Extended Data Fig. 6: Cell non-autonomous effects of an aged immune system in non-lymphoid tissues of Vav-iCre+/−;Ercc1−/fl mice.
From: An aged immune system drives senescence and ageing of solid organs
a, Cyclopurine adducts were measured in the liver and kidneys of 8–11-month-old Vav-iCre+/−;Ercc1−/fl (n = 5) and littermate control Vav-iCre+/− (n = 5 for liver and n = 6 for kidney) by LC–MS/MS/MS (Methods). b, Markers of oxidative stress including HNE protein adducts and the ratio of reduced to oxidized glutathione (GSH/GSSG) measured in the kidneys from 8–11-month-old Vav-iCre+/−;Ercc1−/fl and littermate control Vav-iCre+/− mice (n = 6 mice per group). HNE measure by ELISA. GSH/GSSG measured by chromogenic assay (Methods). c, 8-oxo-guanine DNA adducts measured by ELISA in the spleen, liver and kidney of mice at various ages (n = 5–6/5–6/5 Vav-iCre+/−;Ercc1−/fl; n = 5–6/5–6/5 Vav-iCre+/−; n = 5/5/10 old WT mice for spleen/liver/kidney, respectively) (see Supplementary Table 3 for sample size details by genotype and tissue). d, Urinary levels of pro-geronic factor β2-microglobulin and MCP-1 measured by ELISA in 8–11-month-old Vav-iCre+/−;Ercc1−/fl and littermate controls (n = 9 mice per group). e, Renal Mcp1 (also known as Ccl2) expression in 8–11-month-old Vav-iCre+/−;Ercc1−/fl mice (n = 7 per group) measured by qRT–PCR. f, Representative Coomassie-stained gel of urine samples from 8–11-month-old Vav-iCre+/−;Ercc1−/fl and littermate control mice demonstrating increased proteinuria. Recombinant albumin (Alb) was loaded on the gel as a control (lanes 6, 12) and its approximate molecular mass denoted by a box (marker ladder lanes 1, 5, 7, 11, 13–14). Each lane represents a unique mouse. g, Representative images from tissue sections stained for aggrecan (red) and DAPI (blue) in the nucleus pulposus (NP) of intervertebral discs from 8–11-month-old Vav-iCre+/−;Ercc1−/fl and littermate control mice. h, Quantification of aggrecan staining (n = 4 Vav-iCre+/−; n = 7 Vav-iCre+/−;Ercc1−/fl). i, Measurement of senescence marker expression in the intervertebral discs of 8–11-month-old Vav-iCre+/−;Ercc1−/fl and littermate control mice (n = 4 mice per group) by qRT–PCR. j, Representative images of sections of intervertebral discs from 9-month-old mice stained with haematoxylin and eosin (H&E) and safranin O to detect proteoglycans. Arrows point to the anulus fibrosus. k, Representative images of gastrocnemius muscle sections from 8–11-month-old Vav-iCre+/−;Ercc1−/fl and littermate control mice after cardiotoxin injury (Methods) stained with haematoxylin and eosin or immunostained for M1 (CD68, green) and M2 (CD163, red) macrophages. l, Quantification of the ratio of M2/M1 macrophages (n = 4 Vav-iCre+/−; n = 8 Vav-iCre+/−;Ercc1−/fl mice). m, Body weight and grip strength of Vav-iCre+/−;Ercc1−/fl and littermate controls at the indicated ages (n = 8 Vav-iCre+/−;Ercc1−/fl; n = 4 Vav-iCre+/− at both ages). Data are mean ± s.d. *P < 0.05, ∞P < 0.01, ∇P < 0.001, #P < 0.0001, two-tailed unpaired Student’s t-test (a, b, d, e, h, i, l, m) or two-way ANOVA with Tukey’s test (c).
