Journal: bioRxiv
Article Title: CD4⁺ T cells confer transplantable rejuvenation via Rivers of telomeres
doi: 10.1101/2025.11.14.688504
Figure Lengend Snippet: ( a ) Experimental design for in vivo River generation with or without CD4⁺ T cell depletion. CD4-proficient or CD4-depleted recipient mice were adoptively transferred with telomere-labelled APCs, pre-loaded or not with Fluad vaccine as antigen source, with or without anti-LFA-1α antibody. Serum was collected 15 days later for River collection. Three months mice were used. ( b ) Representative FACS plots showing efficiency of CD4⁺ T cell depletion (top) and detection of telomere Rivers in each experimental group (bottom). Data are from ( n = 5 mice per group). ( c ) Rejuvenating effects of telomere Rivers. Rivers were isolated from mouse serum as in a by FAVS sorting (PKH67⁺TelC⁺GAPDH⁻) and transplanted subcutaneously into 20-month-old congenic recipients. Animals were bred normally and assessed 12 months later. Flow cytometry of different organs demonstrated reduced senescence markers (β-Gal activity) and increased telomere length. For p16, IL-6, sestrin2-p-p38 (sMAC)), Extended Fig. 10) Data are from ( n = 5 mice per group). MFI, mean fluorescence intensity. Young (3-month-old) mice are shown, as control. ( d ) Artificial River generation, experimental design. APCs were purified from total splenocytes of 3-month-old mice and transfected with siRNAs to silence GAPDH or with control siRNA. APC telomeres were then labelled with TelC probe and ionomycin stimulated in vitro to produce vesicles. Telomere vesicles (siCtrl vesicles) or artificial Rivers (siGAPDH vesicles) were sorted and injected into 20-month-old mice to test their rejuvenating effect. ( e ) Elongation of tissue telomeres by artificial Rivers. Telomere length was evaluated by Flow-FISH in multiple target tissues from the same 20-month-old mice as in d . Results are shown as absolute base-pairs (bp). Data are pooled from ( n = 12 organs per group) and expressed as ΔTL relative to the old baseline at the start of the experiment (T0; grey bar), thereby distinguishing anti-aging effects (maintenance of old baseline; left) from rejuvenating effects (progression toward young reference values; right). siCTRL telomere vesicles largely delayed further telomere attrition without substantial rejuvenation, whereas artificial Rivers (siGAPDH) induced marked telomere elongation toward young reference values. Young (3-month-old) reference telomere lengths are indicated in yellow. In parallel, untreated old mice were aged normally to derive telomere attrition trajectories. ( f ) Appearance of representative mice injected with telomere vesicles (siCtrl vesicles) or with artificial telomere Rivers (siGAPDH vesicles). Note fur and body lean differences ( n = 10 mice per group). One-way Anova for repeated measures with Bonferroni post-test correction ( b - e ). **P < 0.01, ***P < 0.001, ****P < 0.0001. Error bars indicate s.e.m.
Article Snippet: Where indicated, cells were additionally treated with anti-LFA-1α blocking antibody (InVivoMAb anti-mouse CD11a, clone M17/4; Bio X Cell, 1 μg per 106 cells) immediately prior to transfer.
Techniques: In Vivo, Isolation, Flow Cytometry, Activity Assay, Fluorescence, Control, Purification, Transfection, In Vitro, Injection
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