Journal: Aging Cell

Article Title: Activation of angiotensin‐converting enzyme 2/angiotensin (1–7)/mas receptor axis triggers autophagy and suppresses microglia proinflammatory polarization via forkhead box class O1 signaling

doi: 10.1111/acel.13480

Figure Lengend Snippet: LPS exposure shifts the balance of brain RAS. (a) The impacts of acute LPS (AcLPS) or repeated LPS (ReLPS) exposure on microglial activation (IBA‐1 staining) and ROS generation (DHE staining) in the brain cortex of C57BL/6 mice. (b) Biomarkers of microglial M1 phenotype mRNA expression. (c) Biomarkers of microglial M2 phenotype mRNA expression. (d–m) Alterations of ACE/AngII/AT1 and ACE2/Ang(1–7)/MasR pathways following LPS treatment. (d) Representative western blots. ACE protein expression (e) and activity (f). (g) AngII concentration. (h) AT1 protein expression. (i) AT2 protein expression. ACE2 protein expression (j) and activity (k). (L) Ang(1–7) concentration. (m) MasR protein expression. Scale bar = 50 μm. Data are means ± SD ( n = 7–9). * p < 0.05, ** p < 0.01 compared to control group

Article Snippet: The sections or fixed cells were then incubated with the following primary antibodies: anti‐MasR (Novus, NBP1‐78444, 1:30), anti‐FOXO1 (Proteintech, 18592‐1‐AP; 1:100), anti‐IBA‐1 (Abcam, ab178847, 1:200), anti‐LC3 (Cell signaling, 4108; 1:200), or anti‐ASC (Novus, NBP1‐78977, 1:50).

Techniques: Activation Assay, Staining, Expressing, Western Blot, Activity Assay, Concentration Assay

Journal: Aging Cell

Article Title: Activation of angiotensin‐converting enzyme 2/angiotensin (1–7)/mas receptor axis triggers autophagy and suppresses microglia proinflammatory polarization via forkhead box class O1 signaling

doi: 10.1111/acel.13480

Figure Lengend Snippet: MasR activation triggers FOXO1 signaling and promotes autophagy. (a–e) The impacts of MasR agonist, AVE, and the ACE2 activator, DIZE, on the expression of FOXOs following repeated LPS treatment. (a) Representative western blots of FOXOs. Statistical graphs of protein expression of FOXO1 (b), FOXO3 (c), and FOXO6 (d). (e) Representative images of immunofluorescence assays of FOXO1. (f–j) The impacts of MasR agonist, AVE, and the ACE2 activator, DIZE, on autophagic genes. (f) mRNA expression of autophagic genes. (g) Representative western blots of autophagic genes. Statistical graphs of protein expression of LC3II (h), Beclin1 (i), and ATG5‐ATG12 (j). mRNA expression (k) and protein activity (l) of FOXO downstream antioxidant enzymes. (m) Representative images of DHE immunofluorescence assays. (n) MDA concentration. Scale bar = 50 μm. Data are means ± SD ( n = 7). ** p < 0.01 compared to control group. ++ p < 0.01 compared to LPS group

Article Snippet: The sections or fixed cells were then incubated with the following primary antibodies: anti‐MasR (Novus, NBP1‐78444, 1:30), anti‐FOXO1 (Proteintech, 18592‐1‐AP; 1:100), anti‐IBA‐1 (Abcam, ab178847, 1:200), anti‐LC3 (Cell signaling, 4108; 1:200), or anti‐ASC (Novus, NBP1‐78977, 1:50).

Techniques: Activation Assay, Expressing, Western Blot, Immunofluorescence, Activity Assay, Concentration Assay

Journal: Aging Cell

Article Title: Activation of angiotensin‐converting enzyme 2/angiotensin (1–7)/mas receptor axis triggers autophagy and suppresses microglia proinflammatory polarization via forkhead box class O1 signaling

doi: 10.1111/acel.13480

Figure Lengend Snippet: MasR activation promotes autophagy via FOXO1 signaling in mice brain cortex. (a, b) The selective FOXO1 inhibitor, AS, suppressed AVE‐induced FOXO1 translocation. (c) AS abrogated AVE‐induced transcriptional status of autophagic genes. (d–g) Representative western blots (d) and statistical graphs of protein expression of LC3II (e), Beclin (f), and ATG5‐ATG12 (g). Data are means ± SD ( n = 7). ** p < 0.01 compared to control group. ++ p < 0.01 compared to LPS group. # p < 0.05, ## p < 0.01 compared to LPS + AVE group

Article Snippet: The sections or fixed cells were then incubated with the following primary antibodies: anti‐MasR (Novus, NBP1‐78444, 1:30), anti‐FOXO1 (Proteintech, 18592‐1‐AP; 1:100), anti‐IBA‐1 (Abcam, ab178847, 1:200), anti‐LC3 (Cell signaling, 4108; 1:200), or anti‐ASC (Novus, NBP1‐78977, 1:50).

Techniques: Activation Assay, Translocation Assay, Western Blot, Expressing

Journal: Aging Cell

Article Title: Activation of angiotensin‐converting enzyme 2/angiotensin (1–7)/mas receptor axis triggers autophagy and suppresses microglia proinflammatory polarization via forkhead box class O1 signaling

doi: 10.1111/acel.13480

Figure Lengend Snippet: Ang(1–7) promotes autophagy via FOXO1 signaling in BV2 cells. (a, b) Knockdown either MasR or FOXO1 by using siRNAs suppressed Ang(1–7)‐induced FOXO1 translocation. (c) MasR siRNA and FOXO1 siRNA both abrogated Ang(1–7)‐induced transcriptional status of autophagic genes. (d–g) Representative western blots (d) and statistical graphs of protein expression of LC3II (e), Beclin (f), and ATG5‐ATG12 (g). (h) Immunofluorescence staining of microglial cells transfected with mRFP‐GFP‐LC3 adenovirus. (i) Calculated numbers of autophagosome (GFP + RFP + yellow puncta) and autolysosome (GFP − RFP + red puncta) numbers. (j, k) mRNA expression (j) and protein activity (k) of FOXO downstream antioxidant enzymes. Scale bar = 5 μm. Data are means ± SD ( n = 6). * p < 0.05, ** p < 0.01 compared to control group. + p < 0.05, ++ p < 0.01 compared to LPS group. # p < 0.05, ## p < 0.01 compared to LPS+Ang(1–7) group

Article Snippet: The sections or fixed cells were then incubated with the following primary antibodies: anti‐MasR (Novus, NBP1‐78444, 1:30), anti‐FOXO1 (Proteintech, 18592‐1‐AP; 1:100), anti‐IBA‐1 (Abcam, ab178847, 1:200), anti‐LC3 (Cell signaling, 4108; 1:200), or anti‐ASC (Novus, NBP1‐78977, 1:50).

Techniques: Translocation Assay, Western Blot, Expressing, Immunofluorescence, Staining, Transfection, Activity Assay