餐后脂质代谢对T细胞应答的持续影响

Fig. 1 | Postprandial metabolism provides human T cells with improved metabolic and functional fitness that persists after in vitro expansion. a, Scheme of fasting and postprandial blood draws from human participants. b, Glucose uptake (left), neutral lipid content (middle; measured by BODIPY-NL (493/503)) and mitochondrial mass (indicated by MitoTracker Green (MTG), right) of CD8+ T cells from human peripheral blood mononuclear cells (PBMCs) ex vivo. GluCy3, 1-amino-Cy3-glucose; hIL-2, human IL-2; MFI, mean fluorescence intensity. c, OCR traces of a mitochondrial stress test of magnetically purified CD3+ T cells from human PBMCs collected in preprandial (fasted) or postprandial (fed) state (top); and tabulation of SRC (bottom). 2DG, 2-deoxy-d-glucose;  AA, antimycin a; FCCP, carbonyl cyanide p-trifluoromethoxyphenylhydrazone; rot., rotenone. d, Extracellular acidification rate (ECAR) of ex vivo CD3+ T cells as in c. Glycolytic capacity was calculated for ex vivo preprandial and postprandial T cells (lower panel). e, Cytokine production of CD8+ T cells stimulated with PMA and ionomycin for 6 h in the presence of a protein transport inhibitor. Representative fluorescence-activated cell sorting (FACS) plot (upper panel) and bar graphs of IFNγ+TNF+ CD8+ T cell frequencies (lower panel, left) and geometric MFI (lower panel, right). f–i, As in b–e, except that T cells from a were activated overnight with anti-CD3 + anti-CD28 mAbs and expanded in hIL-2 for 7 days. In b,e,f and i, data are mean ± s.e.m., n = 10 human donors, two-tailed paired Student’s t-test analysis. In c,d,g and h, data are mean ± s.e.m., n = 6 wells; two-tailed unpaired Student’s t-test analysis. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Data are representative of 31 human donor samples. Human cartoon in panel a adapted from ref. 48, Springer Nature America.

Fig. 2 | Postprandial CD8+ T cells respond more vigorously to infection in vivo. a, Schema: naive CD8+ T cells from Thy1.1 and Thy1.1/1.2 OT-I mice fed ad libitum (ad lib) or fasted 12 h were transferred 1:1 (see preinjection cytogram) to VV-OV-infected mice (106 PFU i.p. (intraperitoneal)). b, Cytogram (upper) and ratios (lower) of transferred splenic CD8+ T cells at day 11 (d11) post-infection (p.i.) (n = 8 mice). c, GluCy3 uptake (left), BODIPY-NL (middle) and MTG (right) of transferred OT-I at d11 p.i. (n = 5 mice). d, OCR (top) and SRC (bottom) of OT-I T cells at d11 p.i. e, ECAR from d. f, Cytokine production in T cells (n = 5 mice). g, Schema: 2 × 105 fasted or fed naive OT-I T cells were separately transferred to VV-OVA-infected mice for 45 d, then rechallenged with DEC-205:OVA (50 μg i.p.). h, Cytogram of T cells from fasted or fed OT-I mice in the blood of recipient mice at d7 p.i. (left), d45 p.i. (middle) and d52 (7 d post-DEC-205:OVA) (n = 6 mice). i, As in a, but mice were preinfected (d−1) or coinfected (d0) or underwent delayed infection (d+1, d+3, d+5). j, Cytogram (upper) and ratios (lower) of transferred OT-I T cells in blood (n = 6 mice). k, As in j, but at d45 p.i. l, Cytogram of postprandial effects on memory T cells (n = 6 mice). Congenically mismatched OT-I T cells from fed mice were separately transferred to VV-OVA infected mice. At memory stage (d45 p.i.), mice were fasted or fed before isolation and cotransfer of OT-I cells into DEC-205:OVA-immunized mice (n = 11 mice). m,n, Differentially expressed genes in fasted versus fed CD8+ T cells ex vivo (m) and 24 h after in vitro TCR stimulation (n) (n = 3 mice). In b,c,f,h and j–n, data are mean ± s.e.m., n = 5 to n = 11 mice. In d and e, n = 6 wells for fasted and n = 3 wells for fed. Two-tailed unpaired (b–e,h,m and n) or two-tailed paired (f and l) Student’s t-test analysis. For j and k, one-way analysis of variance (ANOVA) (statistics with respect to d+5). Data represent three independent experiments. Mouse cartoons in panels a, g, i and l adapted from ref. 48, Springer Nature America.

Fig. 4 | Postprandial metabolic changes induce mTOR-dependent translation of key immune and metabolic genes. a,b, Volcano plot of proteomes from ex vivo (a) and 24 h TCR-activated (b) CD8+ T cells from fasted and fed mice (n = 3 per group). c, Immunoblot of puromycin and actin in 18 h TCR-stimulated CD8+ T cells from fasted and fed mice. Cells were treated with puromycin (10 μg ml−1) for 2 h before collection. d, Flow cytogram of puromycin staining in mice that had been fasted or fed. Some mice received an i.p. injection of rapamycin (rapa.; 5 mg kg−1) during the fast/feed period. Stim., stimulation. e, p-S6, p-4E-BP1 and p-AKT staining in ex vivo CD8+ T cells from fasted and fed mice as in d. f, As in e, but after 18 h of TCR + CD28 activation. g, p-4E-BP1 in naive CD8+ T cells activated for 18 h plus chylomicrons in the presence of cholesterol uptake inhibitor itraconazole (ITRA; 2 μM). h, Puromycin staining of fasted or fed T cells treated with chylomicrons during activation. i,j, OCR (i) and ECAR (j) traces of ex vivo CD8+ T cells from fasted and fed mice that were treated with or without rapamycin in vivo during the fast/feed period (n = 3 to n = 4 wells). k, Relative expansion of OT-I CD8+ T cells at day 11 p.i. with VV-OVA (n = 5) as in Fig. 2a, but with some donor OT-I mice having received rapamycin during the feeding period. In d–h, data are mean ± s.e.m., n = 3 mice per group, one-way ANOVA. In i–k, data are mean ± s.e.m., one-way ANOVA. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Data in c–k are representative of three independent experiments. Mouse cartoon in panel d adapted from ref. 48, Springer Nature America.

Fig. 5 | Postprandial metabolism provides therapeutic T cells with enhanced anti-tumour immunity. a, Scheme showing adoptive transfer of in vitro expanded TCR-Tg Pmel-1 CD8+ T cells (initially isolated from fasted or fed mice) in B16-F10-bearing mice. b, Tumour growth curve (area) of B16-F10-bearing mice. PBS, phosphate-buffered saline. c, Survival curve of mice as in b (n = 10 mice per group; pooled from two independent experiments). d, Experimental scheme for human CAR-T studies. e, OCR trace of human CAR-T cells generated from preprandial or postprandial samples, measured on d12 of their in vitro expansion. SRC was compared among groups. f, ECAR and glycolytic capacity of CAR-T cells. g, Killing assay for d12 expanded human CAR-T cells using A549 hCD19 adherent cells as targets. h, Representative luminescence images of NALM6Luc-engrafted mice that received transfer of human CAR-T cells on d4. Untransduced (UTD, left), pre-CAR (middle) and post-CAR (right) on day 3, day 6, day 25 and day 32. i, NALM6Luc luminescence in mice over time as in h. j, Survival curve for i (n = 23 mice for UTD; n = 16 mice for pre-CAR; and n = 15 mice for post-CAR group). k, Cytogram of cytokine production in preprandial and postprandial CAR-T cells from separate cohorts of NALM6-engrafted NSG mice at day 7 posttransfer (n = 5 mice per group). Splenocytes were cocultured with NALM6Luc cells overnight in the presence of a protein transport inhibitor. For g–k, experiments were performed with three different human donors. In e–g, data are mean ± s.e.m., n = 8 (e and f) or n = 3 (g) wells, one-way ANOVA. In k, data are mean ± s.e.m., n = 5 recipient mice per group, two-tailed unpaired Student’s t-test analysis. Statistical analysis used two-way ANOVA with Tukey’s multiple comparisons (b and i) or log-rank Mantel–Cox test (c and j). Mouse and human cartoons in panels a and d adapted from ref. 48, Springer Nature America.