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. 2017 May 18;2(10):e87032. doi: 10.1172/jci.insight.87032

Figure 2. Adoptively transferred unlicensed NK cells preferentially traffic to draining lymph nodes following viral infection.

Figure 2

(A) Schema of adoptive transfer study where 20 × 106 splenocytes from naive CD45.1 congenic mice were i.v. adoptively transferred to naive C57BL/6 (CD45.2) recipients. Two days after adoptive transfer, recipient mice were challenged with either 100 PFU of APR8 in 40 μl of PBS or 40 μl of PBS (control) intranasally (i.n.). Five days later, the absolute number of donor (CD45.1) NK cells in the mediastinal lymph nodes (mLNs), nondraining lymph nodes (ndLNs), and lung were determined. (B) Absolute number of transferred (CD45.1+) NK cells (CD3-NK1.1+) in the mLNs, ndLNs, or lung of C57BL/6 (CD45.2) mice at 5 days postinfection (d.p.i.). (C) Absolute number of transferred (CD45.1+) licensed or unlicensed NK cell subsets in the mLNs of C57BL/6 (CD45.2) mice at 5 d.p.i. (D) Absolute number of transferred (CD45.1+) licensed or unlicensed NK cell subsets in the lung of C57BL/6 (CD45.2) mice at day 5 d.p.i. (E) Schema of adoptive transfer study where C57BL/6 (CD45.2) mice were challenged with 100 PFU of APR8 in 40 μl of PBS or 40 μl of PBS (control) i.n. One day later, 20 × 106 splenocytes from naive CD45.1 congenic mice were i.v. adoptively transferred to challenged C57BL/6 (CD45.2) recipients. One and 3 days after adoptive transfer, the absolute number of donor (CD45.1) NK cells in the mLNs was determined. (F) Absolute number of transferred (CD45.1+) licensed or unlicensed NK cell subsets in the mLN of C57BL/6 (CD45.2) mice at 1 and 3 d.p.i. (G) Histograms showing the level of Ki-67 of transferred (CD45.1+) NK cells in the mLNs of C57BL/6 (CD45.2) mice at 1 and 3 d.p.i. Three-day concanavalin A (Con A)–stimulated splenocytes were included as a positive control. (H) Absolute number of licensed or unlicensed NK cell subsets that express the chemokine receptor CXCR3 in the spleen, lung, or liver of naive C57BL/6 (H2b) mice. (I) Absolute number of NK cell subsets (CD3–NK1.1+Ly49C/I+G2+A+, Ly49C/I+G2+A–, Ly49C/I+G2–A+, Ly49C/I+G2–A–, Ly49C/I–G2+A+, Ly49C/I–G2+A–, Ly49C/I–G2–A+, or Ly49C/I–G2–A–) that express CXCR3 in the spleen of naive C57BL/6 mice. (J) Absolute number of licensed or unlicensed NK cell subsets that express CXCR3 in the spleen of naive B10.D2 (H2d) mice. n = 3–6 mice per group, representative of 2 to 3 experiments. (C, D, and F) Two-way ANOVA with Tukey post-test used to compare groups. (B, H, and J) Unpaired 2-tailed Student’s t test used to compare groups. *P < 0.05, **P < 0.01, ***P < 0.001.