptor expression frequencies. Some deviations from the PR were similar in mice and humans, such as the decreased presence of NK cell subset lacking inhibitory receptors. Others were different, including a role for NKG2A in determining over- or under-representation of specific subsets in humans but not in mice. Thus, while human and murine inhibitory receptor repertoires differed in details, there may also be shared principles governing NK cell repertoire formation in these two species.

Understanding natural killer cell regulation by mathematical approaches.

The activity of natural killer (NK) cells is regulated by various processes including education/licensing, priming, integration of positive and negative signals through an array of activating and inhibitory receptors, and the development of memory-like functionality. These processes are often very complex due to the large number of different receptors and signaling pathways involved. Understanding these complex mechanisms is therefore a challenge, but is critical for understanding NK cell regulation. Mathematical approaches can facilitate the analysis and understanding of complex systems. Therefore, they may be instrumental for studies in NK cell biology. Here we provide a review of the different mathematical approaches to the analysis of NK cell signal integration, activation, proliferation, and the acquisition of inhibitory receptors. These studies show how mathematical methods can aid the analysis of NK cell regulation.

Models and methods for analysis of lymphocyte repertoire generation, development, selection and evolution.

T and B cell receptor repertoires are diversified by variable region gene rearrangement and selected based on functionality and lack of self-reactivity. Repertoires can also be defined based on phenotype and function rather than receptor specificity - such as the diversity of T helper cell subsets. Natural killer (NK) cell repertoires, in which each cell expresses a randomly chosen subset of its inhibitory receptor genes, and is educated based on self-MHC recognition by yet unknown mechanisms, are also phenotypic repertoires. Studying the generation, development and selection of lymphocyte repertoires, and their functions during immune responses, is essential for understanding the function of the immune system in healthy individuals and in immune deficient, autoimmune or cancer patients. The study of lymphocyte repertoires will enable clinical immunologists to develop better therapeutic monoclonal antibodies, vaccines, transplantation donor-recipient matching protocols, and other immune intervention strategies. The recent development of high-throughput methods for repertoire data collection - from multicolor flow cytometry through single-cell imaging to deep sequencing - presents us now, for the first time, with the ability to analyze and compare large samples of lymphocyte repertoires in health, aging and disease. The exponential growth of these datasets, however, challenges the theoretical immunology community to develop methods for data organization and analysis. Furthermore, the need to test hypotheses regarding immune function, and generate predictions regarding the outcomes of medical interventions, necessitates the development of complex mathematical and computational models, covering processes on multiple scales, from the genetic and molecular to the cellular and system scales.