Skip to main content
. 2022 May 4;25(5):104347. doi: 10.1016/j.isci.2022.104347

Figure 7.

Figure 7

The dynamic NAD+ levels of Cd38−/− OT-1 cells in different models

(A) Left: livers from 5-month-old male WT or Cd38−/− mice were lysed by cold lysis buffer and the NAD+ levels were then measured and quantified according to the standard curve. Right: spleens from indicated mice were harvested and red cells were removed by RBC lysis, then an equal number of indicated cells were collected for NAD+ measurements. Data are shown as Mean ± SD (n = 3), Student’s t test, ∗∗: p < 0.01; ∗: p < 0.05.

(B) Representative FACS plots of mCherry-YFP and mCherry-FiNad fluorescence intensity in Vector or CD38 expressed Cd38−/− OT-1 cells.

(C) Left: comparing fold changes of the normalized mCherry-FiNad (green to red) ratios in indicated mCherry+ OT-1 cells. Data are shown as Mean ± SD (n = 5), Student’s t test, ∗∗: p < 0.01. Right: Vector or CD38 expressed Cd38−/− OT-1 cells were collected and detected ATP levels and ADP/ATP ratio. Calculating and summarizing the relative AXP ([ATP]+[ATP × ADP/ATP ratio]) levels of two groups. Data are shown as Mean ± SD (n = 4), Student’s t test, ns: not significant.

(D) Measurement of NAD+ levels with indicated Cd38−/− OT-1 cells. Data are shown as Mean ± SD (n = 3), Student’s t test, ∗: p < 0.05.

(E) Quantification of the relative NAD+ levels from the chronically stimulated cells, according to the values of AXP and normalized mCherry-FiNad (green to red) ratios as shown in Figure S6F. Data are shown as Mean ± SD (n = 3), Student’s t test, ∗∗∗∗: p < 0.0001; ∗∗: p < 0.01; ns: not significant.

(F) Representative FACS plots of mCherry-FiNad fluorescence intensity in mCherry+ WT or Cd38−/− OT-1 cells from recipient mice spleen or tumor.

(G) Summary of the relative green to red ratios of mCherry-FiNad in indicated mCherry+ OT-1 cells. Data are shown as Mean ± SD (n = 5), Student’s t test, ns: not significant.

(H) The B16-F10 tumors were implanted in WT or Cd38−/− C57BL/6 female mice, tumor infiltrated CD8+ T cells were purified by PE-anti-CD8a antibody and anti-PE beads at day 15 post engraftment. The NAD+ levels were measured as indicated in methods. Data are shown as Mean ± SD (n = 3), Student’s t test, ns: not significant.

(I and J) qPCR experiment showed the relative mRNA levels of Bst1, Sarm1, Slc12a8 and Nampt in WT or Cd38−/− OT-1 TILs. Mean ± SD (n = 3), Student’s t test, ∗∗: p < 0.01; ns: not significant.