Research Paper Volume 11, Issue 15 pp 5628—5645

Rejuvenation of brain, liver and muscle by simultaneous pharmacological modulation of two signaling determinants, that change in opposite directions with age

Figure 2. Alk5i+OT treatment improves hippocampal neurogenesis and attenuates inflammation in the brains of old mice. (A) Immunofluorescence was performed on serial 25-micron brain sections with anti-Ki67 (proliferation marker) and anti-Sox-2 (neural stem cell marker), using Hoechst to stain all nuclei. A representative image of ki67 (red)/Sox2 (green)/Hoechst (blue) triple positive cells in the hippocampal Dentate Gyrus of a young control animal treated with HBSS for 7 consecutive days is shown. Scale bar=50 µm. Isotype-matched IgG negative controls exhibited minimal background fluorescence. (B) Immunofluorescence was performed on serial 25-micron brain sections with anti-Ki67 (proliferation marker), using Hoechst to stain all nuclei, imaging the cells in the SGZ of the hippocampal Dentate Gyrus. Representative images of Ki67 (red)//Hoechst (blue) positive cells in the hippocampal Dentate Gyrus. Arrows point to these double-positive cells. Scale bar=50 µm. (C) Immunofluorescence was performed on serial 25-micron brain sections with anti-CD68 (monocyte/microglia marker), using Hoechst to stain all nuclei. Representative images of CD68 (red)/Hoechst (blue) double positive cells. Scale bar=50 µm. (D) The numbers of Ki67+/Hoechst+ cells in the SGZ of DG were quantified through entire hippocampi of each cohort and were found to decline with age as expected, and to increase in the Alk5i+OT old cohort, as compared to the control vehicle-treated old cohort. Young control (Yc n=5), old control (Oc n=5), Alk5i+OT (OA5iOT n=6) *p Oc & OA5iOT = 0.043, **p Yc & OA5iOT = 0.00159, mean and SE are shown. (E) The number of CD68+ brain cells were quantified in all cohorts and were found to increase with age and to decline in Alk5i+OT-treated old muscle, as compared to the vehicle-treated old control. N young control (Yc n=5), old control (Oc n=5), Alk5i+OT (OA5iOT n=6). ***p<0.001, **p=0.0226.