Fig. 7
Knockout of YKL-40 enhances Aβ induced receptor mediated endocytosis and promotes Aβ lysosomal degradation in primary astrocytes. A Astrocytes were pre-treated with indicated endocytosis inhibitors for 6 h and then incubated with fluorescent labeled Aβ1-42 at a concentration of 0.1 µM for 24 h, the fluorescence intensity of Aβ1-42 up-taken by astrocytes was measured using plate reader (Bars, SD; n = 5; ns, not significant; ***p < 0.001). B Astrocytes were treated with fluorescent labeled Aβ1-42 (0.1 µM) for 24 h. The percentage of Aβ1-42 positive cells was analyzed using image J. Data are mean ± SD; n = 10, *p < 0.05. C Astrocytes were exposed fluorescent labeled Aβ1-42 (0.1 µM) together with or without recombinant YKL-40 protein (250 ng/mL) for 24 h, and the Aβ1-42 positive cells were analyzed with flow cytometry. Data are mean ± SD; n = 3; **p < 0.01, ***p < 0.001. D The sub-cellular localization of Aβ42-555 peptide in WT and YKL-40 KO astrocytes 24 h after addition was revealed by immunostaining with lysosomal protein LAMP1 (green signal), and Aβ1-42 (red signal). E Data are quantified as percentage of Aβ1-42 colocalized with lysosome was analyzed using image J. Data are mean ± SD; n = 10, **p < 0.01. F Astrocytes were pre-treated with fluorescent labeled Aβ1-42 (0.1 µM) for 24 h, and the levels of Aβ1-42 contained in cells after treatment were measured using plate reader every 24 h. Data are mean ± SD; n = 6, **p < 0.01. G Astrocytes were treated with Aβ1-42 (5 µM) for 96 h, the pH of lysosomes were measured using LysoSensor™ Yellow/Blue DND-160. Data are ± SD; n = 6, ***p < 0.001